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Herstine JA, Mensh J, Coffman E, George SM, Herman K, Martin JB, Zatari A, Chandler HL, Kozmik Z, Drysdale TA, Bridgewater D, Plageman TF. Shroom3 facilitates optic fissure closure via tissue alignment and reestablishment of apical-basal polarity during epithelial fusion. Dev Biol 2025; 522:91-105. [PMID: 40113025 DOI: 10.1016/j.ydbio.2025.03.008] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/09/2024] [Revised: 02/24/2025] [Accepted: 03/15/2025] [Indexed: 03/22/2025]
Abstract
Optic cup morphogenesis is a complex process involving cellular behaviors such as epithelial folding, cell shape changes, proliferation, and tissue fusion. Disruptions to these processes can lead to an ocular coloboma, a congenital defect where the optic fissure fails to close. This study investigates the role of Shroom3, a protein implicated in epithelial morphogenesis, in mouse embryos during optic cup development. It was observed that Shroom3 is apically localized in the neural retina and retinal pigmented epithelium, and its deficiency leads to a both a conventional coloboma phenotype characterized by a gap in pigmented tissue as well as a unique type of coloboma where an ectopic ventral fold of neural tissue is present. Increased apical areas of both neural retina and retinal pigmented epithelial cells are present in the absence of Shroom3 leading to a greater apical surface area and disruption of optic fissure alignment. Neural retina specific gene ablation revealed that Shroom3 function in the RPE is likely sufficient to facilitate tissue alignment and permit fusion. However, the fusion process is ultimately disturbed due to a failure of the neural tissue to reestablish apical-basal polarity. Furthermore, it is demonstrated that Shroom3 deficiency also affects other epithelial fusion events in the embryo that rely on polarity reestablishment, such as lens vesicle separation, eyelid formation, and secondary palate closure. These findings highlight the importance of Shroom3 during optic cup morphogenesis, aid our understanding of optic fissure closure and coloboma formation, and implicates a role for Shroom3 in regulating apical-basal polarity.
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Affiliation(s)
| | - Jordyn Mensh
- College of Optometry, The Ohio State University, Columbus, OH, USA
| | - Electra Coffman
- College of Optometry, The Ohio State University, Columbus, OH, USA
| | | | - Kenneth Herman
- College of Optometry, The Ohio State University, Columbus, OH, USA
| | - Jessica B Martin
- College of Optometry, The Ohio State University, Columbus, OH, USA
| | - Ali Zatari
- College of Optometry, The Ohio State University, Columbus, OH, USA
| | | | - Zbynek Kozmik
- Institute of Molecular Genetics, Academy of Sciences of the Czech Republic, Prague, Czech Republic
| | - Thomas A Drysdale
- Department of Physiology and Pharmacology, Schulich School of Medicine & Dentistry, Western University, London, Ontario, Canada
| | - Darren Bridgewater
- Department of Pathology and Molecular Medicine, Faculty of Health Sciences, McMaster University, Hamilton, Ontario, Canada
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2
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Li C, Syed MU, Nimbalkar A, Shen Y, Vieira MD, Fraser C, Inde Z, Qin X, Ouyang J, Kreuzer J, Clark SE, Kelley G, Hensley EM, Morris R, Lazaro R, Belmonte B, Oh A, Walcott M, Nabel CS, Caenepeel S, Saiki AY, Rex K, Lipford JR, Heist RS, Lin JJ, Haas W, Sarosiek K, Hughes PE, Hata AN. LKB1 regulates JNK-dependent stress signaling and apoptotic dependency of KRAS-mutant lung cancers. Nat Commun 2025; 16:4112. [PMID: 40316540 PMCID: PMC12048556 DOI: 10.1038/s41467-025-58753-y] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/15/2025] [Accepted: 04/01/2025] [Indexed: 05/04/2025] Open
Abstract
The efficacy of molecularly targeted therapies may be limited by co-occurring mutations within a tumor. Conversely, these alterations may confer collateral vulnerabilities that can be therapeutically leveraged. KRAS-mutant lung cancers are distinguished by recurrent loss of the tumor suppressor STK11/LKB1. Whether LKB1 modulates cellular responses to therapeutic stress seems unknown. Here we show that in LKB1-deficient KRAS-mutant lung cancer cells, inhibition of KRAS or its downstream effector MEK leads to hyperactivation of JNK due to loss of NUAK-mediated PP1B phosphatase activity. JNK-mediated inhibitory phosphorylation of BCL-XL rewires apoptotic dependencies, rendering LKB1-deficient cells vulnerable to MCL-1 inhibition. These results uncover an unknown role for LKB1 in regulating stress signaling and mitochondrial apoptosis independent of its tumor suppressor activity mediated by AMPK and SIK. Additionally, our study reveals a therapy-induced vulnerability in LKB1-deficient KRAS-mutant lung cancers that could be exploited as a genotype-informed strategy to improve the efficacy of KRAS-targeted therapies.
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Affiliation(s)
- Chendi Li
- Massachusetts General Hospital Cancer Center, Boston, MA, USA
- Department of Medicine, Massachusetts General Hospital and Harvard Medical School, Boston, MA, USA
| | | | | | - Yi Shen
- Massachusetts General Hospital Cancer Center, Boston, MA, USA
| | | | - Cameron Fraser
- John B. Little Center for Radiation Sciences, Harvard T.H. Chan School of Public Health, Boston, MA, USA
- Department of Environmental Health, Harvard T.H. Chan School of Public Health, Boston, MA, USA
| | - Zintis Inde
- John B. Little Center for Radiation Sciences, Harvard T.H. Chan School of Public Health, Boston, MA, USA
- Department of Environmental Health, Harvard T.H. Chan School of Public Health, Boston, MA, USA
| | - Xingping Qin
- John B. Little Center for Radiation Sciences, Harvard T.H. Chan School of Public Health, Boston, MA, USA
- Department of Environmental Health, Harvard T.H. Chan School of Public Health, Boston, MA, USA
| | - Jian Ouyang
- Massachusetts General Hospital Cancer Center, Boston, MA, USA
- Department of Biochemistry & Molecular Biology and Hollings Cancer Center, Medical University of South Carolina, Charleston, SC, USA
| | - Johannes Kreuzer
- Massachusetts General Hospital Cancer Center, Boston, MA, USA
- Department of Medicine, Massachusetts General Hospital and Harvard Medical School, Boston, MA, USA
| | - Sarah E Clark
- Massachusetts General Hospital Cancer Center, Boston, MA, USA
| | - Grace Kelley
- Massachusetts General Hospital Cancer Center, Boston, MA, USA
| | - Emily M Hensley
- Massachusetts General Hospital Cancer Center, Boston, MA, USA
| | - Robert Morris
- Massachusetts General Hospital Cancer Center, Boston, MA, USA
| | - Raul Lazaro
- Amgen Research, Amgen Inc., Thousand Oaks, CA, USA
| | | | - Audris Oh
- Massachusetts General Hospital Cancer Center, Boston, MA, USA
| | - Makeba Walcott
- Massachusetts General Hospital Cancer Center, Boston, MA, USA
| | - Christopher S Nabel
- Massachusetts General Hospital Cancer Center, Boston, MA, USA
- Koch Institute for Cancer Research, Massachusetts Institute of Technology, Cambridge, MA, USA
| | | | - Anne Y Saiki
- Amgen Research, Amgen Inc., Thousand Oaks, CA, USA
| | - Karen Rex
- Amgen Research, Amgen Inc., Thousand Oaks, CA, USA
| | | | - Rebecca S Heist
- Massachusetts General Hospital Cancer Center, Boston, MA, USA
- Department of Medicine, Massachusetts General Hospital and Harvard Medical School, Boston, MA, USA
| | - Jessica J Lin
- Massachusetts General Hospital Cancer Center, Boston, MA, USA
- Department of Medicine, Massachusetts General Hospital and Harvard Medical School, Boston, MA, USA
| | - Wilhelm Haas
- Massachusetts General Hospital Cancer Center, Boston, MA, USA
| | - Kristopher Sarosiek
- John B. Little Center for Radiation Sciences, Harvard T.H. Chan School of Public Health, Boston, MA, USA
- Department of Environmental Health, Harvard T.H. Chan School of Public Health, Boston, MA, USA
- Lab for Systems Pharmacology, Harvard Program in Therapeutics Science, Harvard Medical School, Boston, MA, USA
| | | | - Aaron N Hata
- Massachusetts General Hospital Cancer Center, Boston, MA, USA.
- Department of Medicine, Massachusetts General Hospital and Harvard Medical School, Boston, MA, USA.
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3
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Robinson MA, Kung SHY, Youssef KYM, Scheck KM, Bell RH, Sar F, Haegert AM, Asmae MM, Cheng C, Yeack SV, Mathur BT, Jiang F, Collins CC, Hach F, Willerth SM, Flannigan RK. 3D Bioprinted Coaxial Testis Model Using Human Induced Pluripotent Stem Cells:A Step Toward Bicompartmental Cytoarchitecture and Functionalization. Adv Healthc Mater 2025; 14:e2402606. [PMID: 39955738 PMCID: PMC12004438 DOI: 10.1002/adhm.202402606] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/15/2024] [Revised: 02/04/2025] [Indexed: 02/17/2025]
Abstract
Fertility preservation following pediatric cancer therapy programs has become a major avenue of infertility research. In vitro spermatogenesis (IVS) aims to generate sperm from banked prepubertal testicular tissues in a lab setting using specialized culture conditions. While successful using rodent tissues, progress with human tissues is limited by the scarcity of human prepubertal testicular tissues for research. This study posits that human induced pluripotent stem cells (hiPSCs) can model human prepubertal testicular tissue to facilitate the development of human IVS conditions. Testicular cells derived from hiPSCs are characterized for phenotype markers and profiled transcriptionally. HiPSC-derived testicular cells are bioprinted into core-shell constructs representative of testis cytoarchitecture and found to capture functional aspects of prepubertal testicular tissues within 7 days under xeno-free conditions. Moreover, hiPSC-derived Sertoli cells illustrate the capacity to mature under pubertal-like conditions. The utility of the model is tested by comparing 2 methods of supplementing retinoic acid (RA), the vitamin responsible for inducing spermatogenesis. The model reveals a significant gain in activity under microsphere-released RA compared to RA medium supplementation, indicating that the fragility of free RA in vitro may be a contributing factor to the molecular dysfunction observed in human IVS studies to date.
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Affiliation(s)
| | - Sonia HY Kung
- Vancouver Prostate CentreVancouverBritish ColumbiaV6H 3Z6Canada
| | | | - Kali M Scheck
- Axolotl BiosciencesVictoriaBritish ColumbiaV8W 2Y2Canada
| | - Robert H Bell
- Vancouver Prostate CentreVancouverBritish ColumbiaV6H 3Z6Canada
| | - Funda Sar
- Vancouver Prostate CentreVancouverBritish ColumbiaV6H 3Z6Canada
| | - Anne M Haegert
- Vancouver Prostate CentreVancouverBritish ColumbiaV6H 3Z6Canada
| | - M Mahdi Asmae
- Vancouver Prostate CentreVancouverBritish ColumbiaV6H 3Z6Canada
| | - Changfeng Cheng
- Faculty of ForestryUniversity of British ColumbiaVancouverBritish ColumbiaV6T 1Z4Canada
| | - Salina V Yeack
- Axolotl BiosciencesVictoriaBritish ColumbiaV8W 2Y2Canada
| | - Bhairvi T Mathur
- Faculty of MedicineUniversity of British ColumbiaVancouverBritish ColumbiaV6T 1Z4Canada
| | - Feng Jiang
- Faculty of ForestryUniversity of British ColumbiaVancouverBritish ColumbiaV6T 1Z4Canada
| | - Colin C Collins
- Vancouver Prostate CentreVancouverBritish ColumbiaV6H 3Z6Canada
| | - Faraz Hach
- Vancouver Prostate CentreVancouverBritish ColumbiaV6H 3Z6Canada
| | - Stephanie M Willerth
- Faculty of MedicineUniversity of British ColumbiaVancouverBritish ColumbiaV6T 1Z4Canada
- Department of Mechanical EngineeringUniversity of VictoriaVictoriaBritish ColumbiaV8P 5C2Canada
- Division of Medical SciencesUniversity of VictoriaVictoriaBritish ColumbiaV8P 5C2Canada
| | - Ryan K Flannigan
- Vancouver Prostate CentreVancouverBritish ColumbiaV6H 3Z6Canada
- Department of Urologic SciencesUniversity of British ColumbiaVancouverBritish ColumbiaV6T 1Z4Canada
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4
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Niazi A, Kim JA, Kim DK, Lu D, Sterin I, Park J, Park S. Microvilli control the morphogenesis of the tectorial membrane extracellular matrix. Dev Cell 2025; 60:679-695.e8. [PMID: 39657673 PMCID: PMC11905117 DOI: 10.1016/j.devcel.2024.11.011] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/04/2024] [Revised: 09/17/2024] [Accepted: 11/13/2024] [Indexed: 12/12/2024]
Abstract
The apical extracellular matrix (aECM), organized by polarized epithelial cells, exhibits complex structures. The tectorial membrane (TM), an aECM in the cochlea mediating auditory transduction, exhibits highly ordered domain-specific architecture. α-Tectorin (TECTA), a glycosylphosphatidylinositol (GPI)-anchored ECM protein, is essential for TM organization. Here, we identified that α-tectorin is released by distinct modes: proteolytic shedding by TMPRSS2 and GPI-anchor-dependent release from the microvillus tip in mice. In the medial/limbal domain, proteolytically shed α-tectorin forms dense fibers. In contrast, in the lateral/body domain, where supporting cells exhibit dense microvilli, shedding restricts α-tectorin to the microvillus tip, compartmentalizing collagen-binding sites. Tip-localized α-tectorin is released in a GPI-anchor-dependent manner to form collagen-crosslinking fibers, maintaining the spacing and parallel organization of collagen fibrils. Overall, these distinct release modes of α-tectorin determine domain-specific organization, with the microvillus coordinating release modes along its membrane to assemble the higher-order ECM architecture.
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Affiliation(s)
- Ava Niazi
- Department of Neurobiology, University of Utah, Salt Lake City, UT, USA; Neuroscience Program, University of Utah, Salt Lake City, UT, USA
| | - Ju Ang Kim
- Department of Neurobiology, University of Utah, Salt Lake City, UT, USA
| | - Dong-Kyu Kim
- Department of Neurobiology, University of Utah, Salt Lake City, UT, USA
| | - Di Lu
- Department of Neurobiology, University of Utah, Salt Lake City, UT, USA
| | - Igal Sterin
- Department of Neurobiology, University of Utah, Salt Lake City, UT, USA
| | - Joosang Park
- Department of Neurobiology, University of Utah, Salt Lake City, UT, USA
| | - Sungjin Park
- Department of Neurobiology, University of Utah, Salt Lake City, UT, USA.
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5
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Zwakenberg S, Westland D, van Es RM, Rehmann H, Anink J, Ciapaite J, Bosma M, Stelloo E, Liv N, Sobrevals Alcaraz P, Verhoeven-Duif NM, Jans JJM, Vos HR, Aronica E, Zwartkruis FJT. mTORC1 restricts TFE3 activity by auto-regulating its presence on lysosomes. Mol Cell 2024; 84:4368-4384.e6. [PMID: 39486419 DOI: 10.1016/j.molcel.2024.10.009] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/19/2023] [Revised: 07/10/2024] [Accepted: 10/07/2024] [Indexed: 11/04/2024]
Abstract
To stimulate cell growth, the protein kinase complex mTORC1 requires intracellular amino acids for activation. Amino-acid sufficiency is relayed to mTORC1 by Rag GTPases on lysosomes, where growth factor signaling enhances mTORC1 activity via the GTPase Rheb. In the absence of amino acids, GATOR1 inactivates the Rags, resulting in lysosomal detachment and inactivation of mTORC1. We demonstrate that in human cells, the release of mTORC1 from lysosomes depends on its kinase activity. In accordance with a negative feedback mechanism, activated mTOR mutants display low lysosome occupancy, causing hypo-phosphorylation and nuclear localization of the lysosomal substrate TFE3. Surprisingly, mTORC1 activated by Rheb does not increase the cytoplasmic/lysosomal ratio of mTORC1, indicating the existence of mTORC1 pools with distinct substrate specificity. Dysregulation of either pool results in aberrant TFE3 activity and may explain nuclear accumulation of TFE3 in epileptogenic malformations in focal cortical dysplasia type II (FCD II) and tuberous sclerosis (TSC).
