Yeast Homologues of Tomosyn and lethal giant larvae Function in Exocytosis and Are Associated with the Plasma Membrane Snare, Sec9

Abstract

We have identified a pair of related yeast proteins, Sro7p and Sro77p, based on their ability to bind to the plasma membrane SNARE (SNARE) protein, Sec9p. These proteins show significant similarity to the Drosophila tumor suppressor, lethal giant larvae and to the neuronal syntaxin–binding protein, tomosyn. SRO7 and SRO77 have redundant functions as loss of both gene products leads to a severe cold-sensitive growth defect that correlates with a severe defect in exocytosis. We show that similar to Sec9, Sro7/77 functions in the docking and fusion of post-Golgi vesicles with the plasma membrane. In contrast to a previous report, we see no defect in actin polarity under conditions where we see a dramatic effect on secretion. This demonstrates that the primary function of Sro7/77, and likely all members of the lethal giant larvae family, is in exocytosis rather than in regulating the actin cytoskeleton. Analysis of the association of Sro7p and Sec9p demonstrates that Sro7p directly interacts with Sec9p both in the cytosol and in the plasma membrane and can associate with Sec9p in the context of a SNAP receptor complex. Genetic analysis suggests that Sro7 and Sec9 function together in a pathway downstream of the Rho3 GTPase. Taken together, our studies suggest that members of the lethal giant larvae/tomosyn/Sro7 family play an important role in polarized exocytosis by regulating SNARE function on the plasma membrane.

Tomosyn Interacts with the t-SNAREs Syntaxin4 and SNAP23 and Plays a Role in Insulin-stimulated GLUT4 Translocation

“…This was surprising, as both the family of lethal (2) giant larvae proteins and the yeast homolog of Tomosyn, Sro7/77, do not contain a SNARE motif. However, there is significant sequence conservation in the amino-terminal part among lethal giant larvae proteins, Sro7/77 and Tomosyn, including a series of WD-40 repeats (19,44). We therefore investigated whether this conserved amino-terminal part of Tomosyn could interact with Syntaxin4 independently of the VAMP-2 like domain.…”

The R-SNARE Motif of Tomosyn Forms SNARE Core Complexes with Syntaxin 1 and SNAP-25 and Down-regulates Exocytosis

“…Intriguingly, however, several of the other family members have not only been demonstrated to play a role in membrane traffic of polarized cells but also to directly interact with intracellular SNAREs. In yeast, cells deficient in SRO7 and SRO77 exhibit a defect in vesicle transport from the Golgi apparatus to the plasma membrane, and the protein Sro7p binds to the yeast SNAP-25 homologue Sec9p (9). Furthermore, MglI appears to play a role in polarized membrane trafficking as it becomes associated with the lateral membrane when MDCK cells establish polarity, and membrane binding of MglI seems to be mediated by binding to syntaxin 4 (8).…”

“…As outlined in the introduction, tomosyn is related to the Drosophila tumor suppressor lethal (2) giant larvae (l(2)gl) (7), its mammalian homologue MglI (8) and the yeast proteins Sro7p and Sro77p (9). The homology is confined to the Nterminal domain containing WD40 repeats, protein binding domains with a conserved structure of about 40 residues, each containing a central Trp-Asp motif.…”

“…Tomosyn is a 130-kDa protein that lacks a membrane anchor and in which two domains are distinguished: a small C-terminal region with homology to the R-SNARE subfamily of SNAREs and a large N-terminal region that contains WD40 repeats and shares similarities with the Drosophila tumor suppressor lethal (2) giant larvae (l(2)gl) (7), its recently characterized mammalian homologue MglI (8), and the yeast proteins Sro7p and Sro77p (9). Three splicing variants of tomosyn are known, two of which are predominantly expressed in brain, whereas one is ubiquitously distributed (10).…”

Lateral Membrane Biogenesis in Human Bronchial Epithelial Cells Requires 190-kDa Ankyrin-G

“…Lethal giant larvae has been shown to interact with syntaxin-4, the targetsoluble NSF attachment protein receptors residing in the lateral membrane of epithelial cells (44). Cold-sensitive alleles of the yeast homologues of lethal giant larvae show accumulation of post-Golgi exocytic vesicles under nonpermissive temperatures (46). It is also possible that ankyrin-G accomplishes its function in lateral membrane biogenesis through an as yet unidentified pathway.…”