αPIX nucleotide exchange factor is activated by interaction with phosphatidylinositol 3-kinase

Pathogens can alter epithelial polarity by recruiting polarity proteins to the apical membrane, but how a change in protein localization is linked to polarity disruption is not clear. In this study, we used chemically induced dimerization to rapidly relocalize proteins from the cytosol to the apical surface. We demonstrate that forced apical localization of Par3, which is normally restricted to tight junctions, is sufficient to alter apical membrane identity through its interactions with phosphatidylinositol 3-kinase (PI3K) and the Rac1 guanine nucleotide exchange factor Tiam1. We further show that PI3K activity is required upstream of Rac1, and that simultaneously targeting PI3K and Tiam1 to the apical membrane has a synergistic effect on membrane remodeling. Thus, Par3 coordinates the action of PI3K and Tiam1 to define membrane identity, revealing a signaling mechanism that can be exploited by human mucosal pathogens.

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Insulin Stimulates Phosphatidylinositol 3-Phosphate Production via the Activation of Rab5

Molecular Biology of the Cell ◽

10.1091/mbc.e08-01-0105 ◽

2008 ◽

Vol 19 (7) ◽

pp. 2718-2728 ◽

Cited By ~ 39

Author(s):

Irfan J. Lodhi ◽

Dave Bridges ◽

Shian-Huey Chiang ◽

Yanling Zhang ◽

Alan Cheng ◽

...

Keyword(s):

Plasma Membrane ◽

Glucose Uptake ◽

Critical Role ◽

Small Gtpase ◽

Dominant Negative ◽

Glut4 Translocation ◽

Nucleotide Exchange ◽

Exchange Factor ◽

Nucleotide Exchange Factor ◽

Phosphatidylinositol 3

Phosphatidylinositol 3-phosphate (PI(3)P) plays an important role in insulin-stimulated glucose uptake. Insulin promotes the production of PI(3)P at the plasma membrane by a process dependent on TC10 activation. Here, we report that insulin-stimulated PI(3)P production requires the activation of Rab5, a small GTPase that plays a critical role in phosphoinositide synthesis and turnover. This activation occurs at the plasma membrane and is downstream of TC10. TC10 stimulates Rab5 activity via the recruitment of GAPEX-5, a VPS9 domain–containing guanyl nucleotide exchange factor that forms a complex with TC10. Although overexpression of plasma membrane-localized GAPEX-5 or constitutively active Rab5 promotes PI(3)P formation, knockdown of GAPEX-5 or overexpression of a dominant negative Rab5 mutant blocks the effects of insulin or TC10 on this process. Concomitant with its effect on PI(3)P levels, the knockdown of GAPEX-5 blocks insulin-stimulated Glut4 translocation and glucose uptake. Together, these studies suggest that the TC10/GAPEX-5/Rab5 axis mediates insulin-stimulated production of PI(3)P, which regulates trafficking of Glut4 vesicles.

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Dynamic association of the PI3P-interacting Mon1-Ccz1 GEF with vacuoles is controlled through its phosphorylation by the type 1 casein kinase Yck3

Molecular Biology of the Cell ◽

10.1091/mbc.e13-08-0460 ◽

2014 ◽

Vol 25 (10) ◽

pp. 1608-1619 ◽

Cited By ~ 32

Author(s):

Gus Lawrence ◽

Christopher C. Brown ◽

Blake A. Flood ◽

Surya Karunakaran ◽

Margarita Cabrera ◽

...

Keyword(s):

Fusion Reaction ◽

Casein Kinase ◽

Nucleotide Exchange ◽

Wild Type ◽

Phosphatidylinositol 3 Kinase ◽

Guanine Nucleotide Exchange ◽

Nucleotide Exchange Factor ◽

Phosphatidic Acid Phosphatase ◽

Phosphatidylinositol 3

Maturation of organelles in the endolysosomal pathway requires exchange of the early endosomal GTPase Rab5/Vps21 for the late endosomal Rab7/Ypt7. The Rab exchange depends on the guanine nucleotide exchange factor activity of the Mon1-Ccz1 heterodimer for Ypt7. Here we investigate vacuole binding and recycling of Mon1-Ccz1. We find that Mon1-Ccz1 is absent on vacuoles lacking the phosphatidic acid phosphatase Pah1, which also lack Ypt7, the phosphatidylinositol 3-kinase Vps34, and the lipid phosphatidylinositol 3-phosphate (PI3P). Interaction of Mon1-Ccz1 with wild-type vacuoles requires PI3P, as shown in competition experiments. We also find that Mon1 is released from vacuoles during the fusion reaction and its release requires its phosphorylation by the type 1 casein kinase Yck3. In contrast, Mon1 is retained on vacuoles lacking Yck3 or when Mon1 phosphorylation sites are mutated. Phosphorylation and release of Mon1 is restored with addition of recombinant Yck3. Together the results show that Mon1 is recruited to endosomes and vacuoles by PI3P and, likely after activating Ypt7, is phosphorylated and released from vacuoles for recycling.

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Sbf/MTMR13 coordinates PI(3)P and Rab21 regulation in endocytic control of cellular remodeling

Molecular Biology of the Cell ◽

10.1091/mbc.e12-05-0375 ◽

2012 ◽

Vol 23 (14) ◽

pp. 2723-2740 ◽

Cited By ~ 48

Author(s):

Steve Jean ◽

Sarah Cox ◽

Eric J. Schmidt ◽

Fred L. Robinson ◽

Amy Kiger

Keyword(s):

Endosomal Trafficking ◽

Rab Gtpases ◽

Nucleotide Exchange ◽

Phosphatidylinositol 3 Kinase ◽

Guanine Nucleotide Exchange ◽

Nucleotide Exchange Factor ◽

Membrane Compartment ◽

Endosomal Pathway ◽

Gtpase Activation ◽

Phosphatidylinositol 3

Cells rely on the coordinated regulation of lipid phosphoinositides and Rab GTPases to define membrane compartment fates along distinct trafficking routes. The family of disease-related myotubularin (MTM) phosphoinositide phosphatases includes catalytically inactive members, or pseudophosphatases, with poorly understood functions. We found that Drosophila MTM pseudophosphatase Sbf coordinates both phosphatidylinositol 3-phosphate (PI(3)P) turnover and Rab21 GTPase activation in an endosomal pathway that controls macrophage remodeling. Sbf dynamically interacts with class II phosphatidylinositol 3-kinase and stably recruits Mtm to promote turnover of a PI(3)P subpool essential for endosomal trafficking. Sbf also functions as a guanine nucleotide exchange factor that promotes Rab21 GTPase activation associated with PI(3)P endosomes. Of importance, Sbf, Mtm, and Rab21 function together, along with Rab11-mediated endosomal trafficking, to control macrophage protrusion formation. This identifies Sbf as a critical coordinator of PI(3)P and Rab21 regulation, which specifies an endosomal pathway and cortical control.

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