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Affiliation(s)
- Susan Zwakenberg
- Center for Molecular Medicine, University Medical Center Utrecht, Universiteitsweg 100, 3584 CG Utrecht, the Netherlands; Oncode Institute, Utrecht, the Netherlands
| | - Denise Westland
- Center for Molecular Medicine, University Medical Center Utrecht, Universiteitsweg 100, 3584 CG Utrecht, the Netherlands
| | - Robert M van Es
- Center for Molecular Medicine, University Medical Center Utrecht, Universiteitsweg 100, 3584 CG Utrecht, the Netherlands; Oncode Institute, Utrecht, the Netherlands
| | - Holger Rehmann
- Department of Energy and Life Science, Flensburg University of Applied Sciences, Flensburg, Germany
| | - Jasper Anink
- Department of (Neuro)Pathology, Amsterdam UMC, University of Amsterdam, Amsterdam Neuroscience, Amsterdam, the Netherlands
| | - Jolita Ciapaite
- Department of Genetics, Section Metabolic Diagnostics, University Medical Center Utrecht, 3584 EA Utrecht, the Netherlands
| | - Marjolein Bosma
- Department of Genetics, Section Metabolic Diagnostics, University Medical Center Utrecht, 3584 EA Utrecht, the Netherlands
| | - Ellen Stelloo
- Center for Molecular Medicine, University Medical Center Utrecht, Universiteitsweg 100, 3584 CG Utrecht, the Netherlands
| | - Nalan Liv
- Center for Molecular Medicine, University Medical Center Utrecht, Universiteitsweg 100, 3584 CG Utrecht, the Netherlands
| | - Paula Sobrevals Alcaraz
- Center for Molecular Medicine, University Medical Center Utrecht, Universiteitsweg 100, 3584 CG Utrecht, the Netherlands; Oncode Institute, Utrecht, the Netherlands
| | - Nanda M Verhoeven-Duif
- Department of Genetics, Section Metabolic Diagnostics, University Medical Center Utrecht, 3584 EA Utrecht, the Netherlands
| | - Judith J M Jans
- Department of Genetics, Section Metabolic Diagnostics, University Medical Center Utrecht, 3584 EA Utrecht, the Netherlands
| | - Harmjan R Vos
- Center for Molecular Medicine, University Medical Center Utrecht, Universiteitsweg 100, 3584 CG Utrecht, the Netherlands; Oncode Institute, Utrecht, the Netherlands
| | - Eleonora Aronica
- Department of (Neuro)Pathology, Amsterdam UMC, University of Amsterdam, Amsterdam Neuroscience, Amsterdam, the Netherlands; Stichting Epilepsie Instellingen Nederland (SEIN), Heemstede, the Netherlands
| | - Fried J T Zwartkruis
- Center for Molecular Medicine, University Medical Center Utrecht, Universiteitsweg 100, 3584 CG Utrecht, the Netherlands; Oncode Institute, Utrecht, the Netherlands.
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Huang J, Deng H, Xiao S, Lin Y, Yu Z, Xu X, Peng L, Chao H, Zeng T. CAB39 modulates epithelial-mesenchymal transition through NF-κB signaling activation, enhancing invasion, and metastasis in bladder cancer. ENVIRONMENTAL TOXICOLOGY 2024; 39:4791-4802. [PMID: 39171884 DOI: 10.1002/tox.24333] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 01/16/2024] [Revised: 04/24/2024] [Accepted: 04/29/2024] [Indexed: 08/23/2024]
Abstract
Bladder cancer (BC), the predominant urological malignancy in men, exhibits complex molecular underpinnings contributing to its progression. This investigation aims to elucidate the expression dynamics of calcium-binding protein 39 (CAB39) in both healthy and cancerous tissues and to explore its functional role in the epithelial-mesenchymal transition (EMT) within human bladder cancer contexts. Utilizing immunohistochemistry and quantitative reverse transcription analyses, we assessed CAB39 expression across BC specimens and cell lines. Further, we implemented wound healing, cell invasion, and CCK-8 proliferation assays in CAB39-knockdown cell lines, alongside a nude mouse xenograft model, to gauge the impact of diminished CAB39 expression on the invasive, migratory, and proliferative capacities of BC cells. Our gene set enrichment analysis probed into the repertoire of genes augmented by increased CAB39 expression in BC cells, with subsequent validation via western blotting. Our findings reveal a pronounced overexpression of CAB39 in both BC tissues and cellular models, inversely correlated with disease prognosis. Remarkably, the oncogenic trajectory of bladder cancer was mitigated upon the establishment of shRNA-mediated CAB39 knockdown in vitro and in vivo, effectively reversing the cancer's invasive and metastatic behaviors and curbing tumorigenesis in xenograft models. Hence, CAB39 emerges as a critical biomarker for bladder cancer progression, significantly implicated in facilitating EMT via the upregulation of neural cadherin (N-cadherin) and the suppression of epithelial cadherin through NF-κB signaling pathways. CU-T12-9 effectively overturned the downregulation of p65-NF-kB and N-cadherin, key elements involved in EMT and cell motility, induced by CAB39 knockdown. This study underscores CAB39's pivotal role in bladder cancer pathophysiology and its potential as a therapeutic target.
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Affiliation(s)
- Jianbiao Huang
- Department of Urology, The Second Affiliated Hospital of Nanchang University, Nanchang, People's Republic of China
- Medical College of Nanchang University, Nanchang, Jiangxi, People's Republic of China
| | - Huanhuan Deng
- Medical College of Nanchang University, Nanchang, Jiangxi, People's Republic of China
| | - Shuaiyun Xiao
- Medical College of Nanchang University, Nanchang, Jiangxi, People's Republic of China
| | - Yuanzhen Lin
- Department of Urology, The Second Affiliated Hospital of Nanchang University, Nanchang, People's Republic of China
| | - Zhaojun Yu
- Medical College of Nanchang University, Nanchang, Jiangxi, People's Republic of China
| | - Xiangda Xu
- Department of Urology, The Second Affiliated Hospital of Nanchang University, Nanchang, People's Republic of China
| | - Lifen Peng
- Department of Otolaryngology, Jiangxi Provincial People's Hospital Affiliated Nanchang, People's Republic of China
| | - Haichao Chao
- Department of Urology, The Second Affiliated Hospital of Nanchang University, Nanchang, People's Republic of China
| | - Tao Zeng
- Department of Urology, The Second Affiliated Hospital of Nanchang University, Nanchang, People's Republic of China
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7
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Yu M, Sun F, Xiang G, Zhang Y, Wang X, Liu X, Huang B, Li X, Zhang D. Liver kinase B-1 modulates the activity of dopamine neurons in the ventral tegmental area and regulates social memory formation. Front Mol Neurosci 2024; 17:1289476. [PMID: 38646099 PMCID: PMC11026561 DOI: 10.3389/fnmol.2024.1289476] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/07/2023] [Accepted: 03/19/2024] [Indexed: 04/23/2024] Open
Abstract
Social memory is the ability to discriminate between familiar and unknown conspecifics. It is an important component of social cognition and is therefore essential for the establishment of social relationships. Although the neural circuit mechanisms underlying social memory encoding have been well investigated, little focus has been placed on the regulatory mechanisms of social memory processing. The dopaminergic system, originating from the midbrain ventral tegmental area (VTA), is a key modulator of cognitive function. This study aimed to illustrate its role in modulating social memory and explore the possible molecular mechanisms. Here, we show that the activation of VTA dopamine (DA) neurons is required for the formation, but not the retrieval, of social memory. Inhibition of VTA DA neurons before social interaction, but not 24 h after social interaction, significantly impaired social discrimination the following day. In addition, we showed that the activation of VTA DA neurons was regulated by the serine/threonine protein kinase liver kinase B1 (Lkb1). Deletion of Lkb1 in VTA DA neurons reduced the frequency of burst firing of dopaminergic neurons. Furthermore, Lkb1 plays an important role in regulating social behaviors. Both genetic and virus-mediated deletions of Lkb1 in the VTA of adult mice impaired social memory and subsequently attenuated social familiarization. Altogether, our results provide direct evidence linking social memory formation to the activation of VTA DA neurons in mice and illustrate the crucial role of Lkb1 in regulating VTA DA neuron function.
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Affiliation(s)
- Meng Yu
- Department of Neurosurgery, Qilu Hospital, Cheeloo College of Medicine and Institute of Brain and Brain-Inspired Science, Shandong University, Jinan, China
- Jinan Microecological Biomedicine Shandong Laboratory and Shandong Key Laboratory of Brain Function Remodeling, Jinan, China
| | - Fengjiao Sun
- Institute of Metabolic and Neuropsychiatric Disorders, Binzhou Medical University Hospital, Binzhou, China
| | - Guo Xiang
- Department of Neurosurgery, Qilu Hospital, Cheeloo College of Medicine and Institute of Brain and Brain-Inspired Science, Shandong University, Jinan, China
| | - Yuhan Zhang
- Jinan Microecological Biomedicine Shandong Laboratory and Shandong Key Laboratory of Brain Function Remodeling, Jinan, China
| | - Xuejun Wang
- Jinan Microecological Biomedicine Shandong Laboratory and Shandong Key Laboratory of Brain Function Remodeling, Jinan, China
| | - Xia Liu
- Department of Neurosurgery, Qilu Hospital, Cheeloo College of Medicine and Institute of Brain and Brain-Inspired Science, Shandong University, Jinan, China
| | - Bin Huang
- Department of Neurosurgery, Qilu Hospital, Cheeloo College of Medicine and Institute of Brain and Brain-Inspired Science, Shandong University, Jinan, China
- Jinan Microecological Biomedicine Shandong Laboratory and Shandong Key Laboratory of Brain Function Remodeling, Jinan, China
| | - Xingang Li
- Department of Neurosurgery, Qilu Hospital, Cheeloo College of Medicine and Institute of Brain and Brain-Inspired Science, Shandong University, Jinan, China
- Jinan Microecological Biomedicine Shandong Laboratory and Shandong Key Laboratory of Brain Function Remodeling, Jinan, China
| | - Di Zhang
- Department of Neurosurgery, Qilu Hospital, Cheeloo College of Medicine and Institute of Brain and Brain-Inspired Science, Shandong University, Jinan, China
- Jinan Microecological Biomedicine Shandong Laboratory and Shandong Key Laboratory of Brain Function Remodeling, Jinan, China
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8
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Pasquier N, Jaulin F, Peglion F. Inverted apicobasal polarity in health and disease. J Cell Sci 2024; 137:jcs261659. [PMID: 38465512 PMCID: PMC10984280 DOI: 10.1242/jcs.261659] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 03/12/2024] Open
Abstract
Apicobasal epithelial polarity controls the functional properties of most organs. Thus, there has been extensive research on the molecular intricacies governing the establishment and maintenance of cell polarity. Whereas loss of apicobasal polarity is a well-documented phenomenon associated with multiple diseases, less is known regarding another type of apicobasal polarity alteration - the inversion of polarity. In this Review, we provide a unifying definition of inverted polarity and discuss multiple scenarios in mammalian systems and human health and disease in which apical and basolateral membrane domains are interchanged. This includes mammalian embryo implantation, monogenic diseases and dissemination of cancer cell clusters. For each example, the functional consequences of polarity inversion are assessed, revealing shared outcomes, including modifications in immune surveillance, altered drug sensitivity and changes in adhesions to neighboring cells. Finally, we highlight the molecular alterations associated with inverted apicobasal polarity and provide a molecular framework to connect these changes with the core cell polarity machinery and to explain roles of polarity inversion in health and disease. Based on the current state of the field, failure to respond to extracellular matrix (ECM) cues, increased cellular contractility and membrane trafficking defects are likely to account for most cases of inverted apicobasal polarity.
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Affiliation(s)
- Nicolas Pasquier
- Collective Invasion Team, Inserm U-1279, Gustave Roussy, Villejuif F-94805, France
- Cell Adhesion and Cancer lab, University of Turku, FI-20520 Turku, Finland
| | - Fanny Jaulin
- Collective Invasion Team, Inserm U-1279, Gustave Roussy, Villejuif F-94805, France
| | - Florent Peglion
- Collective Invasion Team, Inserm U-1279, Gustave Roussy, Villejuif F-94805, France
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9
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Niazi A, Kim JA, Kim DK, Lu D, Sterin I, Park J, Park S. Microvilli regulate the release modes of alpha-tectorin to organize the domain-specific matrix architecture of the tectorial membrane. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2024:2024.01.04.574255. [PMID: 38260557 PMCID: PMC10802356 DOI: 10.1101/2024.01.04.574255] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/24/2024]
Abstract
The tectorial membrane (TM) is an apical extracellular matrix (ECM) in the cochlea essential for auditory transduction. The TM exhibits highly ordered domain-specific architecture. Alpha-tectorin/TECTA is a glycosylphosphatidylinositol (GPI)-anchored ECM protein essential for TM organization. Here, we identified that TECTA is released by distinct modes: proteolytic shedding by TMPRSS2 and GPI-anchor-dependent release from the microvillus tip. In the medial/limbal domain, proteolytically shed TECTA forms dense fibers. In the lateral/body domain produced by the supporting cells displaying dense microvilli, the proteolytic shedding restricts TECTA to the microvillus tip and compartmentalizes the collagen-binding site. The tip-localized TECTA, in turn, is released in a GPI-anchor-dependent manner to form collagen-crosslinking fibers, required for maintaining the spacing and parallel organization of collagen fibrils. Overall, we showed that distinct release modes of TECTA determine the domain-specific organization pattern, and the microvillus coordinates the release modes along its membrane to organize the higher-order ECM architecture.
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Affiliation(s)
- Ava Niazi
- Department of Neurobiology, University of Utah, Salt Lake City, Utah, USA
- Neuroscience Program, University of Utah, Salt Lake City, Utah, USA
| | - Ju Ang Kim
- Department of Neurobiology, University of Utah, Salt Lake City, Utah, USA
- Current affiliation: Department of Otorhinolaryngology-Head and Neck Surgery, Seoul National University College of Medicine, Seoul National University Bundang Hospital, Seongnam, South Korea
| | - Dong-Kyu Kim
- Department of Neurobiology, University of Utah, Salt Lake City, Utah, USA
- Current affiliation: Genetics & Genome Biology, The Hospital for Sick Children, Toronto, Ontario, Canada
| | - Di Lu
- Department of Neurobiology, University of Utah, Salt Lake City, Utah, USA
| | - Igal Sterin
- Department of Neurobiology, University of Utah, Salt Lake City, Utah, USA
| | - Joosang Park
- Department of Neurobiology, University of Utah, Salt Lake City, Utah, USA
| | - Sungjin Park
- Department of Neurobiology, University of Utah, Salt Lake City, Utah, USA
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10
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Demouchy F, Nicolle O, Michaux G, Pacquelet A. PAR-4/LKB1 prevents intestinal hyperplasia by restricting endoderm specification in Caenorhabditis elegans embryos. Development 2024; 151:dev202205. [PMID: 38078543 DOI: 10.1242/dev.202205] [Citation(s) in RCA: 3] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/28/2023] [Accepted: 12/04/2023] [Indexed: 01/05/2024]
Abstract
The kinase PAR-4/LKB1 is a major regulator of intestinal homeostasis, which prevents polyposis in humans. Moreover, its ectopic activation is sufficient to induce polarization and formation of microvilli-like structures in intestinal cell lines. Here, we use Caenorhabditis elegans to examine the role of PAR-4 during intestinal development in vivo. We show that it is not required to establish enterocyte polarity and plays only a minor role in brush border formation. By contrast, par-4 mutants display severe deformations of the intestinal lumen as well as supernumerary intestinal cells, thereby revealing a previously unappreciated function of PAR-4 in preventing intestinal hyperplasia. The presence of supernumerary enterocytes in par-4 mutants is not due to excessive cell proliferation, but rather to the abnormal expression of the intestinal cell fate factors end-1 and elt-2 outside the E lineage. Notably, par-4 mutants also display reduced expression of end-1 and elt-2 inside the E lineage. Our work thereby unveils an essential and dual role of PAR-4, which both restricts intestinal specification to the E lineage and ensures its robust differentiation.
