) One of the most foreboding aspects of cancer is its spread, or metastasis, to different tissues throughout the body. This greatly complicates its treatment which might otherwise be cured by surgery. Metastasis occurs when cells break away from the primary tumor, invade adjacent tissues in order to enter the blood stream, later escape from the blood stream (extravasion) and implant into unaffected tissues to seed new tumors. The process of invasion, extravasion and implantation depend on the cells ability to move across and through different tissues. It is essential to understand how cell movements are controlled if we are to develop strategies to prevent and treat cancer metastasis. It is difficult to directly study cell movement in metastasizing tumors in situ. To circumvent this difficulty yet examine cell movements within tissues, the research outlined in this proposal is aimed at understanding how cell movements are controlled using a model system, embryos of the frog, Xenopus laevis, in which cells normally undergo different types of movement in a tissue environment. We have shown that in Xenopus embryos inhibition of PDGF signaling prevents mesoderm cell movement (1). When PDGF binds to its receptor (PDGFR) autophosphorylation of specific tyrosine residues in the receptors intracellular domain generates high affinity binding sites for signaling molecules including RasGAP, Shp-2, c-Src, P13K and PLCy (2).
Three Specific aims are proposed to dissect PDGP signaling in mesoderm cell movement.
Aim 1 : PDGFRs, designed to selectively bind specific signaling molecules, will be used to identify which signaling molecules regulate the movement of cultured mesoderm cells. Time-lapse micrography, confocal microscopy and Boyden chamber assays will be used to monitor cell movement, cell morphology and chemotaxis.
Aim 2 : Once key signaling molecules have been identified in vitro, we will address how these molecules regulate cell movement within the embryo where additional signals from cell adhesion molecules, matrix components and growth factors may modulate PDGP-signaling events.
Aim 3 : PDGF through P13K causes mesoderm cells to spread and undergo rearrangements of the actin cytoskeleton on a fibronectin matrix (3). In other cell types, PDGF signaling through P13K is known to elicit rearrangements of the actin cytoskeleton through Rho small GTP-binding proteins (4). We will determine the role of Rho small GTP-binding proteins in mesoderm cell movement. The cells response to constitutively active or dominant versions of these proteins will be analyzed by time-lapse micrography and confocal microscopy.
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