Expression of a synapsin IIb site 1 phosphorylation mutant in 3T3-L1 adipocytes inhibits basal intracellular retention of Glut4.
Glut4 exocytosis in adipocytes uses protein machinery that is shared with other regulated secretory processes. Synapsins are phosphoproteins that regulate a ;reserve pool' of vesicles clustered behind the active zone in neurons. We found that adipocytes (primary cells and the 3T3-L1 cell line) express synapsin IIb mRNA and protein. Synapsin IIb co-localizes with Glut4 in perinuclear vesicle clusters. To test whether synapsin plays a role in Glut4 traffic, a site 1 phosphorylation mutant (S10A synapsin) was expressed in 3T3-L1 adipocytes. Interestingly, expression of S10A synapsin increased basal cell surface Glut4 almost fourfold (50% maximal insulin effect). Insulin caused a further twofold translocation of Glut4 in these cells. Expression of the N-terminus of S10A synapsin (amino acids 1-118) was sufficient to inhibit basal Glut4 retention. Neither wild-type nor S10D synapsin redistributed Glut4. S10A synapsin did not elevate surface levels of the transferrin receptor in adipocytes or Glut4 in fibroblasts. Therefore, S10A synapsin is inhibiting the specialized process of basal intracellular retention of Glut4 in adipocytes, without affecting general endocytic cycling. While mutant forms of many proteins inhibit Glut4 exocytosis in response to insulin, S10A synapsin is one of only a few that specifically inhibits Glut4 retention in basal adipocytes. These data indicate that the synapsins are important regulators of membrane traffic in many cell types.
Muretta
Joseph M
JM
Department of Biochemistry and Molecular Biology, Mailstop 330, University of Nevada, Reno, NV 89557, USA.
Romenskaia
Irina
I
Cassiday
Patrick A
PA
Mastick
Cynthia Corley
CC
D041721:3T3-L1 Cells; D000256:Adenoviridae / Q000235:genetics; D017667:Adipocytes / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites / Q000235:genetics; D011992:Endosomes / Q000378:metabolism; D005089:Exocytosis; D015870:Gene Expression; D051275:Glucose Transporter Type 4 / Q000037:antagonists & inhibitors*; D006639:Histidine / Q000737:chemistry; D007004:Hypoglycemic Agents / Q000494:pharmacology; D007328:Insulin / Q000494:pharmacology; D051379:Mice; D009154:Mutation; D010766:Phosphorylation; D017434:Protein Structure, Tertiary; D012333:RNA, Messenger / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D013347:Subcellular Fractions / Q000378:metabolism; D016704:Synapsins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
D051275:Glucose Transporter Type 4; D007004:Hypoglycemic Agents; D007328:Insulin; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D016704:Synapsins; D006639:Histidine