|
1
|
Tan M, Pan Q, Yu C, Zhai X, Gu J, Tao L, Xu D. PIGT promotes cell growth, glycolysis, and metastasis in bladder cancer by modulating GLUT1 glycosylation and membrane trafficking. J Transl Med 2024; 22:5. [PMID: 38169393 PMCID: PMC10763284 DOI: 10.1186/s12967-023-04805-0] [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: 10/09/2023] [Accepted: 12/12/2023] [Indexed: 01/05/2024] Open
Abstract
BACKGROUND Bladder cancer is very common worldwide. PIGT is a subunit of the glycosylphosphatidylinositol transamidase which involves in tumorigenesis and invasiveness. m6A modification of mRNA has been linked to cell proliferation, tumor progression and other biological events. However, how PIGT is regulated and what is the function of PIGT in bladder cancer remains to be elucidated. METHODS PIGT was silenced or overexpressed to study its role in regulating bladder cancer. Cell proliferation and invasion were examined with the Cell Counting Kit-8, colony formation and Transwell assay, respectively. Cellular oxygen consumption rates or extracellular acidification rates were detected by a XF24 Analyzer. Quantitative RT-PCR and immunoblots were performed to detect mRNA and protein levels. RESULTS PIGT was overexpressed in bladder cancer. Silencing PIGT inhibited cell proliferation, oxidative phosphorylation, and glycolysis. Overexpressing PIGT promoted cell proliferation, oxidative phosphorylation, glycolysis in vitro and tumor metastasis in vivo by activating glucose transporter 1 (GLUT1). PIGT also promoted GLUT1 glycosylation and membrane trafficking. Wilms' tumor 1-associated protein (WTAP) mediated PIGT m6A modification, and m6A reader, insulin-like growth factor 2 mRNA-binding protein (IGF2BP2), binds to the methylated PIGT to promote the stability of PIGT, leading to up-regulation of PIGT. CONCLUSION WTAP mediates PIGT m6A modification to increase the stability of PIGT via the IGF2BP2, which enhances cell proliferation, glycolysis, and metastasis in bladder cancer by modulating GLUT1 glycosylation and membrane trafficking.
Collapse
Affiliation(s)
- Mingyue Tan
- Urology Center, Shuguang Hospital, Shanghai University of Traditional Chinese Medicine, No. 528, Zhangheng Road, Pudong New Area, Shanghai, 201203, China
| | - Qi Pan
- Department of Urology, Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, 200080, China
| | - Chao Yu
- Department of Urology and Andrology, Longhua Hospital Affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai, 200032, China
| | - Xinyu Zhai
- Urology Center, Shuguang Hospital, Shanghai University of Traditional Chinese Medicine, No. 528, Zhangheng Road, Pudong New Area, Shanghai, 201203, China
| | - Jianyi Gu
- Urology Center, Shuguang Hospital, Shanghai University of Traditional Chinese Medicine, No. 528, Zhangheng Road, Pudong New Area, Shanghai, 201203, China
| | - Le Tao
- Department of Urology, Renji Hospital, Shanghai Jiao Tong University School of Medicine, No.160 Pujian Road, Pudong New Area, Shanghai, 200127, China.
| | - Dongliang Xu
- Urology Center, Shuguang Hospital, Shanghai University of Traditional Chinese Medicine, No. 528, Zhangheng Road, Pudong New Area, Shanghai, 201203, China.
| |
Collapse
|
|
2
|
Aldahmash W, Harrath AH, Aljerian K, Sabr Y, Alwasel S. Expression of Glucose Transporters 1 and 3 in the Placenta of Pregnant Women with Gestational Diabetes Mellitus. Life (Basel) 2023; 13:life13040993. [PMID: 37109521 PMCID: PMC10143906 DOI: 10.3390/life13040993] [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: 03/06/2023] [Revised: 03/29/2023] [Accepted: 04/06/2023] [Indexed: 04/29/2023] Open
Abstract
BACKGROUND The annual prevalence of gestational diabetes mellitus-characterized by an increase in blood glucose in pregnant women-has been increasing worldwide. The goal of this study was to evaluate the expression of glucose transporter 1 (GLUT1) and glucose transporter 3 (GLUT3) in the placenta of women with gestational diabetes mellitus. METHODS Sixty-five placentas from women admitted to the King Saud University Medical City, Riyadh, Saudi Arabia, were analyzed; 34 and 31 placentas were from healthy pregnant women and women with gestational diabetes, respectively. The expressions of GLUT1 and GLUT3 were assessed using RT-PCR, Western blotting, and immunohistochemical methods. The degree of apoptosis in the placental villi was estimated via a TUNEL assay. RESULTS The results of the protein expression assays and immunohistochemical staining showed that the levels of GLUT1 and GLUT3 were significantly higher in the placentas of pregnant women with gestational diabetes than those in the placentas of healthy pregnant women. In addition, the findings showed an increase in apoptosis in the placenta of pregnant women with gestational diabetes compared to that in the placenta of healthy pregnant women. However, the results of gene expression assays showed no significant difference between the two groups. CONCLUSIONS Based on these results, we conclude that gestational diabetes mellitus leads to an increased incidence of apoptosis in the placental villi and alters the level of GLUT1 and GLUT3 protein expressions in the placenta of women with gestational diabetes. Understanding the conditions in which the fetus develops in the womb of a pregnant woman with gestational diabetes may help researchers understand the underlying causes of the development of chronic diseases later in life.
Collapse
Affiliation(s)
- Waleed Aldahmash
- Zoology Department, College of Science, King Saud University, P.O. Box 2455, Riyadh 11451, Saudi Arabia
| | - Abdel Halim Harrath
- Zoology Department, College of Science, King Saud University, P.O. Box 2455, Riyadh 11451, Saudi Arabia
| | - Khaldoon Aljerian
- Pathology Department, College of Medicine, King Saud University, Riyadh 11451, Saudi Arabia
| | - Yasser Sabr
- Obstetrics and Gynaecology Department, College of Medicine, King Saud University, Riyadh 11451, Saudi Arabia
| | - Saleh Alwasel
- Zoology Department, College of Science, King Saud University, P.O. Box 2455, Riyadh 11451, Saudi Arabia
| |
Collapse
|
|
3
|
Feng R, Yin Y, Wei Y, Li Y, Li L, Zhu R, Yu X, Liu Y, Zhao Y, Liu Z. Mutant p53 activates hnRNPA2B1-AGAP1-mediated exosome formation to promote esophageal squamous cell carcinoma progression. Cancer Lett 2023; 562:216154. [PMID: 37030635 DOI: 10.1016/j.canlet.2023.216154] [Citation(s) in RCA: 19] [Impact Index Per Article: 9.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/04/2023] [Revised: 03/28/2023] [Accepted: 03/28/2023] [Indexed: 04/10/2023]
Abstract
p53 mutations predispose cancer cell development, promote their survival and metastasis, and lead to ineffective therapeutic responses and unfavorable prognosis. No drug that abrogates the oncogenic functions of mutant p53 has been approved for cancer treatment. Here, we performed whole-genome sequencing of 663 esophageal squamous cell carcinoma (ESCC) tumor tissues and paired normal tissues. The results indicated that ESCC samples from our cohort had a more dispersed distribution of TP53 mutants and a higher proportion of nonsense mutants than European and American ESCC samples in the International Agency for Research on Cancer (IARC) database. The most frequent p53 mutations disrupt the inhibition of proliferation, migration, and invasion mediated by wild-type p53 in ESCC. Furthermore, p53 mutations alter its protein nucleoplasmic localization and protein stability. The p53 mutation G245S (p53-G245S) interacts with heterogeneous nuclear ribonucleoprotein A2B1 (hnRNPA2B1) to increase protein translation of phosphatidylinositol-dependent Arf GAP (AGAP1) by promoting AGAP1 mRNA stability. AGAP1 promotes cancer cell proliferation and metastasis by enhancing exosome formation. Furthermore, we explored the combination of the HSP90 inhibitor HSP90i and the AGAP1 inhibitor QS11 could inhibit ESCC cell proliferation and metastasis. Thus, the p53-G245S/hnRNPA2B1/AGAP1 axis promotes ESCC progression by enhancing exosome formation, and the combination of an HSP90 inhibitor and an AGAP1 inhibitor may serve as a potential therapeutic strategy.
