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Expression of Iron Regulatory Protein 1 Is Regulated not only by HIF-1 but also pCREB under Hypoxia
The inconsistent of responses of IRP1 and HIF-1 alpha to hypoxia and the similar tendencies in the changes of IRP1 and pCREB contents led us to hypothesize that pCREB might be involved in the regulation of IRP1 under hypoxia. Here, we investigated the role of pCREB in IRP1 expression in HepG2 cells...
Autores principales: | , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
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Ivyspring International Publisher
2016
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5069441/ https://www.ncbi.nlm.nih.gov/pubmed/27766034 http://dx.doi.org/10.7150/ijbs.16437 |
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author | Luo, Qian-Qian Qian, Zhong-Ming Zhou, Yu-Fu Zhang, Meng-Wan Wang, Dang Zhu, Li Ke, Ya |
author_facet | Luo, Qian-Qian Qian, Zhong-Ming Zhou, Yu-Fu Zhang, Meng-Wan Wang, Dang Zhu, Li Ke, Ya |
author_sort | Luo, Qian-Qian |
collection | PubMed |
description | The inconsistent of responses of IRP1 and HIF-1 alpha to hypoxia and the similar tendencies in the changes of IRP1 and pCREB contents led us to hypothesize that pCREB might be involved in the regulation of IRP1 under hypoxia. Here, we investigated the role of pCREB in IRP1 expression in HepG2 cells under hypoxia using quantitative PCR, western blot, immunofluorescence, electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP). We demonstrated that 1) Hypoxia increased pCREB levels inside of the nucleus; 2) Putative CREs were found in the IRP1 gene; 3) Nuclear extracts of HepG2 cells treated with hypoxia could bind to CRE1 and CRE3, and 100-fold competitor of putative CREs could abolish the binding activity to varying degrees; 4) pCREB was found in the CRE1 and CRE3 DNA-protein complexes of EMSA; 5) CRE1 and CRE3 binding activity of IRP1 depended on CREB activation but not on HIF-1; 6) Increased IRP1 expression under hypoxia could be prevented by LY294002; 7) ChIP assays demonstrated that pCREB binds to IRP1 promoter; and 8) HIF-1 and/or HIF-2 siRNA had no effect on the expression of pCREB and IRP1 proteins in cells treated with hypoxia for 8 hours. Our findings evidenced for the involvement of pCREB in IRP1 expression and revealed a dominant role of PI3K/Akt pathway in CREB activation under hypoxia and also suggested that dual-regulation of IRP1 expression by HIF-1 and pCERB or other transcription factor(s) under hypoxia might be a common mechanism in most if not all of hypoxia-inducible genes. |
format | Online Article Text |
id | pubmed-5069441 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2016 |
publisher | Ivyspring International Publisher |
record_format | MEDLINE/PubMed |
spelling | pubmed-50694412016-10-20 Expression of Iron Regulatory Protein 1 Is Regulated not only by HIF-1 but also pCREB under Hypoxia Luo, Qian-Qian Qian, Zhong-Ming Zhou, Yu-Fu Zhang, Meng-Wan Wang, Dang Zhu, Li Ke, Ya Int J Biol Sci Research Paper The inconsistent of responses of IRP1 and HIF-1 alpha to hypoxia and the similar tendencies in the changes of IRP1 and pCREB contents led us to hypothesize that pCREB might be involved in the regulation of IRP1 under hypoxia. Here, we investigated the role of pCREB in IRP1 expression in HepG2 cells under hypoxia using quantitative PCR, western blot, immunofluorescence, electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP). We demonstrated that 1) Hypoxia increased pCREB levels inside of the nucleus; 2) Putative CREs were found in the IRP1 gene; 3) Nuclear extracts of HepG2 cells treated with hypoxia could bind to CRE1 and CRE3, and 100-fold competitor of putative CREs could abolish the binding activity to varying degrees; 4) pCREB was found in the CRE1 and CRE3 DNA-protein complexes of EMSA; 5) CRE1 and CRE3 binding activity of IRP1 depended on CREB activation but not on HIF-1; 6) Increased IRP1 expression under hypoxia could be prevented by LY294002; 7) ChIP assays demonstrated that pCREB binds to IRP1 promoter; and 8) HIF-1 and/or HIF-2 siRNA had no effect on the expression of pCREB and IRP1 proteins in cells treated with hypoxia for 8 hours. Our findings evidenced for the involvement of pCREB in IRP1 expression and revealed a dominant role of PI3K/Akt pathway in CREB activation under hypoxia and also suggested that dual-regulation of IRP1 expression by HIF-1 and pCERB or other transcription factor(s) under hypoxia might be a common mechanism in most if not all of hypoxia-inducible genes. Ivyspring International Publisher 2016-09-15 /pmc/articles/PMC5069441/ /pubmed/27766034 http://dx.doi.org/10.7150/ijbs.16437 Text en © Ivyspring International Publisher. Reproduction is permitted for personal, noncommercial use, provided that the article is in whole, unmodified, and properly cited. See http://ivyspring.com/terms for terms and conditions. |
spellingShingle | Research Paper Luo, Qian-Qian Qian, Zhong-Ming Zhou, Yu-Fu Zhang, Meng-Wan Wang, Dang Zhu, Li Ke, Ya Expression of Iron Regulatory Protein 1 Is Regulated not only by HIF-1 but also pCREB under Hypoxia |
title | Expression of Iron Regulatory Protein 1 Is Regulated not only by HIF-1 but also pCREB under Hypoxia |
title_full | Expression of Iron Regulatory Protein 1 Is Regulated not only by HIF-1 but also pCREB under Hypoxia |
title_fullStr | Expression of Iron Regulatory Protein 1 Is Regulated not only by HIF-1 but also pCREB under Hypoxia |
title_full_unstemmed | Expression of Iron Regulatory Protein 1 Is Regulated not only by HIF-1 but also pCREB under Hypoxia |
title_short | Expression of Iron Regulatory Protein 1 Is Regulated not only by HIF-1 but also pCREB under Hypoxia |
title_sort | expression of iron regulatory protein 1 is regulated not only by hif-1 but also pcreb under hypoxia |
topic | Research Paper |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5069441/ https://www.ncbi.nlm.nih.gov/pubmed/27766034 http://dx.doi.org/10.7150/ijbs.16437 |
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