The Sec6/8 complex in mammalian cells: Characterization of mammalian Sec3, subunit interactions, and expression of subunits in polarized cells

Hugo T Matern; Charles Yeaman; W. James Nelson; Richard H Scheller

doi:10.1073/pnas.171317898

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The Sec6/8 complex in mammalian cells: Characterization of mammalian Sec3, subunit interactions, and expression of subunits in polarized cells

Journal article

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The Sec6/8 complex in mammalian cells: Characterization of mammalian Sec3, subunit interactions, and expression of subunits in polarized cells

Hugo T Matern, Charles Yeaman, W. James Nelson and Richard H Scheller

Proceedings of the National Academy of Sciences - PNAS, Vol.98(17), pp.9648-9653

Inaugural Article

08/14/2001

DOI: 10.1073/pnas.171317898

PMCID: PMC55506

PMID: 11493706

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Abstract

The yeast exocyst complex (also called Sec6/8 complex in higher eukaryotes) is a multiprotein complex essential for targeting exocytic vesicles to specific docking sites on the plasma membrane. It is composed of eight proteins (Sec3, -5, -6, -8, -10, and -15, and Exo70 and -84), with molecular weights ranging from 70 to 144 kDa. Mammalian orthologues for seven of these proteins have been described and here we report the cloning and initial characterization of the remaining subunit, Sec3. Human Sec3 (hSec3) shares 17% sequence identity with yeast Sec3p, interacts in the two-hybrid system with other subunits of the complex (Sec5 and Sec8), and is expressed in almost all tissues tested. In yeast, Sec3p has been proposed to be a spatial landmark for polarized secretion (1), and its localization depends on its interaction with Rho1p (2). We demonstrate here that hSec3 lacks the potential Rho1-binding site and GFP-fusions of hSec3 are cytosolic. Green fluorescent protein (GFP)-fusions of nearly every subunit of the mammalian Sec6/8 complex were expressed in Madin–Darby canine kidney (MDCK) cells, but they failed to assemble into a complex with endogenous proteins and localized in the cytosol. Of the subunits tested, only GFP-Exo70 localized to lateral membrane sites of cell–cell contact when expressed in MDCK cells. Cells overexpressing GFP-Exo70 fail to form a tight monolayer, suggesting the Exo70 targeting interaction is critical for normal development of polarized epithelial cells.

Biological Sciences

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Title: Subtitle: The Sec6/8 complex in mammalian cells: Characterization of mammalian Sec3, subunit interactions, and expression of subunits in polarized cells
Creators: Hugo T Matern - Genentech, Inc., Department of Richard Scheller, 1 DNA Way, South San Francisco, CA 94080-4990; and
Charles Yeaman - Genentech, Inc., Department of Richard Scheller, 1 DNA Way, South San Francisco, CA 94080-4990; and
W. James Nelson - Genentech, Inc., Department of Richard Scheller, 1 DNA Way, South San Francisco, CA 94080-4990; and
Richard H Scheller - Genentech, Inc., Department of Richard Scheller, 1 DNA Way, South San Francisco, CA 94080-4990; and
Resource Type: Journal article
Publication Details: Proceedings of the National Academy of Sciences - PNAS, Vol.98(17), pp.9648-9653
Series: Inaugural Article
DOI: 10.1073/pnas.171317898
PMID: 11493706
PMCID: PMC55506
NLM abbreviation: Proc Natl Acad Sci U S A
ISSN: 0027-8424
eISSN: 1091-6490
Publisher: National Academy of Sciences
Language: English
Date published: 08/14/2001
Academic Unit: Anatomy and Cell Biology; Internal Medicine
Record Identifier: 9984025468002771

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