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1
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Desjardins P, Le-Bel G, Ghio SC, Germain L, Guérin SL. The WNK1 kinase regulates the stability of transcription factors during wound healing of human corneal epithelial cells. J Cell Physiol 2022; 237:2434-2450. [PMID: 35150137 DOI: 10.1002/jcp.30698] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/14/2021] [Revised: 01/19/2022] [Accepted: 01/25/2022] [Indexed: 11/12/2022]
Abstract
Due to its superficial anatomical localization, the cornea is continuously subjected to injuries. Damages to the corneal epithelium trigger important changes in the composition of the extracellular matrix to which the basal human corneal epithelial cells (hCECs) attach. These changes are perceived by membrane-bound integrins and ultimately lead to re-epithelialization of the injured epithelium through intracellular signalin. Among the many downstream targets of the integrin-activated signaling pathways, WNK1 is the kinase whose activity is the most strongly increased during corneal wound healing. We previously demonstrated that pharmacological inhibition of WNK1 prevents proper closure of wounded human tissue-engineered cornea in vitro. In the present study, we investigated the molecular mechanisms by which WNK1 contributes to corneal wound healing. By exploiting transcription factors microarrays, electrophoretic mobility-shift assay, and gene profiling analyses, we demonstrated that the DNA binding properties and expression of numerous transcription factors (TFs), including the well-known, ubiquitous TFs specific protein 1 (Sp1) and activator protein 1 (AP1), were reduced in hCECs upon WNK1 inhibition by WNK463. This process appears to be mediated at least in part by alteration in both the ubiquitination and glycosylation status of these TFs. These changes in TFs activity and expression impacted the transcription of several genes, including that encoding the α5 integrin subunit, a well-known target of both Sp1 and AP1. Gene profiling revealed that only a moderate number of genes in hCECs had their level of expression significantly altered in response to WNK463 exposition. Interestingly, analysis of the microarray data for these deregulated genes using the ingenuity pathway analysis software predicted that hCECs would stop migrating and proliferating but differentiate more when they are grown in the presence of the WNK1 inhibitor. These results demonstrate that WNK1 plays a critical function by orienting hCECs into the appropriate biological response during the process of corneal wound healing.
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Affiliation(s)
- Pascale Desjardins
- Centre Universitaire d'Ophtalmologie - Recherche (CUO-Recherche) et Centre de Recherche du CHU de Québec, Université Laval, Québec City, Québec, Canada.,Centre de Recherche en Organogénèse Expérimentale de l'Université Laval/LOEX, Centre de Recherche du CHU de Québec, Université Laval, Québec City, Québec, Canada.,Département de Chirurgie, Faculté de Médecine, Université Laval, Québec City, Québec, Canada.,Département d'Ophtalmologie, Faculté de Médecine, Université Laval, Québec City, Québec, Canada
| | - Gaëtan Le-Bel
- Centre Universitaire d'Ophtalmologie - Recherche (CUO-Recherche) et Centre de Recherche du CHU de Québec, Université Laval, Québec City, Québec, Canada.,Centre de Recherche en Organogénèse Expérimentale de l'Université Laval/LOEX, Centre de Recherche du CHU de Québec, Université Laval, Québec City, Québec, Canada.,Département de Chirurgie, Faculté de Médecine, Université Laval, Québec City, Québec, Canada.,Département d'Ophtalmologie, Faculté de Médecine, Université Laval, Québec City, Québec, Canada
| | - Sergio C Ghio
- Centre de Recherche en Organogénèse Expérimentale de l'Université Laval/LOEX, Centre de Recherche du CHU de Québec, Université Laval, Québec City, Québec, Canada.,Département de Chirurgie, Faculté de Médecine, Université Laval, Québec City, Québec, Canada
| | - Lucie Germain
- Centre Universitaire d'Ophtalmologie - Recherche (CUO-Recherche) et Centre de Recherche du CHU de Québec, Université Laval, Québec City, Québec, Canada.,Centre de Recherche en Organogénèse Expérimentale de l'Université Laval/LOEX, Centre de Recherche du CHU de Québec, Université Laval, Québec City, Québec, Canada.,Département de Chirurgie, Faculté de Médecine, Université Laval, Québec City, Québec, Canada.,Département d'Ophtalmologie, Faculté de Médecine, Université Laval, Québec City, Québec, Canada
| | - Sylvain L Guérin
- Centre Universitaire d'Ophtalmologie - Recherche (CUO-Recherche) et Centre de Recherche du CHU de Québec, Université Laval, Québec City, Québec, Canada.,Centre de Recherche en Organogénèse Expérimentale de l'Université Laval/LOEX, Centre de Recherche du CHU de Québec, Université Laval, Québec City, Québec, Canada.,Département d'Ophtalmologie, Faculté de Médecine, Université Laval, Québec City, Québec, Canada
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Nakano R, Nakayama T, Sugiya H. Biological Properties of JNK3 and Its Function in Neurons, Astrocytes, Pancreatic β-Cells and Cardiovascular Cells. Cells 2020; 9:cells9081802. [PMID: 32751228 PMCID: PMC7464089 DOI: 10.3390/cells9081802] [Citation(s) in RCA: 24] [Impact Index Per Article: 4.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/27/2020] [Revised: 07/15/2020] [Accepted: 07/27/2020] [Indexed: 12/28/2022] Open
Abstract
JNK is a protein kinase, which induces transactivation of c-jun. The three isoforms of JNK, JNK1, JNK2, and JNK3, are encoded by three distinct genes. JNK1 and JNK2 are expressed ubiquitously throughout the body. By contrast, the expression of JNK3 is limited and observed mainly in the brain, heart, and testes. Concerning the biological properties of JNKs, the contribution of upstream regulators and scaffold proteins plays an important role in the activation of JNKs. Since JNK signaling has been described as a form of stress-response signaling, the contribution of JNK3 to pathophysiological events, such as stress response or cell death including apoptosis, has been well studied. However, JNK3 also regulates the physiological functions of neurons and non-neuronal cells, such as development, regeneration, and differentiation/reprogramming. In this review, we shed light on the physiological functions of JNK3. In addition, we summarize recent advances in the knowledge regarding interactions between JNK3 and cellular reprogramming.
