Mammalian Exocyst Complex Is Required for the Docking Step of InsulinVesicle Exocytosis

Tsuboi, Takashi; Ravier, Magalie A.; Xie, Hao; Ewart, Marie-Ann; Gould, Gwyn W.; Baldwin, Stephen A.; Rutter, Guy A.

doi:10.1074/jbc.m501674200

Cited by 67 publications

(79 citation statements)

References 42 publications

(55 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…However, the incorporation of VSV-G into the PM was significantly affected, as revealed by the 8G5 antibody specifically recognizing the extracellular domain of VSV-G. Similarly the exocyst has been implicated in tethering and fusion of Glut4-containing vesicles in 3T3-L1 adipocytes (Inoue et al, 2003;Ewart et al, 2005;Tsuboi et al, 2005). It is possible that other exocyst components also interact with phospholipids (Moskalenko et al, 2003).…”

Section: Discussionmentioning

confidence: 99%

Phosphatidylinositol 4,5-Bisphosphate Mediates the Targeting of the Exocyst to the Plasma Membrane for Exocytosis in Mammalian Cells

Liu

Zuo

Peng

et al. 2007

MBoC

196

248

View full text Add to dashboard Cite

The exocyst is an evolutionarily conserved octameric protein complex that tethers post-Golgi secretory vesicles at the plasma membrane for exocytosis. To elucidate the mechanism of vesicle tethering, it is important to understand how the exocyst physically associates with the plasma membrane (PM). In this study, we report that the mammalian exocyst subunit Exo70 associates with the PM through its direct interaction with phosphatidylinositol 4,5-bisphosphate (PI(4,5)P 2 ). Furthermore, we have identified key conserved residues at the C-terminus of Exo70 that are crucial for the interaction of Exo70 with PI(4,5)P 2 . Disrupting Exo70-PI(4,5)P 2 interaction abolished the membrane association of Exo70. We have also found that wild-type Exo70 but not the PI(4,5)P 2 -binding-deficient Exo70 mutant is capable of recruiting other exocyst components to the PM. Using the ts045 vesicular stomatitis virus glycoprotein trafficking assay, we demonstrate that Exo70-PI(4,5)P 2 interaction is critical for the docking and fusion of post-Golgi secretory vesicles, but not for their transport to the PM. INTRODUCTIONExocytosis is important for a variety of cellular functions, ranging from the release of hormones to the incorporation of membrane proteins for cell growth and morphogenesis. The late stage of exocytosis is a multistep process that includes directional transport, tethering, docking, and fusion of postGolgi secretory vesicles with the plasma membrane (PM). The tethering step, defined as the initial contact of secretory vesicles with the PM before SNARE-mediated docking and fusion (Pfeffer, 1999;Guo et al., 2000;Waters and Hughson, 2000;Whyte and Munro, 2002), is mediated by the exocyst, an evolutionarily conserved octameric complex composed of Sec3, Sec5, Sec6, Sec8, Sec10, Sec15, Exo70, and Exo84 (for review, see Guo et al., 2000;Hsu et al., 2004;Munson and Novick, 2006;Wang and Hsu, 2006). In budding yeast, the exocyst components localize to the growing end of the daughter cell ("bud"), where active exocytosis and membrane addition take place (TerBush and Novick, 1995;Finger et al., 1998, Guo et al., 1999. This localization pattern contrasts that of the membrane fusion machine, the t-SNAREs, which are evenly distributed along both the mother and daughter cell membrane (Brennwald et al., 1994). In mammalian cells, the exocyst components were found in the cytosol, recycling endosomes and trans-Golgi network Fö lsch et al., 2003;Ang et al., 2004;Langevin et al., 2005). However, they are recruited to the PM during a number of cellular processes. For example, in epithelial cells, the exocyst is recruited to the adherens junction region upon cell-cell contact, where it mediates protein and membrane addition at the basolateral domain (Grindstaff et al., 1998;Yeaman et al., 2001); in developing neurons, the exocyst is localized to the growing neurites, where it mediates membrane expansion (Hazuka et al., 1999;Vega and Hsu, 2001); during cell migration, the exocyst is recruited to the leading edges of the PM (Rosse et al., 2006;...