Collapse
Affiliation(s)
- Riyue Feng
- State Key Laboratory of Molecular Oncology, National Cancer Center, National Clinical Research Center for Cancer, Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, 100021, China
| | - Yin Yin
- State Key Laboratory of Molecular Oncology, National Cancer Center, National Clinical Research Center for Cancer, Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, 100021, China
| | - Yuge Wei
- State Key Laboratory of Molecular Oncology, National Cancer Center, National Clinical Research Center for Cancer, Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, 100021, China
| | - Yang Li
- State Key Laboratory of Molecular Oncology, National Cancer Center, National Clinical Research Center for Cancer, Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, 100021, China
| | - Lei Li
- State Key Laboratory of Molecular Oncology, National Cancer Center, National Clinical Research Center for Cancer, Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, 100021, China
| | - Rui Zhu
- State Key Laboratory of Molecular Oncology, National Cancer Center, National Clinical Research Center for Cancer, Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, 100021, China
| | - Xiao Yu
- State Key Laboratory of Molecular Oncology, National Cancer Center, National Clinical Research Center for Cancer, Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, 100021, China
| | - Yuhao Liu
- State Key Laboratory of Molecular Oncology, National Cancer Center, National Clinical Research Center for Cancer, Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, 100021, China
| | - Yahui Zhao
- State Key Laboratory of Molecular Oncology, National Cancer Center, National Clinical Research Center for Cancer, Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, 100021, China.
| | - Zhihua Liu
- State Key Laboratory of Molecular Oncology, National Cancer Center, National Clinical Research Center for Cancer, Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, 100021, China.
| |
Collapse
|
|
4
|
Li DD, Jawale CV, Zhou C, Lin L, Trevejo-Nunez GJ, Rahman SA, Mullet SJ, Das J, Wendell SG, Delgoffe GM, Lionakis MS, Gaffen SL, Biswas PS. Fungal sensing enhances neutrophil metabolic fitness by regulating antifungal Glut1 activity. Cell Host Microbe 2022; 30:530-544.e6. [PMID: 35316647 PMCID: PMC9026661 DOI: 10.1016/j.chom.2022.02.017] [Citation(s) in RCA: 31] [Impact Index Per Article: 10.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/23/2021] [Revised: 12/10/2021] [Accepted: 02/16/2022] [Indexed: 12/30/2022]
Abstract
Combating fungal pathogens poses metabolic challenges for neutrophils, key innate cells in anti-Candida albicans immunity, yet how host-pathogen interactions cause remodeling of the neutrophil metabolism is unclear. We show that neutrophils mediate renal immunity to disseminated candidiasis by upregulating glucose uptake via selective expression of glucose transporter 1 (Glut1). Mechanistically, dectin-1-mediated recognition of β-glucan leads to activation of PKCδ, which triggers phosphorylation, localization, and early glucose transport by a pool of pre-formed Glut1 in neutrophils. These events are followed by increased Glut1 gene transcription, leading to more sustained Glut1 accumulation, which is also dependent on the β-glucan/dectin-1/CARD9 axis. Card9-deficient neutrophils show diminished glucose incorporation in candidiasis. Neutrophil-specific Glut1-ablated mice exhibit increased mortality in candidiasis caused by compromised neutrophil phagocytosis, reactive oxygen species (ROS), and neutrophil extracellular trap (NET) formation. In human neutrophils, β-glucan triggers metabolic remodeling and enhances candidacidal function. Our data show that the host-pathogen interface increases glycolytic activity in neutrophils by regulating Glut1 expression, localization, and function.
Collapse
Affiliation(s)
- De-Dong Li
- Division of Rheumatology & Clinical Immunology, Department of Medicine, University of Pittsburgh, Pittsburgh, PA, USA
| | - Chetan V Jawale
- Division of Rheumatology & Clinical Immunology, Department of Medicine, University of Pittsburgh, Pittsburgh, PA, USA
| | - Chunsheng Zhou
- Division of Rheumatology & Clinical Immunology, Department of Medicine, University of Pittsburgh, Pittsburgh, PA, USA
| | - Li Lin
- Division of Rheumatology & Clinical Immunology, Department of Medicine, University of Pittsburgh, Pittsburgh, PA, USA
| | - Giraldina J Trevejo-Nunez
- Division of Infectious Diseases, Department of Medicine, University of Pittsburgh, Pittsburgh, PA, USA
| | - Syed A Rahman
- Center for Systems Immunology, Departments of Immunology and Computational & Systems Biology, University of Pittsburgh, Pittsburgh, PA, USA
| | - Steven J Mullet
- Department of Immunology, University of Pittsburgh, Pittsburgh, PA, USA
| | - Jishnu Das
- Center for Systems Immunology, Departments of Immunology and Computational & Systems Biology, University of Pittsburgh, Pittsburgh, PA, USA
| | - Stacy G Wendell
- Department of Immunology, University of Pittsburgh, Pittsburgh, PA, USA
| | - Greg M Delgoffe
- Department of Pharmacology & Chemical Biology, University of Pittsburgh, Pittsburgh, PA, USA
| | - Michail S Lionakis
- Fungal Pathogenesis Section, National Institute of Allergy and Infectious Diseases, Bethesda, MD, USA
| | - Sarah L Gaffen
- Division of Rheumatology & Clinical Immunology, Department of Medicine, University of Pittsburgh, Pittsburgh, PA, USA
| | - Partha S Biswas
- Division of Rheumatology & Clinical Immunology, Department of Medicine, University of Pittsburgh, Pittsburgh, PA, USA.
| |
Collapse
|
|
5
|
Thibault PA, Ganesan A, Kalyaanamoorthy S, Clarke JPWE, Salapa HE, Levin MC. hnRNP A/B Proteins: An Encyclopedic Assessment of Their Roles in Homeostasis and Disease. BIOLOGY 2021; 10:biology10080712. [PMID: 34439945 PMCID: PMC8389229 DOI: 10.3390/biology10080712] [Citation(s) in RCA: 16] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 06/15/2021] [Revised: 07/16/2021] [Accepted: 07/21/2021] [Indexed: 12/13/2022]
Abstract
The hnRNP A/B family of proteins is canonically central to cellular RNA metabolism, but due to their highly conserved nature, the functional differences between hnRNP A1, A2/B1, A0, and A3 are often overlooked. In this review, we explore and identify the shared and disparate homeostatic and disease-related functions of the hnRNP A/B family proteins, highlighting areas where the proteins have not been clearly differentiated. Herein, we provide a comprehensive assembly of the literature on these proteins. We find that there are critical gaps in our grasp of A/B proteins' alternative splice isoforms, structures, regulation, and tissue and cell-type-specific functions, and propose that future mechanistic research integrating multiple A/B proteins will significantly improve our understanding of how this essential protein family contributes to cell homeostasis and disease.
Collapse
Affiliation(s)
- Patricia A. Thibault
- Office of the Saskatchewan Multiple Sclerosis Clinical Research Chair, University of Saskatchewan, Saskatoon, SK S7K 0M7, Canada; (P.A.T.); (J.-P.W.E.C.); (H.E.S.)
- Department of Medicine, Neurology Division, University of Saskatchewan, Saskatoon, SK S7N 0X8, Canada
| | - Aravindhan Ganesan
- ArGan’s Lab, School of Pharmacy, Faculty of Science, University of Waterloo, Waterloo, ON N2L 3G1, Canada;
| | - Subha Kalyaanamoorthy
- Department of Chemistry, Faculty of Science, University of Waterloo, Waterloo, ON N2L 3G1, Canada;
| | - Joseph-Patrick W. E. Clarke
- Office of the Saskatchewan Multiple Sclerosis Clinical Research Chair, University of Saskatchewan, Saskatoon, SK S7K 0M7, Canada; (P.A.T.); (J.-P.W.E.C.); (H.E.S.)
- Department of Health Sciences, College of Medicine, University of Saskatchewan, Saskatoon, SK S7N 5E5, Canada
- Department of Anatomy, Physiology and Pharmacology, University of Saskatchewan, Saskatoon, SK S7N 5E5, Canada
| | - Hannah E. Salapa
- Office of the Saskatchewan Multiple Sclerosis Clinical Research Chair, University of Saskatchewan, Saskatoon, SK S7K 0M7, Canada; (P.A.T.); (J.-P.W.E.C.); (H.E.S.)
- Department of Medicine, Neurology Division, University of Saskatchewan, Saskatoon, SK S7N 0X8, Canada
| | - Michael C. Levin
- Office of the Saskatchewan Multiple Sclerosis Clinical Research Chair, University of Saskatchewan, Saskatoon, SK S7K 0M7, Canada; (P.A.T.); (J.-P.W.E.C.); (H.E.S.)