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Affiliation(s)
- Flora Demouchy
- University of Rennes, CNRS, IGDR (Institut de Génétique et de Développement de Rennes), UMR 6290, F-35000 Rennes, France
| | - Ophélie Nicolle
- University of Rennes, CNRS, IGDR (Institut de Génétique et de Développement de Rennes), UMR 6290, F-35000 Rennes, France
| | - Grégoire Michaux
- University of Rennes, CNRS, IGDR (Institut de Génétique et de Développement de Rennes), UMR 6290, F-35000 Rennes, France
| | - Anne Pacquelet
- University of Rennes, CNRS, IGDR (Institut de Génétique et de Développement de Rennes), UMR 6290, F-35000 Rennes, France
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11
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Xu R, Wang K, Yao Z, Zhang Y, Jin L, Pang J, Zhou Y, Wang K, Liu D, Zhang Y, Sun P, Wang F, Chang X, Liu T, Wang S, Zhang Y, Lin S, Hu C, Zhu Y, Han X. BRSK2 in pancreatic β cells promotes hyperinsulinemia-coupled insulin resistance and its genetic variants are associated with human type 2 diabetes. J Mol Cell Biol 2023; 15:mjad033. [PMID: 37188647 PMCID: PMC10782904 DOI: 10.1093/jmcb/mjad033] [Citation(s) in RCA: 5] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/28/2022] [Revised: 02/20/2023] [Accepted: 05/12/2023] [Indexed: 05/17/2023] Open
Abstract
Brain-specific serine/threonine-protein kinase 2 (BRSK2) plays critical roles in insulin secretion and β-cell biology. However, whether BRSK2 is associated with human type 2 diabetes mellitus (T2DM) has not been determined. Here, we report that BRSK2 genetic variants are closely related to worsening glucose metabolism due to hyperinsulinemia and insulin resistance in the Chinese population. BRSK2 protein levels are significantly elevated in β cells from T2DM patients and high-fat diet (HFD)-fed mice due to enhanced protein stability. Mice with inducible β-cell-specific Brsk2 knockout (βKO) exhibit normal metabolism with a high potential for insulin secretion under chow-diet conditions. Moreover, βKO mice are protected from HFD-induced hyperinsulinemia, obesity, insulin resistance, and glucose intolerance. Conversely, gain-of-function BRSK2 in mature β cells reversibly triggers hyperglycemia due to β-cell hypersecretion-coupled insulin resistance. Mechanistically, BRSK2 senses lipid signals and induces basal insulin secretion in a kinase-dependent manner. The enhanced basal insulin secretion drives insulin resistance and β-cell exhaustion and thus the onset of T2DM in mice fed an HFD or with gain-of-function BRSK2 in β cells. These findings reveal that BRSK2 links hyperinsulinemia to systematic insulin resistance via interplay between β cells and insulin-sensitive tissues in the populations carrying human genetic variants or under nutrient-overload conditions.
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Affiliation(s)
- Rufeng Xu
- Key Laboratory of Human Functional Genomics of Jiangsu Province, Nanjing Medical University, Nanjing 211166, China
| | - Kaiyuan Wang
- Key Laboratory of Human Functional Genomics of Jiangsu Province, Nanjing Medical University, Nanjing 211166, China
| | - Zhengjian Yao
- Key Laboratory of Human Functional Genomics of Jiangsu Province, Nanjing Medical University, Nanjing 211166, China
| | - Yan Zhang
- Key Laboratory of Human Functional Genomics of Jiangsu Province, Nanjing Medical University, Nanjing 211166, China
| | - Li Jin
- Institute for Metabolic Disease, Fengxian Central Hospital Affiliated to Southern Medical University, Shanghai 201499, China
- Shanghai Diabetes Institute, Shanghai Key Laboratory of Diabetes Mellitus, Shanghai Clinical Center for Diabetes, Shanghai Sixth People's Hospital Affiliated to Shanghai Jiao Tong University School of Medicine, Shanghai 200233, China
| | - Jing Pang
- Key Laboratory of Human Functional Genomics of Jiangsu Province, Nanjing Medical University, Nanjing 211166, China
| | - Yuncai Zhou
- Key Laboratory of Human Functional Genomics of Jiangsu Province, Nanjing Medical University, Nanjing 211166, China
| | - Kai Wang
- Key Laboratory of Human Functional Genomics of Jiangsu Province, Nanjing Medical University, Nanjing 211166, China
| | - Dechen Liu
- Key Laboratory of Human Functional Genomics of Jiangsu Province, Nanjing Medical University, Nanjing 211166, China
| | - Yaqin Zhang
- Key Laboratory of Human Functional Genomics of Jiangsu Province, Nanjing Medical University, Nanjing 211166, China
| | - Peng Sun
- Key Laboratory of Human Functional Genomics of Jiangsu Province, Nanjing Medical University, Nanjing 211166, China
| | - Fuqiang Wang
- Analysis Center, Nanjing Medical University, Nanjing 210029, China
| | - Xiaoai Chang
- Key Laboratory of Human Functional Genomics of Jiangsu Province, Nanjing Medical University, Nanjing 211166, China
| | - Tengli Liu
- Organ Transplant Center, Tianjin First Central Hospital, Nankai University, Tianjin 300192, China
| | - Shusen Wang
- Organ Transplant Center, Tianjin First Central Hospital, Nankai University, Tianjin 300192, China
| | - Yalin Zhang
- State Key Laboratory for Cellular Stress Biology, School of Life Sciences, Xiamen University, Xiamen 361102, China
| | - Shuyong Lin
- State Key Laboratory for Cellular Stress Biology, School of Life Sciences, Xiamen University, Xiamen 361102, China
| | - Cheng Hu
- Institute for Metabolic Disease, Fengxian Central Hospital Affiliated to Southern Medical University, Shanghai 201499, China
- Shanghai Diabetes Institute, Shanghai Key Laboratory of Diabetes Mellitus, Shanghai Clinical Center for Diabetes, Shanghai Sixth People's Hospital Affiliated to Shanghai Jiao Tong University School of Medicine, Shanghai 200233, China
| | - Yunxia Zhu
- Key Laboratory of Human Functional Genomics of Jiangsu Province, Nanjing Medical University, Nanjing 211166, China
| | - Xiao Han
- Key Laboratory of Human Functional Genomics of Jiangsu Province, Nanjing Medical University, Nanjing 211166, China
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12
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Naturale VF, Pickett MA, Feldman JL. Persistent cell contacts enable E-cadherin/HMR-1- and PAR-3-based symmetry breaking within a developing C. elegans epithelium. Dev Cell 2023; 58:1830-1846.e12. [PMID: 37552986 PMCID: PMC10592304 DOI: 10.1016/j.devcel.2023.07.008] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/12/2022] [Revised: 05/10/2023] [Accepted: 07/17/2023] [Indexed: 08/10/2023]
Abstract
Tissue-wide patterning is essential to multicellular development, requiring cells to individually generate polarity axes and coordinate them in space and time with neighbors. Using the C. elegans intestinal epithelium, we identified a patterning mechanism that is informed by cell contact lifetime asymmetry and executed via the scaffolding protein PAR-3 and the transmembrane protein E-cadherin/HMR-1. Intestinal cells break symmetry as PAR-3 and HMR-1 recruit apical determinants into punctate "local polarity complexes" (LPCs) at homotypic contacts. LPCs undergo an HMR-1-based migration to a common midline, thereby establishing tissue-wide polarity. Thus, symmetry breaking results from PAR-3-dependent intracellular polarization coupled to HMR-1-based tissue-level communication, which occurs through a non-adhesive signaling role for HMR-1. Differential lifetimes between homotypic and heterotypic cell contacts are created by neighbor exchanges and oriented divisions, patterning where LPCs perdure and thereby breaking symmetry. These cues offer a logical and likely conserved framework for how epithelia without obvious molecular asymmetries can polarize.
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Affiliation(s)
| | - Melissa A Pickett
- Department of Biology, Stanford University, Stanford, CA 94305, USA; Department of Biological Sciences, San José State University, San José, CA 95192, USA
| | - Jessica L Feldman
- Department of Biology, Stanford University, Stanford, CA 94305, USA.
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13
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Tan I, Xu S, Huo J, Huang Y, Lim HH, Lam KP. Identification of a novel mitochondria-localized LKB1 variant required for the regulation of the oxidative stress response. J Biol Chem 2023; 299:104906. [PMID: 37302555 PMCID: PMC10404683 DOI: 10.1016/j.jbc.2023.104906] [Citation(s) in RCA: 4] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/15/2023] [Revised: 05/31/2023] [Accepted: 06/01/2023] [Indexed: 06/13/2023] Open
Abstract
The tumor suppressor Liver Kinase B1 (LKB1) is a multifunctional serine/threonine protein kinase that regulates cell metabolism, polarity, and growth and is associated with Peutz-Jeghers Syndrome and cancer predisposition. The LKB1 gene comprises 10 exons and 9 introns. Three spliced LKB1 variants have been documented, and they reside mainly in the cytoplasm, although two possess a nuclear-localization sequence (NLS) and are able to shuttle into the nucleus. Here, we report the identification of a fourth and novel LKB1 isoform that is, interestingly, targeted to the mitochondria. We show that this mitochondria-localized LKB1 (mLKB1) is generated from alternative splicing in the 5' region of the transcript and translated from an alternative initiation codon encoded by a previously unknown exon 1b (131 bp) hidden within the long intron 1 of LKB1 gene. We found by replacing the N-terminal NLS of the canonical LKB1 isoform, the N-terminus of the alternatively spliced mLKB1 variant encodes a mitochondrial transit peptide that allows it to localize to the mitochondria. We further demonstrate that mLKB1 colocalizes histologically with mitochondria-resident ATP Synthase and NAD-dependent deacetylase sirtuin-3, mitochondrial (SIRT3) and that its expression is rapidly and transiently upregulated by oxidative stress. We conclude that this novel LKB1 isoform, mLKB1, plays a critical role in regulating mitochondrial metabolic activity and oxidative stress response.
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Affiliation(s)
- Ivan Tan
- Singapore Immunology Network (SIgN), Agency for Science, Technology and Research (A∗STAR), Singapore, Singapore
| | - Shengli Xu
- Singapore Immunology Network (SIgN), Agency for Science, Technology and Research (A∗STAR), Singapore, Singapore; Department of Physiology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore, Singapore
| | - Jianxin Huo
- Singapore Immunology Network (SIgN), Agency for Science, Technology and Research (A∗STAR), Singapore, Singapore
| | - Yuhan Huang
- Singapore Immunology Network (SIgN), Agency for Science, Technology and Research (A∗STAR), Singapore, Singapore
| | - Hong-Hwa Lim
- Singapore Immunology Network (SIgN), Agency for Science, Technology and Research (A∗STAR), Singapore, Singapore; Institute of Molecular and Cell Biology, Agency for Science, Technology and Research, Singapore, Singapore
| | - Kong-Peng Lam
- Singapore Immunology Network (SIgN), Agency for Science, Technology and Research (A∗STAR), Singapore, Singapore; Department of Microbiology and Immunology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore, Singapore; School of Biological Sciences, Nanyang Technological University, Singapore, Singapore.
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14
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Thüring EM, Hartmann C, Schwietzer YA, Ebnet K. TMIGD1: Emerging functions of a tumor supressor and adhesion receptor. Oncogene 2023:10.1038/s41388-023-02696-5. [PMID: 37087524 DOI: 10.1038/s41388-023-02696-5] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/10/2023] [Revised: 04/11/2023] [Accepted: 04/17/2023] [Indexed: 04/24/2023]
Abstract
The development of multicellular organisms depends on cell adhesion molecules (CAMs) that connect cells to build tissues. The immunoglobulin superfamily (IgSF) constitutes one of the largest families of CAMs. Members of this family regulate such diverse processes like synapse formation, spermatogenesis, leukocyte-endothelial interactions, or epithelial cell-cell adhesion. Through their extracellular domains, they undergo homophilic and heterophilic interactions in cis and trans. Their cytoplasmic domains frequently bind scaffolding proteins to assemble signaling complexes. Transmembrane and immunoglobulin domain-containing protein 1 (TMIGD1) is a IgSF member with two Ig-like domains and a short cytoplasmic tail that contains a PDZ domain-binding motif. Recent observations indicate that TMIGD1 has pleiotropic functions in epithelial cells and has a critical role in suppressing malignant cell behavior. Here, we review the molecular characteristics of TMIGD1, its interaction with cytoplasmic scaffolding proteins, the regulation of its expression, and its downregulation in colorectal and renal cancers.
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Affiliation(s)
- Eva-Maria Thüring
- Institute-associated Research Group "Cell adhesion and cell polarity", Institute of Medical Biochemistry, ZMBE, University of Münster, Münster, Germany
| | - Christian Hartmann
- Institute-associated Research Group "Cell adhesion and cell polarity", Institute of Medical Biochemistry, ZMBE, University of Münster, Münster, Germany
| | - Ysabel A Schwietzer
- Institute-associated Research Group "Cell adhesion and cell polarity", Institute of Medical Biochemistry, ZMBE, University of Münster, Münster, Germany
| | - Klaus Ebnet
- Institute-associated Research Group "Cell adhesion and cell polarity", Institute of Medical Biochemistry, ZMBE, University of Münster, Münster, Germany.
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15
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Gaeta IM, Tyska MJ. BioID2 screening identifies KIAA1671 as an EPS8 proximal factor that marks sites of microvillus growth. Mol Biol Cell 2023; 34:ar31. [PMID: 36790915 PMCID: PMC10092648 DOI: 10.1091/mbc.e22-11-0498] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/16/2023] Open
Abstract
Microvilli are defining morphological features of the apical surfaces in diverse epithelial tissues. To develop our understanding of microvillus biogenesis, we used a biotin proximity-labeling approach to uncover new molecules enriched near EPS8, a well-studied marker of the microvillus distal tip compartment. Mass spectrometry of biotinylated hits identified KIAA1671, a large (∼200 kDa), disordered, and previously uncharacterized protein. Based on immunofluorescent staining and expression of fluorescent protein-tagged constructs, we found that KIAA1671 localizes to the base of the brush border in native intestinal tissue and polarized epithelial-cell culture models, as well as dynamic actin-rich structures in unpolarized, nonepithelial cell types. Live imaging also revealed that during the early stages of microvillar growth, KIAA1671 colocalizes with EPS8 in diffraction-limited puncta. However, once elongation of the core bundle begins, these two factors separate, with EPS8 tracking the distal end and KIAA1671 remaining behind at the base of the structure. These results suggest that KIAA1671 cooperates with EPS8 and potentially other assembly factors to initiate growth of microvilli on the apical surface. These findings offer new details on how transporting epithelial cells builds the brush border and may inform our understanding of how apical specializations are assembled in other epithelial contexts.
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Affiliation(s)
- Isabella M Gaeta
- Department of Cell and Developmental Biology, Vanderbilt University School of Medicine, Nashville, TN 37232
| | - Matthew J Tyska
- Department of Cell and Developmental Biology, Vanderbilt University School of Medicine, Nashville, TN 37232
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16
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Hu L, Liu M, Tang B, Li Q, Pan BS, Xu C, Lin HK. Posttranslational regulation of liver kinase B1 (LKB1) in human cancer. J Biol Chem 2023; 299:104570. [PMID: 36870679 PMCID: PMC10068580 DOI: 10.1016/j.jbc.2023.104570] [Citation(s) in RCA: 2] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/02/2023] [Revised: 02/20/2023] [Accepted: 02/22/2023] [Indexed: 03/06/2023] Open
Abstract
Liver kinase B1 (LKB1) is a serine-threonine kinase that participates in multiple cellular and biological processes, including energy metabolism, cell polarity, cell proliferation, cell migration, and many others. LKB1 is initially identified as a germline-mutated causative gene in Peutz-Jeghers syndrome (PJS) and is commonly regarded as a tumor suppressor due to frequent inactivation in a variety of cancers. LKB1 directly binds and activates its downstream kinases including the AMP-activated protein kinase (AMPK) and AMPK-related kinases by phosphorylation, which has been intensively investigated for the past decades. An increasing number of studies has uncovered the posttranslational modifications (PTMs) of LKB1 and consequent changes in its localization, activity, and interaction with substrates. The alteration in LKB1 function as a consequence of genetic mutations and aberrant upstream signaling regulation leads to tumor development and progression. Here, we review current knowledge about the mechanism of LKB1 in cancer and the contributions of PTMs, such as phosphorylation, ubiquitination, SUMOylation, acetylation, prenylation, and others, to the regulation of LKB1 function, offering new insights into the therapeutic strategies in cancer.
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Affiliation(s)
- Lanlin Hu
- Department of Oncology & Cancer Institute, Sichuan Academy of Medical Sciences, Sichuan Provincial People's Hospital, University of Electronic Science and Technology of China, Chengdu, China; Department of Laboratory Medicine and Sichuan Provincial Key Laboratory for Human Disease Gene Study, Sichuan Provincial People's Hospital, University of Electronic Science and Technology of China, Chengdu, China; Sichuan Cancer Hospital & Institute, Sichuan Cancer Center, University of Electronic Science and Technology of China, Chengdu, China
| | - Mingxin Liu
- Department of Oncology & Cancer Institute, Sichuan Academy of Medical Sciences, Sichuan Provincial People's Hospital, University of Electronic Science and Technology of China, Chengdu, China; Sichuan Cancer Hospital & Institute, Sichuan Cancer Center, University of Electronic Science and Technology of China, Chengdu, China
| | - Bo Tang
- Department of Oncology & Cancer Institute, Sichuan Academy of Medical Sciences, Sichuan Provincial People's Hospital, University of Electronic Science and Technology of China, Chengdu, China; Department of Laboratory Medicine and Sichuan Provincial Key Laboratory for Human Disease Gene Study, Sichuan Provincial People's Hospital, University of Electronic Science and Technology of China, Chengdu, China; Sichuan Cancer Hospital & Institute, Sichuan Cancer Center, University of Electronic Science and Technology of China, Chengdu, China
| | - Qiang Li
- Sichuan Cancer Hospital & Institute, Sichuan Cancer Center, University of Electronic Science and Technology of China, Chengdu, China
| | - Bo-Syong Pan
- Department of Cancer Biology, Wake Forest School of Medicine, Winston-Salem, North Carolina, USA
| | - Chuan Xu
- Department of Oncology & Cancer Institute, Sichuan Academy of Medical Sciences, Sichuan Provincial People's Hospital, University of Electronic Science and Technology of China, Chengdu, China; Department of Laboratory Medicine and Sichuan Provincial Key Laboratory for Human Disease Gene Study, Sichuan Provincial People's Hospital, University of Electronic Science and Technology of China, Chengdu, China; Department of Cancer Biology, Wake Forest School of Medicine, Winston-Salem, North Carolina, USA.
| | - Hui-Kuan Lin
- Department of Cancer Biology, Wake Forest School of Medicine, Winston-Salem, North Carolina, USA.
