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Roles of proline‐rich regions of the actin patch protein App1p in its phosphatidate phosphatase activity and membrane localization
Author(s) -
FAKAS STYLIANOS,
CARMAN GEORGE
Publication year - 2013
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.27.1_supplement.585.8
Subject(s) - phosphatidate , diacylglycerol kinase , phosphatase , actin , microbiology and biotechnology , biochemistry , mutant , biology , chemistry , actin binding protein , enzyme , actin cytoskeleton , cytoskeleton , gene , protein kinase c , cell
Actin patch protein App1p is a component of cortical actin patches. It has been recently identified as a Mg 2+ ‐dependent phosphatidate phosphatase (PAP) that catalyzes the conversion of phosphatidate (PA) to diacylglycerol (DAG) and Pi. The protein contains a canonical D X D X (T/V) catalytic motif that is typical of Mg 2+ ‐dependent phosphatase enzymes and several proline‐rich regions (P XX P motifs). These proline‐rich regions are known to interact with Src homology 3 (SH3) domains that are characteristic of actin patch proteins (e.g., Abp1p and Rvs167p) involved in endocytosis. In fact, App1p has been shown to physically interact with several actin patch proteins. The fact that App1p possesses PAP activity indicates that it may regulate the local concentrations of PA and DAG found in cortical actin patches. PA and DAG are known to facilitate membrane fission/fusion events in model systems, and they are also known to interact with and regulate enzymes that play important roles in vesicular trafficking. In this work, the proline residues in the P XX P motifs in App1p were mutated to glycine by site‐specific mutagenesis. The mutant App1p enzymes were expressed in app1 Δ and app1 Δ pah1 Δ dpp1 Δ lpp1 Δ quadruple mutant cells to assess the roles of P XX P motifs in the interaction with actin patch proteins, PAP activity, and the association of App1p with membranes. Supported by NIH grant GM‐28140.