Mts1: A Molecular Link Between the Cytoskeleton and Breast Tumor Metastasis.

摘要

The objectives of this grant are: (1) examine how mts1 expression alters directed cellular motility in vitro; (2) generate myosin-IIA antibodies that mimic mts1 binding to examine how the regulation of myosin-IIA affects directed motility in vitro; and (3) utilize an intravital imaging system to evaluate the impact of mts1 expression on metastasis in live animal models. We have established MTC cell lines with high, reliable and heritable mts1 expression and have made significant progress on our in vitro analysis evaluating the effects of mts1 expression on chemoallractant-stimulated motility. In conjunction with these in vitro analyses, our intravital imaging studies, which visualize the motile behavior of mts1 expressing tumor cells within the primary tumor in situ, are in progress. These combined analyses Nill allow us to obtain a comprehensive behavioral phenotype and will identify which aspects of directed motility are sensitive to the expression of mts1. In addition, we identified 16 residues in the myosin-IIA heavy chain that comprise the mtsi binding site and have generated a rabbit polyclonal antibody to the mts1 binding site on the myosin-IIA heavy chain. Immunoblot and immunoprecipitation analyses indicate that the mts1 binding site antibody specifically reacts with nonmuscle myosin-IIA. In vitro assays demonstrate that the mts1 binding site antibody has the same affect on the assembly and actin- activated ATPase activity of myosin-IIA as ntsl. In vivo studies are now direcuy testing if alterations in the directed motility of MTO-mts1 cells are due to the regulation of myosin-IIA function by mts1.