EFFECT OF A DOMINANT-NEGATIVE RAS ON MYOCARDIAL HYPERTROPHY BY USING ADENOVIRAL-MEDIATED GENE-TRANSFER

Background. The small guanosine triphosphate-binding protein ras regul ates a signal transduction cascade linking cell surface receptors to m itogen-activated protein kinase (MAPK). Because the molecular signalin g mechanisms underlying cardiac hypertrophy remain unclear, the curren t study examined the regulatory role of ras in both the biochemical an d morphologic aspects of hypertrophy. Methods. Adenoviral-mediated gen e transfer was used to express a dominant negative mutant of ras (rasN 17) at high efficiency in primary neonatal ventricular myocytes. Beta- galactosidase staining and Western blot analysis confirmed successful transfection and expression of the rasN17 gene product. MAPK activity was measured by an in vitro kinase assay resulting in radioactive Phos phorus labeled product. Morphologic hypertrophy was assessed by fluore scein-conjugated phalloidin. Results. Compared with uninfected or cont rol adenoviral-infected cells, myocytes infected with rasN17 demonstra ted attenuated basal MAPK activity. In contrast, rasN17 expression did not affect endothelin 1-induced MAPK activation. Morphologic studies showed that although rasN17 produced a phenotypic difference in the ba sal state, the ability of cardiac myocytes to morphologically respond to endothelin 1 stimulation, as manifested by sarcomeric reorganizatio n remained unaltered by the expression of the rasN17 gene product. Con clusions. Endothelin 1-stimulated MAPK activation and endothelin 1-ind uced morphologic hypertrophy are ras-independent processes.