Abstract 167 Cardiology: Mouse Models and Basic Mechanisms of Cardiac Disease Platform, Tuesday, 5/4

dHAND is a basic helix-loop-helix transcription factor that, in animal models, is expressed in the developing right ventricle and is required for right ventricular development. It has been proposed that one of its functions is to inhibit apoptosis in the myocytes of the developing right ventricle enabling this population of cardiomyocytes to proliferate and allowing proper growth of the right ventricular chamber. To identify genes regulated by dHAND which may be responsible for these developmental effects, the dHAND gene was transiently overexpressed in NIH-3T3 mouse embryonic fibroblasts. Twenty four hours after liposome-mediated (Lipofectamine, Gibco BRL) transfection, the RNA was isolated using the Trizol reagent. After conversion of the mRNA to cDNA, cDNAs from control vector-transfected NIH-3T3 cultures were subtracted from the cDNA from dHAND-transfected cultures using a PCR-based subtraction process. The resultant, differentially-expressed, cDNA transcripts were cloned and transferred to a membrane. Using the subtracted cDNA pools as probes, twenty four differentially-expressed cDNA clones were identified. Of the twenty clones that demonstrated increased expression in response to dHAND overexpression, ten encoded for the mouse L41 ribosomal binding protein. This protein binds to the ribosomal complex and has been demonstrated to prevent cyclohexamide-induced cytotoxicity. It has also been associated with substrate-specific activation of casien kinase II, an important regulator of calcium signaling in the cardiac myocyte. Therefore, dHAND expression in the developing right ventricle may, either directly or indirectly, regulate L41 expression which, in turn, stabilizes the ribosome complex and modulates intracellular calcium signaling. In other models, stabilization of the ribosomal complex has been associated with inhibition of apoptosis and calcium signaling is an important regulator of myocyte proliferation and hypertrophy. These findings suggest that L41 may be a important element in a dHAND-dependent regulatory pathway, increasing cardiocyte resistence to apoptosis and allowing appropriate myocyte proliferation.