- Department of Medicine, Neurology Division, University of Saskatchewan, Saskatoon, SK S7N 0X8, Canada
- Department of Health Sciences, College of Medicine, University of Saskatchewan, Saskatoon, SK S7N 5E5, Canada
- Department of Anatomy, Physiology and Pharmacology, University of Saskatchewan, Saskatoon, SK S7N 5E5, Canada
- Correspondence:
| |
Collapse
|
|
6
|
Jiang F, Tang X, Tang C, Hua Z, Ke M, Wang C, Zhao J, Gao S, Jurczyszyn A, Janz S, Beksac M, Zhan F, Gu C, Yang Y. HNRNPA2B1 promotes multiple myeloma progression by increasing AKT3 expression via m6A-dependent stabilization of ILF3 mRNA. J Hematol Oncol 2021; 14:54. [PMID: 33794982 PMCID: PMC8017865 DOI: 10.1186/s13045-021-01066-6] [Citation(s) in RCA: 106] [Impact Index Per Article: 26.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/24/2020] [Accepted: 03/23/2021] [Indexed: 12/20/2022] Open
Abstract
N6-methyladenosine (m6A) modification is the most prevalent modification in eukaryotic RNAs while accumulating studies suggest that m6A aberrant expression plays an important role in cancer. HNRNPA2B1 is a m6A reader which binds to nascent RNA and thus affects a perplexing array of RNA metabolism exquisitely. Despite unveiled facets that HNRNPA2B1 is deregulated in several tumors and facilitates tumor growth, a clear role of HNRNPA2B1 in multiple myeloma (MM) remains elusive. Herein, we analyzed the function and the regulatory mechanism of HNRNPA2B1 in MM. We found that HNRNPA2B1 was elevated in MM patients and negatively correlated with favorable prognosis. The depletion of HNRNPA2B1 in MM cells inhibited cell proliferation and induced apoptosis. On the contrary, the overexpression of HNRNPA2B1 promoted cell proliferation in vitro and in vivo. Mechanistic studies revealed that HNRNPA2B1 recognized the m6A sites of ILF3 and enhanced the stability of ILF3 mRNA transcripts, while AKT3 downregulation by siRNA abrogated the cellular proliferation induced by HNRNPA2B1 overexpression. Additionally, the expression of HNRNPA2B1, ILF3 and AKT3 was positively associated with each other in MM tissues tested by immunohistochemistry. In summary, our study highlights that HNRNPA2B1 potentially acts as a therapeutic target of MM through regulating AKT3 expression mediated by ILF3-dependent pattern.
Collapse
Affiliation(s)
- Fengjie Jiang
- The Third Affiliated Hospital of Nanjing University of Chinese Medicine, Nanjing, 210001, China.,School of Medicine and Holistic Integrative Medicine, Nanjing University of Chinese Medicine, 138 Xianlin Road, Nanjing, 210023, China
| | - Xiaozhu Tang
- School of Medicine and Holistic Integrative Medicine, Nanjing University of Chinese Medicine, 138 Xianlin Road, Nanjing, 210023, China
| | - Chao Tang
- School of Medicine and Holistic Integrative Medicine, Nanjing University of Chinese Medicine, 138 Xianlin Road, Nanjing, 210023, China
| | - Zhen Hua
- School of Medicine and Holistic Integrative Medicine, Nanjing University of Chinese Medicine, 138 Xianlin Road, Nanjing, 210023, China
| | - Mengying Ke
- School of Medicine and Holistic Integrative Medicine, Nanjing University of Chinese Medicine, 138 Xianlin Road, Nanjing, 210023, China
| | - Chen Wang
- School of Medicine and Holistic Integrative Medicine, Nanjing University of Chinese Medicine, 138 Xianlin Road, Nanjing, 210023, China
| | - Jiamin Zhao
- School of Medicine and Holistic Integrative Medicine, Nanjing University of Chinese Medicine, 138 Xianlin Road, Nanjing, 210023, China
| | - Shengyao Gao
- School of Medicine and Holistic Integrative Medicine, Nanjing University of Chinese Medicine, 138 Xianlin Road, Nanjing, 210023, China
| | - Artur Jurczyszyn
- Department of Hematology, Jagiellonian University Medical College, Cracow, Poland
| | - Siegfried Janz
- Division of Hematology and Oncology, Medical College of Wisconsin, Milwaukee, USA
| | - Meral Beksac
- Department of Hematology, School of Medicine, Ankara University, Ankara, Turkey
| | - Fenghuang Zhan
- Internal Medicine, University of Iowa, Iowa City, USA.,Myeloma Center, University of Arkansas, Little Rock, USA
| | - Chunyan Gu
- The Third Affiliated Hospital of Nanjing University of Chinese Medicine, Nanjing, 210001, China. .,School of Medicine and Holistic Integrative Medicine, Nanjing University of Chinese Medicine, 138 Xianlin Road, Nanjing, 210023, China.
| | - Ye Yang
- School of Medicine and Holistic Integrative Medicine, Nanjing University of Chinese Medicine, 138 Xianlin Road, Nanjing, 210023, China. .,Internal Medicine, University of Iowa, Iowa City, USA.
| |
Collapse
|
|
7
|
Pifferi F, Cunnane SC, Guesnet P. Evidence of the Role of Omega-3 Polyunsaturated Fatty Acids in Brain Glucose Metabolism. Nutrients 2020; 12:nu12051382. [PMID: 32408634 PMCID: PMC7285025 DOI: 10.3390/nu12051382] [Citation(s) in RCA: 11] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/14/2020] [Revised: 05/11/2020] [Accepted: 05/11/2020] [Indexed: 11/30/2022] Open
Abstract
In mammals, brain function, particularly neuronal activity, has high energy needs. When glucose is supplemented by alternative oxidative substrates under different physiological conditions, these fuels do not fully replace the functions fulfilled by glucose. Thus, it is of major importance that the brain is almost continuously supplied with glucose from the circulation. Numerous studies describe the decrease in brain glucose metabolism during healthy or pathological ageing, but little is known about the mechanisms that cause such impairment. Although it appears difficult to determine the exact role of brain glucose hypometabolism during healthy ageing or during age-related neurodegenerative diseases such as Alzheimer’s disease, uninterrupted glucose supply to the brain is still of major importance for proper brain function. Interestingly, a body of evidence suggests that dietary n-3 polyunsaturated fatty acids (PUFAs) might play significant roles in brain glucose regulation. Thus, the goal of the present review is to summarize this evidence and address the role of n-3 PUFAs in brain energy metabolism. Taken together, these data suggest that ensuring an adequate dietary supply of n-3 PUFAs could constitute an essential aspect of a promising strategy to promote optimal brain function during both healthy and pathological ageing.
Collapse
Affiliation(s)
- Fabien Pifferi
- Unité Mixte de Recherche (UMR), Centre Nationnal de la Recherche Scientifique (CNRS), Museum National d’Histoire Naturelle (MNHN) 7179, Mécanismes Adaptatifs et Evolution (MECADEV), 1 Avenue du Petit Château, 91800 Brunoy, France
- Correspondence:
| | - Stephen C. Cunnane
- Department of Medicine, Université de Sherbrooke, Sherbrooke, QC J1H 5N4, Canada;
- Research Center on Aging, Sherbrooke, QC J1H 4C4, Canada
- Department of Pharmacology and Physiology, Université de Sherbrooke, Sherbrooke, QC J1H 5N4, Canada
| | | |
Collapse
|
|
8
|
Yuan J, Peng G, Xiao G, Yang Z, Huang J, Liu Q, Yang Z, Liu D. Xanthohumol suppresses glioblastoma via modulation of Hexokinase 2 -mediated glycolysis. J Cancer 2020; 11:4047-4058. [PMID: 32368287 PMCID: PMC7196271 DOI: 10.7150/jca.33045] [Citation(s) in RCA: 19] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/11/2019] [Accepted: 10/26/2019] [Indexed: 12/16/2022] Open
Abstract
Deregulation of aerobic glycolysis is a common phenomenon in human cancers, including glioblastoma (GBM). In the present study, we demonstrated that the natural compound xanthohumol has a profound anti-tumor effect on GBM via direct inhibition of glycolysis. Xanthohumol suppressed cell proliferation and colony formation of GBM cells, and significantly impaired glucose metabolism via inhibiting Hexokinase 2 (HK2) expression. We demonstrated that down-regulation of c-Myc was required for xanthohumol-induced decrease of HK2. Xanthohumol destabilization of c-Myc, and promoted FBW7-mediated ubiquitination of c-Myc. Xanthohumol attenuated Akt activity and inhibited the activation of GSK3β, resulted in c-Myc degradation. Overexpression of Myr-Akt1 significantly rescued xanthohumol-mediated c-Myc inhibition and glycolysis suppression. Finally, the xanthohumol-mediated down-regulation of the PI3-K/Akt-GSK3beta-FBW7 signaling axis promoted the destabilization of c-Myc. Finally, the animal results demonstrated that xanthohumol substantially inhibited tumor growth in vivo. Collectively, xanthohumol appears to be a promising new anti-tumor agent with the therapeutic potential for GBM.