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Affiliation(s)
- Rei Nakano
- Laboratory for Cellular Function Conversion Technology, RIKEN Center for Integrative Medical Sciences (IMS), 1-7-22 Suehiro-cho, Tsurumi-ku, Yokohama 230-0045, Japan
- Laboratory of Veterinary Radiology, College of Bioresource Sciences, Nihon University, 1866 Kameino, Fujisawa 252-0880, Japan; (T.N.); (H.S.)
- Correspondence:
| | - Tomohiro Nakayama
- Laboratory of Veterinary Radiology, College of Bioresource Sciences, Nihon University, 1866 Kameino, Fujisawa 252-0880, Japan; (T.N.); (H.S.)
| | - Hiroshi Sugiya
- Laboratory of Veterinary Radiology, College of Bioresource Sciences, Nihon University, 1866 Kameino, Fujisawa 252-0880, Japan; (T.N.); (H.S.)
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3
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Das A, Durrant D, Mitchell C, Dent P, Batra SK, Kukreja RC. Sildenafil (Viagra) sensitizes prostate cancer cells to doxorubicin-mediated apoptosis through CD95. Oncotarget 2016; 7:4399-413. [PMID: 26716643 PMCID: PMC4826214 DOI: 10.18632/oncotarget.6749] [Citation(s) in RCA: 38] [Impact Index Per Article: 4.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/14/2015] [Accepted: 11/26/2015] [Indexed: 01/16/2023] Open
Abstract
We previously reported that Sildenafil enhances apoptosis and antitumor efficacy of doxorubicin (DOX) while attenuating its cardiotoxic effect in prostate cancer. In the present study, we investigated the mechanism by which sildenafil sensitizes DOX in killing of prostate cancer (PCa) cells, DU145. The death receptor Fas (APO-1 or CD95) induces apoptosis in many carcinoma cells, which is negatively regulated by anti-apoptotic molecules such as FLIP (Fas-associated death domain (FADD) interleukin-1-converting enzyme (FLICE)-like inhibitory protein). Co-treatment of PCa cells with sildenafil and DOX for 48 hours showed reduced expression of both long and short forms of FLIP (FLIP-L and -S) as compared to individual drug treatment. Over-expression of FLIP-s with an adenoviral vector attentuated the enhanced cell-killing effect of DOX and sildenafil. Colony formation assays also confirmed that FLIP-S over-expression inhibited the DOX and sildenafil-induced synergistic killing effect as compared to the cells infected with an empty vector. Moreover, siRNA knock-down of CD95 abolished the effect of sildenafil in enhancing DOX lethality in cells, but had no effect on cell killing after treatment with a single agent. Sildenafil co-treatment with DOX inhibited DOX-induced NF-κB activity by reducing phosphorylation of IκB and nuclear translocation of the p65 subunit, in addition to down regulation of FAP-1 (Fas associated phosphatase-1, a known inhibitor of CD95-mediated apoptosis) expression. This data provides evidence that the CD95 is a key regulator of sildenafil and DOX mediated enhanced cell death in prostate cancer.
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Affiliation(s)
- Anindita Das
- Department of Internal Medicine, Pauley Heart Center, Virginia Commonwealth University, Richmond, VA, USA
| | - David Durrant
- Department of Internal Medicine, Pauley Heart Center, Virginia Commonwealth University, Richmond, VA, USA
| | - Clint Mitchell
- Department of Biochemistry and Molecular Biology, Virginia Commonwealth University, Richmond, VA, USA
| | - Paul Dent
- Department of Biochemistry and Molecular Biology, Virginia Commonwealth University, Richmond, VA, USA
| | - Surinder K. Batra
- Department of Biochemistry and Molecular Biology, Buffett Cancer Center, University of Nebraska Medical Center, Omaha, NE, USA
| | - Rakesh C. Kukreja
- Department of Internal Medicine, Pauley Heart Center, Virginia Commonwealth University, Richmond, VA, USA
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Hendriks WJAJ, Böhmer FD. Non-transmembrane PTPs in Cancer. PROTEIN TYROSINE PHOSPHATASES IN CANCER 2016:47-113. [DOI: 10.1007/978-1-4939-3649-6_3] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/06/2025]
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5
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Qiu T, Zhou X, Wang J, Du Y, Xu J, Huang Z, Zhu W, Shu Y, Liu P. MiR-145, miR-133a and miR-133b inhibit proliferation, migration, invasion and cell cycle progression via targeting transcription factor Sp1 in gastric cancer. FEBS Lett 2014; 588:1168-77. [PMID: 24613927 DOI: 10.1016/j.febslet.2014.02.054] [Citation(s) in RCA: 146] [Impact Index Per Article: 13.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/18/2013] [Revised: 02/14/2014] [Accepted: 02/26/2014] [Indexed: 01/06/2023]
Abstract
MicroRNAs have recently emerged as key regulators of gastric cancers. Here we found that miR-145, miR-133a and miR-133b were down-regulated in gastric cancer tissues and cell lines. Overexpression of miR-145, miR-133a and miR-133b induced G1 cell cycle arrest and inhibited cell proliferation, migration and invasion in vitro. MiR-145, miR-133a and miR-133b targeted the transcription factor SP1, knockdown of which reduced the expression of MMP-9 and Cyclin D1 that were involved in cell growth and invasion. Thus, our findings demonstrated for the first time that miR-145, miR-133a and miR-133b suppressed the proliferation, migration, invasion and cell cycle progression of gastric cancer cells through decreasing expression of Sp1 and its downstream proteins.
