The objective of this research program is to characterize the role of the proteinase inhibitor alpha2-macroglobulin (alpha2M) and the alpha2M receptor in the regulation of cellular growth. Alpha2M inhibits proteinases from all four major classes; however, the function of alpha2M remains unclear. Recent studies have demonstrated that alpha2M binds many cytokines including transforming growth factor-beta1 (TGF-beta1), TGF- beta2, interleukin-1beta and tumor necrosis factor-alpha. After reaction with proteinase, alpha2M undergoes conformational change and expresses altered cytokine binding affinity. Work from this laboratory demonstrated that the alpha2M receptor mediates cellular uptake of alpha2M-TGF-beta1 complexes. The significance of this process is unknown.
The first aim of this proposal is to study the formation and activity of alpha2M-cytokine complexes. Purified alpha2M-proteinase complexes and structurally modified alpha2M variants will be used to analyze the role of alpha2M conformation in cytokine binding. Anti-alpha2M monoclonal antibodies will also be applied in this analysis. Model systems (in vitro and in vivo) will be developed to determine the significance of alpha2M-cytokine interactions. Does alpha2M serve to clear cytokines from the plasma and related fluids or is alpha2M a carrier that targets active signalling molecules to specific cells? Experiments using cell culture and animal model systems will address this question and provide information regarding the role of alpha2M in biological processes such as hematopoiesis and neoplasia. The cytokine carrier activity of other proteins including thrombospondin will be evaluated.
A final aim will be to characterize the modulation of alpha2M receptor expression by cytokines including some that bind to alpha2M. Receptor expression will be determined using 125I-alpha2M-methylamine binding studies, a specific anti-alpha2M receptor monoclonal antibody, and a cDNA probe that hybridizes with murine or human alpha2M receptor mRNA. The goal of this program is to advance our understanding of the function of alpha2M and the alpha2M receptor in cellular growth, differentiation and neoplastic transformation. Ultimately, these studies will provide information regarding growth factor-carrier interactions in general.
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