[Malignant transformation and alteration of actin-related proteins]

Numerous genes and their products are involved in the expression of the transformed phenotype. Chemical carcinogens transform cells by altering the structure and function of these macromolecules. Some of them have been identified as oncogenes and oncogene products whereas most of the rest are unknown. Since the expressions of transformed phenotype such as alteration of cell morphology, motility and growth are closely correlated to the alteration of cytoskeletal structures, we have investigated the genetic and post-translational alterations of actin and actin-binding proteins in transformed cells. We have found that the expression of a point-mutated beta-actin correlates with the expression of transformed phenotype in one of these chemically transformed human cell lines. Mutated beta-actin showed reduced ability for self-polymerization in vitro, which coincides with the observation in vivo in reduced incorporation of the mutated actin into the cytoskeletal fraction. On the other hand, actin-binding proteins, caldesmon and calspectin which also bind to calmodulin, and 36K protein which also binds to calspectin, were examined using antibodies specific for each protein. Their binding to counterproteins was dependent on the Ca++ concentration. Compared with the untransformed NIH3T3 cells, the NIH3T3 cells transformed by various oncogenes showed considerably smaller amounts of the three proteins and an increase in their phosphorylated forms. The degradation rate of caldesmon and calspectin did not differ between untransformed and transformed cells. Phosphoamino acid analysis showed that the serine residue was the major site for phosphorylation in calspectin whereas calspectin was a good substrate in vitro for both src tyrosine protein kinase and protein kinase C, while 36K protein was phosphorylated at the tyrosine residue as well as the serine residue. The addition of tumor promoter to cultured cells also caused changes in actin-binding proteins simultaneously with changes in morphology, motility and microfilament structure and with the induction of transcription of the beta-actin gene. The role of actin and actin-binding proteins in the cascade reactions induced by oncogene products was discussed.