Hydrogen peroxide depletes phosphatidylinositol-3-phosphate from endosomes in a p38 MAPK-dependent manner and perturbs endocytosis.

Phosphatidylinositol-3-phosphate (PI3P) is a lipid that is enriched specifically in early endosomes. Given that early endosomes containing PI3P act as a microdomain to recruit proteins that contain a PI3P-binding domain (FYVE domain), the equilibrium between the production and degradation of PI3P influences a variety of processes, including endocytosis and signal transduction via endosomes. In the study reported herein, we have developed a novel analytical method to quantify the amount of PI3P in endosomes by introducing a GST-2xFYVE protein probe into semi-intact cells. The GST-2xFYVE probe was targeted specifically to intracellular PI3P-containing endosomes, which retained their small punctate structure, and allowed the semi-quantitative measurement of intracellular PI3P. Using the method, we found that treatment of HeLa cells with H(2)O(2) decreased the amount of PI3P in endosomes in a p38 MAPK-dependent manner. In addition, H(2)O(2) treatment delayed transport through various endocytic pathways, especially post-early endosome transport; the retrograde transport of cholera toxin was especially dependent on the amount of PI3P in endosomes. This article is part of a Special Issue entitled: 11th European Symposium on Calcium.