Cardiac morphogenesis and myofibrillogenesis

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1. Mechanisms of myofibrillar protein isoform changes during developmentVarious combinations of two different myosin alkali light chain (LC) isoform cDNAs tagged with either VSV or mT antigen were co-exapressed in cultured cardiomyocytes from rat and chicken ventricles. Expressed isoproteins were detected using antibodies against the tags and their sorting patterns were analyzed by conforcal microscopy. The sorting specificity of LC isoforms to sarcomeres was shown to increase in the order of following developmental expression sequence : nonmuscle, slow skeletal muscle, ventricular, and fast skeletal muscle type. This suggests that during myofibrillogenesis isoforms with progressively higher binding affinities to myofibrils efficiently replace previous proteins, ensuring rapid exchange of isoforms without an extraordinary increase in protein synthesis. Expression of chimeric cDNAs revealed that the second EF-hand in the N-terminal lobe of each isoprotein is responsible for the isoform- … More specific sorting pattern. Although this part is not included in the key contact (s) with the myosin heavy chain, amino acid differences in this domain might affect the relative disposition of neighboring domains, thereby influencing the precise association of the key contact (s).2. Behavior of cardiac and skeletal muscle type isoproteins during myofibrillogenesisCardiac and skeletal (breast) muscle troponin Is (CTnI and FTnI,respectively) and their mutants (chimeric and truncated TnIs) were force expressed in cultured muscle cells to examine incorporation mechanism of TnI isoforms into myofibrils (Mfs). Since CTnI was not incorporated into breast Mfs, sorting mechanism of CTnI appeared to exist in breast MFs. Analysis of chimeric and truncated TnIs showed that TnIs were composed of two functionally distinct domains : C-terminal domain which binds to Mfs and N-terminal domain regulating the strength of binding affinity and specificity to cardiac and breast MFs.3. Morphogenesis and teratogenesis of the heartThe cardiac wall in embryos was investigated with the scanning electron microscope. The different surface characteristics of the four heterotypic cells that constitute the embryonic heart were demonstrated. The epicardial cells migrate from the mesothelium of the sinus venousus, encircled the ventricular wall, then extended cranically/caudally to unsheathe the entire heart usrface. Incidence of cardiac malformation was very high if retionic acid was administered into pregnant mice on 8ed day at the dose of 20 mg/kg body weight. Abnormalities observed are : TGA,DORV,VSD,A-loop and L-loop. Less