show abstract

Section: Discussionmentioning

confidence: 99%

Phosphatidylinositol 4,5-Bisphosphate Mediates the Targeting of the Exocyst to the Plasma Membrane for Exocytosis in Mammalian Cells

Liu

Zuo

Peng

et al. 2007

MBoC

196

248

View full text Add to dashboard Cite

show abstract

“…32 Therefore, we hypothesized that TNFAIP2 could promote angiogenesis by upregulating VEGF secretion. However, we did not observe any decrease in VEGF secretion by TNFAIP2-knockdown HK1 cells (Figure 3d), suggesting that TNFAIP2 is not involved in the regulation of VEGF secretion.…”

Section: Discussionmentioning

confidence: 99%

“…Finally, we tested for a potential interaction between TNFAIP2 and VEGF, an important secreted angiogenesis-initiating factor that has also been correlated with microvessel density in nasopharyngeal carcinoma patients. 30 As the protein sequence of TNFAIP2 (amino acids 91-651) is highly similar to that of SEC6 (based on a publicly available website http://www.sanger.ac.uk/), an exocyst complex component involved in exocytosis 31 and secretion, 32 we examined whether TNFAIP2 affected the secretion of VEGF by HK1 cells. As shown in Figure 3d, our results revealed that VEGF secretion by HK1 cells was not affected by TNFAIP2 knockdown.…”

Section: Promotion Of Cell Migration and Invasion By Tnfaip2mentioning

confidence: 99%

A novel role for TNFAIP2: its correlation with invasion and metastasis in nasopharyngeal carcinoma

Chen

et al. 2011

Modern Pathology

View full text Add to dashboard Cite

Tumor necrosis factor alpha (TNFa) is an inflammatory cytokine that is present in the microenvironment of many tumors and is known to promote tumor progression. To examine how TNFa modulates the progression and metastasis of nasopharyngeal carcinoma, we used Affymetrix chips to identify TNFa-inducible genes that are dysregulated in this tumor. Elevated expression of TNFAIP2, which encodes TNFa-inducible protein 2 and not previously known to be associated with cancer, was found and confirmed by quantitative RT-PCR of TNFAIP2 expression in nasopharyngeal carcinoma and adjacent normal tissues. Immunohistochemical analysis showed that the TNFAIP2 protein was highly expressed in tumor cells. Analysis of 95 nasopharyngeal carcinoma biopsy specimens revealed that high TNFAIP2 expression was significantly correlated with highlevel intratumoral microvessel density (P ¼ 0.005) and low distant metastasis-free survival (P ¼ 0.001). A multivariate analysis further confirmed that TNFAIP2 was an independent prognostic factor for nasopharyngeal carcinoma (P ¼ 0.002). In vitro, TNFa treatment of nasopharyngeal carcinoma HK1 cells was found to induce TNFAIP2 expression, and siRNA-based knockdown of TNFAIP2 dramatically reduced the migration and invasion of nasopharyngeal carcinoma HK1 cells. These results collectively suggest for the first time that TNFAIP2 is a cell migration-promoting protein and its expression predicts distant metastasis. Our data suggest that TNFAIP2 may serve as an independent prognostic indicator for nasopharyngeal carcinoma.

show abstract

“…Among its proposed roles, the exocyst is thought to function as a vesicle tether at the plasma membrane and contribute to the specificity of membrane fusion (5). Recent studies in the ␤-cell demonstrated that the exocyst regulates the ability of insulin granules to dock with the plasma membrane (10).…”

Section: Snaresmentioning

confidence: 99%

“…Exo70 Interacts with a Helical Domain in the C Terminus of Snapin-Snapin contains an N-terminal hydrophobic domain (amino acids [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17][18][19][20] and two helical domains (amino acids 37-65 and 83-119), the second of which is the major binding site for SNAP23 (22). To map the Exo70 binding domain in Snapin, a series of FLAG-Snapin truncation mutants were generated ( Fig.…”

