Molecular Identification of Two Novel Munc-18 Isoforms Expressed in Non-neuronal Tissues

Abstract

Munc-18, also known as n-Sec1 or rbSec1, is a syntaxin-binding protein thought to play a role in regulating synaptic vesicle exocytosis. Although a gene family of syntaxins has been identified, only a limited subset bind to Munc-18. This implicates the existence of other mammalian Munc-18 homologues that may be involved in a range of vesicle transport reactions. The purpose of the present study was to identify other members of the Munc-18 family by cDNA cloning. Three distinct Munc-18 isoforms, Munc-18a, previously identified in neuronal tissue, and two novel isoforms, Munc-18b and Munc-18c, were isolated from a 3T3-L1 adipocyte cDNA library by screening with a rat brain Munc-18 DNA probe. Munc-18a is identical to Munc-18 and by Northern analysis is expressed predominantly in brain and to a lesser extent in testis and 3T3-L1 cells. Munc-18b is 62% identical to Munc-18 at the amino acid level and is expressed in testis, intestine, kidney, rat adipose tissue, and 3T3-L1 cells. Munc-18c is 51% identical to Munc-18 and is ubiquitously expressed. It is likely, based on these findings, that unique Munc-18/syntaxin interactions may play an important role in generating a combinatorial mechanism for the regulation of vesicle transport in mammalian cells.

Syntaxin-3 regulates newcomer insulin granule exocytosis and compound fusion in pancreatic beta cells

“…This is of importance, as first-phase GSIS is much reduced in patients with type 2 diabetes, and this has been attributed in part to greatly reduced islet SYN-1A levels [23], along with reduction of cognate SNAREs (SNAP25 and VAMP2) and the Sec1/Munc18-like protein (SM) Munc18a. SYN-3 was shown in pancreatic acinar cells to preferentially form complexes with VAMP8 [20] and Munc18b [38,39], which are also present in beta cells [40][41][42]. We recently reported, employing the Vamp8-knockout mice [43], that VAMP8 also mediated the recruitment and fusion of newcomer SGs, and was the putative cognate v-SNARE that binds SYN-3.…”

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

“…pcDNA3-Munc18c was constructed by subcloning a 2.5-kb EcoRI fragment from pMEXneo-Munc18c into the EcoRI site of pcDNA3 (Invitrogen) (21). The pBluescript KG-Munc18b and pRSETA-Munc18c constructs have been described (21,30). pET20b-Syntaxin4 was kindly donated by Dr. Jenny Martin (Institute of Molecular Bioscience, Brisbane, Australia).…”

Supramaximal cholecystokinin displaces Munc18c from the pancreatic acinar basal surface, redirecting apical exocytosis to the basal membrane

“…In vitro studies of recombinant proteins indeed demonstrated stable binding of VAMP-2, SNAP-23, and syntaxin-4 (22), raising the possibility that a complex of VAMP-2/ SNAP-23/syntaxin-4 could mediate basolateral exocytosis of digestive enzymes within the pancreas. Non-neuronal Munc18 isoforms were also identified, including Munc18b and Munc18c (21)(22)(23)(24)(25)(26). Munc18c binds to syntaxin-4 and blocks its binding to SNAP-23 and VAMP-2 (24,25), and overexpression of Munc18c in adipocytes inhibits insulin-mediated GLUT-4 translocation to the plasma membrane and glucose transport (26).…”