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17
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Tzavlaki K, Ohata Y, Morén A, Watanabe Y, Eriksson J, Tsuchiya M, Kubo Y, Yamamoto K, Sellin ME, Kato M, Caja L, Heldin CH, Moustakas A. The liver kinase B1 supports mammary epithelial morphogenesis by inhibiting critical factors that mediate epithelial-mesenchymal transition. J Cell Physiol 2023; 238:790-812. [PMID: 36791282 DOI: 10.1002/jcp.30975] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/01/2022] [Revised: 01/24/2023] [Accepted: 01/31/2023] [Indexed: 02/17/2023]
Abstract
The liver kinase B1 (LKB1) controls cellular metabolism and cell polarity across species. We previously established a mechanism for negative regulation of transforming growth factor β (TGFβ) signaling by LKB1. The impact of this mechanism in the context of epithelial polarity and morphogenesis remains unknown. After demonstrating that human mammary tissue expresses robust LKB1 protein levels, whereas invasive breast cancer exhibits significantly reduced LKB1 levels, we focused on mammary morphogenesis studies in three dimensional (3D) acinar organoids. CRISPR/Cas9-introduced loss-of-function mutations of STK11 (LKB1) led to profound defects in the formation of 3D organoids, resulting in amorphous outgrowth and loss of rotation of young organoids embedded in matrigel. This defect was associated with an enhanced signaling by TGFβ, including TGFβ auto-induction and induction of transcription factors that mediate epithelial-mesenchymal transition (EMT). Protein marker analysis confirmed a more efficient EMT response to TGFβ signaling in LKB1 knockout cells. Accordingly, chemical inhibition of the TGFβ type I receptor kinase largely restored the morphogenetic defect of LKB1 knockout cells. Similarly, chemical inhibition of the bone morphogenetic protein pathway or the TANK-binding kinase 1, or genetic silencing of the EMT factor SNAI1, partially restored the LKB1 knockout defect. Thus, LKB1 sustains mammary epithelial morphogenesis by limiting pathways that promote EMT. The observed downregulation of LKB1 expression in breast cancer is therefore predicted to associate with enhanced EMT induced by SNAI1 and TGFβ family members.
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Affiliation(s)
- Kalliopi Tzavlaki
- Science for Life Laboratory, Department of Medical Biochemistry and Microbiology, Biomedical Center, Uppsala University, Uppsala, Sweden
| | - Yae Ohata
- Science for Life Laboratory, Department of Medical Biochemistry and Microbiology, Biomedical Center, Uppsala University, Uppsala, Sweden
| | - Anita Morén
- Science for Life Laboratory, Department of Medical Biochemistry and Microbiology, Biomedical Center, Uppsala University, Uppsala, Sweden
| | - Yukihide Watanabe
- Department of Experimental Pathology and Transborder Medical Research Center, Faculty of Medicine, University of Tsukuba, Tsukuba, Ibaraki, Japan
| | - Jens Eriksson
- Science for Life Laboratory, Department of Medical Biochemistry and Microbiology, Biomedical Center, Uppsala University, Uppsala, Sweden
| | - Maiko Tsuchiya
- Department of Oral Pathology, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Tokyo, Japan.,Department of Pathology, Teikyo University School of Medicine, Tokyo, Japan
| | - Yuki Kubo
- Department of Comprehensive Pathology, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Tokyo, Japan
| | - Kouhei Yamamoto
- Department of Comprehensive Pathology, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Tokyo, Japan
| | - Mikael E Sellin
- Science for Life Laboratory, Department of Medical Biochemistry and Microbiology, Biomedical Center, Uppsala University, Uppsala, Sweden
| | - Mitsuyasu Kato
- Department of Experimental Pathology and Transborder Medical Research Center, Faculty of Medicine, University of Tsukuba, Tsukuba, Ibaraki, Japan
| | - Laia Caja
- Science for Life Laboratory, Department of Medical Biochemistry and Microbiology, Biomedical Center, Uppsala University, Uppsala, Sweden
| | - Carl-Henrik Heldin
- Science for Life Laboratory, Department of Medical Biochemistry and Microbiology, Biomedical Center, Uppsala University, Uppsala, Sweden
| | - Aristidis Moustakas
- Science for Life Laboratory, Department of Medical Biochemistry and Microbiology, Biomedical Center, Uppsala University, Uppsala, Sweden
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18
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Chiraphapphaiboon W, Thongnoppakhun W, Limjindaporn T, Sawasdichai S, Roothumnong E, Prangphan K, Pamornpol B, Limwongse C, Pithukpakorn M. STK11 Causative Variants and Copy Number Variations Identified in Thai Patients With Peutz-Jeghers Syndrome. Cureus 2023; 15:e34495. [PMID: 36874343 PMCID: PMC9983355 DOI: 10.7759/cureus.34495] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 01/31/2023] [Indexed: 02/04/2023] Open
Abstract
Introduction Peutz-Jeghers syndrome (PJS) is a rare autosomal dominant inherited disorder caused by germline mutations in the serine-threonine kinase 11 (STK11) tumor suppressor gene. This syndrome is characterized by hamartomatous gastrointestinal polyps, mucocutaneous melanin pigmentation, and a higher risk of developing various cancers. Methods We summarized the clinical and molecular characteristics of five unrelated Thai patients with PJS. Denaturing high-performance liquid chromatography (DHPLC) screening, coupled with direct DNA sequencing and multiplex ligation-dependent probe amplification (MLPA), were applied for the molecular analysis of STK11. Results A total of four STK11 pathogenic changeswere identified in the five PJS patients, including two frameshift variants (a novel c.199dup, p.Leu67ProfsTer96 and a known c.834_835del, p.Cys278TrpfsTer6) and two types of copy number variations (CNV), exon 1 deletion and exons 2-3 deletion. Among reported STK11 exonic deletions, exon 1 and exons 2-3 deletions were found to be the two most commonly deleted exons. Conclusion All identified STK11 mutations were null mutations that were associated with more severe PJS phenotypes and cancers. This study broadens the phenotypic and mutational spectrum of STK11 in PJS.
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Affiliation(s)
| | - Wanna Thongnoppakhun
- Siriraj Genomics, Office of the Dean, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, THA
| | | | - Sunisa Sawasdichai
- Siriraj Genomics, Office of the Dean, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, THA
| | - Ekkapong Roothumnong
- Division of Medical Genetics, Department of Medicine, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, THA
| | - Kanjana Prangphan
- Siriraj Genomics, Office of the Dean, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, THA
| | - Benjaporn Pamornpol
- Department of Anatomy, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, THA
| | - Chanin Limwongse
- Siriraj Genomics, Office of the Dean, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, THA.,Division of Medical Genetics, Department of Medicine, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, THA
| | - Manop Pithukpakorn
- Siriraj Genomics, Office of the Dean, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, THA.,Division of Medical Genetics, Department of Medicine, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, THA
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19
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Haydinger CD, Ferreira LB, Williams KA, Smith JR. Mechanisms of macular edema. Front Med (Lausanne) 2023; 10:1128811. [PMID: 36960343 PMCID: PMC10027768 DOI: 10.3389/fmed.2023.1128811] [Citation(s) in RCA: 34] [Impact Index Per Article: 17.0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/21/2022] [Accepted: 02/16/2023] [Indexed: 03/09/2023] Open
Abstract
Macular edema is the pathological accumulation of fluid in the central retina. It is a complication of many retinal diseases, including diabetic retinopathy, retinal vascular occlusions and uveitis, among others. Macular edema causes decreased visual acuity and, when chronic or refractory, can cause severe and permanent visual impairment and blindness. In most instances, it develops due to dysregulation of the blood-retinal barrier which permits infiltration of the retinal tissue by proteins and other solutes that are normally retained in the blood. The increase in osmotic pressure in the tissue drives fluid accumulation. Current treatments include vascular endothelial growth factor blockers, corticosteroids, and non-steroidal anti-inflammatory drugs. These treatments target vasoactive and inflammatory mediators that cause disruption to the blood-retinal barrier. In this review, a clinical overview of macular edema is provided, mechanisms of disease are discussed, highlighting processes targeted by current treatments, and areas of opportunity for future research are identified.
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20
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Yu Z, Liu L, Jiang F, Ji Y, Wang X, Liu L. A novel missense mutation of the STK11 gene in a Chinese family with Peutz-Jeghers syndrome. BMC Gastroenterol 2022; 22:536. [PMID: 36550395 PMCID: PMC9784088 DOI: 10.1186/s12876-022-02617-y] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/26/2022] [Accepted: 12/13/2022] [Indexed: 12/24/2022] Open
Abstract
BACKGROUND Peutz-Jeghers syndrome (PJS) is a rare autosomal dominant inherited disease caused by mutations in the Serine-Threonine Kinase 11 (STK11) gene. This study aimed to diagnose a Chinese pedigree with PJS and to expand the spectrum of STK11 variants. METHODS We performed an inductive analysis of clinical features, gastrointestinal endoscopy, radiologic imaging, and pathological findings in a Chinese family with PJS. Whole-exome sequencing (WES), Sanger sequencing, and STK11 protein 3D structure prediction were performed for establishing a molecular diagnosis. RESULTS The proband, her mother, and grandfather presented with pigmentation spots on lips, oral mucosa, and fingers. Her mother and grandfather also had pigmentation spots on face and feet, while her brother had pigmentation spots only on the lower lip. On endoscopy, polyps were discovered in the proband, her mother, and grandfather. A novel heterozygous mutation (c.521A > C) in exon 4 of STK11 was identified in all four patients, leading to a change from histidine to proline in amino acid 174. The variable site p.H174 was highly conserved in different species on multiple sequence alignment analysis. CONCLUSIONS We diagnosed a Chinese pedigree with PJS based on clinical features, gastrointestinal endoscopy, and genetic testing results. Our results expanded the spectrum of STK11 variants, which will be helpful for genetic counseling.
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Affiliation(s)
- Zhen Yu
- grid.27255.370000 0004 1761 1174Department of Pediatrics, Qilu Hospital, Shandong University, 107 Wen Hua Xi Road, Jinan, 250012 Shandong People’s Republic of China
| | - Lin Liu
- grid.27255.370000 0004 1761 1174Shandong Provincial Maternal and Child Health Care Hospital, Shandong University, 238 Jing Shi Dong Road, Jinan, 250012 Shandong People’s Republic of China
| | - Fang Jiang
- grid.27255.370000 0004 1761 1174Department of Pediatrics, Qilu Hospital, Shandong University, 107 Wen Hua Xi Road, Jinan, 250012 Shandong People’s Republic of China
| | - Yimin Ji
- grid.27255.370000 0004 1761 1174Department of Pediatrics, Qilu Hospital, Shandong University, 107 Wen Hua Xi Road, Jinan, 250012 Shandong People’s Republic of China
| | - Xiao Wang
- grid.27255.370000 0004 1761 1174Department of Pediatrics, Qilu Hospital, Shandong University, 107 Wen Hua Xi Road, Jinan, 250012 Shandong People’s Republic of China
| | - Lili Liu
- grid.27255.370000 0004 1761 1174Department of Pediatrics, Qilu Hospital, Shandong University, 107 Wen Hua Xi Road, Jinan, 250012 Shandong People’s Republic of China
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21
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Barriga EH, Alasaadi DN, Mencarelli C, Mayor R, Pichaud F. RanBP1 plays an essential role in directed migration of neural crest cells during development. Dev Biol 2022; 492:79-86. [PMID: 36206829 DOI: 10.1016/j.ydbio.2022.09.010] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/29/2021] [Revised: 09/15/2022] [Accepted: 09/23/2022] [Indexed: 11/21/2022]
Abstract
Collective cell migration is essential for embryonic development, tissue regeneration and repair, and has been implicated in pathological conditions such as cancer metastasis. It is, in part, directed by external cues that promote front-to-rear polarity in individual cells. However, our understanding of the pathways that underpin the directional movement of cells in response to external cues remains incomplete. To examine this issue we made use of neural crest cells (NC), which migrate as a collective during development to generate vital structures including bones and cartilage. Using a candidate approach, we found an essential role for Ran-binding protein 1 (RanBP1), a key effector of the nucleocytoplasmic transport pathway, in enabling directed migration of these cells. Our results indicate that RanBP1 is required for establishing front-to-rear polarity, so that NCs are able to chemotax. Moreover, our work suggests that RanBP1 function in chemotaxis involves the polarity kinase LKB1/PAR4. We envisage that regulated nuclear export of LKB1 through Ran/RanBP1 is a key regulatory step required for establishing front-to-rear polarity and thus chemotaxis, during NC collective migration.
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Affiliation(s)
- Elias H Barriga
- Cell and Developmental Biology, University College London, London, WC1E 6BT, United Kingdom; Mechanisms of Morphogenesis Lab, Instituto Gulbenkian de Ciencia, Oeiras, 2780-156, Portugal
| | - Delan N Alasaadi
- Cell and Developmental Biology, University College London, London, WC1E 6BT, United Kingdom
| | - Chiara Mencarelli
- Laboratory for Molecular Cell Biology, University College London, London, WC1E 6BT, United Kingdom
| | - Roberto Mayor
- Cell and Developmental Biology, University College London, London, WC1E 6BT, United Kingdom.
| | - Franck Pichaud
- Laboratory for Molecular Cell Biology, University College London, London, WC1E 6BT, United Kingdom.
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22
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Cartón-García F, Brotons B, Anguita E, Dopeso H, Tarragona J, Nieto R, García-Vidal E, Macaya I, Zagyva Z, Dalmau M, Sánchez-Martín M, van Ijzendoorn SCD, Landolfi S, Hernandez-Losa J, Schwartz Jr S, Matias-Guiu X, Ramón y Cajal S, Martínez-Barriocanal Á, Arango D. Myosin Vb as a tumor suppressor gene in intestinal cancer. Oncogene 2022; 41:5279-5288. [DOI: 10.1038/s41388-022-02508-2] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/05/2022] [Revised: 10/08/2022] [Accepted: 10/11/2022] [Indexed: 11/07/2022]
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23
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Huang E, Li S. Liver Kinase B1 Functions as a Regulator for Neural Development and a Therapeutic Target for Neural Repair. Cells 2022; 11:cells11182861. [PMID: 36139438 PMCID: PMC9496952 DOI: 10.3390/cells11182861] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/29/2022] [Revised: 09/02/2022] [Accepted: 09/10/2022] [Indexed: 11/16/2022] Open
Abstract
The liver kinase B1 (LKB1), also known as serine/threonine kinase 11 (STK11) and Par-4 in C. elegans, has been identified as a master kinase of AMPKs and AMPK-related kinases. LKB1 plays a crucial role in cell growth, metabolism, polarity, and tumor suppression. By interacting with the downstream signals of SAD, NUAK, MARK, and other kinases, LKB1 is critical to regulating neuronal polarization and axon branching during development. It also regulates Schwann cell function and the myelination of peripheral axons. Regulating LKB1 activity has become an attractive strategy for repairing an injured nervous system. LKB1 upregulation enhances the regenerative capacity of adult CNS neurons and the recovery of locomotor function in adult rodents with CNS axon injury. Here, we update the major cellular and molecular mechanisms of LKB1 in regulating neuronal polarization and neural development, and the implications thereof for promoting neural repair, axon regeneration, and functional recovery in adult mammals.