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Affiliation(s)
- Tianzhu Qiu
- Department of Oncology, First Affiliated Hospital of Nanjing Medical University, 300 Guangzhou Road, Nanjing 210029, China
| | - Xin Zhou
- Department of Oncology, First Affiliated Hospital of Nanjing Medical University, 300 Guangzhou Road, Nanjing 210029, China
| | - Jian Wang
- Department of Oncology, First Affiliated Hospital of Nanjing Medical University, 300 Guangzhou Road, Nanjing 210029, China
| | - Yiping Du
- Department of Oncology, First Affiliated Hospital of Nanjing Medical University, 300 Guangzhou Road, Nanjing 210029, China
| | - Jun Xu
- Department of Oncology, First Affiliated Hospital of Nanjing Medical University, 300 Guangzhou Road, Nanjing 210029, China
| | - Zebo Huang
- Department of Oncology, First Affiliated Hospital of Nanjing Medical University, 300 Guangzhou Road, Nanjing 210029, China
| | - Wei Zhu
- Department of Oncology, First Affiliated Hospital of Nanjing Medical University, 300 Guangzhou Road, Nanjing 210029, China
| | - Yongqian Shu
- Department of Oncology, First Affiliated Hospital of Nanjing Medical University, 300 Guangzhou Road, Nanjing 210029, China
| | - Ping Liu
- Department of Oncology, First Affiliated Hospital of Nanjing Medical University, 300 Guangzhou Road, Nanjing 210029, China.
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6
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Huang CC, Chang WSW. Cooperation between NRF-2 and YY-1 transcription factors is essential for triggering the expression of the PREPL-C2ORF34 bidirectional gene pair. BMC Mol Biol 2009; 10:67. [PMID: 19575798 PMCID: PMC2713978 DOI: 10.1186/1471-2199-10-67] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/06/2008] [Accepted: 07/03/2009] [Indexed: 11/11/2022] Open
Abstract
Background Many mammalian genes are organized as bidirectional (head-to-head) gene pairs with the two genes separated only by less than 1 kb. The transcriptional regulation of these bidirectional gene pairs remains largely unclear, but a few studies have suggested that the two closely adjacent genes in divergent orientation can be co-regulated by a single transcription factor binding to a specific regulatory fragment. Here we report an evolutionarily conserved bidirectional gene pair, known as the PREPL-C2ORF34 gene pair, whose transcription relies on the synergic cooperation of two transcription factors binding to an intergenic bidirectional minimal promoter. Results While PREPL is present primarily in brain and heart, C2ORF34 is ubiquitously and abundantly expressed in almost all tissues. Genomic analyses revealed that these two non-homologous genes are adjacent in a head-to-head configuration on human chromosome 2p21 and separated by only 405 bp. Within this short intergenic region, a 243-bp GC-rich segment was demonstrated to function as a bidirectional minimal promoter to initiate the transcription of both flanking genes. Two key transcription factors, NRF-2 and YY-1, were further identified to coordinately participate in driving both gene expressions in an additive manner. The functional cooperation between these two transcription factors, along with their genomic binding sites and some cis-acting repressive elements, are essential for the transcriptional activation and tissue distribution of the PREPL-C2ORF34 bidirectional gene pair. Conclusion This study provides new insights into the complex transcriptional mechanism of a mammalian head-to-head gene pair which requires cooperative binding of multiple transcription factors to a bidirectional minimal promoter of the shared intergenic region.
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Affiliation(s)
- Chien-Chang Huang
- 1Institute of Life Sciences, National Defense Medical Center, Taipei 114, Taiwan.
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Abstract
Protein tyrosine phosphatase, PTPL1, (also known as PTPN13, FAP-1, PTP-BAS, PTP1E) is a non-receptor type PTP and, at 270 kDa, is the largest phosphatase within this group. In addition to the well-conserved PTP domain, PTPL1 contains at least 7 putative macromolecular interaction domains. This structural complexity indicates that PTPL1 may modulate diverse cellular functions, perhaps exerting both positive and negative effects. In accordance with this idea, while certain studies suggest that PTPL1 can act as a tumor-promoting gene other experimental studies have suggested that PTPL1 may function as a tumor suppressor. The role of PTPL1 in the cancer cell is therefore likely to be both complex and context dependent with possible roles including the modulation of growth, stress-response, and cytoskeletal remodeling pathways. Understanding the nature of molecular complexes containing PTPL1, its interaction partners, substrates, regulation and subcellular localization are key to unraveling the complex personality of this protein phosphatase.
