The first DEP domain of the RhoGEF P-Rex1 autoinhibits activity and contributes to membrane binding

Ravala, Sandeep K.; Hopkins, Jesse B.; Plescia, Caroline B.; Allgood, Samantha R.; Kane, Madison A.; Cash, Jennifer N.; Stahelin, Robert V.; Tesmer, John G.

doi:10.1074/jbc.ra120.014534

Title: The first DEP domain of the RhoGEF P-Rex1 autoinhibits activity and contributes to membrane binding

Journal Article · Mon Jul 13 00:00:00 EDT 2020 · Journal of Biological Chemistry

DOI:https://doi.org/10.1074/jbc.ra120.014534· OSTI ID:1673789

Ravala, Sandeep K. ^[1]; Hopkins, Jesse B. ^[2]; Plescia, Caroline B. ^[1]; Allgood, Samantha R. ^[1]; Kane, Madison A. ^[3];

^[4];

^[1];

^[1]

Purdue Univ., West Lafayette, IN (United States)
Argonne National Lab. (ANL), Lemont, IL (United States)
California State Univ. (CalState), Long Beach, CA (United States)
Univ. of Michigan, Ann Arbor, MI (United States)

Phosphatidylinositol (3,4,5)-trisphosphate (PIP₃)-dependent Rac exchanger 1 (P-Rex1) catalyzes the exchange of GDP for GTP on Rac GTPases, thereby triggering changes in the actin cytoskeleton and in transcription. Its overexpression is highly correlated with the metastasis of certain cancers. P-Rex1 recruitment to the plasma membrane and its activity are regulated via interactions with heterotrimeric Gβγ subunits, PIP₃, and protein kinase A (PKA). Deletion analysis has further shown that domains C-terminal to its catalytic Dbl homology (DH) domain confer autoinhibition. Among these, the first dishevelled, Egl-10, and pleckstrin domain (DEP1) remains to be structurally characterized. DEP1 also harbors the primary PKA phosphorylation site, suggesting that an improved understanding of this region could substantially increase our knowledge of P-Rex1 signaling and open the door to new selective chemotherapeutics. Here we show that the DEP1 domain alone can autoinhibit activity in context of the DH/PH-DEP1 fragment of P-Rex1 and interacts with the DH/PH domains in solution. The 3.1 Å crystal structure of DEP1 features a domain swap, similar to that observed previously in the Dvl2 DEP domain, involving an exposed basic loop that contains the PKA site. Using purified proteins, we show that although DEP1 phosphorylation has no effect on the activity or solution conformation of the DH/PH-DEP1 fragment, it inhibits binding of the DEP1 domain to liposomes containing phosphatidic acid. Thus, we propose that PKA phosphorylation of the DEP1 domain hampers P-Rex1 binding to negatively charged membranes in cells, freeing the DEP1 domain to associate with and inhibit the DH/PH module.

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Research Organization:: Argonne National Lab. (ANL), Argonne, IL (United States)

Sponsoring Organization:: National Institutes of Health (NIH); USDOE

OSTI ID:: 1673789

Journal Information:: Journal of Biological Chemistry, Vol. 295, Issue 36; ISSN 0021-9258

Publisher:: American Society for Biochemistry and Molecular BiologyCopyright Statement

Country of Publication:: United States

Language:: ENGLISH

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