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A New Dimension to Ras Function: A Novel Role for Nucleotide-Free Ras in Class II Phosphatidylinositol 3-Kinase Beta (PI3KC2 beta) Regulation

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posted on 06.12.2013, 00:00 by Katy A. Wong, Angela Russo, Xuerong Wang, Yun-Ju Chen, Arnon Lavie, John P. O’Bryan
The intersectin 1 (ITSN1) scaffold stimulates Ras activation on endocytic vesicles without activating classic Ras effectors. The identification of Class II phosphatidylinositol 3-kinase beta, PI3KC2 beta, as an ITSN1 target on vesicles and the presence of a Ras binding domain (RBD) in PI3KC2 beta suggests a role for Ras in PI3KC2 beta activation. Here, we demonstrate that nucleotide-free Ras negatively regulates PI3KC2 beta activity. PI3KC2 beta preferentially interacts in vivo with dominant-negative (DN) Ras, which possesses a low affinity for nucleotides. PI3KC2 beta interaction with DN Ras is disrupted by switch 1 domain mutations in Ras as well as RBD mutations in PI3KC2 beta. Using purified proteins, we demonstrate that the PI3KC2 beta-RBD directly binds nucleotide-free Ras in vitro and that this interaction is not disrupted by nucleotide addition. Finally, nucleotide-free Ras but not GTP-loaded Ras inhibits PI3KC2 beta lipid kinase activity in vitro. Our findings indicate that PI3KC2 beta interacts with and is regulated by nucleotide-free Ras. These data suggest a novel role for nucleotide-free Ras in cell signaling in which PI3KC2 beta stabilizes nucleotide-free Ras and that interaction of Ras and PI3KC2 beta mutually inhibit one another.

Funding

These studies were supported by grants to J.P.O from the United States National Institutes of Health (HL090651), Department of Defense (PR080428), the Foundation Jerome Lejeune, and the St. Baldrick’s Foundation. K.A.W. was supported by a grant from the Foundation Jerome Lejeune.

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Publisher Statement

© 2012 Wong et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. The original version is available through Public Library of Science at DOI: 10.1371/journal.pone.0045360

Citation

Wong KA, Russo A, Wang XR, Chen YJ, Lavie A, O'Bryan JP. A New Dimension to Ras Function: A Novel Role for Nucleotide-Free Ras in Class II Phosphatidylinositol 3-Kinase Beta (PI3KC2 beta) Regulation. Plos One. Sep 2012;7(9). DOI: 10.1371/journal.pone.0045360

Publisher

Public Library of Science

Language

en_US

issn

1932-6203

Issue date

01/09/2012

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