Collapse
Affiliation(s)
- Jian Yuan
- Department of Neurosurgery, Xiangya Hospital, Central South University, 87 Xiangya Road, Changsha, Hunan 410008, China
- The Institute of Skull Base Surgery and Neuro-oncology at Hunan, 87 Xiangya Road, Changsha, Hunan, 410008, China
| | - Gang Peng
- Department of Neurosurgery, Xiangya Hospital, Central South University, 87 Xiangya Road, Changsha, Hunan 410008, China
- The Institute of Skull Base Surgery and Neuro-oncology at Hunan, 87 Xiangya Road, Changsha, Hunan, 410008, China
| | - Gelei Xiao
- Department of Neurosurgery, Xiangya Hospital, Central South University, 87 Xiangya Road, Changsha, Hunan 410008, China
| | - Zhuanyi Yang
- Department of Neurosurgery, Xiangya Hospital, Central South University, 87 Xiangya Road, Changsha, Hunan 410008, China
| | - Jun Huang
- Department of Neurosurgery, Xiangya Hospital, Central South University, 87 Xiangya Road, Changsha, Hunan 410008, China
| | - Qing Liu
- Department of Neurosurgery, Xiangya Hospital, Central South University, 87 Xiangya Road, Changsha, Hunan 410008, China
- The Institute of Skull Base Surgery and Neuro-oncology at Hunan, 87 Xiangya Road, Changsha, Hunan, 410008, China
| | - Zhiquan Yang
- Department of Neurosurgery, Xiangya Hospital, Central South University, 87 Xiangya Road, Changsha, Hunan 410008, China
| | - Dingyang Liu
- Department of Neurosurgery, Xiangya Hospital, Central South University, 87 Xiangya Road, Changsha, Hunan 410008, China
- The Institute of Skull Base Surgery and Neuro-oncology at Hunan, 87 Xiangya Road, Changsha, Hunan, 410008, China
| |
Collapse
|
|
9
|
Masterson JC, Biette KA, Hammer JA, Nguyen N, Capocelli KE, Saeedi BJ, Harris RF, Fernando SD, Hosford LB, Kelly CJ, Campbell EL, Ehrentraut SF, Ahmed FN, Nakagawa H, Lee JJ, McNamee EN, Glover LE, Colgan SP, Furuta GT. Epithelial HIF-1α/claudin-1 axis regulates barrier dysfunction in eosinophilic esophagitis. J Clin Invest 2019; 129:3224-3235. [PMID: 31264974 PMCID: PMC6668670 DOI: 10.1172/jci126744] [Citation(s) in RCA: 50] [Impact Index Per Article: 8.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/29/2019] [Accepted: 05/16/2019] [Indexed: 12/19/2022] Open
Abstract
Epithelial barrier dysfunction is a significant factor in many allergic diseases, including eosinophilic esophagitis (EoE). Infiltrating leukocytes and tissue adaptations increase metabolic demands and decrease oxygen availability at barrier surfaces. Understanding of how these processes impact barrier is limited, particularly in allergy. Here, we identified a regulatory axis whereby the oxygen-sensing transcription factor HIF-1α orchestrated epithelial barrier integrity, selectively controlling tight junction CLDN1 (claudin-1). Prolonged experimental hypoxia or HIF1A knockdown suppressed HIF-1α-dependent claudin-1 expression and epithelial barrier function, as documented in 3D organotypic epithelial cultures. L2-IL5OXA mice with EoE-relevant allergic inflammation displayed localized eosinophil oxygen metabolism, tissue hypoxia, and impaired claudin-1 barrier via repression of HIF-1α/claudin-1 signaling, which was restored by transgenic expression of esophageal epithelial-targeted stabilized HIF-1α. EoE patient biopsy analysis identified a repressed HIF-1α/claudin-1 axis, which was restored via pharmacologic HIF-1α stabilization ex vivo. Collectively, these studies reveal HIF-1α's critical role in maintaining barrier and highlight the HIF-1α/claudin-1 axis as a potential therapeutic target for EoE.
Collapse
Affiliation(s)
- Joanne C. Masterson
- Allergy, Inflammation and Remodeling Research Laboratory, Human Health Research Institute, Department of Biology, Maynooth University, Maynooth, Co. Kildare, Ireland
- Gastrointestinal Eosinophilic Diseases Program, Department of Pediatrics, University of Colorado School of Medicine; Digestive Health Institute, Children’s Hospital Colorado; Aurora, Colorado, USA
- Mucosal Inflammation Program, Department of Medicine, University of Colorado, Aurora, Colorado, USA
| | - Kathryn A. Biette
- Gastrointestinal Eosinophilic Diseases Program, Department of Pediatrics, University of Colorado School of Medicine; Digestive Health Institute, Children’s Hospital Colorado; Aurora, Colorado, USA
- Mucosal Inflammation Program, Department of Medicine, University of Colorado, Aurora, Colorado, USA
| | - Juliet A. Hammer
- Gastrointestinal Eosinophilic Diseases Program, Department of Pediatrics, University of Colorado School of Medicine; Digestive Health Institute, Children’s Hospital Colorado; Aurora, Colorado, USA
- Mucosal Inflammation Program, Department of Medicine, University of Colorado, Aurora, Colorado, USA
| | - Nathalie Nguyen
- Gastrointestinal Eosinophilic Diseases Program, Department of Pediatrics, University of Colorado School of Medicine; Digestive Health Institute, Children’s Hospital Colorado; Aurora, Colorado, USA
- Mucosal Inflammation Program, Department of Medicine, University of Colorado, Aurora, Colorado, USA
| | - Kelley E. Capocelli
- Department of Pathology, Children’s Hospital Colorado, Aurora, Colorado, USA
| | - Bejan J. Saeedi
- Mucosal Inflammation Program, Department of Medicine, University of Colorado, Aurora, Colorado, USA
| | - Rachel F. Harris
- Gastrointestinal Eosinophilic Diseases Program, Department of Pediatrics, University of Colorado School of Medicine; Digestive Health Institute, Children’s Hospital Colorado; Aurora, Colorado, USA
- Mucosal Inflammation Program, Department of Medicine, University of Colorado, Aurora, Colorado, USA
| | - Shahan D. Fernando
- Gastrointestinal Eosinophilic Diseases Program, Department of Pediatrics, University of Colorado School of Medicine; Digestive Health Institute, Children’s Hospital Colorado; Aurora, Colorado, USA
- Mucosal Inflammation Program, Department of Medicine, University of Colorado, Aurora, Colorado, USA
| | - Lindsay B. Hosford
- Gastrointestinal Eosinophilic Diseases Program, Department of Pediatrics, University of Colorado School of Medicine; Digestive Health Institute, Children’s Hospital Colorado; Aurora, Colorado, USA
- Mucosal Inflammation Program, Department of Medicine, University of Colorado, Aurora, Colorado, USA
| | - Caleb J. Kelly
- Mucosal Inflammation Program, Department of Medicine, University of Colorado, Aurora, Colorado, USA
| | - Eric L. Campbell
- Mucosal Inflammation Program, Department of Medicine, University of Colorado, Aurora, Colorado, USA
| | - Stefan F. Ehrentraut
- Mucosal Inflammation Program, Department of Medicine, University of Colorado, Aurora, Colorado, USA
| | - Faria N. Ahmed
- Gastrointestinal Eosinophilic Diseases Program, Department of Pediatrics, University of Colorado School of Medicine; Digestive Health Institute, Children’s Hospital Colorado; Aurora, Colorado, USA
| | - Hiroshi Nakagawa
- Division of Gastroenterology, Department of Medicine, University of Pennsylvania Perelman School of Medicine, Philadelphia, Pennsylvania, USA
| | | | - Eóin N. McNamee
- Allergy, Inflammation and Remodeling Research Laboratory, Human Health Research Institute, Department of Biology, Maynooth University, Maynooth, Co. Kildare, Ireland
- Mucosal Inflammation Program, Department of Medicine, University of Colorado, Aurora, Colorado, USA
| | - Louise E. Glover
- Mucosal Inflammation Program, Department of Medicine, University of Colorado, Aurora, Colorado, USA
| | - Sean P. Colgan
- Mucosal Inflammation Program, Department of Medicine, University of Colorado, Aurora, Colorado, USA
| | - Glenn T. Furuta
- Gastrointestinal Eosinophilic Diseases Program, Department of Pediatrics, University of Colorado School of Medicine; Digestive Health Institute, Children’s Hospital Colorado; Aurora, Colorado, USA
- Mucosal Inflammation Program, Department of Medicine, University of Colorado, Aurora, Colorado, USA
| |
Collapse
|
|
10
|
Mechanism of the natural product moracin-O derived MO-460 and its targeting protein hnRNPA2B1 on HIF-1α inhibition. Exp Mol Med 2019; 51:1-14. [PMID: 30755586 PMCID: PMC6372683 DOI: 10.1038/s12276-018-0200-4] [Citation(s) in RCA: 24] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/09/2018] [Revised: 10/12/2018] [Accepted: 10/16/2018] [Indexed: 02/06/2023] Open
Abstract
Hypoxia-inducible factor-1α (HIF-1α) mediates tumor cell adaptation to hypoxic conditions and is a potentially important anticancer therapeutic target. We previously developed a method for synthesizing a benzofuran-based natural product, (R)-(-)-moracin-O, and obtained a novel potent analog, MO-460 that suppresses the accumulation of HIF-1α in Hep3B cells. However, the molecular target and underlying mechanism of action of MO-460 remained unclear. In the current study, we identified heterogeneous nuclear ribonucleoprotein A2B1 (hnRNPA2B1) as a molecular target of MO-460. MO-460 inhibits the initiation of HIF-1α translation by binding to the C-terminal glycine-rich domain of hnRNPA2B1 and inhibiting its subsequent binding to the 3’-untranslated region of HIF-1α mRNA. Moreover, MO-460 suppresses HIF-1α protein synthesis under hypoxic conditions and induces the accumulation of stress granules. The data provided here suggest that hnRNPA2B1 serves as a crucial molecular target in hypoxia-induced tumor survival and thus offer an avenue for the development of novel anticancer therapies. A synthetic analog of a chemical found in fruit suppresses tumor growth by targeting an RNA-binding protein (hnRNPA2B1) and preventing the production of a pro-cancer regulatory factor. Nak-Kyun Soung from the Korea Research Institute of Bioscience and Biotechnology, Cheongju, South Korea, and coworkers built on their previous discovery that a compound derived from a medicinal plant metabolite can suppress the activity of hypoxia-inducible factor-1α (HIF-1α). This protein, which is involved in many aspects of cancer biology, is activated in the low-oxygen microenvironments found inside tumors. The researchers show that the compound binds to a protein that helps with the conversion of HIF-1α–encoding RNA transcripts into HIF-1α proteins. Liver cancer cells treated with the compound grew slowly and produced less HIF-1α under both normal and low-oxygen culture conditions, highlighting the potential of this anti-cancer strategy.