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Affiliation(s)
- Ogan D Abaan
- Department of Oncology, Lombardi Comprehensive Cancer Center, Georgetown University, Washington, DC 20057, USA
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8
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Konta T, Emi M, Toriyama S, Ariumi H, Ishii M, Takasaki S, Ikeda A, Ichikawa K, Shibata Y, Takabatake N, Takeishi Y, Kato T, Kawata S, Kubota I. Association of CC chemokine ligand 5 genotype with urinary albumin excretion in the non-diabetic Japanese general population: the Takahata study. J Hum Genet 2008; 53:267. [PMID: 18217191 DOI: 10.1007/s10038-008-0246-6] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/09/2007] [Accepted: 12/26/2007] [Indexed: 10/22/2022]
Abstract
Albuminuria is an early marker of vascular damage, and its development in diabetic nephropathy is associated with genotype of inflammatory CC chemokine ligand 5 (CCL5). This study investigated whether the association of CCL5 and albuminuria is a general phenomenon. We characterized a Japanese population consisting of 2,749 non-diabetic individuals over 40 years in Takahata, Japan. The urine albumin-creatinine ratio (UACR) was obtained from morning spot urine. We genotyped SNPs within the CCL5 gene that displayed frequent minor allele frequencies in Japanese (i.e., rs2107538, rs2280789, rs3817655 and rs9909416). Assessment of possible association and linkage disequilibrium (LD) revealed that all four SNP genotypes are correlated significantly with UACR (P = 0.004-0.005), and these four SNPs variations showed an obvious consistency of genotypes by detecting almost complete linkage disequilibrium (D' = 1 and r (2) > 0.95). We found two exclusive haplotypes in the CCL5 gene (haplotype1: rs2107538G/rs2280789T/rs3817655T/rs9909416G, frequency 0.64 and haplotype2: rs2107538A/rs2280789C/rs3817655A/rs9909416A, frequency 0.35) among the population. A significant association with elevated UACR was identified with haplotype1 (P = 0.002). Homozygotes for haplotype1 displayed strikingly-elevated UACR (48.5 +/- 6.6 mg/g, n = 1,116) compared to the rest (28.6 +/- 1.6 mg/g, n = 1,530) (P = 0.001). In conclusion, these results suggested that genetic variation of CCL5 might be an important risk factor for albuminuria in the non-diabetic Japanese general population.
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Affiliation(s)
- Tsuneo Konta
- Department of Cardiology, Pulmonology, and Nephrology, Yamagata University School of Medicine, 2-2-2, Iida-Nishi, Yamagata, 990-9585, Japan.
| | - Mitsuru Emi
- HuBit Genomix Research Institute, Tokyo, Japan
| | | | | | - Miho Ishii
- HuBit Genomix Research Institute, Tokyo, Japan
| | - Satoshi Takasaki
- Department of Cardiology, Pulmonology, and Nephrology, Yamagata University School of Medicine, 2-2-2, Iida-Nishi, Yamagata, 990-9585, Japan
| | - Ami Ikeda
- Department of Cardiology, Pulmonology, and Nephrology, Yamagata University School of Medicine, 2-2-2, Iida-Nishi, Yamagata, 990-9585, Japan
| | - Kazunobu Ichikawa
- Department of Cardiology, Pulmonology, and Nephrology, Yamagata University School of Medicine, 2-2-2, Iida-Nishi, Yamagata, 990-9585, Japan
| | - Yoko Shibata
- Department of Cardiology, Pulmonology, and Nephrology, Yamagata University School of Medicine, 2-2-2, Iida-Nishi, Yamagata, 990-9585, Japan
| | - Noriaki Takabatake
- Department of Cardiology, Pulmonology, and Nephrology, Yamagata University School of Medicine, 2-2-2, Iida-Nishi, Yamagata, 990-9585, Japan
| | - Yasuchika Takeishi
- Department of Cardiology, Pulmonology, and Nephrology, Yamagata University School of Medicine, 2-2-2, Iida-Nishi, Yamagata, 990-9585, Japan
| | - Takeo Kato
- Department of Neurology, Hematology, Metabolism, Endocrinology and Diabetes, Yamagata University School of Medicine, Yamagata, Japan
| | - Sumio Kawata
- Department of Gastroenterology, Yamagata University School of Medicine, Yamagata, Japan
| | - Isao Kubota
- Department of Cardiology, Pulmonology, and Nephrology, Yamagata University School of Medicine, 2-2-2, Iida-Nishi, Yamagata, 990-9585, Japan
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Huang W, Zhu C, Wang H, Horvath E, Eklund EA. The interferon consensus sequence-binding protein (ICSBP/IRF8) represses PTPN13 gene transcription in differentiating myeloid cells. J Biol Chem 2008; 283:7921-35. [PMID: 18195016 DOI: 10.1074/jbc.m706710200] [Citation(s) in RCA: 47] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/06/2022] Open
Abstract
The interferon consensus sequence-binding protein (ICSBP/IRF8) is an interferon regulatory factor that is expressed in myeloid and B-cells. ICSBP-deficient mice develop a myeloproliferative disorder characterized by cytokine hypersensitivity and apoptosis resistance. To identify ICSBP target genes involved in these effects, we screened a CpG island microarray with chromatin that co-immunoprecipitated with ICSBP from myeloid cells. Using this technique, we identified PTPN13 as an ICSBP target gene. PTPN13 encodes Fas-associated phosphatase 1 (Fap-1), a ubiquitously expressed protein-tyrosine phosphatase. This was of interest because interaction of Fap-1 with Fas results in Fas dephosphorylation and inhibition of Fas-induced apoptosis. In this study, we found that ICSBP influenced Fas-induced apoptosis in a Fap-1-dependent manner. We also found that ICSBP interacted with a cis element in the proximal PTPN13 promoter and repressed transcription. This interaction increased during myeloid differentiation and was regulated by phosphorylation of conserved tyrosine residues in the interferon regulatory factor domain of ICSBP. ICSBP deficiency was present in human myeloid malignancies, including chronic myeloid leukemia. Therefore, these studies identified a mechanism for increased survival of mature myeloid cells in the ICSBP-deficient murine model and in human myeloid malignancies with decreased ICSBP expression.
