A Positive GATA Element and a Negative Vitamin D Receptor-Like Element Control Atrial Chamber-Specific Expression of a Slow Myosin Heavy-Chain Gene during Cardiac Morphogenesis

ABSTRACT

We have used the slow myosin heavy chain (MyHC) 3 gene to study the molecular mechanisms that control atrial chamber-specific gene expression. Initially, slow MyHC 3 is uniformly expressed throughout the tubular heart of the quail embryo. As cardiac development proceeds, an anterior-posterior gradient of slow MyHC 3 expression develops, culminating in atrial chamber-restricted expression of this gene following chamberization. Two cis elements within the slow MyHC 3 gene promoter, a GATA-binding motif and a vitamin D receptor (VDR)-like binding motif, control chamber-specific expression. The GATA element of the slow MyHC 3 is sufficient for expression of a heterologous reporter gene in both atrial and ventricular cardiomyocytes, and expression of GATA-4, but not Nkx2-5 or myocyte enhancer factor 2C, activates reporter gene expression in fibroblasts. Equivalent levels of GATA-binding activity were found in extracts of atrial and ventricular cardiomyocytes from embryonic chamberized hearts. These observations suggest that GATA factors positively regulate slow MyHC 3 gene expression throughout the tubular heart and subsequently in the atria. In contrast, an inhibitory activity, operating through the VDR-like element, increased in ventricular cardiomyocytes during the transition of the heart from a tubular to a chambered structure. Overexpression of the VDR, acting via the VDR-like element, duplicates the inhibitory activity in ventricular but not in atrial cardiomyocytes. These data suggest that atrial chamber-specific expression of the slow MyHC 3 gene is achieved through the VDR-like inhibitory element in ventricular cardiomyocytes at the time distinct atrial and ventricular chambers form.

ACKNOWLEDGMENTS

We are grateful to Everet Bandman for providing us with the NA8 monoclonal antibody, David B. Wilson for GATA antiserum and the pMT2-GATA-4 expression vector, Eric Olson for the MEF2C expression vector, Robert Schwartz for the Nkx2-5 expression vector, David Feldman for the VDR expression plasmid, Ronald Evans for the RARα and RXRα expression plasmids, Elizabeth Allegretto for rabbit polyclonal anti-RXRα and anti-RARα antibodies, and David G. Gardner for the hANF-CAT construct. Sandra Conlon provided excellent technical assistance, and Gordon Cann provided helpful discussions.