Collapse
|
|
11
|
Brandi J, Cecconi D, Cordani M, Torrens-Mas M, Pacchiana R, Dalla Pozza E, Butera G, Manfredi M, Marengo E, Oliver J, Roca P, Dando I, Donadelli M. The antioxidant uncoupling protein 2 stimulates hnRNPA2/B1, GLUT1 and PKM2 expression and sensitizes pancreas cancer cells to glycolysis inhibition. Free Radic Biol Med 2016; 101:305-316. [PMID: 27989750 DOI: 10.1016/j.freeradbiomed.2016.10.499] [Citation(s) in RCA: 52] [Impact Index Per Article: 5.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/11/2016] [Revised: 10/12/2016] [Accepted: 10/24/2016] [Indexed: 01/09/2023]
Abstract
Several evidence indicate that metabolic alterations play a pivotal role in cancer development. Here, we report that the mitochondrial uncoupling protein 2 (UCP2) sustains the metabolic shift from mitochondrial oxidative phosphorylation (mtOXPHOS) to glycolysis in pancreas cancer cells. Indeed, we show that UCP2 sensitizes pancreas cancer cells to the treatment with the glycolytic inhibitor 2-deoxy-D-glucose. Through a bidimensional electrophoresis analysis, we identify 19 protein species differentially expressed after treatment with the UCP2 inhibitor genipin and, by bioinformatic analyses, we show that these proteins are mainly involved in metabolic processes. In particular, we demonstrate that the antioxidant UCP2 induces the expression of hnRNPA2/B1, which is involved in the regulation of both GLUT1 and PKM2 mRNAs, and of lactate dehydrogenase (LDH) increasing the secretion of L-lactic acid. We further demonstrate that the radical scavenger N-acetyl-L-cysteine reverts hnRNPA2/B1 and PKM2 inhibition by genipin indicating a role for reactive oxygen species in the metabolic reprogramming of cancer cells mediated by UCP2. We also observe an UCP2-dependent decrease in mtOXPHOS complex I (NADH dehydrogenase), complex IV (cytochrome c oxidase), complex V (ATPase) and in mitochondrial oxygen consumption, suggesting a role for UCP2 in the counteraction of pancreatic cancer cellular respiration. All these results reveal novel mechanisms through which UCP2 promotes cancer cell proliferation with the concomitant metabolic shift from mtOXPHOS to the glycolytic pathway.
Collapse
Affiliation(s)
- Jessica Brandi
- Department of Biotechnology, Proteomics and Mass Spectrometry Laboratory, University of Verona, Verona, Italy
| | - Daniela Cecconi
- Department of Biotechnology, Proteomics and Mass Spectrometry Laboratory, University of Verona, Verona, Italy
| | - Marco Cordani
- Department of Neuroscience, Biomedicine and Movement, Biochemistry Section, University of Verona, Verona, Italy
| | - Margalida Torrens-Mas
- Physiopathology of Obesity and Nutrition, CIBERobn (CB06/03), Carlos III Health Research Institute (ISCIII), Madrid, Spain; Palma Institute for Health Research (IdISPa), E07010 Palma, Spain; Multidisciplinar Group of Translational Oncology, University Research Institute on Health Sciences (IUNICS), University of the Balearic Islands, E07122 Palma, Spain
| | - Raffaella Pacchiana
- Department of Neuroscience, Biomedicine and Movement, Biochemistry Section, University of Verona, Verona, Italy
| | - Elisa Dalla Pozza
- Department of Neuroscience, Biomedicine and Movement, Biochemistry Section, University of Verona, Verona, Italy
| | - Giovanna Butera
- Department of Neuroscience, Biomedicine and Movement, Biochemistry Section, University of Verona, Verona, Italy
| | - Marcello Manfredi
- Department of Sciences and Technological Innovation, University of Eastern Piedmont, Alessandria, Italy; ISALIT, Novara, Italy
| | - Emilio Marengo
- Department of Sciences and Technological Innovation, University of Eastern Piedmont, Alessandria, Italy
| | - Jordi Oliver
- Physiopathology of Obesity and Nutrition, CIBERobn (CB06/03), Carlos III Health Research Institute (ISCIII), Madrid, Spain; Palma Institute for Health Research (IdISPa), E07010 Palma, Spain; Multidisciplinar Group of Translational Oncology, University Research Institute on Health Sciences (IUNICS), University of the Balearic Islands, E07122 Palma, Spain
| | - Pilar Roca
- Physiopathology of Obesity and Nutrition, CIBERobn (CB06/03), Carlos III Health Research Institute (ISCIII), Madrid, Spain; Palma Institute for Health Research (IdISPa), E07010 Palma, Spain; Multidisciplinar Group of Translational Oncology, University Research Institute on Health Sciences (IUNICS), University of the Balearic Islands, E07122 Palma, Spain
| | - Ilaria Dando
- Department of Neuroscience, Biomedicine and Movement, Biochemistry Section, University of Verona, Verona, Italy.
| | - Massimo Donadelli
- Department of Neuroscience, Biomedicine and Movement, Biochemistry Section, University of Verona, Verona, Italy.
| |
Collapse
|
|
12
|
Khabar KSA. Hallmarks of cancer and AU-rich elements. WILEY INTERDISCIPLINARY REVIEWS-RNA 2016; 8. [PMID: 27251431 PMCID: PMC5215528 DOI: 10.1002/wrna.1368] [Citation(s) in RCA: 58] [Impact Index Per Article: 6.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 04/06/2016] [Revised: 05/05/2016] [Accepted: 05/09/2016] [Indexed: 12/14/2022]
Abstract
Post‐transcriptional control of gene expression is aberrant in cancer cells. Sustained stabilization and enhanced translation of specific mRNAs are features of tumor cells. AU‐rich elements (AREs), cis‐acting mRNA decay determinants, play a major role in the posttranscriptional regulation of many genes involved in cancer processes. This review discusses the role of aberrant ARE‐mediated posttranscriptional processes in each of the hallmarks of cancer, including sustained cellular growth, resistance to apoptosis, angiogenesis, invasion, and metastasis. WIREs RNA 2017, 8:e1368. doi: 10.1002/wrna.1368 For further resources related to this article, please visit the WIREs website.
Collapse
Affiliation(s)
- Khalid S A Khabar
- King Faisal Specialist Hospital and Research Centre, Riyadh, Saudi Arabia
| |
Collapse
|
|
13
|
Ishida YI, Takeshita M, Kataoka H. Functional foods effective for hepatitis C: Identification of oligomeric proanthocyanidin and its action mechanism. World J Hepatol 2014; 6:870-879. [PMID: 25544874 PMCID: PMC4269906 DOI: 10.4254/wjh.v6.i12.870] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/26/2014] [Revised: 10/03/2014] [Accepted: 10/27/2014] [Indexed: 02/06/2023] Open
Abstract
Hepatitis C virus (HCV) is a major cause of viral hepatitis and currently infects approximately 170 million people worldwide. An infection by HCV causes high rates of chronic hepatitis (> 75%) and progresses to liver cirrhosis and hepatocellular carcinoma ultimately. HCV can be eliminated by a combination of pegylated α-interferon and the broad-spectrum antiviral drug ribavirin; however, this treatment is still associated with poor efficacy and tolerability and is often accompanied by serious side-effects. While some novel direct-acting antivirals against HCV have been developed recently, high medical costs limit the access to the therapy in cost-sensitive countries. To search for new natural anti-HCV agents, we screened local agricultural products for their suppressive activities against HCV replication using the HCV replicon cell system in vitro. We found a potent inhibitor of HCV RNA expression in the extracts of blueberry leaves and then identified oligomeric proanthocyanidin as the active ingredient. Further investigations into the action mechanism of oligomeric proanthocyanidin suggested that it is an inhibitor of heterogeneous nuclear ribonucleoproteins (hnRNPs) such as hnRNP A2/B1. In this review, we presented an overview of functional foods and ingredients efficient for HCV infection, the chemical structural characteristics of oligomeric proanthocyanidin, and its action mechanism.