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Affiliation(s)
- Weiqi Huang
- The Feinberg School of Medicine and The Robert H. Lurie Comprehensive Cancer Center, Northwestern University, Chicago, Illinois 60611, USA
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Bernardi A, Bavaresco L, Wink MR, Jacques-Silva MC, Delgado-Cañedo A, Lenz G, Battastini AMO. Indomethacin stimulates activity and expression of ecto-5'-nucleotidase/CD73 in glioma cell lines. Eur J Pharmacol 2007; 569:8-15. [PMID: 17568578 DOI: 10.1016/j.ejphar.2007.04.058] [Citation(s) in RCA: 16] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/10/2006] [Revised: 04/16/2007] [Accepted: 04/21/2007] [Indexed: 01/23/2023]
Abstract
Gliomas are the most common and devastating primary tumors of the central nervous system. Ecto-NTPDases and ecto-5'-nucleotidase/CD73 can control extracellular ATP/adenosine levels, which have been described as proliferation factors. Here, we investigate the influence of indomethacin on the enzyme cascade that catalyses the interconversion of purine nucleotides in U138-MG and C6 glioma cell lines. Exposure of glioma cells to 100 microM indomethacin for 48 h caused increases of 52% (P < 0.05) and 62% (P < 0.05) in the AMP hydrolysis rate in C6 and U138-MG cell lines, respectively. Indomethacin treatments also increased ATP hydrolysis. Significant increase in ecto-5'-nucleotidase/CD73 mRNA and protein levels were observed after treatment with indomethacin. Pretreatment of glioma cells with a specific antagonist of the adenosine A(3) receptor, MRS1220 (1 microM; 9-Chloro-2-(2-furanyl)-5-((phenylacetyl)amino)-[1,2,4]triazolo[1,5-c]quinazoline), significantly reduced the inhibition of cell proliferation induced by indomethacin. In addition, a significant increase in mRNA levels of the adenosine A(3) receptor was observed after treatment with indomethacin. In conclusion, our data indicate that adenosine A(3) receptors and the enzyme, ecto-5'-nucleotidase/CD73, are involved in the anti-proliferative effect of indomethacin in glioma cells.
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Affiliation(s)
- Andressa Bernardi
- Departamento de Bioquímica, Instituto de Ciências Básicas da Saúde, Universidade Federal do Rio Grande do Sul, Av. Ramiro Barcelos, 2600-anexo, CEP 90035-003, Porto Alegre, RS, Brazil
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Ivanov VN, Ronai Z, Hei TK. Opposite roles of FAP-1 and dynamin in the regulation of Fas (CD95) translocation to the cell surface and susceptibility to Fas ligand-mediated apoptosis. J Biol Chem 2005; 281:1840-52. [PMID: 16306044 PMCID: PMC4376329 DOI: 10.1074/jbc.m509866200] [Citation(s) in RCA: 52] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/13/2022] Open
Abstract
Human melanoma is the most aggressive form of skin cancer and is extremely resistant to radiation and chemotherapy. One of the critical parameters of this resistance is down-regulation of Fas (CD95) cell-surface expression. Using TIG3 normal human fibroblasts and human melanoma cell lines, we investigated transcriptional regulation of FAP-1, a regulator of Fas translocation in the cell. Protein-tyrosine phosphatase FAP-1 (PTPN13, PTP-BAS) interacts with human Fas protein and prevents its export from the cytoplasm to the cell surface. In contrast, dynamin-2 facilitates Fas protein translocation from the Golgi apparatus via the trans-Golgi network to the cell surface. Suppression of dynamin functions by dominant negative dynamin K44A blocks Fas export, whereas the down-regulation of FAP-1 expression by specific RNA interference restores Fas export (a phenomenon that could still be down-regulated in the presence of dominant-negative dynamin). Based on the FAP-1- and dynamin-dependent regulation of Fas translocation, we have created human melanoma lines with different levels of surface expression of Fas. Treatment of these melanoma lines with soluble Fas ligand resulted in programmed cell death that was proportional to the pre-existing levels of surface Fas. Taking into consideration the well known observations that FAP-1 expression is often up-regulated in metastatic tumors, we have established a causal connection between high basal NF-kappaB transcription factor activity (which is a hallmark of many types of metastatic tumors) and NF-kappaB-dependent transcriptional regulation of FAP-1 gene expression that finally restricts Fas protein trafficking, thereby, facilitating the survival of cancer cells.
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Affiliation(s)
- Vladimir N Ivanov
- Center for Radiological Research, College of Physicians and Surgeons, Columbia University, New York, New York 10032, USA.
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12
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Zhang J, Zhu ZG, Ji J, Yuan F, Yu YY, Liu BY, Lin YZ. Transcription factor Sp1 expression in gastric cancer and its relationship to long-term prognosis. World J Gastroenterol 2005; 11:2213-7. [PMID: 15818728 PMCID: PMC4305801 DOI: 10.3748/wjg.v11.i15.2213] [Citation(s) in RCA: 38] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
AIM: To explore the expression of Sp1 in gastric carcinoma as well as its association with other clinicopathologic features, and to evaluate the role of Sp1 as a prognostic indicator of gastric carcinoma.
METHODS: By using immunohistochemistry, we examined the Sp1 expression patterns in 65 cases of human gastric cancer, and 40 normal gastric mucosa specimens. Simultaneously, the correlation between Sp1 expression and clinical outcome or clinicopathologic features was investigated.
