Cardiac ankyrin repeat protein attenuates cardiomyocyte apoptosis by upregulation of Bcl-2 expression

Biochim Biophys Acta. 2016 Dec;1863(12):3040-3049. doi: 10.1016/j.bbamcr.2016.09.024. Epub 2016 Oct 3.

Abstract

Cardiac ankyrin repeat protein (CARP) is a nuclear transcriptional co-factor that has additional functions in the myoplasm as a component of the muscle sarcomere. Previous studies have demonstrated increased expression of CARP in cardiovascular diseases, however, its role in cardiomyocyte apoptosis is unclear and controversial. In the present study, we investigated possible roles of CARP in hypoxia/reoxygenation (H/R) -induced cardiomyocyte apoptosis and the underlying mechanisms. Neonatal mouse ventricular cardiomyocytes were isolated and infected with adenovirus encoding Flag-tagged CARP (Ad-CARP) and lentivirus encoding CARP targeted shRNA (sh-CARP), respectively. Cardiomyocyte apoptosis induced by exposure to H/R conditions was evaluated by TUNEL staining and western blot analysis of cleaved caspase-3. The results showed that H/R-induced apoptosis was significantly decreased in Ad-CARP cardiomyocytes and increased in sh-CARP cardiomyocytes, suggesting a protective anti-apoptosis role for CARP. Interestingly, over-expressed CARP was mainly distributed in the nucleus, consistent with its role in regulating transcriptional activity. qPCR analysis showed that Bcl-2 transcripts were significantly increased in Ad-CARP cardiomyocytes. ChIP and co-IP assays confirmed the binding of CARP to the Bcl-2 promoter through interaction with transcription factor GATA4. Collectively, our results suggest that CARP can protect against H/R induced cardiomyocyte apoptosis, possibly through increasing anti-apoptosis Bcl-2 gene expression.

Keywords: Apoptosis; Bcl-2 family; Cardiac ankyrin repeat protein; Cardiomyocyte; Hypoxia/reoxygenation; Transcription co-factor.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenoviridae / genetics
  • Adenoviridae / metabolism
  • Animals
  • Animals, Newborn
  • Apoptosis
  • Caspase 3 / genetics
  • Caspase 3 / metabolism
  • Cell Nucleus / metabolism
  • GATA4 Transcription Factor / genetics
  • GATA4 Transcription Factor / metabolism
  • Gene Expression Regulation
  • Genetic Vectors / chemistry
  • Genetic Vectors / metabolism
  • Lentivirus / genetics
  • Lentivirus / metabolism
  • Male
  • Mice
  • Mice, Inbred C57BL
  • Muscle Proteins / antagonists & inhibitors
  • Muscle Proteins / genetics*
  • Muscle Proteins / metabolism
  • Myocardial Ischemia / genetics*
  • Myocardial Ischemia / metabolism
  • Myocardial Ischemia / pathology
  • Myocytes, Cardiac / metabolism*
  • Myocytes, Cardiac / pathology
  • Nuclear Proteins / antagonists & inhibitors
  • Nuclear Proteins / genetics*
  • Nuclear Proteins / metabolism
  • Primary Cell Culture
  • Promoter Regions, Genetic
  • Protein Binding
  • Proto-Oncogene Proteins c-bcl-2 / agonists
  • Proto-Oncogene Proteins c-bcl-2 / genetics*
  • Proto-Oncogene Proteins c-bcl-2 / metabolism
  • RNA, Small Interfering / genetics
  • RNA, Small Interfering / metabolism
  • Reperfusion Injury / genetics*
  • Reperfusion Injury / metabolism
  • Reperfusion Injury / pathology
  • Repressor Proteins / antagonists & inhibitors
  • Repressor Proteins / genetics*
  • Repressor Proteins / metabolism
  • Signal Transduction
  • Transcription, Genetic

Substances

  • Ankrd1 protein, mouse
  • GATA4 Transcription Factor
  • Gata4 protein, mouse
  • Muscle Proteins
  • Nuclear Proteins
  • Proto-Oncogene Proteins c-bcl-2
  • RNA, Small Interfering
  • Repressor Proteins
  • Bcl2 protein, mouse
  • Casp3 protein, mouse
  • Caspase 3