Neuronal specific ßPix-b stimulates actin-dependent processes via the interaction between its PRD and WH1 domain of N-WASP.

ABSTRACT

ßPix, a Pak-interacting nucleotide exchange factor (Cool-1/p85SPR), is a Cdc42/Rac1-specific guanine nucleotide exchange factor (GEF) involved in various actin-related processes. Many previous studies have focused on ubiquitously expressed ßPix-a, while the role of the neuronal-specific isoform ßPix-b is still unknown, especially whether its role is distinct from or similar to ßPix-a. Here we show that unlike ßPix-a, overexpression of ßPix-b stimulates actin-dependent comet formation in BHK21 cells. This effect is attributed to the interaction between its proline-rich domain (PRD) and the WH1 domain of N-WASP. In addition, we show that overexpression of ßPix-b stimulates actin-dependent dendritic spine formation in rat hippocampal neurons in culture, a formation that is blocked by co-expression of the WH1 domain of N-WASP or the PRD of ßPix-b. Knocking-down endogenous expression of ßPix-b by shRNA reduced the number of dendritic spines, which were rescued only by PRD-containing ßPix-b mutants. GEF activity of ßPix-b is also required for these effects. The results show that neuronal-specific ßPix-b stimulates actin-dependent processes in cells via the interaction between its PRD and the WH1 domain of N-WASP. Our results identify N-WASP as the first protein shown to interact with the PRD of ßPix-b, raising the possibility that, as an N-WASP WH1-binding protein, ßPix-b may regulate N-WASP's activity in cells.