RESULTS: The percentage of Sp1 expression was 12.5% (5/40) in normal gastric mucosa, and the Sp1 protein was mainly expressed in the nuclei of cells located in the mucous neck region. In sharp contrast, strong Sp1 expression was detected in tumor cells, whereas no or faint Sp1 staining was detected in stromal cells and normal glandular cells surrounding the tumors. The expression rate of Sp1 in gastric cancer lesions was 53.85% (35/65). The medium survival duration in patients who had a tumor with negative, weak and strong Sp1 expressions was 1700, 1560 and 1026 d, respectively (P<0.05). Sp1 protein expression was closely related to the depth of tumor infiltration (χ2 = 13.223, P<0.01) and TNM stage (χ2 = 11.009, P<0.05), but had no relationship with the number of lymph nodes and Lauren’s classification (P>0.05). Cox regression model for multivariate analysis revealed that high Sp1 expression (P<0.05) and advanced stage (P<0.01) were independent predictors of poor survival.
CONCLUSION: Normal and malignant gastric tissues have unique Sp1 expression patterns. Sp1 might serve as an independent prognostic factor, by influencing the tumor infiltration and progression.
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Affiliation(s)
- Jun Zhang
- Department of Surgery, Rui Jin Hospital, Shanghai Institute of Digestive Surgery, Shanghai Second Medical University, Shanghai 200025, China.
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13
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Doerwald L, van Rheede T, Dirks RP, Madsen O, Rexwinkel R, van Genesen ST, Martens GJ, de Jong WW, Lubsen NH. Sequence and Functional Conservation of the Intergenic Region Between the Head-to-Head Genes Encoding the Small Heat Shock Proteins αB-Crystallin and HspB2 in the Mammalian Lineage. J Mol Evol 2004; 59:674-86. [PMID: 15693623 DOI: 10.1007/s00239-004-2659-y] [Citation(s) in RCA: 17] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/30/2022]
Abstract
An unexpected feature of the large mammalian genome is the frequent occurrence of closely linked head-to-head gene pairs. Close apposition of such gene pairs has been suggested to be due to sharing of regulatory elements. We show here that the head-to-head gene pair encoding two small heat shock proteins, alphaB-crystallin and HspB2, is closely linked in all major mammalian clades, suggesting that this close linkage is of selective advantage. Yet alphaB-crystallin is abundantly expressed in lens and muscle and in response to a heat shock, while HspB2 is abundant only in muscle and not upregulated by a heat shock. The intergenic distance between the genes for these two proteins in mammals ranges from 645 bp (platypus) to 1069 bp (opossum), with an average of about 900 bp; in chicken the distance was the same as in duck (1.6 kb). Phylogenetic footprinting and sequence alignment identified a number of conserved sequence elements close to the HspB2 promoter and two farther upstream. All known regulatory elements of the mouse alphaB-crystallin promoter are conserved, except in platypus and birds. The lens-specific region 1 (LSR1) and the heat shock elements (HSEs) lack in birds; in platypus the LSR1 is reduced to a Pax-6 site, while the Pax-6 site in LSR2 and a HSE are absent. Most likely the primordial mammalian alphaB-crystallin promoter had two LSRs and two HSEs. In transfection experiments the platypus alphaB-crystallin promoter retained heat shock responsiveness and lens expression. It also directed lens expression in Xenopus laevis transgenes, as did the HspB2 promoter of rat or blind mole rat. Deletion of the middle of the intergenic region including the upstream enhancer affected the activity of both the rat alphaB-crystallin and the HspB2 promoters, suggesting sharing of the enhancer region by the two promoters.
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Affiliation(s)
- Linda Doerwald
- Department of Biochemistry, Faculty of Science, University of Nijmegen, Nijmegen, The Netherlands
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14
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Herr DR, Harris GL. Close head-to-head juxtaposition of genes favors their coordinate regulation inDrosophila melanogaster. FEBS Lett 2004; 572:147-53. [PMID: 15304339 DOI: 10.1016/j.febslet.2004.07.026] [Citation(s) in RCA: 17] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/21/2004] [Revised: 07/12/2004] [Accepted: 07/13/2004] [Indexed: 11/23/2022]
Abstract
This report identifies a large number of gene-pairs in Drosophila melanogaster that share a common upstream region. 877 gene-pairs (approximately 12% of the genome) are separated by less than 350 bp in a head-to-head orientation. This positional relationship is more highly favored in flies than in other organisms. These gene pairs have a higher correlation of expression than similarly spaced genes that have head-to-tail or tail-to-tail orientations. Thus, the positional arrangement of genes appears to play a significant role in coordinating relative expression patterns and may provide clues for identifying the functions of unknown genes.
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Affiliation(s)
- Deron R Herr
- Department of Biology and Molecular Biology Institute, San Diego State University, San Diego, CA 92182-4614, USA.