Collapse
|
|
14
|
Zuccotti P, Colombrita C, Moncini S, Barbieri A, Lunghi M, Gelfi C, De Palma S, Nicolin A, Ratti A, Venturin M, Riva P. hnRNPA2/B1 and nELAV proteins bind to a specific U-rich element in CDK5R1 3'-UTR and oppositely regulate its expression. BIOCHIMICA ET BIOPHYSICA ACTA-GENE REGULATORY MECHANISMS 2014; 1839:506-16. [PMID: 24792867 DOI: 10.1016/j.bbagrm.2014.04.018] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Subscribe] [Scholar Register] [Received: 11/08/2013] [Revised: 04/09/2014] [Accepted: 04/23/2014] [Indexed: 01/27/2023]
Abstract
Cyclin-dependent kinase 5 regulatory subunit 1 (CDK5R1) encodes p35, a specific activator of cyclin-dependent kinase 5 (CDK5). CDK5 and p35 have a fundamental role in neuronal migration and differentiation during CNS development. Both the CDK5R1 3'-UTR's remarkable size and its conservation during evolution strongly indicate an important role in post-transcriptional regulation. We previously validated different regulatory elements in the 3'-UTR of CDK5R1, which affect transcript stability, p35 levels and cellular migration through the binding with nELAV proteins and miR-103/7 miRNAs. Interestingly, a 138 bp-long region, named C2.1, was identified as the most mRNA destabilizing portion within CDK5R1 3'-UTR. This feature was maintained by a shorter region of 73 bp, characterized by two poly-U stretches. UV-CL experiments showed that this region interacts with protein factors. UV-CLIP assays and pull-down experiments followed by mass spectrometry analysis demonstrated that nELAV and hnRNPA2/B1 proteins bind to the same U-rich element. These RNA-binding proteins (RBPs) were shown to oppositely control CDK5R1 mRNA stability and p35 protein content at post-trascriptional level. While nELAV proteins have a positive regulatory effect, hnRNPA2/B1 has a negative action that is responsible for the mRNA destabilizing activity both of the C2.1 region and of the full-length 3'-UTR. In co-expression experiments of hnRNPA2/B1 and nELAV RBPs we observed an overall decrease of p35 content. We also demonstrated that hnRNPA2/B1 can downregulate nELAV protein content but not vice versa. This study, by providing new insights on the combined action of different regulatory factors, contributes to clarify the complex post-transcriptional control of CDK5R1 gene expression.
Collapse
Affiliation(s)
- Paola Zuccotti
- Dipartimento di Biotecnologie Mediche e Medicina Traslazionale, Università degli Studi di Milano, Milan, Italy
| | - Claudia Colombrita
- Dipartimento di Fisiopatologia Medico-Chirurgica e dei Trapianti, Università degli Studi di Milano, Milan, Italy; IRCCS Istituto Auxologico Italiano, Cusano, Milan, Italy
| | - Silvia Moncini
- Dipartimento di Biotecnologie Mediche e Medicina Traslazionale, Università degli Studi di Milano, Milan, Italy
| | - Andrea Barbieri
- Dipartimento di Biotecnologie Mediche e Medicina Traslazionale, Università degli Studi di Milano, Milan, Italy
| | - Marta Lunghi
- Dipartimento di Biotecnologie Mediche e Medicina Traslazionale, Università degli Studi di Milano, Milan, Italy
| | - Cecilia Gelfi
- Dipartimento di Scienze Biomediche per la Salute, Università degli Studi di Milano, Milan, Italy; IBFM-CNR, Segrate, Milan, Italy
| | - Sara De Palma
- Dipartimento di Scienze Biomediche per la Salute, Università degli Studi di Milano, Milan, Italy; IBFM-CNR, Segrate, Milan, Italy
| | - Angelo Nicolin
- Dipartimento di Biotecnologie Mediche e Medicina Traslazionale, Università degli Studi di Milano, Milan, Italy
| | - Antonia Ratti
- Dipartimento di Fisiopatologia Medico-Chirurgica e dei Trapianti, Università degli Studi di Milano, Milan, Italy; IRCCS Istituto Auxologico Italiano, Cusano, Milan, Italy
| | - Marco Venturin
- Dipartimento di Biotecnologie Mediche e Medicina Traslazionale, Università degli Studi di Milano, Milan, Italy.
| | - Paola Riva
- Dipartimento di Biotecnologie Mediche e Medicina Traslazionale, Università degli Studi di Milano, Milan, Italy.
| |
Collapse
|
|
15
|
Fellows A, Mierke DF, Nichols RC. AUF1-RGG peptides up-regulate the VEGF antagonist, soluble VEGF receptor-1 (sFlt-1). Cytokine 2013; 64:337-42. [PMID: 23769804 DOI: 10.1016/j.cyto.2013.05.019] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/20/2013] [Revised: 04/16/2013] [Accepted: 05/17/2013] [Indexed: 01/17/2023]
Abstract
The macrophage is essential to the innate immune response, but also contributes to human disease by aggravating inflammation. Under severe inflammation, macrophages and other immune cells over-produce immune mediators, including vascular endothelial growth factor (VEGF). The VEGF protein stimulates macrophage activation and induces macrophage migration. A natural inhibitor of VEGF, the soluble VEGF receptor (sFlt-1) is also produced by macrophages and sFlt-1 has been used clinically to block VEGF. In macrophages, we have shown that the mRNA regulatory protein AUF1/hnRNP D represses VEGF gene expression by inhibiting translation of AURE-regulated VEGF mRNA. Peptides (AUF1-RGG peptides) that are modeled on the arginine-glycine-glycine (RGG) motif in AUF1 also block VEGF expression. This report shows that the AUF1-RGG peptides reduce two other AURE-regulated genes, TNF and GLUT1. Three alternative splice variants of sFlt-1 contain AURE in their 3'UTR, and in an apparent paradox, AUF1-RGG peptides stimulate expression of these three sFlt-1 Variants. The AUF1-RGG peptides likely act by distinct mechanisms with complimentary effects to repress VEGF gene expression and over-express the endogenous VEGF blocking agent, sFlt-1. The AUF1-RGG peptides are novel reagents that reduce VEGF and other inflammatory mediators, and may be useful tools to suppress severe inflammation.
Collapse
Affiliation(s)
- Abigail Fellows
- Veterans Administration Research Service, White River Junction, VT 05009, USA; Department of Medicine, Geisel School of Medicine at Dartmouth, Hanover, NH 03755, USA
| | | | | |
Collapse
|
|
16
|
Fellows A, Deng B, Mierke DF, Robey RB, Nichols RC. Peptides modeled on the RGG domain of AUF1/hnRNP-D regulate 3' UTR-dependent gene expression. Int Immunopharmacol 2013; 17:132-41. [PMID: 23747316 DOI: 10.1016/j.intimp.2013.05.014] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/07/2013] [Revised: 05/13/2013] [Accepted: 05/14/2013] [Indexed: 11/27/2022]
Abstract
Messenger RNA binding proteins control post-transcriptional gene expression of targeted mRNAs. The RGG (arginine-glycine-glycine) domain of the AUF1/hnRNP-D mRNA binding protein is a regulatory region that is essential for protein function. The AUF1-RGG peptide, modeled on the RGG domain of AUF1, represses expression of the macrophage cytokine, VEGF. This report expands studies on the AUF1-RGG peptide and evaluates the role of post-translational modifications of the AUF1 protein. Results show that a minimal 31-amino acid AUF1-RGG peptide that lacks poly-glutamine and nuclear localization motifs retains suppressive activity on a VEGF-3'UTR reporter. Arginine residues in RGG motifs may be methylated with resulting changes in protein function. Mass spectroscopy analysis was performed on AUF1 expressed in RAW-264.7 cells. In resting cells, arginines in the first and second RGG motifs are monomethylated. Following activation with lipopolysaccharide, the arginines are dimethylated. To evaluate if the arginine residues are essential for AUF1-RGG activity, the methylatable arginines in the AUF1-3RGG peptide were mutated to lysine or alanine. The R→K and R→A mutants lack activity. We also demonstrate that PI3K/AKT inhibitors reduce VEGF gene expression. Although immunoscreening of AUF1 suggests that LPS and PI3K inhibitors alter the phosphorylation status of AUF1-p37, mass spectroscopy results show that the p37 AUF1 isoform is not phosphorylated with or without lipopolysaccharide stimulation. In summary, arginines in the RGG domain of AUF1 are methylated, and AUF1-RGG peptides may be novel reagents that reduce macrophage activation in inflammation.
