TYROSINE PHOSPHORYLATION REGULATES THE SH3-MEDIATED BINDING OF THE WISKOTT-ALDRICH-SYNDROME PROTEIN TO PSTPIP, A CYTOSKELETAL-ASSOCIATED PROTEIN

Wiskott-Aldrich syndrome is an X-linked hematopoietic disease that man ifests itself in platelet deficiency and a compromised immune system. Analysis of hematopoietic cells from affected individuals reveals that mutations in the Wiskott-Aldrich syndrome protein (WASP) result in st ructural and functional abnormalities in the cell cortex, consistent w ith the suggestion that WASP is involved with regulation of the actin- rich cortical cytoskeleton. Here we report that WASP interacts with a recently described cytoskeletal-associated protein, PSTPIP, a molecule that is related to the Schizosaccharomyces pombe cleavage furrow regu latory protein, CDC15p. This association is mediated by an interaction between the PSTPIP SH3 domain and two polyproline-rich regions in WAS P. Co-expression of PSTPIP with WASP in vivo results in a loss of WASP -induced actin bundling activity and co-localization of the two protei ns, which requires the PSTPIP SH3 domain. Analysis of tyrosine phospho rylation of PSTPIP reveals that two sites are modified in response to v-Src co-transfection or pervanadate incubation. One of these tyrosine s is found in the SH3 domain poly-proline recognition site, and mutati on of this tyrosine to aspartate or glutamate to mimic this phosphoryl ation state results in a loss of WASP binding in vitro and a dissoluti on of co-localization in vivo. In addition, PSTPIP that is tyrosine ph osphorylated in the SH3 domain interacts poorly with WASP in vitro. Th ese data suggest that the PSTPIP and WASP interaction is regulated by tyrosine phosphorylation of the PSTPIP SH3 domain, and this binding ev ent may control aspects of the actin cytoskeleton.