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15
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Nobe Y, Sato K, Emi M, Ezura Y, Fujita Y, Takada D, Ishigami T, Umemura S, Xin Y, Wu LL, Larrinaga-Shum S, Stephenson SH, Hunt SC, Hopkins PN. G-substrate gene promoter SNP (-1323T>C) modifies plasma total cholesterol and triglyceride phenotype in familial hypercholesterolemia: Intra-familial association study in an eight-generation hyperlipidemic kindred. Geriatr Gerontol Int 2004. [DOI: 10.1111/j.1447-0594.2004.00126.x] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/27/2022]
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16
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Ishihara M, Iwasaki T, Nagano M, Ishii J, Takano M, Kujiraoka T, Tsuji M, Hattori H, Emi M. Functional impairment of two novel mutations detected in lipoprotein-associated phospholipase A2 (Lp-PLA2) deficiency patients. J Hum Genet 2004; 49:302-307. [PMID: 15148590 DOI: 10.1007/s10038-004-0151-6] [Citation(s) in RCA: 34] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/10/2004] [Accepted: 03/10/2004] [Indexed: 11/30/2022]
Abstract
Plasma lipoprotein-associated phospholipase A2 (Lp-PLA2), also known as platelet-activating factor (PAF) acetylhydrolase (PAF-AH), is a member of the serine-dependent class of A2 phospholipases that hydrolyze sn2-ester bonds of fragmented or oxidized phospholipids at sites where atherosclerotic plaques are forming. Most circulating Lp-PLA2 is bound to low-density lipoprotein (LDL) particles in plasma and the rest to high-density lipoprotein (HDL). Deficiency of Lp-PLA2 is a predisposing factor for cardiovascular diseases in the Japanese population. We describe here two novel mutations of the gene encoding Lp-PLA2, InsA191 and I317N in Japanese subjects. The first patient, with partial Lp-PLA2 deficiency, was heterozygous for the InsA191 mutation; macrophages from this patient secreted only half the normal amount of Lp-PLA2 in vitro. The other patient, who showed complete Lp-PLA2 deficiency, was a compound heterozygote for the novel I317N mutation and a common V279F mutation; macrophages from that patient failed to secrete any Lp-PLA2. Measurement of Lp-PLA2 mass, activity and Western blotting verified impaired production and secretion of the enzyme after transfection of mutant construct into COS-7 cells. These results indicated that both novel mutants, InsA191 and I317N, impair function of the Lp-PLA2 gene.
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Affiliation(s)
- Mitsuaki Ishihara
- Department of Advanced Medical Technology and Development, BML, Inc., Kawagoe, Japan
| | - Tadao Iwasaki
- Department of Advanced Medical Technology and Development, BML, Inc., Kawagoe, Japan
| | - Makoto Nagano
- Department of Advanced Medical Technology and Development, BML, Inc., Kawagoe, Japan
| | - Jun Ishii
- Internal Medicine, Hokkaido Hospital for Social Health Insurance, Sapporo, Japan
| | - Mayumi Takano
- Department of Advanced Medical Technology and Development, BML, Inc., Kawagoe, Japan
| | - Takeshi Kujiraoka
- Department of Advanced Medical Technology and Development, BML, Inc., Kawagoe, Japan
| | - Masahiro Tsuji
- Institute of Medical Science, Health Sciences University of Hokkaido, Sapporo, Japan
| | - Hiroaki Hattori
- Department of Advanced Medical Technology and Development, BML, Inc., Kawagoe, Japan
| | - Mitsuru Emi
- Department of Molecular Biology, Institute of Gerontology, Nippon Medical School, 1-396 Kosugi-cho, Nakahara-ku, Kawasaki 211-8533, Japan.
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17
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Sato K, Emi M, Ezura Y, Fujita Y, Takada D, Ishigami T, Umemura S, Xin Y, Wu LL, Larrinaga-Shum S, Stephenson SH, Hunt SC, Hopkins PN. Soluble epoxide hydrolase variant (Glu287Arg) modifies plasma total cholesterol and triglyceride phenotype in familial hypercholesterolemia: intrafamilial association study in an eight-generation hyperlipidemic kindred. J Hum Genet 2003; 49:29-34. [PMID: 14673705 DOI: 10.1007/s10038-003-0103-6] [Citation(s) in RCA: 60] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/25/2003] [Accepted: 10/16/2003] [Indexed: 10/26/2022]
Abstract
Plasma lipid and lipoprotein in general reflect the complex influences of multiple genetic loci, for instance, even familial hypercholesterolemia (FH), a representative example of monogenic hyperlipidemia, often presents with phenotypic heterogeneity. In the course of investigating familial coronary artery disease in Utah, we studied 160 members of an eight-generation extended family of FH in which 69 members were affected with type IIa hyperlipoproteinemia (HLPIIa; high plasma cholesterol) and ten with type IIb hyperlipoproteinemia (HLPIIb; high plasma cholesterol as well as plasma triglyceride). Soluble epoxide hydrolase ( EPHX2, sEH) plays a role in disposition of epoxides in plasma lipoprotein particles. Intrafamilial correlation analysis of the modifier effect of Glu287Arg substitution in the EPHX2 gene was carried out among 79 LDLR mutation carriers and 81 noncarriers. In the carriers, plasma cholesterol levels were elevated among carriers of the 287Arg allele (mean +/- SD=358 +/- 72 mg/dl) in comparison with 287Glu homozygotes (mean +/- SD=302 +/- 72 mg/dl) (p=0.0087). Similarly, in the LDLR mutation carriers, the plasma triglyceride levels were elevated among carriers of the 287Arg allele (mean +/- SD=260 +/- 100 mg/dl) in comparison with 287Glu homozygotes (mean +/- SD=169 +/- 83 mg/dl) (p=0.020). No such gene-interactive effect was observed among noncarriers of the LDLR mutation. Half of the patients who presented with HLPIIb had inherited a defective LDLR allele as well as an EPHX2-287Arg allele, whereas the majority who presented with HLPIIa had a defective LDLR allele but not an EPHX2-287Arg allele. These results indicate a significant modification of the phenotype of FH with defective LDLR allele by EPHX2-287Arg variation in our studied kindred.
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Affiliation(s)
- Keiko Sato
- Department of Molecular Biology, Institute of Gerontology, Nippon Medical School, 1-396, Kosugi-cho, Kawasaki 211-8533, Japan
- Department of Second Internal Medicine, Yokohama City University School of Medicine, Kanazawa-ku, Yokohama 236-0004, Japan
| | - Mitsuru Emi
- Department of Molecular Biology, Institute of Gerontology, Nippon Medical School, 1-396, Kosugi-cho, Kawasaki 211-8533, Japan.
