DOMINANT-NEGATIVE EFFECT OF CYTOPLASMIC ACTIN ISOPROTEINS ON CARDIOMYOCYTE CYTOARCHITECTURE AND FUNCTION

The intracompartmental sorting and functional consequences of ectopic expression of the six vertebrate actin isoforms was investigated in di fferent types of cultured cells. In transfected fibroblasts all isoact in species associated with the endogenous microfilament cytoskeleton, even though cytoplasmic actins also showed partial localization to per ipheral submembranous sites. Functional and structural studies were pe rformed in neonatal and adult rat cardiomyocytes. All the muscle isoac tin constructs sorted preferentially to sarcomeric sites and, to a les ser extent, also to stress-fiber-like structures. The expression of mu scle actins did not interfere with cell contractility, and did not dis turb the localization of endogenous sarcomeric proteins, In sharp cont rast, ectopic expression of the two cytoplasmic actin isoforms resulte d in rapid cessation of cellular contractions and induced severe morph ological alterations characterized by an exceptional outgrowth of filo podia and cell flattening. Quantitative analysis in neonatal cardiomyo cytes indicated that the levels of accumulation of the different isoac tins are very similar and cannot be responsible for the observed isopr oteins-specific effects. Structural analysis revealed a remodeling of the cytoarchitecture including a specific alteration of sarcomeric org anization; proteins constituting the sarcomeric thin filaments relocat ed to nonmyofibrillar sites while thick filaments and titin remained u naffected, Experiments with chimeric proteins strongly suggest that is oform specific residues in the carboxy-terminal portion of the cytopla smic actins are responsible for the dominant negative effects on funct ion and morphology.