Protein kinase C binding protein 1 inhibits hypoxia-inducible factor-1 in the heart
| Autor: | Ralph V. Shohet, Chrisy Mafnas, Allison L Williams, Cynthia D. Anderson, David R Veal, Chad B. Walton, Kathryn J Schunke |
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| Jazyk: | angličtina |
| Rok vydání: | 2018 |
| Předmět: |
0301 basic medicine
Transcription Genetic Physiology Immunoprecipitation Transgene Primary Cell Culture Myocardial Infarction Mice Transgenic 030204 cardiovascular system & hematology 03 medical and health sciences 0302 clinical medicine Species Specificity Physiology (medical) Protein Kinase C-Binding Protein 1 Animals Humans Myocytes Cardiac Hypoxia Transcription factor Protein Kinase C Gene knockdown Chemistry Tumor Suppressor Proteins Binding protein Original Articles DNA Methylation Hypoxia-Inducible Factor 1 alpha Subunit Cell Hypoxia Chromatin Cell biology Mice Inbred C57BL Disease Models Animal HEK293 Cells 030104 developmental biology Gene Expression Regulation Hypoxia-Inducible Factor 1 Cardiology and Cardiovascular Medicine Chromatin immunoprecipitation Signal Transduction |
| Zdroj: | Cardiovasc Res |
| Popis: | Aims Hypoxia-inducible factor-1 alpha (HIF-1α) is a key transcription factor responsible for the induction of genes that facilitate adaptation to hypoxia. To study HIF-1 signalling in the heart, we developed a mouse model in which an oxygen-stable form of HIF-1α can be inducibly expressed in cardiac myocytes, under the regulation of tetracycline. Methods and results Remarkably, expression of the transgene in mice generated two distinct phenotypes. One was the expected expression of HIF-regulated transcripts and associated changes in cardiac angiogenesis and contractility. The other was an unresponsive phenotype with much less expression of typical HIF-response genes and substantial expression of a zinc-finger protein, Protein Kinase C Binding Protein 1 (PRKCBP1). We have demonstrated that this second phenotype is due to an insertion of a fragment of DNA upstream of the PRKCBP1 gene that contains two additional canonical HIF binding sites and leads to substantial HIF binding, assessed by chromatin immunoprecipitation, and transcriptional activation. This insertion is found only in the FVB strain of mice that contributed the αMHC-tet binding protein transgene to these biallelic mice. In HEK293 cells transfected with oxygen-stable HIF-1α and PRKCBP1, we demonstrated inhibition of HIF-1 activity by a luciferase reporter assay. Using mouse primary cells and cell lines, we show that transfection with oxygen-stable HIF-1α and PRKCBP1 reduced expression of direct HIF-1 gene targets and that knockdown of PRKCBP1 removes that negative inhibition. Consistent with previous reports suggesting that PRKCBP1 modulates the chromatin landscape, we found that HL-1 cells transfected with oxygen-stable HIF-1α and PRKCBP1 have reduced global 5-methyl cytosine compared to HIF-1 alone. Conclusion We show genetic, transcriptional, biochemical, and physiological evidence that PRKCBP1 inhibits HIF activity. Identification of a new oxygen-dependent and previously unsuspected regulator of HIF may provide a target for new therapeutic approaches to ischaemic heart disease. |
| Databáze: | OpenAIRE |
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