- Department of Second Internal Medicine, Yokohama City University School of Medicine, Kanazawa-ku, Yokohama 236-0004, Japan.
| | - Yoichi Ezura
- Department of Molecular Biology, Institute of Gerontology, Nippon Medical School, 1-396, Kosugi-cho, Kawasaki 211-8533, Japan
| | - Yuko Fujita
- Department of Molecular Biology, Institute of Gerontology, Nippon Medical School, 1-396, Kosugi-cho, Kawasaki 211-8533, Japan
| | - Daisuke Takada
- Department of Molecular Biology, Institute of Gerontology, Nippon Medical School, 1-396, Kosugi-cho, Kawasaki 211-8533, Japan
| | - Tomoaki Ishigami
- Department of Second Internal Medicine, Yokohama City University School of Medicine, Kanazawa-ku, Yokohama 236-0004, Japan
| | - Satoshi Umemura
- Department of Second Internal Medicine, Yokohama City University School of Medicine, Kanazawa-ku, Yokohama 236-0004, Japan
| | - Yunpei Xin
- Cardiovascular Genetics Research Clinic, University of Utah Hearth Science Center, 410 Chipeta Way, Salt Lake City, UT, 84108, USA
| | - Lily L Wu
- Cardiovascular Genetics Research Clinic, University of Utah Hearth Science Center, 410 Chipeta Way, Salt Lake City, UT, 84108, USA
| | - Stacey Larrinaga-Shum
- Cardiovascular Genetics Research Clinic, University of Utah Hearth Science Center, 410 Chipeta Way, Salt Lake City, UT, 84108, USA
| | - Susan H Stephenson
- Cardiovascular Genetics Research Clinic, University of Utah Hearth Science Center, 410 Chipeta Way, Salt Lake City, UT, 84108, USA
| | - Steven C Hunt
- Cardiovascular Genetics Research Clinic, University of Utah Hearth Science Center, 410 Chipeta Way, Salt Lake City, UT, 84108, USA
| | - Paul N Hopkins
- Cardiovascular Genetics Research Clinic, University of Utah Hearth Science Center, 410 Chipeta Way, Salt Lake City, UT, 84108, USA
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18
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Fujita Y, Ezura Y, Emi M, Sato K, Takada D, Iino Y, Katayama Y, Takahashi K, Kamimura K, Bujo H, Saito Y. Hypercholesterolemia associated with splice-junction variation of inter-α-trypsin inhibitor heavy chain 4 (ITIH4) gene. J Hum Genet 2003; 49:24-28. [PMID: 14661079 DOI: 10.1007/s10038-003-0101-8] [Citation(s) in RCA: 25] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/20/2003] [Accepted: 10/15/2003] [Indexed: 11/26/2022]
Abstract
Factors predisposing to the phenotypic features of higher total cholesterol (T-Cho) have not been clearly defined. Here we report an association between a C/T single nucleotide polymorphism at IVS17+8 in the inter-alpha-trypsin inhibitor heavy chain 4 gene (ITIH4) and plasma total cholesterol levels in 351 adult individuals from an east-central area of Japan. Age and gender-adjusted levels of plasma T-Cho, LDL-cholesterol, triglyceride, and HDL-cholesterol were analyzed. When we separate the subjects into two genotypic groups regarding this single nucleotide polymorphism (SNP), those who lack the T-allele had significantly higher plasma T-Cho levels than the others who bear T-allele (mean 252.3 mg/dl versus 241.7 mg/dl; p=0.009). Of the 309 individuals without the T-allele, approximately 90% presented with hypercholesterolemia, whereas only 10% were hypercholesterolemic among 42 individuals with the T-allele (p <0.0001). These data suggest that genetic variation at ITIH4 locus is one of the likely candidate determinants for plasma cholesterol metabolisms.
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Affiliation(s)
- Yuko Fujita
- Department of Molecular Biology-Institute of Gerontology and Department of Internal Medicine II, Nippon Medical School, Kawasaki, Japan
| | - Yoichi Ezura
- Department of Molecular Biology-Institute of Gerontology and Department of Internal Medicine II, Nippon Medical School, Kawasaki, Japan
| | - Mitsuru Emi
- Department of Molecular Biology-Institute of Gerontology and Department of Internal Medicine II, Nippon Medical School, Kawasaki, Japan.
- Department of Molecular Biology-Institute of Gerontology, Nippon Medical School, 1-396 Kosugi-cho, Nakahara-ku, Kawasaki 211-8533, Japan.
| | - Keiko Sato
- Department of Molecular Biology-Institute of Gerontology and Department of Internal Medicine II, Nippon Medical School, Kawasaki, Japan
| | - Daisuke Takada
- Department of Molecular Biology-Institute of Gerontology and Department of Internal Medicine II, Nippon Medical School, Kawasaki, Japan
| | - Yasuhiko Iino
- Department of Molecular Biology-Institute of Gerontology and Department of Internal Medicine II, Nippon Medical School, Kawasaki, Japan
| | - Yasuo Katayama
- Department of Molecular Biology-Institute of Gerontology and Department of Internal Medicine II, Nippon Medical School, Kawasaki, Japan
| | | | | | - Hideaki Bujo
- Department of Genome Research and Clinical Application and Department of Clinical Cell Biology, Chiba University Graduate School, Chiba, Japan
| | - Yasushi Saito
- Department of Genome Research and Clinical Application and Department of Clinical Cell Biology, Chiba University Graduate School, Chiba, Japan
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