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24
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Hartmann C, Thüring EM, Greune L, Michels BE, Pajonczyk D, Leußink S, Brinkmann F, Glaesner-Ebnet M, Wardelmann E, Zobel T, Schmidt MA, Janssen KP, Gerke V, Ebnet K. Intestinal brush border formation requires a TMIGD1-based intermicrovillar adhesion complex. Sci Signal 2022; 15:eabm2449. [PMID: 36099341 DOI: 10.1126/scisignal.abm2449] [Citation(s) in RCA: 12] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/02/2022]
Abstract
Intestinal epithelial cells absorb nutrients through the brush border, composed of dense arrays of highly ordered microvilli at their apical membranes. A protocadherin-based intermicrovillar adhesion complex localized at microvilli tips mediates microvilli packing and organization. Here, we identified a second adhesion complex localized at the proximal base region of microvilli. This complex contained the immunoglobulin superfamily member TMIGD1, which directly interacted with the microvillar scaffolding proteins EBP50 and E3KARP. Complex formation with EBP50 required the activation of EBP50 by the actin-binding protein ezrin and was enhanced by the dephosphorylation of Ser162 in the PDZ2 domain of EBP50 by the phosphatase PP1α. Binding of the EBP50-ezrin complex to TMIGD1 enhanced the dynamic turnover of EBP50 at microvilli. Enterocyte-specific inactivation of Tmigd1 in mice resulted in microvillar blebbing, loss of intermicrovillar adhesion, and perturbed brush border formation. Thus, we identified a second adhesion complex in microvilli and propose a mechanism that promotes microvillar formation and dynamics.
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Affiliation(s)
- Christian Hartmann
- Institute-associated Research Group "Cell adhesion and cell polarity", ZMBE, University of Münster, D-48149 Münster, Germany.,Institute of Medical Biochemistry, ZMBE, University of Münster, D-48149 Münster, Germany
| | - Eva-Maria Thüring
- Institute-associated Research Group "Cell adhesion and cell polarity", ZMBE, University of Münster, D-48149 Münster, Germany.,Institute of Medical Biochemistry, ZMBE, University of Münster, D-48149 Münster, Germany
| | - Lilo Greune
- Institute of Infectiology, ZMBE, University of Münster, D-48149 Münster, Germany
| | - Birgitta E Michels
- Institute-associated Research Group "Cell adhesion and cell polarity", ZMBE, University of Münster, D-48149 Münster, Germany.,Institute of Medical Biochemistry, ZMBE, University of Münster, D-48149 Münster, Germany
| | - Denise Pajonczyk
- Institute-associated Research Group "Cell adhesion and cell polarity", ZMBE, University of Münster, D-48149 Münster, Germany.,Institute of Medical Biochemistry, ZMBE, University of Münster, D-48149 Münster, Germany
| | - Sophia Leußink
- Institute-associated Research Group "Cell adhesion and cell polarity", ZMBE, University of Münster, D-48149 Münster, Germany.,Institute of Medical Biochemistry, ZMBE, University of Münster, D-48149 Münster, Germany
| | - Frauke Brinkmann
- Institute-associated Research Group "Cell adhesion and cell polarity", ZMBE, University of Münster, D-48149 Münster, Germany.,Institute of Medical Biochemistry, ZMBE, University of Münster, D-48149 Münster, Germany
| | - Mark Glaesner-Ebnet
- Institute-associated Research Group "Cell adhesion and cell polarity", ZMBE, University of Münster, D-48149 Münster, Germany.,Institute of Medical Biochemistry, ZMBE, University of Münster, D-48149 Münster, Germany
| | - Eva Wardelmann
- Gerhard-Domagk-Institute of Pathology, University Hospital Münster, D-48149 Münster, Germany
| | - Thomas Zobel
- Imaging Network Microscopy, University of Münster, D-48149 Münster, Germany
| | - M Alexander Schmidt
- Institute of Infectiology, ZMBE, University of Münster, D-48149 Münster, Germany
| | | | - Volker Gerke
- Institute of Medical Biochemistry, ZMBE, University of Münster, D-48149 Münster, Germany.,Cells-in-Motion Interfaculty Center (CiMIC), University of Münster, D-48419 Münster, Germany
| | - Klaus Ebnet
- Institute-associated Research Group "Cell adhesion and cell polarity", ZMBE, University of Münster, D-48149 Münster, Germany.,Institute of Medical Biochemistry, ZMBE, University of Münster, D-48149 Münster, Germany.,Cells-in-Motion Interfaculty Center (CiMIC), University of Münster, D-48419 Münster, Germany.,Interdisciplinary Center for Clinical Research (IZKF), University of Münster, D-48149 Münster, Germany
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25
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Targeting of microvillus protein Eps8 by the NleH effector kinases from enteropathogenic E. coli. Proc Natl Acad Sci U S A 2022; 119:e2204332119. [PMID: 35976880 PMCID: PMC9407544 DOI: 10.1073/pnas.2204332119] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/17/2023] Open
Abstract
Attaching and effacing (AE) lesion formation on enterocytes by enteropathogenic Escherichia coli (EPEC) requires the EPEC type III secretion system (T3SS). Two T3SS effectors injected into the host cell during infection are the atypical kinases, NleH1 and NleH2. However, the host targets of NleH1 and NleH2 kinase activity during infection have not been reported. Here phosphoproteomics identified Ser775 in the microvillus protein Eps8 as a bona fide target of NleH1 and NleH2 phosphorylation. Both kinases interacted with Eps8 through previously unrecognized, noncanonical "proline-rich" motifs, PxxDY, that bound the Src Homology 3 (SH3) domain of Eps8. Structural analysis of the Eps8 SH3 domain bound to a peptide containing one of the proline-rich motifs from NleH showed that the N-terminal part of the peptide adopts a type II polyproline helix, and its C-terminal "DY" segment makes multiple contacts with the SH3 domain. Ser775 phosphorylation by NleH1 or NleH2 hindered Eps8 bundling activity and drove dispersal of Eps8 from the AE lesion during EPEC infection. This finding suggested that NleH1 and NleH2 altered the cellular localization of Eps8 and the cytoskeletal composition of AE lesions during EPEC infection.
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26
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Khuntia P, Rawal S, Marwaha R, Das T. Actin-driven Golgi apparatus dispersal during collective migration of epithelial cells. Proc Natl Acad Sci U S A 2022; 119:e2204808119. [PMID: 35749357 PMCID: PMC9245705 DOI: 10.1073/pnas.2204808119] [Citation(s) in RCA: 10] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/22/2022] [Accepted: 04/20/2022] [Indexed: 12/26/2022] Open
Abstract
As a sedentary epithelium turns motile during wound healing, morphogenesis, and metastasis, the Golgi apparatus moves from an apical position, above the nucleus, to a basal position. This apical-to-basal repositioning of Golgi is critical for epithelial cell migration. Yet the molecular mechanism underlying it remains elusive, although microtubules are believed to play a role. Using live-cell and super-resolution imaging, we show that at the onset of collective migration of epithelial cells, Golgi stacks get dispersed to create an unpolarized transitional structure, and surprisingly, this dispersal process depends not on microtubules but on actin cytoskeleton. Golgi-actin interaction involves Arp2/3-driven actin projections emanating from the actin cortex, and a Golgi-localized actin elongation factor, MENA. While in sedentary epithelial cells, actin projections intermittently interact with the apically located Golgi, and the frequency of this event increases before the dispersion of Golgi stacks, at the onset of cell migration. Preventing Golgi-actin interaction with MENA-mutants eliminates Golgi dispersion and reduces the persistence of cell migration. Taken together, we show a process of actin-driven Golgi dispersion that is mechanistically different from the well-known Golgi apparatus fragmentation during mitosis and is essential for collective migration of epithelial cells.
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Affiliation(s)
- Purnati Khuntia
- Tata Institute of Fundamental Research Hyderabad, Hyderabad 500 046, India
| | - Simran Rawal
- Tata Institute of Fundamental Research Hyderabad, Hyderabad 500 046, India
| | - Rituraj Marwaha
- Tata Institute of Fundamental Research Hyderabad, Hyderabad 500 046, India
| | - Tamal Das
- Tata Institute of Fundamental Research Hyderabad, Hyderabad 500 046, India
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27
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Pasapera AM, Heissler SM, Eto M, Nishimura Y, Fischer RS, Thiam HR, Waterman CM. MARK2 regulates directed cell migration through modulation of myosin II contractility and focal adhesion organization. Curr Biol 2022; 32:2704-2718.e6. [PMID: 35594862 DOI: 10.1016/j.cub.2022.04.088] [Citation(s) in RCA: 18] [Impact Index Per Article: 6.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/18/2021] [Revised: 02/23/2022] [Accepted: 04/28/2022] [Indexed: 12/11/2022]
Abstract
Cancer cell migration during metastasis is mediated by a highly polarized cytoskeleton. MARK2 and its invertebrate homolog Par1B are kinases that regulate the microtubule cytoskeleton to mediate polarization of neurons in mammals and embryos in invertebrates. However, the role of MARK2 in cancer cell migration is unclear. Using osteosarcoma cells, we found that in addition to its known localizations on microtubules and the plasma membrane, MARK2 also associates with the actomyosin cytoskeleton and focal adhesions. Cells depleted of MARK proteins demonstrated that MARK2 promotes phosphorylation of both myosin II and the myosin phosphatase targeting subunit MYPT1 to synergistically drive myosin II contractility and stress fiber formation in cells. Studies with isolated proteins showed that MARK2 directly phosphorylates myosin II regulatory light chain, while its effects on MYPT1 phosphorylation are indirect. Using a mutant lacking the membrane-binding domain, we found that membrane association is required for focal adhesion targeting of MARK2, where it specifically enhances cell protrusion by promoting FAK phosphorylation and formation of focal adhesions oriented in the direction of migration to mediate directionally persistent cell motility. Together, our results define MARK2 as a master regulator of the actomyosin and microtubule cytoskeletal systems and focal adhesions to mediate directional cancer cell migration.
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Affiliation(s)
- Ana M Pasapera
- Cell Biology and Physiology Center, National Heart, Lung and Blood Institute, National Institutes of Health, Building 50, South Drive, Room 4537, MSC 8019, Bethesda, MD 20892, USA
| | - Sarah M Heissler
- Cell Biology and Physiology Center, National Heart, Lung and Blood Institute, National Institutes of Health, Building 50, South Drive, Room 4537, MSC 8019, Bethesda, MD 20892, USA; Department of Physiology and Cell Biology, The Ohio State University College of Medicine, 370 W. 9th Avenue, Columbus, OH 43210, USA
| | - Masumi Eto
- Department of Veterinary Medicine, Okayama University of Science, 1-3 Ikoino-oka, Imabari, Ehime 794-8555, Japan
| | - Yukako Nishimura
- Cell Biology and Physiology Center, National Heart, Lung and Blood Institute, National Institutes of Health, Building 50, South Drive, Room 4537, MSC 8019, Bethesda, MD 20892, USA; Division of Developmental Physiology, Institute for Genetic Medicine, Hokkaido University, Kita 15, Nishi 7, Kita-Ku, Sapporo, Hokkaido 060-0815, Japan
| | - Robert S Fischer
- Cell Biology and Physiology Center, National Heart, Lung and Blood Institute, National Institutes of Health, Building 50, South Drive, Room 4537, MSC 8019, Bethesda, MD 20892, USA
| | - Hawa R Thiam
- Cell Biology and Physiology Center, National Heart, Lung and Blood Institute, National Institutes of Health, Building 50, South Drive, Room 4537, MSC 8019, Bethesda, MD 20892, USA
| | - Clare M Waterman
- Cell Biology and Physiology Center, National Heart, Lung and Blood Institute, National Institutes of Health, Building 50, South Drive, Room 4537, MSC 8019, Bethesda, MD 20892, USA.
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28
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Kazmi S, Khan MA, Shamma T, Altuhami A, Assiri AM, Broering DC. Therapeutic nexus of T cell immunometabolism in improving transplantation immunotherapy. Int Immunopharmacol 2022; 106:108621. [PMID: 35189469 DOI: 10.1016/j.intimp.2022.108621] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/06/2021] [Revised: 02/03/2022] [Accepted: 02/10/2022] [Indexed: 11/26/2022]
Abstract
Immunometabolism is a therapeutic strategy to tune immune cells through metabolic reprogramming, which allows immune cells to be differentiated according to their energy requirements. Recent therapeutic strategies targeting immunometabolism suggest that intracellular metabolic reprogramming controls T cell activation, proliferation, and differentiation into effector (Teff) or regulatory (Treg) cells. Immunometabolism is being studied for the treatment of inflammatory diseases, including those associated with solid organ transplantation (SOT). Here, we review immunometabolic regulation of immune cells, with a particular focus on Treg metabolic regulation and liver kinase B1 (LKB1) signaling, which stabilize Tregs and prevent inflammation-associated tissue injuries. All in all, here we discussed how targeting T cell immunometabolism modulates Teff and Treg-mediated immune responses, which can be used to boost Treg differentiation, stability, and ultimately favor immunotolerance in clinical transplants.
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Affiliation(s)
- Shadab Kazmi
- Transplant Research and Innovation Department, Organ Transplant Centre of Excellence, King Faisal Specialist Hospital and Research Centre, Riyadh 12713, Saudi Arabia.
| | - Mohammad Afzal Khan
- Transplant Research and Innovation Department, Organ Transplant Centre of Excellence, King Faisal Specialist Hospital and Research Centre, Riyadh 12713, Saudi Arabia.
| | - Talal Shamma
- Transplant Research and Innovation Department, Organ Transplant Centre of Excellence, King Faisal Specialist Hospital and Research Centre, Riyadh 12713, Saudi Arabia.
| | - Abdullah Altuhami
- Transplant Research and Innovation Department, Organ Transplant Centre of Excellence, King Faisal Specialist Hospital and Research Centre, Riyadh 12713, Saudi Arabia.
| | - Abdullah Mohammed Assiri
- Comparative Medicine Department, King Faisal Specialist Hospital and Research Centre, Riyadh 12713, Saudi Arabia; College of Medicine, Alfaisal University, Riyadh 11533, Saudi Arabia.
| | - Dieter Clemens Broering
- Transplant Research and Innovation Department, Organ Transplant Centre of Excellence, King Faisal Specialist Hospital and Research Centre, Riyadh 12713, Saudi Arabia.
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29
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Morales EA, Arnaiz C, Krystofiak ES, Zanic M, Tyska MJ. Mitotic Spindle Positioning (MISP) is an actin bundler that selectively stabilizes the rootlets of epithelial microvilli. Cell Rep 2022; 39:110692. [PMID: 35443169 PMCID: PMC9097542 DOI: 10.1016/j.celrep.2022.110692] [Citation(s) in RCA: 16] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/18/2021] [Revised: 01/20/2022] [Accepted: 03/24/2022] [Indexed: 11/11/2022] Open
Abstract
Microvilli are conserved actin-based surface protrusions that have been repurposed throughout evolution to fulfill diverse cell functions. In the case of transporting epithelia, microvilli are supported by a core of actin filaments bundled in parallel by villin, fimbrin, and espin. Remarkably, microvilli biogenesis persists in mice lacking all three of these factors, suggesting the existence of unknown bundlers. We identified Mitotic Spindle Positioning (MISP) as an actin-binding factor that localizes specifically to the rootlet end of the microvillus. MISP promotes rootlet elongation in cells, and purified MISP exhibits potent filament bundling activity in vitro. MISP-bundled filaments also recruit fimbrin, which further elongates and stabilizes bundles. MISP confinement to the rootlet is enforced by ezrin, which prevents decoration of the membrane-wrapped distal end of the core bundle. These discoveries reveal how epithelial cells optimize apical membrane surface area and offer insight on the remarkable robustness of microvilli biogenesis. Morales et al. identify Mitotic Spindle Positioning (MISP) as an actin bundler in the rootlets of epithelial microvilli. MISP cooperates with other bundlers, and its rootlet-specific localization is enforced by membrane-actin linker ezrin. These findings illuminate mechanisms that drive the assembly and compartmentalization of actin bundle-supported protrusions.
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Affiliation(s)
- E Angelo Morales
- Department of Cell and Developmental Biology, Vanderbilt University, Nashville, TN 37232, USA
| | - Cayetana Arnaiz
- Department of Cell and Developmental Biology, Vanderbilt University, Nashville, TN 37232, USA
| | - Evan S Krystofiak
- Cell Imaging Shared Resource, Vanderbilt University, Nashville, TN 37232, USA
| | - Marija Zanic
- Department of Cell and Developmental Biology, Vanderbilt University, Nashville, TN 37232, USA
| | - Matthew J Tyska
- Department of Cell and Developmental Biology, Vanderbilt University, Nashville, TN 37232, USA.
