Jt. Tellam et al., CHARACTERIZATION OF MUNC-18C AND SYNTAXIN-4 IN 3T3-L1 ADIPOCYTES - PUTATIVE ROLE IN INSULIN-DEPENDENT MOVEMENT OF GLUT-4, The Journal of biological chemistry, 272(10), 1997, pp. 6179-6186
We have previously identified three mammalian Seel/Munc-18 homologues
in adipocytes (Tellam, J. T., McIntosh, S., and James, D. E. (1995) J.
Biol, Chem, 270, 5857-5863). These proteins are thought to modulate t
he interaction between vesicle membrane and target membrane soluble N-
ethylmaleimide-sensitive factor attachment protein receptors (SNAREs)
and thus regulate intracellular vesicular transport, This study aimed
to further characterize these Munc-18 isoforms and to define their pot
ential role in the trafficking of GLUT-4 in adipocytes, a process repo
rted to involve the vesicle membrane SNARE, VAMP-8, Using an in vitro
binding assay with recombinant fusion proteins, we show that Munc-18a
and Munc-18b bind to syntaxin-1A, -2, and -3, while Munc-18c binds onl
y to syntaxin-2 and -4, The specific interaction between Munc-18c and
syntaxin 4 is of interest because aside from syntaxin-1A, which is not
expressed in adipocytes, syntaxin-4 is the only syntaxin that binds t
o VAMP S. Using a three-way binding assay, it was shown that Munc-18c
inhibits the binding of syntaxin-4 to VAMP-S, The subcellular distribu
tion of syntaxin-4 and Munc-18c was almost identical, both being enric
hed in the plasma membrane, and both exhibiting an insulin-dependent m
ovement out of an intracellular membrane fraction similar to that obse
rved for GLUT-4, Munc-18b had a similar distribution to Munc-18c and s
o may also be involved in vesicle transport to the cell surface, where
as Munc-18a was undetectable by immunoblotting; in adipocytes, Microin
jection of a syntaxin-4 antibody into 3T3-L1 adipocytes blocked the in
sulin-dependent recruitment of GLUT 4 to the cell surface, These data
suggest that syntaxin-4/Munc-18c/VAMP-2 may play a role in the docking
/fusion of intracellular GLUT-4-containing vesicles with the cell surf
ace in adipocytes.