Collapse
Affiliation(s)
- Abigail Fellows
- Veterans Administration Research Service, White River Junction, VT 05009, USA
| | | | | | | | | |
Collapse
|
|
17
|
Fellows A, Griffin ME, Petrella BL, Zhong L, Parvin-Nejad FP, Fava R, Morganelli P, Robey RB, Nichols RC. AUF1/hnRNP D represses expression of VEGF in macrophages. Mol Biol Cell 2012; 23:1414-22. [PMID: 22379108 PMCID: PMC3327320 DOI: 10.1091/mbc.e11-06-0545] [Citation(s) in RCA: 21] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/13/2023] Open
Abstract
Vascular endothelial growth factor (VEGF) expression is regulated by sequence elements in the 3′ UTR of VEGF mRNA. AUF1/hnRNP D suppresses VEGF 3′ UTR–dependent expression. Peptides with arginine–glycine–glycine motifs derived from AUF1 also suppress VEGF expression. Vascular endothelial growth factor (VEGF) is a regulator of vascularization in development and is a key growth factor in tissue repair. In disease, VEGF contributes to vascularization of solid tumors and arthritic joints. This study examines the role of the mRNA-binding protein AUF1/heterogeneous nuclear ribonucleoprotein D (AUF1) in VEGF gene expression. We show that overexpression of AUF1 in mouse macrophage-like RAW-264.7 cells suppresses endogenous VEGF protein levels. To study 3′ untranslated region (UTR)–mediated regulation, we introduced the 3′ UTR of VEGF mRNA into a luciferase reporter gene. Coexpression of AUF1 represses VEGF-3′ UTR reporter expression in RAW-264.7 cells and in mouse bone marrow–derived macrophages. The C-terminus of AUF1 contains arginine–glycine–glycine (RGG) repeat motifs that are dimethylated. Deletion of the RGG domain of AUF1 eliminated the repressive effects of AUF1. Surprisingly, expression of an AUF1-RGG peptide reduced endogenous VEGF protein levels and repressed VEGF-3′ UTR reporter activity in RAW-264.7 cells. These findings demonstrate that AUF1 regulates VEGF expression, and this study identifies an RGG peptide that suppresses VEGF gene expression.
Collapse
Affiliation(s)
- Abigail Fellows
- Veterans Administration Research Service, White River Junction, VT 05009, USA
| | | | | | | | | | | | | | | | | |
Collapse
|
|
18
|
Herling A, König M, Bulik S, Holzhütter HG. Enzymatic features of the glucose metabolism in tumor cells. FEBS J 2011; 278:2436-59. [PMID: 21564549 DOI: 10.1111/j.1742-4658.2011.08174.x] [Citation(s) in RCA: 50] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/20/2023]
Abstract
Many tumor types exhibit an impaired Pasteur effect, i.e. despite the presence of oxygen, glucose is consumed at an extraordinarily high rate compared with the tissue from which they originate - the so-called 'Warburg effect'. Glucose has to serve as the source for a diverse array of cellular functions, including energy production, synthesis of nucleotides and lipids, membrane synthesis and generation of redox equivalents for antioxidative defense. Tumor cells acquire specific enzyme-regulatory mechanisms to direct the main flux of glucose carbons to those pathways most urgently required under challenging external conditions such as varying substrate availability, presence of anti-cancer drugs or different phases of the cell cycle. In this review we summarize the currently available information on tumor-specific expression, activity and kinetic properties of enzymes involved in the main pathways of glucose metabolism with due regard to the explanation of the regulatory basis and physiological significance of the Warburg effect. We conclude that, besides the expression level of the metabolic enzymes involved in the glucose metabolism of tumor cells, the unique tumor-specific pattern of isozymes and accompanying changes in the metabolic regulation below the translation level enable tumor cells to drain selfishly the blood glucose pool that non-transformed cells use as sparingly as possible.
Collapse
Affiliation(s)
- Anique Herling
- University Medicine Berlin (Charité), Institute of Biochemistry, Berlin, Germany
| | | | | | | |
Collapse
|
|
19
|
von Roretz C, Di Marco S, Mazroui R, Gallouzi IE. Turnover of AU-rich-containing mRNAs during stress: a matter of survival. WILEY INTERDISCIPLINARY REVIEWS-RNA 2010; 2:336-47. [PMID: 21957021 DOI: 10.1002/wrna.55] [Citation(s) in RCA: 103] [Impact Index Per Article: 6.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/13/2022]
Abstract
Cells undergo various adaptive measures in response to stress. Among these are specific changes in the posttranscriptional regulation of various genes. In particular, the turnover of mRNA is modified to either increase or decrease the abundance of certain target messages. Some of the best-studied mRNAs that are affected by stress are those that contain adenine/uridine-rich elements (AREs) in their 3'-untranslated regions. ARE-containing mRNAs are involved in many important cellular processes and are normally labile, but in response to stress they are differentially regulated through the concerted efforts of ARE-binding proteins (AUBPs) such as HuR, AUF1, tristetraprolin, BRF1, and KSRP, along with microRNA-mediated effects. Additionally, the fate of ARE-containing mRNAs is modified by inducing their localization to stress granules or mRNA processing bodies. Coordination of these various mechanisms controls the turnover of ARE-containing mRNAs, and thereby enables proper responses to cellular stress. In this review, we discuss how AUBPs regulate their target mRNAs in response to stress, along with the involvement of cytoplasmic granules in this process.
Collapse
|
|
20
|
Liu Y, Gao Y, Wu Y, Wu Y, Wang H, Zhang C. Histochemical mapping of hnRNP A2/B1 in rat brain after ischemia-reperfusion insults. J Histochem Cytochem 2010; 58:695-705. [PMID: 20421594 DOI: 10.1369/jhc.2010.955021] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/22/2022] Open
Abstract
Cerebral ischemia-reperfusion (I/R) insults result in neuronal cell death, brain tissue loss, and severe neurological deficits. However, the underlying mechanism is still not fully understood. Heterogeneous nuclear ribonucleoprotein (hnRNP) A2/B1 belongs to a family of RNA-binding proteins that plays a central role in pre-mRNA processing. Recent studies have revealed that hnRNP A2/B1 may be involved in the progress of I/R; therefore, the present study aimed to examine expression patterns of hnRNP A2/B1 to better understand posttranscriptional regulations in cerebral I/R insults. Focal cerebral I/R models were induced by right middle cerebral artery occlusion (MCAO) for 120 min followed by 3, 6, 12, 24, 48, and 72 hr of reperfusion in male Sprague-Dawley rats. We employed immunohistochemistry to examine expression of hnRNP A2/B1 in rat cerebral cortex (including cingulate cortex, striate cortex, temporal cortex, and piriform cortex) and hippocampus after I/R insults. Results showed that expression of hnRNP A2/B1 was significantly downregulated in cerebral cortex and hippocampus from 3 to 24 hr of reperfusion after MCAO for 120 min, but significantly upregulated at 48 hr of reperfusion. Unexpectedly, translocation of hnRNP A2/B1 from nucleus to cytoplasm and even to neurites was observed in cerebral cortex at 3 hr of reperfusion, reaching a peak at 24 hr of reperfusion, but not in hippocampus, indicating different posttranscriptional regulation patterns in different brain regions. Interestingly, translocation of hnRNP A2/B1 was only observed in cerebral cortex with MCAO but not in the opposite side, suggesting an I/R-specific expression pattern in the brain. Our data suggest that hnRNP A2/B1 participates in posttranscriptional regulation of neurons in cerebral cortex and hippocampus that suffered I/R insults, although posttranscriptional regulation is more extensive in neuronal cells of cerebral cortex than in hippocampus.
Collapse
Affiliation(s)
- Yichen Liu
- Beijing Institute of Radiation Medicine, State Key Laboratory of Proteomics, Beijing, China
| | | | | | | | | | | |
Collapse
|
|
21
|
Wang W, Zhu JQ, Yang WX. Molecular cloning and characterization of KIFC1-like kinesin gene (ot-kifc1) from Octopus tankahkeei. Comp Biochem Physiol B Biochem Mol Biol 2010; 156:174-82. [PMID: 20304088 DOI: 10.1016/j.cbpb.2010.03.004] [Citation(s) in RCA: 15] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/10/2010] [Revised: 03/12/2010] [Accepted: 03/15/2010] [Indexed: 01/31/2023]
Abstract
Spermiogenesis in Octopus tankahkeei involves striking cellular reorganization to generate a mature spermatozoon. This process may require spermatid-specific adaptation of cytoskeleton and associated molecular motor proteins. KIFC1 is a C-terminal kinesin motor with important roles in acrosome biogenesis and nuclear reshaping during spermiogenesis in rat. Here, we have cloned and characterized the gene encoding a homologue of rat KIFC1, termed as ot-kifc1, from the testis of O. tankahkeei. The 2229 bp complete cDNA contains a 75 bp 5'-untranslated region, a 1992 bp open reading frame and a 162 bp 3'-untranslated region. The deduced protein shares an overall identity of 40%, 41%, 39% and 41% with its counterpart from human, rat, mouse and African clawed frog, respectively. Tissue expression analysis revealed ot-kifc1 was expressed in testis, gill and hepatopancreas, but not in other tissues examined. In situ hybridization result showed the ot-kifc1 message was hardly detectable in early spermatid, concentrated at the tail region of intermediate spermatid, abundant in spermatid undergoing dramatic elongation and compression, enriched at one end in late spermatids and disappeared in mature sperm. In conclusion, the expression of ot-kifc1 at specific stages of spermiogenesis suggests a role for this motor in major cytological transformations.