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30
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Rahman M, Ravichandran R, Bansal S, Sanborn K, Bowen S, Eschbacher J, Sureshbabu A, Fleming T, Bharat A, Walia R, Hachem R, Bremner RM, Smith MA, Mohanakumar T. Novel role for tumor suppressor gene, liver kinase B1, in epithelial-mesenchymal transition leading to chronic lung allograft dysfunction. Am J Transplant 2022; 22:843-852. [PMID: 34859569 DOI: 10.1111/ajt.16903] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/04/2021] [Revised: 11/16/2021] [Accepted: 11/16/2021] [Indexed: 01/25/2023]
Abstract
Epithelial-mesenchymal transition (EMT) has been implicated to play a role in chronic lung allograft dysfunction (CLAD). Liver kinase B1 (LKB1), a tumor suppressor gene, can regulate EMT. However, its role in CLAD development following lung transplantation remains unknown. Using qRT-PCR, biopsies from lung transplant recipients with bronchiolitis obliterans syndrome (BOS) demonstrated significant downregulation of LKB1 (p = .0001), compared to stable biopsies. To determine the role of LKB1 in EMT development, we analyzed EMT in human bronchial epithelial cell line BEAS-2B. Knockdown of LKB1 by siRNA significantly dysregulated mesenchymal markers expression in BEAS-2B cells. Following incubation of human primary bronchial epithelial cell or BEAS-2B cells with exosomes isolated from BOS or stable lung transplant recipients, LKB1 expression was inhibited when incubated with BOS-exosome. Incubation with BOS-exosomes also decreased LKB1 expression and induced EMT markers in air-liquid interface culture method. Our results provide novel evidence that exosomes released from transplanted lungs undergoing chronic rejection are associated with inactivated tumor suppressor gene LKB1 and this loss induces EMT leading to the pathogenesis of CLAD following human lung transplantation.
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Affiliation(s)
- Mohammad Rahman
- St. Joseph's Hospital and Medical Center, Norton Thoracic Institute, Phoenix, Arizona
| | | | - Sandhya Bansal
- St. Joseph's Hospital and Medical Center, Norton Thoracic Institute, Phoenix, Arizona
| | - Kristina Sanborn
- St. Joseph's Hospital and Medical Center, Norton Thoracic Institute, Phoenix, Arizona
| | - Sara Bowen
- St. Joseph's Hospital and Medical Center, Norton Thoracic Institute, Phoenix, Arizona
| | - Jennifer Eschbacher
- St. Joseph's Hospital and Medical Center, Barrow Neurological Institute, Phoenix, Arizona
| | - Angara Sureshbabu
- St. Joseph's Hospital and Medical Center, Norton Thoracic Institute, Phoenix, Arizona
| | - Timothy Fleming
- St. Joseph's Hospital and Medical Center, Norton Thoracic Institute, Phoenix, Arizona
| | | | - Rajat Walia
- St. Joseph's Hospital and Medical Center, Norton Thoracic Institute, Phoenix, Arizona
| | - Ramsey Hachem
- Washington University School of Medicine, St. Louis, Missouri
| | - Ross M Bremner
- St. Joseph's Hospital and Medical Center, Norton Thoracic Institute, Phoenix, Arizona
| | - Michael A Smith
- St. Joseph's Hospital and Medical Center, Norton Thoracic Institute, Phoenix, Arizona
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31
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Gaeta IM, Meenderink LM, Tyska MJ. A protocol for imaging microvilli biogenesis on the surface of cultured porcine kidney epithelial cell monolayers. STAR Protoc 2021; 2:100998. [PMID: 34950883 PMCID: PMC8672049 DOI: 10.1016/j.xpro.2021.100998] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/23/2022] Open
Abstract
A key facet of epithelial differentiation is the assembly of actin-based protrusions known as microvilli, which amplify apical membrane surface area for various cell functions. To probe mechanisms of microvillus assembly, we developed a protocol using spinning disk confocal microscopy to directly visualize microvillus biogenesis on the surface of cultured porcine kidney epithelial cell monolayers engineered to express fluorescent proteins. This protocol offers access to the molecular details of individual protrusion growth events at high spatiotemporal resolution. For complete details on the use and execution of this protocol, please refer to Gaeta et al. (2021).
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Affiliation(s)
- Isabella M. Gaeta
- Department of Cell and Developmental Biology, Vanderbilt University, Nashville, TN 37232, USA
| | - Leslie M. Meenderink
- Department of Medicine, Division of Infectious Diseases, Vanderbilt University Medical Center, Nashville, TN 37232, USA
- Veterans Affairs Tennessee Valley Health Care System, Nashville, TN 37212, USA
| | - Matthew J. Tyska
- Department of Cell and Developmental Biology, Vanderbilt University, Nashville, TN 37232, USA
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STK25 and MST3 Have Overlapping Roles to Regulate Rho GTPases during Cortical Development. J Neurosci 2021; 41:8887-8903. [PMID: 34518307 DOI: 10.1523/jneurosci.0523-21.2021] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/10/2021] [Revised: 08/13/2021] [Accepted: 09/06/2021] [Indexed: 11/21/2022] Open
Abstract
Precise control of neuronal migration is required for the laminar organization of the neocortex and critical for brain function. We previously reported that the acute disruption of the Stk25 gene (Stk25 conditional knock-out; cKO) during mouse embryogenesis causes anomalous neuronal migration in the neocortex, but paradoxically the Stk25 cKO did not have a cortical phenotype, suggesting some forms of compensation exist. In this study, we report that MST3, another member of the GCKIII subgroup of the Ste20-like kinase family, compensates for loss of Stk25 and vice versa with sex independent manner. MST3 overexpression rescued neuronal migration deficit and abnormal axonogenesis in Stk25 cKO brains. Mechanistically, STK25 leads to Rac1 activation and reduced RhoA levels in the developing brain, both of which are required to fully restore neuronal migration in the Stk25 cKO brain. Abnormal migration phenotypes are also rescued by overexpression of Bacurd1and Cul3, which target RhoA for degradation, and activate Rac1. This study reveals that MST3 upregulation is capable of rescuing acute Stk25 deficiency and resolves details of signaling downstream STK25 required for corticogenesis both common to and distinct from MST3 signaling.SIGNIFICANCE STATEMENT Proper neuronal migration during cortical development is required for normal neuronal function. Here, we show that STK25 and MST3 kinases regulate neuronal migration and polarization in a mutually compensatory manner. Furthermore, STK25 balances Rac1 activity and RhoA level through forming complexes with α-PIX and β-PIX, GTPase regulatory enzymes, and Cullin3-Bacurd1/Kctd13, a pair of RhoA ubiquitination molecules in a kinase activity-independent manner. Our findings demonstrate the importance of overlapping and unique roles of STK25 and MST3 to regulate Rho GTPase activities in cortical development.
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González-González L, Gallego-Gutiérrez H, Martin-Tapia D, Avelino-Cruz JE, Hernández-Guzmán C, Rangel-Guerrero SI, Alvarez-Salas LM, Garay E, Chávez-Munguía B, Gutiérrez-Ruiz MC, Hernández-Melchor D, López-Bayghen E, González-Mariscal L. ZO-2 favors Hippo signaling, and its re-expression in the steatotic liver by AMPK restores junctional sealing. Tissue Barriers 2021; 10:1994351. [PMID: 34689705 DOI: 10.1080/21688370.2021.1994351] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/05/2023] Open
Abstract
ZO-2 is a peripheral tight junction (TJ) protein whose silencing in renal epithelia induces cell hypertrophy. Here, we found that in ZO-2 KD MDCK cells, in compensatory renal hypertrophy triggered in rats by a unilateral nephrectomy and in liver steatosis of obese Zucker (OZ) rats, ZO-2 silencing is accompanied by the diminished activity of LATS, a kinase of the Hippo pathway, and the nuclear concentration of YAP, the final effector of this signaling route. ZO-2 appears to function as a scaffold for the Hippo pathway as it associates to LATS1. ZO-2 silencing in hypertrophic tissue is due to a diminished abundance of ZO-2 mRNA, and the Sp1 transcription factor is critical for ZO-2 transcription in renal cells. Treatment of OZ rats with metformin, an activator of AMPK that blocks JNK activity, augments ZO-2 and claudin-1 expression in the liver, reduces the paracellular permeability of hepatocytes, and serum bile acid content. Our results suggest that ZO-2 silencing is a common feature of hypertrophy, and that ZO-2 is a positive regulator of the Hippo pathway that regulates cell size. Moreover, our observations highlight the importance of AMPK, JNK, and ZO-2 as therapeutic targets for blood-bile barrier dysfunction.
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Affiliation(s)
- Laura González-González
- Department of Physiology, Biophysics, and Neurosciences, Center for Research and Advanced Studies (Cinvestav), Mexico City, Mexico
| | - Helios Gallego-Gutiérrez
- Department of Physiology, Biophysics, and Neurosciences, Center for Research and Advanced Studies (Cinvestav), Mexico City, Mexico
| | - Dolores Martin-Tapia
- Department of Physiology, Biophysics, and Neurosciences, Center for Research and Advanced Studies (Cinvestav), Mexico City, Mexico
| | - José Everardo Avelino-Cruz
- Laboratory of Molecular Cardiology, Institute of Physiology, Benemérita Universidad Autónoma de Puebla, Puebla, Mexico
| | - Christian Hernández-Guzmán
- Department of Physiology, Biophysics, and Neurosciences, Center for Research and Advanced Studies (Cinvestav), Mexico City, Mexico
| | - Sergio Israel Rangel-Guerrero
- Department of Genetics and Molecular Biology, Center for Research and Advanced Studies (Cinvestav), Mexico City, Mexico
| | - Luis Marat Alvarez-Salas
- Department of Genetics and Molecular Biology, Center for Research and Advanced Studies (Cinvestav), Mexico City, Mexico
| | - Erika Garay
- Department of Physiology, Biophysics, and Neurosciences, Center for Research and Advanced Studies (Cinvestav), Mexico City, Mexico
| | - Bibiana Chávez-Munguía
- Department of Infectomics and Molecular Pathogenesis, Center for Research and Advanced Studies (Cinvestav), Mexico City, Mexico
| | - María Concepción Gutiérrez-Ruiz
- Department of Health Sciences, Autonomous Metropolitan University- Iztapalapa (UAM-I), Mexico City, Mexico; Laboratory of Experimental Medicine, Unit of Translational Medicine, Institute of Biomedical Research, Unam, National Institute of Cardiology "Ignacio Chávez", Mexico City, Mexico
| | | | - Esther López-Bayghen
- Department of Toxicology, Center for Research and Advanced Studies (Cinvestav), Mexico City, Mexico
| | - Lorenza González-Mariscal
- Department of Physiology, Biophysics, and Neurosciences, Center for Research and Advanced Studies (Cinvestav), Mexico City, Mexico
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Ruebel ML, Zambelli F, Schall PZ, Barragan M, VandeVoort CA, Vassena R, Latham KE. Shared aspects of mRNA expression associated with oocyte maturation failure in humans and rhesus monkeys indicating compromised oocyte quality. Physiol Genomics 2021; 53:137-149. [PMID: 33554756 DOI: 10.1152/physiolgenomics.00155.2020] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/08/2023] Open
Abstract
Oocyte maturation failure observed in assisted reproduction technology (ART) cycles can limit the number of quality oocytes obtained and present a pronounced barrier for some patients. The potential exists to use unmatured oocytes for ART through in vitro maturation. Understanding the molecular basis of oocyte maturation failure is pertinent to minimizing this loss of oocytes and considerations of whether such oocytes can be used safely for ART. We identified shared transcriptome abnormalities for rhesus monkey and human failed-to-mature (FTM) oocytes relative to healthy matured MII stage oocytes. We discovered that, although the number of shared affected genes was comparatively small, FTM oocytes in both species shared effects for several pathways and functions, including predicted activation of oxidative phosphorylation (OxPhos) with additional effects on mitochondrial function, lipid metabolism, transcription, nucleotide excision repair, endoplasmic reticulum stress, unfolded protein response, and cell viability. RICTOR emerged as a prominent upstream regulator with predicted inhibition across all analyses. Alterations in KDM5A, MTOR, MTORC1, INSR, CAB39L, and STK11 activities were implicated along with RICTOR in modulating mitochondrial activity and OxPhos. Defects in cell cycle progression were not a prominent feature of FTM oocytes. These results identify a common set of transcriptome abnormalities associated with oocyte maturation failure. While our results do not demonstrate causality, they indicate that fundamental aspects of cellular function are abnormal in FTM oocytes and raise significant concerns about the potential risks of using FTM oocytes for ART.
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Affiliation(s)
- Meghan L Ruebel
- Department of Animal Science and Reproductive and Developmental Sciences Program, Michigan State University, East Lansing, Michigan
| | | | - Peter Z Schall
- Department of Animal Science and Reproductive and Developmental Sciences Program, Michigan State University, East Lansing, Michigan
| | | | - Catherine A VandeVoort
- California National Primate Research Center, University of California, Davis, California.,Department of Obstetrics and Gynecology, University of California, Davis, California
| | | | - Keith E Latham
- Department of Animal Science and Reproductive and Developmental Sciences Program, Michigan State University, East Lansing, Michigan
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35
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Chinowsky CR, Pinette JA, Meenderink LM, Lau KS, Tyska MJ. Nonmuscle myosin-2 contractility-dependent actin turnover limits the length of epithelial microvilli. Mol Biol Cell 2020; 31:2803-2815. [PMID: 33026933 PMCID: PMC7851865 DOI: 10.1091/mbc.e20-09-0582] [Citation(s) in RCA: 26] [Impact Index Per Article: 5.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/10/2020] [Revised: 09/24/2020] [Accepted: 09/28/2020] [Indexed: 12/14/2022] Open
Abstract
Brush border microvilli enable functions that are critical for epithelial homeostasis, including solute uptake and host defense. However, the mechanisms that regulate the assembly and morphology of these protrusions are poorly understood. The parallel actin bundles that support microvilli have their pointed-end rootlets anchored in a filamentous meshwork referred to as the "terminal web." Although classic electron microscopy studies revealed complex ultrastructure, the composition and function of the terminal web remain unclear. Here we identify nonmuscle myosin-2C (NM2C) as a component of the terminal web. NM2C is found in a dense, isotropic layer of puncta across the subapical domain, which transects the rootlets of microvillar actin bundles. Puncta are separated by ∼210 nm, the expected size of filaments formed by NM2C. In intestinal organoid cultures, the terminal web NM2C network is highly dynamic and exhibits continuous remodeling. Using pharmacological and genetic perturbations in cultured intestinal epithelial cells, we found that NM2C controls the length of growing microvilli by regulating actin turnover in a manner that requires a fully active motor domain. Our findings answer a decades-old question on the function of terminal web myosin and hold broad implications for understanding apical morphogenesis in diverse epithelial systems.
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Affiliation(s)
- Colbie R Chinowsky
- Department of Cell and Developmental Biology, Vanderbilt University School of Medicine, Nashville, TN 37232
| | - Julia A Pinette
- Department of Molecular Physiology and Biophysics, Vanderbilt University School of Medicine, Nashville, TN 37232
| | - Leslie M Meenderink
- Division of Infectious Diseases, Department of Medicine, Vanderbilt University Medical Center, Nashville, TN 37232
| | - Ken S Lau
- Department of Cell and Developmental Biology, Vanderbilt University School of Medicine, Nashville, TN 37232
| | - Matthew J Tyska
- Department of Cell and Developmental Biology, Vanderbilt University School of Medicine, Nashville, TN 37232
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36
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Yang W, Wang L, Wang F, Yuan S. Roles of AMP-Activated Protein Kinase (AMPK) in Mammalian Reproduction. Front Cell Dev Biol 2020; 8:593005. [PMID: 33330475 PMCID: PMC7710906 DOI: 10.3389/fcell.2020.593005] [Citation(s) in RCA: 26] [Impact Index Per Article: 5.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/09/2020] [Accepted: 10/23/2020] [Indexed: 12/01/2022] Open
Abstract
Reproduction is an energy demanding function and only take place in case of sufficient available energy status in mammals. Metabolic diseases such as anorexia nervosa are clinically associated with reduced fertility. AMP-activated protein kinase (AMPK), as a major regulator of cellular energy homeostasis, is activated in limited energy reserves to ensure the orderly progress of various physiological activities. In recent years, mounting evidence shows that AMPK is involved in the regulation of reproductive function through multiple mechanisms. AMPK is likely to be a metabolic sensor integrating central and peripheral signals. In this review, we aim to explore the preclinical studies published in the last decade that investigate the role of AMP-activated protein kinase in the reproductive field, and its role as a target for drug therapy of reproductive system-related diseases. We also emphasized the emerging roles of AMPK in transcriptional regulation of reproduction processes and metabolisms, which are tightly related to the energy state and fertility of an organism.
