Journal of Cell Science, Vol 114, Issue 1 219-227, Copyright © 2001 by Company of Biologists

JOURNAL ARTICLES

Localization of cellubrevin-related peptide, endobrevin, in the early endosome in pancreatic beta cells and its physiological function in exo-endocytosis of secretory granules

S Nagamatsu, Y Nakamichi, T Watanabe, S Matsushima, S Yamaguchi, J Ni, E Itagaki and H Ishida
Departments of Biochemistry, Clinical Pathology, Internal Medicine (III), Kyorin University School of Medicine, Mitaka, Tokyo 181-8611, Japan. shinya@kyorin-u.ac.jp

Cellubrevins are integral membrane proteins expressed in a wide variety of tissues and usually localized in recycling vesicles. Here, we investigated the cellular localization of a cellubrevin-related peptide, endobrevin, in pancreatic (beta) cells and its implication in the exo-endocytosis of insulin and (gamma)-amino butyric acid (GABA). Immunocytochemistry showed that endobrevin is associated with tubulo-vesicular structures, which are colocalized with early endosomes labeled by early endosome antigen (EEA)-1 in insulinoma MIN6 cells. To determine the cellular localization of endobrevin, we appended the green fluorescent protein (GFP) to endobrevin and the fusion protein was introduced into MIN6 cells. The subcellular localization of GFP-endobrevin was visualized by confocal laser microscopy. Colocalization study based on the expressed GFP-endobrevin and endocytosed Texas-Red(Tx-R) labeled transferrin receptor and immunocytochemistry with anti-EEA1 antibody revealed that endobrevin was preferentially localized in the early endosome. Then, we examined the functional role of endobrevin in the exocytosis of insulin and GABA from pancreatic (beta) cells. Endobrevin overexpression increased the amount of GABA released from MIN6 cells; in contrast, it decreased the glucose-stimulated insulin release from rat islets, MIN6 and INS1-D cells to approximately 50% of the control levels. Both in vitro and in vivo binding studies showed that endobrevin binds to syntaxin 1. Finally, using the fluorescent probe FM4-64, it was revealed that endobrevin overexpression accelerates vesicle recycling. We conclude that (1) endobrevin is localized in the early endosome in pancreatic (beta) cells and (2) endobrevin plays a physiological role in the exo-endocytosis of insulin and GABA from pancreatic (beta) cells, probably via an interaction between endocytic vesicles and the endosome.
CiteULike    Complore    Connotea    Del.icio.us    Digg    Reddit    Technorati    Twitter    What's this?

This article has been cited by other articles:

				T. Kimura, Y. Kaneko, S. Yamada, H. Ishihara, T. Senda, A. Iwamatsu, and I. Niki The GDP-dependent Rab27a effector coronin 3 controls endocytosis of secretory membrane in insulin-secreting cell lines J. Cell Sci., September 15, 2008; 121(18): 3092 - 3098. [Abstract] [Full Text] [PDF]

				H. Mashima, J. Suzuki, T. Hirayama, Y. Yoshikumi, H. Ohno, H. Ohnishi, H. Yasuda, T. Fujita, and M. Omata Involvement of Vesicle-Associated Membrane Protein 7 in Human Gastric Epithelial Cell Vacuolation Induced by Helicobacter pylori-Produced VacA Infect. Immun., June 1, 2008; 76(6): 2296 - 2303. [Abstract] [Full Text] [PDF]

				Y. Brunner, Y. Coute, M. Iezzi, M. Foti, M. Fukuda, D. F. Hochstrasser, C. B. Wollheim, and J.-C. Sanchez Proteomics Analysis of Insulin Secretory Granules Mol. Cell. Proteomics, June 1, 2007; 6(6): 1007 - 1017. [Abstract] [Full Text] [PDF]

				R. D. Pooley, S. Reddy, V. Soukoulis, J. T. Roland, J. R. Goldenring, and D. M. Bader CytLEK1 Is a Regulator of Plasma Membrane Recycling through Its Interaction with SNAP-25 Mol. Biol. Cell, July 1, 2006; 17(7): 3176 - 3186. [Abstract] [Full Text] [PDF]

				L. Ma, V. P. Bindokas, A. Kuznetsov, C. Rhodes, L. Hays, J. M. Edwardson, K. Ueda, D. F. Steiner, and L. H. Philipson Direct imaging shows that insulin granule exocytosis occurs by complete vesicle fusion PNAS, June 22, 2004; 101(25): 9266 - 9271. [Abstract] [Full Text] [PDF]

				R. Kuliawat, E. Kalinina, J. Bock, L. Fricker, T. E. McGraw, S. R. Kim, J. Zhong, R. Scheller, and P. Arvan Syntaxin-6 SNARE Involvement in Secretory and Endocytic Pathways of Cultured Pancreatic {beta}-Cells Mol. Biol. Cell, April 1, 2004; 15(4): 1690 - 1701. [Abstract] [Full Text] [PDF]

				J. K. Jaiswal, N. W. Andrews, and S. M. Simon Membrane proximal lysosomes are the major vesicles responsible for calcium-dependent exocytosis in nonsecretory cells J. Cell Biol., November 25, 2002; 159(4): 625 - 635. [Abstract] [Full Text] [PDF]

				L. Nadav, A. Eldor, O. Yacoby-Zeevi, E. Zamir, I. Pecker, N. Ilan, B. Geiger, I. Vlodavsky, and B.-Z. Katz Activation, processing and trafficking of extracellular heparanase by primary human fibroblasts J. Cell Sci., May 15, 2002; 115(10): 2179 - 2187. [Abstract] [Full Text] [PDF]