SYNTAXIN-4 IN 3T3-L1 ADIPOCYTES - REGULATION BY INSULIN AND PARTICIPATION IN INSULIN-DEPENDENT GLUCOSE-TRANSPORT

Syntaxins are thought to be membrane receptors that bind proteins of t he synaptobrevin/vesicle-associated membrane protein (VAMP) family fou nd on transport vesicles. Recently, we detected synaptobrevin II and c ellubrevin on immunopurified vesicles containing the glucose transport er 4 (GLUT4) in insulin-responsive cells. In an effort to identify the plasma membrane receptors for these vesicles, we now examine the expr ession of syntaxins in the 3T3-L1 adipocyte cell line. Neither syntaxi n 1A nor 1B was found, in keeping with the neuronal restriction of the se isoforms. In contrast, syntaxins 2 and 4 were readily detectable. B y subcellular fractionation and estimation of protein yields, 67% of s yntaxin 4 was localized to the plasma membrane, 24% to the low-density microsomes, and 9% to the high-density microsomes. Interestingly, acu te insulin treatment decreased the content of syntaxin 4 in low-densit y microsomes and caused a corresponding gain in the plasma membrane fr action, reminiscent of the recruitment of GLUT4 glucose transporters. In contrast, there was no change in the distribution of syntaxin 2, wh ich was mostly associated in the plasma membrane. A fraction of the in tracellular syntaxin 4 was recovered with immunopurified GLUT4-contain ing vesicles. Moreover, anti-syntaxin 4 antibodies introduced into per meabilized 3T3-L1 adipocytes significantly reduced the insulin-depende nt stimulation of glucose transport, in contrast to the introduction o f irrelevant immunoglobulin G, which was without consequence. We propo se that either the plasma membrane and/or the vesicular syntaxin 4 are involved in docking and/or fusion of GLUT4 vesicles at the cell surfa ce of 3T3-L1 adipocytes.