The Ras/Rac guanine nucleotide exchange factor mammalian son-of-sevenless interacts with PACSIN 1/syndapin I, a regulator of endocytosis and the actin cytoskeleton

Mammalian Son-of-sevenless (mSos) functions as a guanine nucleotide exchang e factor for Res and Rac, thus regulating signaling to mitogen-activated pr otein kinases and actin dynamics. In the current study, we have identified a new mSos-binding protein of 50 kDa (p50) that interacts with the mSos1 pr oline-rich domain. Mass spectrometry analysis and immunodepletion studies r eveal p50 as PACSIN 1/syndapin I, a Src homology 3 domain-containing protei n functioning in endocytosis and regulation of actin dynamics. In addition to PACSIN 1, which is neuron-specific, mSos also interacts with PACSIN 2, w hich is expressed in neuronal and nonneuronal tissues. PACSIN 2 shows enhan ced binding to the mSos proline-rich domain in pull-down assays from brain extracts as compared with lung extracts, suggesting a tissue-specific regul ation of the interaction. Proline to leucine mutations within the Src homol ogy 3 domains of PACSIN 1 and 2 abolish their binding to mSos, demonstratin g the specificity of the interactions. In situ, PACSIN 1 and mSos1 are co-e xpressed in growth cones and actin-rich filopodia in hippocampal and dorsal root ganglion neurons, and the two proteins coimmunoprecipitate from brain extracts. Moreover, epidermal growth factor treatment of COS-7 cells cause s co-localization of PACSIN 1 and mSos1 in actin-rich membrane ruffles, and their interaction is regulated through epidermal growth factor-stimulated mSos1 phosphorylation, These data suggest that PACSINs may function with mS os1 in regulation of actin dynamics.