The Role of BetaPIX in RET-mediated Cell Migration
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RET is a transmembrane receptor that is implicated in a variety of processes such as cell proliferation, differentiation, migration, survival, and death. One of the proteins that is activated downstream of RET is the guanine nucleotide exchange factor (GEF), BetaPIX. BetaPIX removes GDP bound to inactive Cdc42/Rac, freeing a space for GTP-binding and transformation of Cdc42/Rac to the active GTPase state. BetaPIX is vital in cytoskeletal rearrangements and the regulation of focal adhesion complex assembly and disassembly. Although our lab has previously shown that RET and BetaPIX do not directly interact, the mechanism through which BetaPIX is activated downstream of RET is not yet known. In the current study, we confirmed that RET activation mediates BetaPIX phosphorylation. We showed that mutations in the SH3 and Dbl-homology domains of BetaPIX result in slower rates and in some cases impairment of wound healing and cell migration downstream of RET, indicating a role for BetaPIX in RET-mediated cell migration. In contrast, we have shown that GTPase mutants of Cdc42 and Rac were not sufficient to impair wound healing, but did result in less cell migration. This suggests that similar, yet distinct roles exist for Cdc42 and Rac in the regulation of cytoskeletal dynamics. We have shown that SH3 and Dbl-homology domain mutations in BetaPIX result in changes in cell morphology, suggesting a role of BetaPIX in the development of cell extensions downstream of RET. Alternatively, we have shown that GTPase mutants of Cdc42 or Rac alone do not induce cell morphology changes, further emphasizing the similar, yet distinct roles for Cdc42 and Rac in RET-mediated cell migration. Lastly, we have shown that BetaPIX is re-localized from the focal adhesion to the cytoplasm upon RET activation, suggesting that following focal adhesion complex formation, BetaPIX may be recycled.