Prostate cancer is a disease with considerable biological heterogeneity and clinical aggressiveness. Although prostate cancer is prevalent among elderly population, only a small proportion of prostate cancer become invasive, metastatic and life-threatening. Thus, identifying genes and the underlying mechanisms that convert prostate cancer from a relatively indolent disease to an aggressive one hold the key for reducing the mortality of the disease. Recently, by using """"""""differential subtraction chain"""""""", we identified a novel gene named """"""""myopodin"""""""" that is frequently deleted in aggressive type of prostate cancer. Regardless of Gleason grade, deletions of myopodin, either partial or complete, correlate with high rate of invasion, metastasis or clinical relapse of prostate cancer. Sequence analysis indicates that myopodin shares significant homology with synaptopodin, a protein expressed in neurons and kidney podocytes and responsible for forming physiological cell-cell contact for these cells. Interestingly, over 90% of the partial deletions of myopodin are located in the region sharing homology with synaptopodin. Overexpression of myopodin in PC-3 or LNCaP cells reduced the invasiveness of these cell lines by 3 fold or more in matrigel traverse analysis, and by 10 fold in tumor growth in vivo. Overexpression of myopodin induces actin bundling and retards migration in PC-3. Yeast two-hybrid and co-immunoprecipitation analyses indicate that myopodin binds zyxin, a critical regulatory protein in actin cytoskeleton reorganization, cell motility and mitosis. In our proposal, we hypothesize that myopodin plays an important role in regulating cell motility, maintaining the integrity of normal cell-cell contacts of prostate epithelium, and inhibits the invasiveness of prostate cancer cells, and propose the following specific aims: 1) to overexpress myopodin in PC-3, Du145 and LNCaP cells and subsequently to test the changes of invasiveness of these cells through anchorage independent growth assay, matrigel traverse analysis, diaphragm invasion analysis, metastasis assay and metalloproteinase assays; 2) to define the motifs and minimal domain of myopodin essential for regulation of invasion by constructing arrays of deletion mutants and test their ability to regulate invasiveness; 3) to identify the functional significance of myopodin/zyxin interaction by focusing on cell motility and cytoskeleton re-organization related analysis on myopodin transformed cells; and 4) To identify potential effector genes that might mediate the biological function of myopodin through a comprehensive analysis of gene expression of myopodin expressing cells. ? ?
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