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.The vertebrate heart, which initially forms as a linear, tubular structure, undergoes a complex series of morphogenetic movements to form a multichambered organ. The low-pressure atrial and high-pressure ventricular chambers which are formed differ in morphology, electrophysiology, and the repertoire of muscle contractile protein genes which are expressed (8,30). The developmental and molecular mechanisms responsible for establishing and maintaining chamber-specific differences are largely unknown.In adult animals, several members of contractile protein gene families show chamber-restricted or strong chamber-preferential expression. The myosin heavy-chain (MyHC) AMHC1 and slow MyHC 3 genes (56, 60), the myosin light-chain 1a (MLC-1a) and MLC-2a genes (21,31), and the atrial natriuretic factor (ANF) gene (61) show atrial specificity. In small mammals, the ␣-MyHC gene exhibits an atrial chamber preference of expression before birth (31). Conversely, MLC-2v

The quail slow myosin heavy chain 3 (slow MyHC 3) gene is expressed in the developing heart and in slow muscles of the developing limb. It is first expressed in the pulsatile cardiac tube in the embryo, and as the heart chamberizes its expression becomes restricted to the atria. To identify regulatory elements responsible for atrial-specific expression, the 5 upstream region of slow MyHC 3 gene was investigated. An atrial regulatory domain (ARD1) between ؊840 and ؊680 acts as an atrial cell-specific enhancer in primary cardiocyte cultures. ARD1 also specifies atrial-specific expression in vivo when the ARD1/heterologous promoter was introduced into developing chick embryos by a replication-competent retroviral vector. ARD1 is the first atrial cell-specific enhancer to be identified. Fine deletion and mutation analysis within ARD1 defined a 40-base pair vitamin D 3 receptor-like element that controls atrial cell-specific expression of the slow MyHC 3 gene by inhibiting its expression in ventricular cardiocytes.