BRUSH-BORDER MYOSIN-I MICROINJECTED INTO CULTURED-CELLS IS TARGETED TO ACTIN-CONTAINING SURFACE-STRUCTURES

The isolated intestinal microvillus cytoskeleton (core) consists of fo ur major proteins: actin, villin, fimbrin and brush border myosin-I. T hese proteins can assemble in vitro into structures resembling native microvillus cores. Of these components, villin and brush border myosin -I show tissue-specific expression, so they may be involved in the mor phogenesis of intestinal microvilli. When introduced into cultured cel ls that normally lack the protein, villin induces a reorganization of the actin filaments to generate large surface microvilli. Here we exam ine the consequences of microinjecting brush border myosin-I either al one or together with villin into cultured fibroblasts. Injection of br ush border myosin-I has no discernible effect on the overall morpholog y of the cells, but does become localized to either normal or villin-i nduced microvilli and other surface structures containing an actin cyt oskeleton. Since some endogenous myosin-Is have been found associated with cytoplasmic vesicles, these results show that brush border myosin -I has a domain that specifically targets it to the plasma membrane in both intestinal and cultured cell systems. Ultrastructural examinatio n of microvilli on control cultured cells revealed that they contain a far more highly ordered bundle of microfilaments than had been previo usly appreciated. The actin filaments in microvilli of villin-injected cells appeared to be more tightly cross-linked when examined by thin- section electron microscopy. In intestinal microvilli, the core bundle is separated from the plasma membrane by about 30 nm due to the prese nce of brush border myosin-I. As this same 30 nm separation is seen in microvilli of control cells and we see more prominent links in the my osin-I-injected cells, it seems likely that cultured cells contain an immunologically distinct, but functionally similar, myosin-I that link s the microfilament core bundle to the plasma membrane.