The guanine nucleotide exchange factor Trio activates the phagocyte NADPH oxidase in the absence of GDP to GTP exchange on Rac: "The emperor's new clothes"

Natalia Sigal, Yara Gorzalczany, Rive Sarfstein, Carolyn Weinbaum, Yi Zheng, Edgar Pick*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

Abstract

The superoxide-generating NADPH oxidase complex of phagocytes consists of a membrane-associated flavocytochrome b559 and four cytosolic components as follows: p47phox, p67phox, p40phox, and the small GTPase Rac (1 or 2). Activation of the oxidase is the result of assembly of the cytosolic components with cytochrome b559 and can be mimicked in vitro by mixtures of membrane and cytosolic components exposed to an anionic amphiphile, serving as activator. We reported that prenylation of Rac1 endows it with the ability to support oxidase activation in conjunction with p67phox but in the absence of amphiphile and p47phox. We now show the following 6 points. 1) The Rac guanine nucleotide exchange factor Trio markedly potentiates oxidase activation by prenylated Rac1-GDP. 2) This occurs in the absence of exogenous GTP or any other source of GTP generation, demonstrating that the effect of Trio does not involve GDP to GTP exchange on Rac1. 3) Trio does not potentiate oxidase activation by prenylated Rac1-GTP, by nonprenylated Rac1-GDP in the presence or absence of amphiphile, and by a prenylated [p67phox-Rac1] chimera in GDP-bound form. 4) Rac1 mutants defective in the ability to bind Trio or to respond to Trio by nucleotide exchange fail to respond to Trio by enhanced oxidase activation. 5) A Trio mutant with conserved Rac1-binding ability but lacking nucleotide exchange activity fails to enhance oxidase activation. 6) The effect of Trio is mimicked by displacement of Mg2+ from Rac1-GDP. These results reveal the existence of a novel mechanism of Rac activation by a guanine nucleotide exchange factor and suggest that the induction by Trio of a conformational change in Rac1, in the absence of nucleotide exchange, is sufficient for enhancing its effector function.

Original languageEnglish
Pages (from-to)4854-4861
Number of pages8
JournalJournal of Biological Chemistry
Volume278
Issue number7
DOIs
StatePublished - 14 Feb 2003

Funding

FundersFunder number
National Institute of General Medical SciencesR01GM060523

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