Rabs constitute the largest branch of the Ras GTPase superfamily, with ten members in yeast and more then 60 in mammalian cells. They serve as key nodes in the regulation of membrane traffic, each typically controlling several different aspects of a specific stage of membrane traffic by recruiting diverse effector proteins such as cytoskeletal motors, vesicle tethering proteins and regulators of SNARE complex assembly. Rabs are activated by specific guanine nucleotide exchange factors (GEFs) that catalyze the displacement of GDP and binding of GTP and are inactivated by GTPase activating proteins (GAPs) that stimulate the slow intrinsic rate of GTP hydrolysis. We have proposed that adjacent Rabs on a traffic pathway are networked to one another through their regulators. The postulated effect of these counter-current cascades is a programmed series of abrupt Rab conversions that lead to changes in the functional identity of the membrane as it flows along a pathway. Rabs are also thought to define the anterograde versus retrograde directionality of vesicular transport. In addition, Rab GEF-effector interactions are believed to generate positive feedback loops that promote vesicle maturation. While the roles of individual elements of the regulatory network have been examined, these proposals have not yet been tested at a more global, integrated level. We propose three specific aims that will address important aspects of the Rab regulatory network: 1. We will test the effects of rewiring a Rab regulatory circuit to evaluate its role in establishing organelle identity and pursue a novel inhibitory role for a coiled-coil vesicle tether that has emerged from these studies 2. We will directly test the role of Rabs in the control of the directionality of membrane traffic 3. We will determine the role of a Rab GEF-effector interaction in secretory vesicle maturation

Public Health Relevance

Membrane traffic is required for a broad range of essential cellular functions and the regulation of membrane traffic by Rab GTPases is therefore relevant to major human diseases, including cancer, diabetes and neural degeneration. Additional diseases have been directly attributed to specific defects in Rab expression, Rab modification, Rab regulation and Rab effectors and a number of clinically important human pathogens have evolved to exploit and disrupt our Rab regulatory pathways to promote their own intracellular agenda and to evade host defenses. We will analyze how Rab proteins are functionally linked to one another through regulatory networks to control membrane traffic.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM082861-11
Application #
9565614
Study Section
Nuclear and Cytoplasmic Structure/Function and Dynamics Study Section (NCSD)
Program Officer
Flicker, Paula F
Project Start
2008-07-01
Project End
2021-06-30
Budget Start
2018-07-01
Budget End
2019-06-30
Support Year
11
Fiscal Year
2018
Total Cost
Indirect Cost
Name
University of California, San Diego
Department
Other Basic Sciences
Type
Schools of Medicine
DUNS #
804355790
City
La Jolla
State
CA
Country
United States
Zip Code
92093
Chen, Shuliang; Cui, Yixian; Parashar, Smriti et al. (2018) ER-phagy requires Lnp1, a protein that stabilizes rearrangements of the ER network. Proc Natl Acad Sci U S A 115:E6237-E6244
Liu, Dongmei; Li, Xia; Shen, David et al. (2018) Two subunits of the exocyst, Sec3p and Exo70p, can function exclusively on the plasma membrane. Mol Biol Cell 29:736-750
Yuan, Hua; Davis, Saralin; Ferro-Novick, Susan et al. (2017) Rewiring a Rab regulatory network reveals a possible inhibitory role for the vesicle tether, Uso1. Proc Natl Acad Sci U S A 114:E8637-E8645
Stalder, Danièle; Novick, Peter J (2016) The casein kinases Yck1p and Yck2p act in the secretory pathway, in part, by regulating the Rab exchange factor Sec2p. Mol Biol Cell 27:686-701
Novick, Peter (2016) Regulation of membrane traffic by Rab GEF and GAP cascades. Small GTPases 7:252-256
Chen, Shuliang; Desai, Tanvi; McNew, James A et al. (2015) Lunapark stabilizes nascent three-way junctions in the endoplasmic reticulum. Proc Natl Acad Sci U S A 112:418-23
Sánchez-León, Eddy; Bowman, Barry; Seidel, Constanze et al. (2015) The Rab GTPase YPT-1 associates with Golgi cisternae and Spitzenkörper microvesicles in Neurospora crassa. Mol Microbiol 95:472-90
Stalder, Danièle; Novick, Peter J (2015) Assaying the interaction of the Rab guanine nucleotide exchange protein Sec2 with the upstream Rab, a downstream effector, and a phosphoinositide. Methods Mol Biol 1298:85-98
Ling, Yading; Hayano, Scott; Novick, Peter (2014) Osh4p is needed to reduce the level of phosphatidylinositol-4-phosphate on secretory vesicles as they mature. Mol Biol Cell 25:3389-400
Novick, Peter J (2014) A pathway of a hundred genes starts with a single mutant: isolation of sec1-1. Proc Natl Acad Sci U S A 111:9019-20

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