Collapse
Affiliation(s)
- Wei Wang
- Faculty of Life Science and Bioengineering, Ningbo University, Zhejiang 315211, PR China
| | | | | |
Collapse
|
|
22
|
Hrytsenko O, Pohajdak B, Xu BY, Morrison C, van Tol B, Wright JR. Cloning and molecular characterization of the glucose transporter 1 in tilapia (Oreochromis niloticus). Gen Comp Endocrinol 2010; 165:293-303. [PMID: 19651126 DOI: 10.1016/j.ygcen.2009.07.017] [Citation(s) in RCA: 26] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 04/15/2009] [Revised: 07/13/2009] [Accepted: 07/18/2009] [Indexed: 11/21/2022]
Abstract
Facilitative glucose transporters (GLUTs) are responsible for passively transporting monosaccharides across the plasma membrane. We sequenced and characterized the Nile tilapia (Oreochromis niloticus) GLUT-1 (tGLUT-1) cDNA and genomic DNA. Using rapid amplification of the cDNA ends (RACE), two tGLUT-1 transcripts were detected differing in the length of the 3' untranslated region, 2851 and 4577 bp. Translated tGLUT-1 is a 490 amino acid product, which shares 74% homology with that of humans. Computer analysis of the amino acid sequence predicted 12 transmembrane domains, which are conserved in the GLUT-1 of various species. The tGLUT-1 gene spans more than 11 kb, and similar to the mammalian GLUT-1 genes has a 10 exon, 9 intron organization. Potential promoter regulatory elements have some similarity to those recorded for human, mouse, and rat GLUT-1 genes. Tissue expression studies revealed both GLUT-1 transcripts in liver, Brockmann bodies (BB), heart, small intestine, adipose tissue, white and red muscle, gill, spleen, pituitary gland, and brain. The highest level of expression was detected in tilapia heart, followed by BB, brain, and muscle. Protein based food and glucose had minor or no effects on the level of tGLUT-1 expression in most tissues. The tGLUT-1 mRNA level was significantly induced by glucose and food only in white muscle. Current results suggest that tGLUT-1 is similar to the GLUT-1 of other teleost species and mammals at the genomic, mRNA, and amino acid levels, supporting the concept that tGLUT-1 functions as a ubiquitous basal level glucose transporter.
Collapse
Affiliation(s)
- Olga Hrytsenko
- Department of Biology, Dalhousie University, Halifax, NS, Canada
| | | | | | | | | | | |
Collapse
|
|
23
|
Tauler J, Mulshine JL. Lung cancer and inflammation: interaction of chemokines and hnRNPs. Curr Opin Pharmacol 2009; 9:384-8. [DOI: 10.1016/j.coph.2009.06.004] [Citation(s) in RCA: 34] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/25/2009] [Accepted: 06/02/2009] [Indexed: 12/16/2022]
|
|
24
|
He Y, Rothnagel JA, Epis MR, Leedman PJ, Smith R. Downstream targets of heterogeneous nuclear ribonucleoprotein A2 mediate cell proliferation. Mol Carcinog 2009; 48:167-79. [PMID: 18680105 DOI: 10.1002/mc.20467] [Citation(s) in RCA: 22] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/10/2022]
Abstract
Over-expression of heterogeneous nuclear ribonucleoprotein (hnRNP) A2/B1 is regarded as an early marker for several cancers. This protein is associated with proto-oncogenes and tumor suppressor genes and has itself been described as a proto-oncogene. Our earlier experiments drew a connection between hnRNP A2/B1 levels and cell proliferation and raised the possibility that this protein contributes to the uncontrolled cell division that characterizes cancer. Limited knowledge of the downstream targets of hnRNP A2/B1 has, however, precluded a clear understanding of their roles in cancer cell growth. To define the pathways in which this protein acts we have now carried out microarray experiments with total RNA from Colo16 epithelial cells transfected with an shRNA that markedly suppresses hnRNP A2/B1 expression. The microarray data identified 123 genes, among 22 283 human gene probe sets, with altered expression levels in hnRNP A2/B1-depleted cells. Ontological analysis showed that many of these downstream targets are involved in regulation of the cell cycle and cell proliferation and that this group of proteins is significantly over-represented amongst the affected proteins. The changes detected in the microarray experiments were confirmed by real-time PCR for a subset of proliferation-related genes. Immunoprecipitation-RT-PCR demonstrated that hnRNP A2/B1 formed complexes with the transcripts of many of the verified downstream genes, suggesting that hnRNP A2/B1 contributes to the regulation of these genes. These results reinforce the conclusion that hnRNP A2/B1 is associated with cellular processes that affect the cell cycle and proliferation.
Collapse
Affiliation(s)
- Yaowu He
- School of Molecular and Microbial Sciences, University of Queensland, Brisbane, Queensland, Australia
| | | | | | | | | |
Collapse
|
|
25
|
Söderberg M, Lang MA. Megaprimer-based methodology for deletion of a large fragment within a repetitive polypyrimidine-rich DNA. Mol Biotechnol 2006; 32:65-71. [PMID: 16382183 DOI: 10.1385/mb:32:1:065] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/11/2022]
Abstract
Site-directed mutagenesis is often a prerequisite for elucidation of the functional significance of cis- and trans-factors involved in gene regulation. The aim of this study was to delete the primary binding site for heterogeneous nuclear ribonucleoprotein I (hnRNPI) within the inducible nitric oxide synthase (iNOS) 3' untranslated region mRNA. The binding site consists of a 53-nucleotide CU-rich region within a long stretch of polypyrimidines. As a result of primer pair annealing, the repetitive sequence limited the use of several deletion methods based on polymerase chain reaction. Therefore, a megaprimer approach was chosen. The megaprimer was produced by a forward primer outside the polypyrimidine-rich region, and a mutagenic reverse primer annealing to flanking regions of the desired deletion, thereby looping out the target sequence. Subsequently, this megaprimer was used to create the final deletion recombinant. The deletion was verified by sequencing and by ultraviolet cross-linking mouse liver protein extracts with radiolabeled mutant and wild-type RNAs. In conclusion, the megaprimer method offers a solution for generating large internal deletions in repetitive sequences, which facilitates investigations on large repetitive DNA or RNA regions interacting with trans-factors.
Collapse
Affiliation(s)
- Malin Söderberg
- Department of Pharmaceutical Biosciences, Division of Biochemistry, Uppsala University, Biomedicum, Box 578, SE-751 23 Uppsala, Sweden.
| | | |
Collapse
|
|
26
|
Du M, Roy KM, Zhong L, Shen Z, Meyers HE, Nichols RC. VEGF gene expression is regulated post-transcriptionally in macrophages. FEBS J 2006; 273:732-45. [PMID: 16441660 DOI: 10.1111/j.1742-4658.2006.05106.x] [Citation(s) in RCA: 23] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
Abstract
The macrophage is critical to the innate immune response and contributes to human diseases, including inflammatory arthritis and plaque formation in atherosclerosis. Vascular endothelial growth factor (VEGF) is an angiogenic cytokine that is produced by macrophages. To study the regulation of VEGF production in macrophages we show that stimulation of monocyte-macrophage-like RAW-264.7 cells by lipopolysaccharide (LPS) increases expression of VEGF mRNA and protein. Three alternative splicing VEGF mRNA isoforms are produced, and the stability of VEGF mRNA increases following cellular activation. To study post-transcriptional regulation of the VEGF gene the 3'-untranslated region (3' UTR) was introduced into the 3' UTR of the luciferase gene in a reporter construct. In both RAW-264.7 cells and thioglycollate-elicited macrophages, the 3' UTR sequence dramatically reduces reporter expression. Treatment with activators of macrophages, including LPS, lipoteichoic acid, and VEGF protein, stimulates expression of 3' UTR reporters. Finally, mapping studies of the 3' UTR of VEGF mRNA show that deletion of the heterogeneous nuclear ribonucleoprotein l binding site affects basal reporter expression in RAW-264.7 cells, but does not affect reporter activation with LPS. Together these results demonstrate that a post-transcriptional mechanism contributes to VEGF gene expression in activated macrophage cells.
Collapse
Affiliation(s)
- Min Du
- Department of Microbiology and Immunology, Dartmouth Medical School, Hanover, NH, and Veterans Administration Research Service, White River Junction, VT 05009-0001, USA
| | | | | | | | | | | |
Collapse
|