Adenovirus E1A Inhibits Cardiac Myocyte-specific Gene Expression through Its Amino Terminus

Abstract

Adenovirus E1A oncoproteins inhibit muscle-specific gene expression and myogenic differentiation by suppressing the transcriptional activating functions of basic helix-loop-helix proteins. As one approach to identifying cardiac-specific gene regulatory proteins, we analyzed the functional regions of E1A proteins that are required for muscle gene repression in cardiac cells. Myocyte-specific promoters, including the ␣-actins and ␣-myosin heavy chain, were selectively and potently inhibited (>90%) by E1A, while the ubiquitously expressed ␤-actin promoter was only partially (ϳ30%) repressed; endogenous gene expression was also affected. Distinct E1A protein binding sites mediated repression of muscle-specific and ubiquitous actin promoters. E1A-mediated inhibition of ␤-actin required both an intact binding site for the tumor repressor proteins pRb and p107 and a second E1A domain (residues 15-35). In contrast, cardiac-specific promoter repression required the E1A amino-terminal residues 2-36. The proximal skeletal actin promoter (3 to base pair ؊153) was a target for repression by E1A. Although E1A binding to p300 was not required for inhibition of either promoter, co-expression of p300 partially reversed E1A-mediated transcriptional repression. We conclude that cardiac-specific and general promoter inhibition by E1A occurs by distinct mechanisms and that cardiac-specific gene expression is modulated by cellular factors interacting with the E1A p300/CBP-binding domain.

Hypoxia-activated apoptosis of cardiac myocytes requires reoxygenation or a pH shift and is independent of p53

“…The bar graphs indicate the percentage of apoptotic nuclei scored for each condition. Control aerobic cultures contained 5-7% apoptotic cells, similar to previous reports (51). This increased to 44% after 48 hours of hypoxia with metabolite buildup, and to 60% after 72 hours of hypoxia.…”

“…Cells were examined for morphological evidence of apoptosis or necro-sis after staining with the fluorescent DNA-binding dyes HOECHST 33342 and PI as described previously (51). Treated and control cell monolayers grown on uncoated Nunc 2-well coverslip dishes were rinsed with PBS, stained with 5 µg/mL HOECHST 33342 and 5 µg/mL PI for 15 minutes, and viewed ×400 on a Zeiss IM fluorescence microscope (Carl Zeiss Inc., Thornwood, New York, USA).…”

FOG-2 Competes with GATA-4 for Transcriptional Coactivator p300 and Represses Hypertrophic Responses in Cardiac Myocytes

“…In the hearts of these mice, the expression of cardiac muscle structural proteins, such as myosin heavy chain and ␣-actin, is clearly reduced. These findings are compatible with in vitro data showing that p300 is required for cardiac-specific transcription and for differentiation of cardiac myocytes (15)(16)(17)(18). p300 protein also serves as a cofactor of hypertrophy-responsive transcription factors such as MEF-2 and AP-1 as well as GATA-4 and is involved in the activation of the embryonic gene program during myocardial cell hypertrophy (11, 19 -21).…”

The Transcription Factors GATA4 and dHAND Physically Interact to Synergistically Activate Cardiac Gene Expression through a p300-dependent Mechanism

“…7C). The E1A protein was previously shown to down-regulate expression of muscle-specific genes, presumably by inhibiting p300 (42,43). In addition, overexpression of the p300 deletion mutant lacking the CH3 interaction domain (missing amino acids 1737-1836) blocked GATA4-dHAND functional synergy (Fig.…”