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Séminaire impromptu - Liangyi Chen Exocytosis and Endocytosis in Insulin-secreting Cells

Abstract :

 Exocytosis-coupled clathrin mediated endocytosis:
 In eukaryotic cells, many receptor-mediated endocytic processes are mediated by constitutively budding of clathrin-coated pits (CCPs) at spatially randomized sites before slowly pinching-off from the plasma membrane (60~100 s). In contrast, clathrin-mediated endocytosis (CME) coupled with regulated exocytosis in excitable cells occurs at peri-exocytic sites shortly after vesicle fusion (~10 s). The molecular mechanism underlying this spatiotemporal coupling remains elusive. Here, we show that coupled endocytosis makes use of pre-formed CCPs whose trafficking is orchestrated by cortical microtubular dynamics and local lipid gradients. Following an exocytic event, a nearby CCP hops to the fusion site along with cargo proteins to be internalized. A dynamic cortical microtubular network, anchored at the cell surface by the cytoplasmic linker-associated protein on microtubules and the LL5b/ELKS complex on the plasma membrane, provides the track for CCP hopping. Local diacylglycerol gradients generated upon exocytosis guide the direction of hopping. Overall, the CCP-cytoskeleton-lipid interaction demonstrated here mediates exocytosis-coupled fast recycling of both plasma membrane and vesicular proteins, and may participate in the regulation of diverse functions in all eukaryotic cells.

 Numbers of SNARE proteins required for different types of exocytosis:  SNARE proteins provide the energy for the merger of two opposing lipid bilayers. Although the number of SNARE proteins required for the fusion of one secretory vesicle has been studied using a variety of methods, directly visualization of such process in live cells have never been achieved. Here we have combined spatiotemporal averaging of fluorescence images with single molecule tracking to count the number of SNARE proteins required for a vesicle fusion. We show that different types of vesicle fusion require different number of SNARE complexes. 

Selected publications

[1] Zong W, Zhao J, Chen X, Lin Y, Ren H, Zhang Y, Fan M, Zhou Z, Cheng H, Sun Y, Chen L (2015) Large-field high-resolution two-photon digital scanned light-sheet microscopy. Cell Res 25: 254-257

[2] Yuan T, Lu J, Zhang J, Zhang Y, Chen L (2015) Spatiotemporal detection and analysis of exocytosis reveal fusion "hotspots" organized by the cytoskeleton in endocrine cells. Biophys J 108: 251-260

[3] Hao Z, Wei L, Feng Y, Chen X, Du W, Ma J, Zhou Z, Chen L, Li W (2015) Impaired maturation of large dense-core vesicles in muted-deficient adrenal chromaffin cells. J Cell Sci 128: 1365-1374

[4] Dou H, Wang C, Wu X, Yao L, Zhang X, Teng S, Xu H, Liu B, Wu Q, Zhang Q, Hu M, Wang Y, Wang L, Wu Y, Shang S, Kang X, Zheng L, Zhang J, Raoux M, Lang J, Li Q, Su J, Yu X, Chen L, Zhou Z (2015) Calcium influx activates adenylyl cyclase 8 for sustained insulin secretion in rat pancreatic beta cells. Diabetologia 58: 324-333

[5] Zong W, Huang X, Zhang C, Yuan T, Zhu LL, Fan M, Chen L (2014) Shadowless-illuminated variable-angle TIRF (siva-TIRF) microscopy for the observation of spatial-temporal dynamics in live cells. Biomed Opt Express 5: 1530-1540

[6] Shang W, Lu F, Sun T, Xu J, Li LL, Wang Y, Wang G, Chen L, Wang X, Cannell MB, Wang SQ, Cheng H (2014) Imaging Ca2+ nanosparks in heart with a new targeted biosensor. Circ Res 114: 412-420

Scientific focus :

Dr. Liangyi Chen is a professor and principal investigator in Institute of Molecular Medicine, Peking University. His current research interests are focused on development of novel high spatiotemporal fluorescence imaging techniques for in vivo studies. His lab also use these new technologies to study the exocytosis-endocytosis coupling in pancreatic b-cells and its association with blood glucose regulation and diabetes, using cell lines, Zebrafish and mouse models. As the corresponding author, Dr. Liangyi Chen has published papers in peer-reviewed journals such as Cell Res., Proc Natl Acad Sci U S A., Biophys J., Biomed Opt Express., Diabetes and Diabetologia, etc. He has been served as a senior faculty member in the Neuronal Signaling Mechanisms Section in Faculty of 1000 Biology since 2012. He has been invited to give talks in international conferences hosted by academic societies such as OSA, SPIE and Biophysical Society, etc.