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Affiliation(s)
- Weina Yang
- Institute of Reproductive Health, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China
| | - Lingjuan Wang
- Institute of Reproductive Health, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China
| | - Fengli Wang
- Institute of Reproductive Health, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China
| | - Shuiqiao Yuan
- Institute of Reproductive Health, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China
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37
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Díaz-Díaz C, Martín-Belmonte F. Apical poles without neighbouring cells. NATURE MATERIALS 2020; 19:935-937. [PMID: 32820290 DOI: 10.1038/s41563-020-0766-9] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 06/11/2023]
Affiliation(s)
- Covadonga Díaz-Díaz
- Centro de Biología Molecular Severo Ochoa (CBMSO), Tissue and Organ Homeostasis Program, Consejo Superior de Investigaciones Científicas (CSIC) and Universidad Autónoma de Madrid (UAM), Madrid, Spain
| | - Fernando Martín-Belmonte
- Centro de Biología Molecular Severo Ochoa (CBMSO), Tissue and Organ Homeostasis Program, Consejo Superior de Investigaciones Científicas (CSIC) and Universidad Autónoma de Madrid (UAM), Madrid, Spain.
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38
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Zhang Y, De Mets R, Monzel C, Acharya V, Toh P, Chin JFL, Van Hul N, Ng IC, Yu H, Ng SS, Tamir Rashid S, Viasnoff V. Biomimetic niches reveal the minimal cues to trigger apical lumen formation in single hepatocytes. NATURE MATERIALS 2020; 19:1026-1035. [PMID: 32341512 DOI: 10.1038/s41563-020-0662-3] [Citation(s) in RCA: 21] [Impact Index Per Article: 4.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 05/10/2019] [Accepted: 03/11/2020] [Indexed: 06/11/2023]
Abstract
The symmetry breaking of protein distribution and cytoskeleton organization is an essential aspect for the development of apicobasal polarity. In embryonic cells this process is largely cell autonomous, while differentiated epithelial cells collectively polarize during epithelium formation. Here, we demonstrate that the de novo polarization of mature hepatocytes does not require the synchronized development of apical poles on neighbouring cells. De novo polarization at the single-cell level by mere contact with the extracellular matrix and immobilized cadherin defining a polarizing axis. The creation of these single-cell liver hemi-canaliculi allows unprecedented imaging resolution and control and over the lumenogenesis process. We show that the density and localization of cadherins along the initial cell-cell contact act as key triggers of the reorganization from lateral to apical actin cortex. The minimal cues necessary to trigger the polarization of hepatocytes enable them to develop asymmetric lumens with ectopic epithelial cells originating from the kidney, breast or colon.
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Affiliation(s)
- Yue Zhang
- Mechanobiology Institute, National University of Singapore, Singapore, Singapore
| | - Richard De Mets
- Mechanobiology Institute, National University of Singapore, Singapore, Singapore
| | - Cornelia Monzel
- Experimental Medical Physics, Heinrich-Heine University, Düsseldorf, Germany
| | | | - Pearlyn Toh
- Mechanobiology Institute, National University of Singapore, Singapore, Singapore
| | - Jasmine Fei Li Chin
- Mechanobiology Institute, National University of Singapore, Singapore, Singapore
| | - Noémi Van Hul
- Department of Biosciences and Nutrition, Karolinska Institutet, Stockholm, Sweden
- Institute of Molecular and Cell Biology, A*STAR, Singapore, Singapore
| | - Inn Chuan Ng
- Department of Physiology, Yong Loo Lin School of Medicine, National University Health System, Singapore, Singapore
| | - Hanry Yu
- Mechanobiology Institute, National University of Singapore, Singapore, Singapore
- Department of Physiology, Yong Loo Lin School of Medicine, National University Health System, Singapore, Singapore
- Institute of Bioengineering and Nanotechnology, Agency for Science, Technology and Research, Singapore, Singapore
| | - Soon Seng Ng
- Centre for Stem Cells and Regenerative Medicine, King's College London, London, UK
| | - S Tamir Rashid
- Centre for Stem Cells and Regenerative Medicine, King's College London, London, UK
- Institute for Liver Studies, King's College Hospital, King's College London, London, UK
| | - Virgile Viasnoff
- Mechanobiology Institute, National University of Singapore, Singapore, Singapore.
- Department of Biological Science, National University of Singapore, Singapore, Singapore.
- Centre National de la Recherche Scientifique Unité Mixte Internationale, Singapore, Singapore.
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39
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Jossin Y. Molecular mechanisms of cell polarity in a range of model systems and in migrating neurons. Mol Cell Neurosci 2020; 106:103503. [PMID: 32485296 DOI: 10.1016/j.mcn.2020.103503] [Citation(s) in RCA: 21] [Impact Index Per Article: 4.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/06/2020] [Revised: 04/20/2020] [Accepted: 05/23/2020] [Indexed: 01/09/2023] Open
Abstract
Cell polarity is defined as the asymmetric distribution of cellular components along an axis. Most cells, from the simplest single-cell organisms to highly specialized mammalian cells, are polarized and use similar mechanisms to generate and maintain polarity. Cell polarity is important for cells to migrate, form tissues, and coordinate activities. During development of the mammalian cerebral cortex, cell polarity is essential for neurogenesis and for the migration of newborn but as-yet undifferentiated neurons. These oriented migrations include both the radial migration of excitatory projection neurons and the tangential migration of inhibitory interneurons. In this review, I will first describe the development of the cerebral cortex, as revealed at the cellular level. I will then define the core molecular mechanisms - the Par/Crb/Scrib polarity complexes, small GTPases, the actin and microtubule cytoskeletons, and phosphoinositides/PI3K signaling - that are required for asymmetric cell division, apico-basal and front-rear polarity in model systems, including C elegans zygote, Drosophila embryos and cultured mammalian cells. As I go through each core mechanism I will explain what is known about its importance in radial and tangential migration in the developing mammalian cerebral cortex.
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Affiliation(s)
- Yves Jossin
- Laboratory of Mammalian Development & Cell Biology, Institute of Neuroscience, Université Catholique de Louvain, Brussels, Belgium.
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40
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Chauhan AS, Zhuang L, Gan B. Spatial control of AMPK signaling at subcellular compartments. Crit Rev Biochem Mol Biol 2020; 55:17-32. [PMID: 32069425 DOI: 10.1080/10409238.2020.1727840] [Citation(s) in RCA: 27] [Impact Index Per Article: 5.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
Abstract
AMP-activated protein kinase (AMPK) is a master regulator of energy homeostasis that functions to restore the energy balance by phosphorylating its substrates during altered metabolic conditions. AMPK activity is tightly controlled by diverse regulators including its upstream kinases LKB1 and CaMKK2. Recent studies have also identified the localization of AMPK at different intracellular compartments as another key mechanism for regulating AMPK signaling in response to specific stimuli. This review discusses the AMPK signaling associated with different subcellular compartments, including lysosomes, endoplasmic reticulum, mitochondria, Golgi apparatus, nucleus, and cell junctions. Because altered AMPK signaling is associated with various pathologic conditions including cancer, targeting AMPK signaling in different subcellular compartments may present attractive therapeutic approaches for treatment of disease.
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Affiliation(s)
- Anoop Singh Chauhan
- Department of Experimental Radiation Oncology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA.,Institute of Cancer and Genomic Sciences, University of Birmingham, Birmingham, UK
| | - Li Zhuang
- Department of Experimental Radiation Oncology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA
| | - Boyi Gan
- Department of Experimental Radiation Oncology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA.,Department of Molecular and Cellular Oncology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA.,Health Graduate School of Biomedical Sciences, The University of Texas MD Anderson UT, Houston, TX, USA
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41
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Roehlen N, Roca Suarez AA, El Saghire H, Saviano A, Schuster C, Lupberger J, Baumert TF. Tight Junction Proteins and the Biology of Hepatobiliary Disease. Int J Mol Sci 2020; 21:ijms21030825. [PMID: 32012812 PMCID: PMC7038100 DOI: 10.3390/ijms21030825] [Citation(s) in RCA: 38] [Impact Index Per Article: 7.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/04/2019] [Revised: 01/10/2020] [Accepted: 01/21/2020] [Indexed: 12/24/2022] Open
Abstract
Tight junctions (TJ) are intercellular adhesion complexes on epithelial cells and composed of integral membrane proteins as well as cytosolic adaptor proteins. Tight junction proteins have been recognized to play a key role in health and disease. In the liver, TJ proteins have several functions: they contribute as gatekeepers for paracellular diffusion between adherent hepatocytes or cholangiocytes to shape the blood-biliary barrier (BBIB) and maintain tissue homeostasis. At non-junctional localizations, TJ proteins are involved in key regulatory cell functions such as differentiation, proliferation, and migration by recruiting signaling proteins in response to extracellular stimuli. Moreover, TJ proteins are hepatocyte entry factors for the hepatitis C virus (HCV)—a major cause of liver disease and cancer worldwide. Perturbation of TJ protein expression has been reported in chronic HCV infection, cholestatic liver diseases as well as hepatobiliary carcinoma. Here we review the physiological function of TJ proteins in the liver and their implications in hepatobiliary diseases.
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Affiliation(s)
- Natascha Roehlen
- Institut de Recherche sur les Maladies Virales et Hépatiques, Inserm UMR1110, F-67000 Strasbourg, France; (N.R.); (A.A.R.S.); (H.E.S.); (A.S.); (C.S.); (J.L.)
- Université de Strasbourg, F-67000 Strasbourg, France
| | - Armando Andres Roca Suarez
- Institut de Recherche sur les Maladies Virales et Hépatiques, Inserm UMR1110, F-67000 Strasbourg, France; (N.R.); (A.A.R.S.); (H.E.S.); (A.S.); (C.S.); (J.L.)
- Université de Strasbourg, F-67000 Strasbourg, France
| | - Houssein El Saghire
- Institut de Recherche sur les Maladies Virales et Hépatiques, Inserm UMR1110, F-67000 Strasbourg, France; (N.R.); (A.A.R.S.); (H.E.S.); (A.S.); (C.S.); (J.L.)
- Université de Strasbourg, F-67000 Strasbourg, France
| | - Antonio Saviano
- Institut de Recherche sur les Maladies Virales et Hépatiques, Inserm UMR1110, F-67000 Strasbourg, France; (N.R.); (A.A.R.S.); (H.E.S.); (A.S.); (C.S.); (J.L.)
- Université de Strasbourg, F-67000 Strasbourg, France
- Pôle Hepato-digestif, Institut Hopitalo-universitaire, Hôpitaux Universitaires de Strasbourg, F-67000 Strasbourg, France
| | - Catherine Schuster
- Institut de Recherche sur les Maladies Virales et Hépatiques, Inserm UMR1110, F-67000 Strasbourg, France; (N.R.); (A.A.R.S.); (H.E.S.); (A.S.); (C.S.); (J.L.)
- Université de Strasbourg, F-67000 Strasbourg, France
| | - Joachim Lupberger
- Institut de Recherche sur les Maladies Virales et Hépatiques, Inserm UMR1110, F-67000 Strasbourg, France; (N.R.); (A.A.R.S.); (H.E.S.); (A.S.); (C.S.); (J.L.)
- Université de Strasbourg, F-67000 Strasbourg, France
| | - Thomas F. Baumert
- Institut de Recherche sur les Maladies Virales et Hépatiques, Inserm UMR1110, F-67000 Strasbourg, France; (N.R.); (A.A.R.S.); (H.E.S.); (A.S.); (C.S.); (J.L.)
- Université de Strasbourg, F-67000 Strasbourg, France
- Pôle Hepato-digestif, Institut Hopitalo-universitaire, Hôpitaux Universitaires de Strasbourg, F-67000 Strasbourg, France
- Correspondence: ; Tel.: +33-3688-53703
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Reciprocal Association between the Apical Junctional Complex and AMPK: A Promising Therapeutic Target for Epithelial/Endothelial Barrier Function? Int J Mol Sci 2019; 20:ijms20236012. [PMID: 31795328 PMCID: PMC6928779 DOI: 10.3390/ijms20236012] [Citation(s) in RCA: 18] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/08/2019] [Revised: 11/26/2019] [Accepted: 11/26/2019] [Indexed: 12/17/2022] Open
Abstract
Epithelial/endothelial cells adhere to each other via cell–cell junctions including tight junctions (TJs) and adherens junctions (AJs). TJs and AJs are spatiotemporally and functionally integrated, and are thus often collectively defined as apical junctional complexes (AJCs), regulating a number of spatiotemporal events including paracellular barrier, selective permeability, apicobasal cell polarity, mechano-sensing, intracellular signaling cascades, and epithelial morphogenesis. Over the past 15 years, it has been acknowledged that adenosine monophosphate (AMP)-activated protein kinase (AMPK), a well-known central regulator of energy metabolism, has a reciprocal association with AJCs. Here, we review the current knowledge of this association and show the following evidences: (1) as an upstream regulator, AJs activate the liver kinase B1 (LKB1)–AMPK axis particularly in response to applied junctional tension, and (2) TJ function and apicobasal cell polarization are downstream targets of AMPK and are promoted by AMPK activation. Although molecular mechanisms underlying these phenomena have not yet been completely elucidated, identifications of novel AMPK effectors in AJCs and AMPK-driven epithelial transcription factors have enhanced our knowledge. More intensive studies along this line would eventually lead to the development of AMPK-based therapies, enabling us to manipulate epithelial/endothelial barrier function.
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Motegi F, Plachta N, Viasnoff V. Novel approaches to link apicobasal polarity to cell fate specification. Curr Opin Cell Biol 2019; 62:78-85. [PMID: 31731147 DOI: 10.1016/j.ceb.2019.09.003] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/09/2019] [Revised: 09/19/2019] [Accepted: 09/26/2019] [Indexed: 12/26/2022]
Abstract
Understanding the development of apicobasal polarity (ABP) is a long-standing problem in biology. The molecular components involved in the development and maintenance of APB have been largely identified and are known to have ubiquitous roles across organisms. Our knowledge of the functional consequences of ABP establishment and maintenance is far less comprehensive. Recent studies using novel experimental approaches and cellular models have revealed a growing link between ABP and the genetic program of cell lineage. This mini-review describes some of the most recent advances in this new field, highlighting examples from Caenorhabditis elegans and mouse embryos, human pluripotent stem cells, and epithelial cells. We also speculate on the most interesting and challenging avenues that can be explored.
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Affiliation(s)
- Fumio Motegi
- Department of Biological Sciences, National University of Singapore, 117583, Singapore; Mechanobiology Institute, National University of Singapore, 117 411, Singapore; Temasek Life-sciences Laboratory, 117604, Singapore; Contributed equally
| | - Nicolas Plachta
- Institute of Molecular and Cell Biology, ASTAR, Singapore; Contributed equally
| | - Virgile Viasnoff
- Department of Biological Sciences, National University of Singapore, 117583, Singapore; Mechanobiology Institute, National University of Singapore, 117 411, Singapore; CNRS, 117411, Singapore; Contributed equally.
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44
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Lammert E, Thorn P. The Role of the Islet Niche on Beta Cell Structure and Function. J Mol Biol 2019; 432:1407-1418. [PMID: 31711959 DOI: 10.1016/j.jmb.2019.10.032] [Citation(s) in RCA: 46] [Impact Index Per Article: 7.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/20/2019] [Revised: 10/25/2019] [Accepted: 10/29/2019] [Indexed: 01/15/2023]
Abstract
The islets of Langerhans or pancreatic islets are pivotal in the control of blood glucose and are complex microorgans embedded within the larger volume of the exocrine pancreas. Humans can have ~3.2 million islets [1] which, to our current knowledge, function in a similar manner to sense circulating blood glucose levels and respond with the secretion of a mix of different hormones that act to maintain glucose concentrations around a specific set point [2]. At a cellular level, the control of hormone secretion by glucose and other secretagogues is well-understood [3]. The key signal cascades have been identified and many details of the secretory process are known. However, if we shift focus from single cells and consider cells within intact islets, we do not have a comprehensive model as to how the islet environment influences cell function and how the islets work as a whole. This is important because there is overwhelming evidence that the structure and function of the individual endocrine cells are dramatically affected by the islet environment [4,5]. Uncovering the influence of this islet niche might drive future progress in treatments for Type 2 diabetes [6] and cell replacement therapies for Type 1 diabetes [7]. In this review, we focus on the insulin secreting beta cells and their interactions with the immediate environment that surrounds them including endocrine-endocrine interactions and contacts with capillaries.
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Affiliation(s)
- Eckhard Lammert
- Institute of Metabolic Physiology, Heinrich Heine University, Düsseldorf, Germany; Institute for Vascular and Islet Cell Biology, German Diabetes Center, Leibniz Center for Diabetes Research, Heinrich Heine University, Düsseldorf, Germany; German Center for Diabetes Research (DZD e.V.), Neuherberg, Germany
| | - Peter Thorn
- Charles Perkins Centre, School of Medical Sciences, University of Sydney, Camperdown, NSW 2006, Australia.
