A functional role for VAP-33 in insulin-stimulated GLUT4 traffic

Traffic. 2000 Jun;1(6):512-21. doi: 10.1034/j.1600-0854.2000.010609.x.

Abstract

Soluble N-ethylmaleimide sensitive factor attachment protein receptors (SNAREs) are critical proteins in membrane fusion, in both regulated and constitutive vesicular traffic. In addition, proteins that interact with the SNAREs are thought to regulate fusion. Vesicle-associated membrane protein-2 (VAMP-2) is a SNARE protein involved in insulin-dependent glucose transporter 4 (GLUT4) traffic. VAMP-2 is required for productive GLUT4 incorporation into the plasma membrane. VAMP-associated protein of 33 kDa (VAP-33) is an integral membrane protein that binds VAMPs in vitro, and is hypothesized to be a regulator of VAMPs. In L6 skeletal myoblasts, which display insulin-dependent traffic of GLUT4, we show that VAP-33 colocalized significantly with VAMP-2 using indirect confocal immunofluorescence and biochemical cosegregation. Overexpression of wild-type VAP-33 in L6 myoblasts attenuated the insulin-dependent incorporation of myc-tagged GLUT4 into the plasma membrane, and this response was restored by co-overexpression of VAMP-2 linked to green fluorescent protein. Antibodies to VAP-33 microinjected into 3T3-L1 adipocytes abrogated the insulin-stimulated translocation of GLUT4 to the plasma membrane, as measured in adhered plasma membrane lawns. Immunopurified VAMP-2-containing compartments from L6 myotubes and 3T3-L1 adipocytes showed significant levels of VAP-33. We propose that VAP-33 may be a regulator of VAMP-2 availability for GLUT4 traffic and other vesicle fusion events.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Biological Transport, Active / drug effects
  • Carrier Proteins / antagonists & inhibitors
  • Carrier Proteins / genetics
  • Carrier Proteins / metabolism*
  • Clone Cells
  • Endoplasmic Reticulum / metabolism
  • Endosomes / metabolism
  • Glucose Transporter Type 4
  • Golgi Apparatus / metabolism
  • Insulin / pharmacology
  • Mannose-Binding Lectins*
  • Membrane Proteins / antagonists & inhibitors
  • Membrane Proteins / genetics
  • Membrane Proteins / metabolism*
  • Monosaccharide Transport Proteins / metabolism*
  • Muscle Proteins*
  • R-SNARE Proteins
  • Rats
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Transfection
  • Vesicular Transport Proteins*

Substances

  • Carrier Proteins
  • Glucose Transporter Type 4
  • Insulin
  • Lman1 protein, rat
  • Mannose-Binding Lectins
  • Membrane Proteins
  • Monosaccharide Transport Proteins
  • Muscle Proteins
  • R-SNARE Proteins
  • Recombinant Proteins
  • Slc2a4 protein, rat
  • VAPA protein, human
  • Vapa protein, rat
  • Vesicular Transport Proteins