NM23s are members of a family of homologous proteins with key roles in the regulation of cell function. Alterations in the expression levels of modifications in the structure of NM23 affect profoundly processes such as growth, development, tumor progression, metastasis suppression and apoptosis. We found that NM23 also participates in the G protein-coupled activation of K+ channels by muscarinic receptors by an unknown mechanism that does not require its NDP kinase activity, suggesting that another function of NM23 is involved in this process of signal transduction. The overall objective of this proposal is to understand the molecular mechanism by which NM23 contributes to the muscarinic K+ channel pathway in particular and to signal transduction in general. The experimental approach uses cells with a known deficiency in NM23 as ell as cells carrying point mutants of NM23. Channels and muscarinic receptors will be expressed in these cells and their function will be assayed by patch clamp methods, to detect effects of reduced expression of NM23 and of mutations that affect differentially NM23 activities. Since both NM23 and G protein- coupled K+ channel subunits can associate with vimentin intermediate filaments, we will examine the hypothesis that NM23 acts to stabilize intermediate filaments during, receptor activation, and thus helps to preserve the structure of macromolecular assemblies necessary for efficient transmembrane signalling. We expect that results from these experiments will allow us to define the molecular mechanisms by which alterations in NM23 modify muscarinic K+ channel function. Furthermore, given the known connection of both NM23 and vimentin intermediate filaments to cell motility and tumor invasiveness, the work proposed may provide new insight into the signal transduction pathways that modulate cytoskeletal organization in tumor cells and are central to cell migration during metastatic spread.
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