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Olivier S, Leclerc J, Grenier A, Foretz M, Tamburini J, Viollet B. AMPK Activation Promotes Tight Junction Assembly in Intestinal Epithelial Caco-2 Cells. Int J Mol Sci 2019; 20:E5171. [PMID: 31635305 PMCID: PMC6829419 DOI: 10.3390/ijms20205171] [Citation(s) in RCA: 42] [Impact Index Per Article: 7.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/17/2019] [Revised: 10/16/2019] [Accepted: 10/17/2019] [Indexed: 12/13/2022] Open
Abstract
The AMP-activated protein kinase (AMPK) is principally known as a major regulator of cellular energy status, but it has been recently shown to play a key structural role in cell-cell junctions. The aim of this study was to evaluate the impact of AMPK activation on the reassembly of tight junctions in intestinal epithelial Caco-2 cells. We generated Caco-2 cells invalidated for AMPK α1/α2 (AMPK dKO) by CRISPR/Cas9 technology and evaluated the effect of the direct AMPK activator 991 on the reassembly of tight junctions following a calcium switch assay. We analyzed the integrity of the epithelial barrier by measuring the trans-epithelial electrical resistance (TEER), the paracellular permeability, and quantification of zonula occludens 1 (ZO-1) deposit at plasma membrane by immunofluorescence. Here, we demonstrated that AMPK deletion induced a delay in tight junction reassembly and relocalization at the plasma membrane during calcium switch, leading to impairments in the establishment of TEER and paracellular permeability. We also showed that 991-induced AMPK activation accelerated the reassembly and reorganization of tight junctions, improved the development of TEER and paracellular permeability after calcium switch. Thus, our results show that AMPK activation ensures a better recovery of epithelial barrier function following injury.
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Affiliation(s)
- Séverine Olivier
- Université de Paris, Institut Cochin, CNRS UMR8104, INSERM U1016, F-75014 Paris, France.
| | - Jocelyne Leclerc
- Université de Paris, Institut Cochin, CNRS UMR8104, INSERM U1016, F-75014 Paris, France.
| | - Adrien Grenier
- Université de Paris, Institut Cochin, CNRS UMR8104, INSERM U1016, F-75014 Paris, France.
| | - Marc Foretz
- Université de Paris, Institut Cochin, CNRS UMR8104, INSERM U1016, F-75014 Paris, France.
| | - Jérôme Tamburini
- Université de Paris, Institut Cochin, CNRS UMR8104, INSERM U1016, F-75014 Paris, France.
| | - Benoit Viollet
- Université de Paris, Institut Cochin, CNRS UMR8104, INSERM U1016, F-75014 Paris, France.
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46
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Faust JJ, Millis BA, Tyska MJ. Profilin-Mediated Actin Allocation Regulates the Growth of Epithelial Microvilli. Curr Biol 2019; 29:3457-3465.e3. [PMID: 31607529 DOI: 10.1016/j.cub.2019.08.051] [Citation(s) in RCA: 15] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/29/2019] [Revised: 07/17/2019] [Accepted: 08/20/2019] [Indexed: 01/18/2023]
Abstract
Transporting epithelial cells, like those that line the intestinal tract, are specialized for solute processing and uptake. One defining feature is the brush border, an array of microvilli that serves to amplify apical membrane surface area and increase functional capacity. During differentiation, upon exit from stem-cell-containing crypts, enterocytes build thousands of microvilli, each supported by a parallel bundle of actin filaments several microns in length. Given the high concentration of actin residing in mature brush borders, we sought to determine whether enterocytes were resource (i.e., actin monomer) limited in assembling this domain. To examine this possibility, we inhibited Arp2/3, the ubiquitous branched actin nucleator, to increase G-actin availability during brush border assembly. In native intestinal tissues, Arp2/3 inhibition led to increased microvilli length on the surface of crypt, but not villus, enterocytes. In a cell culture model of brush border assembly, Arp2/3 inhibition accelerated the growth and increased the length of microvilli; it also led to a redistribution of F-actin from cortical lateral networks into the brush border. Effects on brush border growth were rescued by treatment with the G-actin sequestering drug, latrunculin A. G-actin binding protein, profilin-1, colocalized in the terminal web with G-actin, and knockdown of this factor compromised brush border growth in a concentration-dependent manner. Finally, the acceleration in brush border assembly induced by Arp2/3 inhibition was abrogated by profilin-1 knockdown. Thus, brush border assembly is limited by G-actin availability, and profilin-1 directs unallocated actin monomers into microvillar core bundles during enterocyte differentiation.
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Affiliation(s)
- James J Faust
- Department of Cell and Developmental Biology, Vanderbilt University School of Medicine, Nashville, TN 37232, USA
| | - Bryan A Millis
- Department of Cell and Developmental Biology, Vanderbilt University School of Medicine, Nashville, TN 37232, USA; Department of Biomedical Engineering, Vanderbilt University School of Engineering, Nashville, TN 37232, USA; Cell Imaging Shared Resource, Vanderbilt University School of Medicine, Nashville, TN 37232, USA; Vanderbilt Biophotonics Center, Vanderbilt University, Nashville, TN 37232, USA
| | - Matthew J Tyska
- Department of Cell and Developmental Biology, Vanderbilt University School of Medicine, Nashville, TN 37232, USA.
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Abstract
Small GTPases are organizers of a plethora of cellular processes. The time and place of their activation are tightly controlled by the localization and activation of their regulators, guanine-nucleotide exchange factors (GEFs) and GTPase-activating proteins (GAPs). Remarkably, in some systems, the upstream regulators of GTPases are also found downstream of their activity. Resulting feedback loops can generate complex spatiotemporal dynamics of GTPases with important functional consequences. Here we discuss the concept of positive autoregulation of small GTPases by the GEF-effector feedback modules and survey recent developments in this exciting area of cell biology.
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Affiliation(s)
- Andrew B. Goryachev
- Centre for Synthetic and Systems Biology, Institute for Cell Biology, University of Edinburgh, Edinburgh, EH9 3BF, UK
| | - Marcin Leda
- Centre for Synthetic and Systems Biology, Institute for Cell Biology, University of Edinburgh, Edinburgh, EH9 3BF, UK
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48
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Postema MM, Grega-Larson NE, Meenderink LM, Tyska MJ. PACSIN2-dependent apical endocytosis regulates the morphology of epithelial microvilli. Mol Biol Cell 2019; 30:2515-2526. [PMID: 31390291 PMCID: PMC6743356 DOI: 10.1091/mbc.e19-06-0352] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/27/2022] Open
Abstract
Apical microvilli are critical for the homeostasis of transporting epithelia, yet mechanisms that control the assembly and morphology of these protrusions remain poorly understood. Previous studies in intestinal epithelial cell lines suggested a role for the F-BAR domain protein PACSIN2 in normal microvillar assembly. Here we report the phenotype of PACSIN2 KO mice and provide evidence that through its role in promoting apical endocytosis, this molecule plays a role in controlling microvillar morphology. PACSIN2 KO enterocytes exhibit reduced numbers of microvilli and defects in the microvillar ultrastructure, with membranes lifting away from rootlets of core bundles. Dynamin2, a PACSIN2 binding partner, and other endocytic factors were also lost from their normal localization near microvillar rootlets. To determine whether loss of endocytic machinery could explain defects in microvillar morphology, we examined the impact of PACSIN2 KD and endocytosis inhibition on live intestinal epithelial cells. These assays revealed that when endocytic vesicle scission fails, tubules are pulled into the cytoplasm and this, in turn, leads to a membrane-lifting phenomenon reminiscent of that observed at PACSIN2 KO brush borders. These findings lead to a new model where inward forces generated by endocytic machinery on the plasma membrane control the membrane wrapping of cell surface protrusions.
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Affiliation(s)
- Meagan M Postema
- Department of Cell and Developmental Biology, Vanderbilt University School of Medicine, University Medical Center, Nashville, TN 37232
| | - Nathan E Grega-Larson
- Department of Cell and Developmental Biology, Vanderbilt University School of Medicine, University Medical Center, Nashville, TN 37232
| | - Leslie M Meenderink
- Division of Infectious Diseases, Department of Medicine, Vanderbilt University Medical Center, Nashville, TN 37232
| | - Matthew J Tyska
- Department of Cell and Developmental Biology, Vanderbilt University School of Medicine, University Medical Center, Nashville, TN 37232
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49
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Amaya E, Alarcón L, Martín-Tapia D, Cuellar-Pérez F, Cano-Cortina M, Ortega-Olvera JM, Cisneros B, Rodriguez AJ, Gamba G, González-Mariscal L. Activation of the Ca 2+ sensing receptor and the PKC/WNK4 downstream signaling cascade induces incorporation of ZO-2 to tight junctions and its separation from 14-3-3. Mol Biol Cell 2019; 30:2377-2398. [PMID: 31318316 PMCID: PMC6741067 DOI: 10.1091/mbc.e18-09-0591] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
Zonula occludens-2 (ZO-2) is a tight junction (TJ) cytoplasmic protein, whose localization varies according to cell density and Ca2+ in the media. In cells cultured in low calcium (LC), ZO-2 displays a diffuse cytoplasmic distribution, but activation of the Ca2+ sensing receptor (CaSR) with Gd3+ triggers the appearance of ZO-2 at the cell borders. CaSR downstream signaling involves activation of protein kinase C, which phosphorylates and activates with no lysine kinase-4 that phosphorylates ZO-2 inducing its concentration at TJs. In LC, ZO-2 is protected from degradation by association to 14-3-3 proteins. When monolayers are transferred to normal calcium, the complexes ZO-2/14-3-3ζ and ZO-2/14-3-3σ move to the cell borders and dissociate. The 14-3-3 proteins are then degraded in proteosomes, whereas ZO-2 integrates to TJs. From the plasma membrane residual ZO-2 is endocyted and degradaded in lysosomes. The unique region 2 of ZO-2, and S261 located within a nuclear localization signal, are critical for the interaction with 14-3-3 ζ and σ and for the efficient nuclear importation of ZO-2. These results explain the molecular mechanism through which extracellular Ca2+ triggers the appearance of ZO-2 at TJs in epithelial cells and reveal the novel interaction between ZO-2 and 14-3-3 proteins, which is critical for ZO-2 protection and intracellular traffic.
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Affiliation(s)
- Elida Amaya
- Center for Research and Advanced Studies (Cinvestav), Department of Physiology, Biophysics and Neuroscience, Mexico City 07360, Mexico
| | - Lourdes Alarcón
- Center for Research and Advanced Studies (Cinvestav), Department of Physiology, Biophysics and Neuroscience, Mexico City 07360, Mexico
| | - Dolores Martín-Tapia
- Center for Research and Advanced Studies (Cinvestav), Department of Physiology, Biophysics and Neuroscience, Mexico City 07360, Mexico
| | - Francisco Cuellar-Pérez
- Center for Research and Advanced Studies (Cinvestav), Department of Physiology, Biophysics and Neuroscience, Mexico City 07360, Mexico
| | - Misael Cano-Cortina
- Center for Research and Advanced Studies (Cinvestav), Department of Physiology, Biophysics and Neuroscience, Mexico City 07360, Mexico
| | - Jose Mario Ortega-Olvera
- Center for Research and Advanced Studies (Cinvestav), Department of Physiology, Biophysics and Neuroscience, Mexico City 07360, Mexico
| | - Bulmaro Cisneros
- Department of Genetics and Molecular Biology, Mexico City 07360, Mexico
| | - Alexis J Rodriguez
- Department of Biological Science, Rutgers, The State University of New Jersey, Newark, NJ 07102
| | - Gerardo Gamba
- Molecular Physiology Unit, Instituto de Investigaciones Biomédicas, Universidad Nacional Autónoma de México, Mexico City 14080, México.,Department of Nephrology and Mineral Metabolism, Instituto Nacional de Ciencias Médicas y Nutrición Salvador Zubirán, Mexico City 14080, Mexico.,Tecnológico de Monterrey, Escuela de Medicina y Ciencias de la Salud, 64710 Monterrey, Nuevo Leon, México
| | - Lorenza González-Mariscal
- Center for Research and Advanced Studies (Cinvestav), Department of Physiology, Biophysics and Neuroscience, Mexico City 07360, Mexico
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50
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Radu AG, Torch S, Fauvelle F, Pernet-Gallay K, Lucas A, Blervaque R, Delmas V, Schlattner U, Lafanechère L, Hainaut P, Tricaud N, Pingault V, Bondurand N, Bardeesy N, Larue L, Thibert C, Billaud M. LKB1 specifies neural crest cell fates through pyruvate-alanine cycling. SCIENCE ADVANCES 2019; 5:eaau5106. [PMID: 31328154 PMCID: PMC6636984 DOI: 10.1126/sciadv.aau5106] [Citation(s) in RCA: 16] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 06/25/2018] [Accepted: 06/10/2019] [Indexed: 05/08/2023]
Abstract
Metabolic processes underlying the development of the neural crest, an embryonic population of multipotent migratory cells, are poorly understood. Here, we report that conditional ablation of the Lkb1 tumor suppressor kinase in mouse neural crest stem cells led to intestinal pseudo-obstruction and hind limb paralysis. This phenotype originated from a postnatal degeneration of the enteric nervous ganglia and from a defective differentiation of Schwann cells. Metabolomic profiling revealed that pyruvate-alanine conversion is enhanced in the absence of Lkb1. Mechanistically, inhibition of alanine transaminases restored glial differentiation in an mTOR-dependent manner, while increased alanine level directly inhibited the glial commitment of neural crest cells. Treatment with the metabolic modulator AICAR suppressed mTOR signaling and prevented Schwann cell and enteric defects of Lkb1 mutant mice. These data uncover a link between pyruvate-alanine cycling and the specification of glial cell fate with potential implications in the understanding of the molecular pathogenesis of neural crest diseases.
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Affiliation(s)
- Anca G. Radu
- Institute for Advanced Biosciences, INSERM U1209, CNRS UMR5309, Université Grenoble Alpes, 38000 Grenoble, France
| | - Sakina Torch
- Institute for Advanced Biosciences, INSERM U1209, CNRS UMR5309, Université Grenoble Alpes, 38000 Grenoble, France
| | - Florence Fauvelle
- Univ. Grenoble Alpes, INSERM, U1216, Grenoble Institute of Neurosciences GIN, 38000 Grenoble, France
- Univ. Grenoble Alpes, INSERM, US17, MRI facility IRMaGe, 38000 Grenoble, France
| | - Karin Pernet-Gallay
- Univ. Grenoble Alpes, INSERM, U1216, Grenoble Institute of Neurosciences GIN, 38000 Grenoble, France
| | - Anthony Lucas
- Institute for Advanced Biosciences, INSERM U1209, CNRS UMR5309, Université Grenoble Alpes, 38000 Grenoble, France
| | - Renaud Blervaque
- Institute for Advanced Biosciences, INSERM U1209, CNRS UMR5309, Université Grenoble Alpes, 38000 Grenoble, France
| | - Véronique Delmas
- Institut Curie, Normal and Pathological Development of Melanocytes, CNRS UMR3347; INSERM U1021; Equipe Labellisée–Ligue Nationale Contre le Cancer, Orsay, France
| | - Uwe Schlattner
- Laboratory of Fundamental and Applied Bioenergetics, Univ Grenoble Alpes, 38185 Grenoble, France
- INSERM U1055, 38041 Grenoble France
| | - Laurence Lafanechère
- Institute for Advanced Biosciences, INSERM U1209, CNRS UMR5309, Université Grenoble Alpes, 38000 Grenoble, France
| | - Pierre Hainaut
- Institute for Advanced Biosciences, INSERM U1209, CNRS UMR5309, Université Grenoble Alpes, 38000 Grenoble, France
| | - Nicolas Tricaud
- INSERM U1051, Institut des Neurosciences de Montpellier (INM), Université de Montpellier, Montpellier, France
| | | | | | - Nabeel Bardeesy
- Cancer Center, Massachusetts General Hospital, 185 Cambridge Street, Boston, MA 02114, USA
- Center for Regenerative Medicine, Massachusetts General Hospital, 185 Cambridge Street, Boston, MA 02114, USA
- Department of Medicine, Harvard Medical School, Boston, MA 02114, USA
| | - Lionel Larue
- Institut Curie, Normal and Pathological Development of Melanocytes, CNRS UMR3347; INSERM U1021; Equipe Labellisée–Ligue Nationale Contre le Cancer, Orsay, France
| | - Chantal Thibert
- Institute for Advanced Biosciences, INSERM U1209, CNRS UMR5309, Université Grenoble Alpes, 38000 Grenoble, France
- Corresponding author. (M.B.); (C.T.)
| | - Marc Billaud
- Institute for Advanced Biosciences, INSERM U1209, CNRS UMR5309, Université Grenoble Alpes, 38000 Grenoble, France
- “Clinical and experimental model of lymphomagenesis” Univ Lyon, Université Claude Bernard Lyon1, INSERM 1052, CNRS 5286, Centre Léon Bérard, Centre de recherche en cancérologie de Lyon, Lyon France
- Corresponding author. (M.B.); (C.T.)
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