Regulated exocytosis is a stimulus-dependent membrane fusion event of fundamental importance to a range of physiological processes. The membrane fusion reaction involves substantial lipid rearrangements and is opposed by the powerful hydrophobic force. For fusion to occur, a high energy barrier must be overcome. The overall goal of this proposed research is to determine the functional roles of membrane bilayer remodeling in overcoming the energy barrier of exocytic vesicle fusion, using the trafficking of the glucose transporter GLUT4 as a model system. To achieve this goal, we will employ a unique combination of complementary approaches including biochemical reconstitution, electron microscopic imaging, and biophysical measurements. First, we will define how two membrane bilayers are brought into close apposition by the SNARE-SM complex to prepare for the subsequent steps of membrane remodeling and merging. Next, we will assess the functional role of vesicle membrane bending in the fusion reaction. Finally, we will examine how the fusion reaction is regulated by local remodeling of the plasma membrane. We hypothesize that the fusion of exocytic vesicles with the plasma membrane involves a novel dual-curvature-induction mechanism: while the amphipathic motif of the v-SNARE bends the vesicle membrane, the hydrophobic loops of C2-domain molecules penetrate into the plasma membrane and induce local membrane curvature. Together, these membrane-bending activities are expected to create curvature stresses at the fusion sites to overcome the energy barrier for the fusion reaction. Successful completion of this proposed research will provide key insights into the molecular mechanisms of exocytic vesicle fusion. This work will also serve as a paradigm for understanding the general principles of intracellular membrane fusion. Ultimately, our findings will facilitate the development of novel therapeutic strategies for diseases associated with dysfunctional regulated exocytosis including diabetes, epilepsy, and immune disorders.
Regulated exocytosis is critical to the cell surface localization of receptors and transporters that are key to human physiology. We will investigate membrane dynamics and remodeling in the exocytic vesicle fusion process. Imbalances in receptor/transporter exocytosis give rise to severe forms of human disease. This study will provide key insights into these diseases and may offer new strategies for therapeutic intervention.
|Gulbranson, Daniel R; Davis, Eric M; Demmitt, Brittany A et al. (2017) RABIF/MSS4 is a Rab-stabilizing holdase chaperone required for GLUT4 exocytosis. Proc Natl Acad Sci U S A 114:E8224-E8233|
|Yu, Haijia; Liu, Yinghui; Gulbranson, Daniel R et al. (2016) Extended synaptotagmins are Ca2+-dependent lipid transfer proteins at membrane contact sites. Proc Natl Acad Sci U S A 113:4362-7|
|Zhang, Conggang; Lee, Schuyler; Peng, Yinghua et al. (2015) A chemical genetic approach to probe the function of PINK1 in regulating mitochondrial dynamics. Cell Res 25:394-7|
|Yu, Haijia; Rathore, Shailendra S; Shen, Chong et al. (2015) Reconstituting Intracellular Vesicle Fusion Reactions: The Essential Role of Macromolecular Crowding. J Am Chem Soc 137:12873-83|
|Davis, Eric M; Kim, Jihye; Menasche, Bridget L et al. (2015) Comparative Haploid Genetic Screens Reveal Divergent Pathways in the Biogenesis and Trafficking of Glycophosphatidylinositol-Anchored Proteins. Cell Rep 11:1727-36|
|Shen, Chong; Rathore, Shailendra S; Yu, Haijia et al. (2015) The trans-SNARE-regulating function of Munc18-1 is essential to synaptic exocytosis. Nat Commun 6:8852|
|Yu, Haijia; Rathore, Shailendra S; Gulbranson, Daniel R et al. (2014) The N- and C-terminal domains of tomosyn play distinct roles in soluble N-ethylmaleimide-sensitive factor attachment protein receptor binding and fusion regulation. J Biol Chem 289:25571-80|
|Yu, Haijia; Rathore, Shailendra S; Lopez, Jamie A et al. (2013) Comparative studies of Munc18c and Munc18-1 reveal conserved and divergent mechanisms of Sec1/Munc18 proteins. Proc Natl Acad Sci U S A 110:E3271-80|
|Yu, Haijia; Rathore, Shailendra S; Davis, Eric M et al. (2013) Doc2b promotes GLUT4 exocytosis by activating the SNARE-mediated fusion reaction in a calcium- and membrane bending-dependent manner. Mol Biol Cell 24:1176-84|
|Yu, Haijia; Rathore, Shailendra S; Shen, Jingshi (2013) Synip arrests soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE)-dependent membrane fusion as a selective target membrane SNARE-binding inhibitor. J Biol Chem 288:18885-93|