Insulin stimulates a profound shift in GLUT4
trafficking to the PM. We have recently developed a GLUT4 construct
(rGLUTpHluor), that alongside a specialised workflow allows quantitative
stepwise analysis of GLUT4 trafficking and provides a powerful tool for
pinpointing regulatory nodes that contribute to insulin regulation and insulin
resistance. We have used this methodology to investigate the involvement of the
b isoform of Double C2 domain protein (Doc2B) in insulin-stimulated GLUT4
trafficking. Doc2b has been implicated in the fusion of GLUT4 vesicles with the
PM. Based on its domain structure Doc2B is thought to promote membrane fusion
by inducing positive membrane curvature in a calcium dependant manner (in a
manner analogous to synaptotagmins in other cell types). In our hands, Doc2B
null mice were mildly glucose intolerant when challenged with a glucose bolus.
Insulin-stimulated glucose uptake was compromised in adipose tissue but not
muscle of Doc2b KO mice. This suggests that there is an adipose specific defect
in these mice that is partially responsible for the observed glucose
intolerance. GLUT4 delivery to the PM was impaired in adipocytes isolated from
Doc2B KO mice . Detailed analysis of GLUT4 exocytosis suggests this is due to
defects at two distinct sites within the GLUT4 trafficking pathway. These data
suggest that Doc2b is a key regulator of insulin stimulated GLUT4 exocytosis in
adipocytes.