JHIBRG:Abstract June 06 2007

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Identification of Nedd4 as a βPix binding protein and Studies on Ubiquitination of βPix

βPix, a Pak-interacting exchange factor is known as a GEF for Rac/Cdc42. In this study, I present evidence that Nedd4, a HECT domain-containing ubiquitin ligase, is a novel binding protein to the Proline-rich (PxxP) domain of βPix. Interaction of Nedd4 with βPix required both the C2 and WW domains of Nedd4, and this interaction was not observed with βPix-c which lacks the PxxP domain. Interestingly, the E3 ligase inactive Nedd4 mutant co-immunoprecipited more efficiently with βPix. This result suggests that the interaction of βPix with Nedd4 may be transient in the cells. I also found that Nedd4 mediates the ubiquitination of βPix. Accumulation of the ubiquitinated βPix by proteasome inhibitor, MG132, treatment suggests that ubiquitinated βPix is destined to proteasomal degradation. Nedd4-induced ubiquitination destabilized βPix-b which contains a PEST motif, while the turnover of βPix-a which lacks this motif was not affected. On the other hand, the cellular localization of βPix-a was affected by Nedd4. E3 ligase activity was not crucial for the change in βPix-a localization, implying a role of Nedd4 in targeting βPix. In this study, the Nedd4-mediated ubiquitination of βPix was demonstrated for the first time. Evidences presented here also suggest that ubiquitination of βPix may play a role in the regulation of the stability and/or cellular localization of βPix.


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Seyun Kim, Seung-Hye Lee, and Dongeun Park. (2000), Leucine zipper-mediated homodimerization of the p21-activated kinase-interacting factor, beta Pix. Implication for a role in cytoskeletal reorganization. J Biol. Chem. 276(14):10581-4.