Section: Identification Of Snapin As An Exo70-binding Protein-mentioning

confidence: 99%

Snapin Interacts with the Exo70 Subunit of the Exocyst and Modulates GLUT4 Trafficking

Bao

Lopez

James

et al. 2008

Journal of Biological Chemistry

View full text Add to dashboard Cite

The exocyst is a multisubunit complex that has been implicated in the transport of vesicles from the Golgi complex to the plasma membrane, possibly acting as a vesicle tether and contributing to the specificity of membrane fusion. Here we characterize a novel interaction between the Exo70 subunit of the exocyst and Snapin, a ubiquitous protein known to associate with at least two t-SNAREs, SNAP23 and SNAP25. The interaction between Exo70 and Snapin is mediated via an N-terminal coil-coil domain in Exo70 and a C-terminal helical region in Snapin. Exo70 competes with SNAP23 for Snapin binding, suggesting that Snapin does not provide a direct link between the exocyst and the SNARE complex but, rather, mediates cross-talk between the two complexes by sequential interactions. The insulin-regulated trafficking of GLUT4 to the plasma membrane serves to facilitate glucose uptake in adipocytes, and both SNAP23 and the exocyst have been implicated in this process. In this study, depletion of Snapin in adipocytes using RNA interference inhibits insulin-stimulated glucose uptake. Thus, Snapin interacts with the exocyst and plays a modulatory role in GLUT4 vesicle trafficking. SNAREs4 are central components of the vesicle docking and fusion machinery (1, 2). In vitro liposome fusion experiments have shown that SNAREs alone are sufficient to catalyze fusion but at a much slower rate than occurs in vivo (3). In vivo, vesicle docking and fusion likely requires additional components to tightly regulate the specificity and kinetics of membrane fusion.The seminal work of Novick and Scheckman (4) identified several genes necessary for secretion in yeast, collectively named the SEC genes. Temperature-sensitive mutations in each of these genes block secretion of invertase and accumulate membrane-bound organelles within the yeast cell. Six of the SEC genes, SEC3, SEC5, SEC6, SEC8, SEC10, and SEC15, encode subunits of an evolutionarily conserved octameric protein complex known as the exocyst. Two additional components, Exo70p and Exo84p, were subsequently biochemically purified in complex with the exocyst proteins (5-7). Similar to their yeast homologues, the mammalian exocyst proteins have been implicated in the transport of vesicles from the Golgi to the plasma membrane (8). The exocyst complex has also been implicated in the trafficking of GLUT4 vesicles to the plasma membrane (9). Among its proposed roles, the exocyst is thought to function as a vesicle tether at the plasma membrane and contribute to the specificity of membrane fusion (5). Recent studies in the ␤-cell demonstrated that the exocyst regulates the ability of insulin granules to dock with the plasma membrane (10).Within the cell, each step of membrane trafficking is coupled to maintain the sequential nature and ensure the high fidelity of the process. Emerging evidence suggests a link between the membrane tethering and the SNARE-mediated docking/fusion steps (11,12). Genetic experiments in yeast are consistent with the SNAREs acting directly downstream of the exocyst. ...

show abstract

Mammalian Exocyst Complex Is Required for the Docking Step of InsulinVesicle Exocytosis

Cited by 67 publications

References 42 publications

Phosphatidylinositol 4,5-Bisphosphate Mediates the Targeting of the Exocyst to the Plasma Membrane for Exocytosis in Mammalian Cells

Phosphatidylinositol 4,5-Bisphosphate Mediates the Targeting of the Exocyst to the Plasma Membrane for Exocytosis in Mammalian Cells

A novel role for TNFAIP2: its correlation with invasion and metastasis in nasopharyngeal carcinoma

Snapin Interacts with the Exo70 Subunit of the Exocyst and Modulates GLUT4 Trafficking

Contact Info

Product

Resources

About