Increases in phosphorylation of the myosin II heavy chain but not regulatory light chains correlate with insulin secretion in rat pancreatic islets and RINm5F cells

Although cytoskeletal proteins such as myosin II are implicated in the cont rol of insulin secretion, their precise role is poorly understood. In other secretory cells myosin II is predominantly regulated via the phosphorylati on of the regulatory light chains (RLC). The current study was aimed at inv estigating RLC phosphorylation in p-cells. In both the insulin-secreting ce ll line RINm5F and rat pancreatic islets, the RLC was basally phosphorylate d on the myosin light chain kinase sites (Ser(19)/Thr(18)). Phosphorylation at these sites was not consistently increased by either metabolic stimuli (glyceraldehyde/glucose) or the depolarizing agent KCl. The RLC sites phosp horylated by protein kinase C (PKC) (Ser(1)/Ser(2)) mere unphosphorylated i n the basal state, not affected by nutrients or KCl, and only slightly incr eased by the PKC activator phorbol 12-myristate 13-acetate (PMA). Like the other insulin secretagogues, however, PMA did promote serine phosphorylatio n of the myosin heavy chain (MHC) in RINm5F cells. Phosphopeptide mapping s uggested that the same peptide was phosphorylated under both PMA and glycer aldehyde stimulation, which further extends our previous study of the Ca2+- dependent phosphorylation of this protein (Wilson JR, Ludowyke RI, Biden TJ : Nutrient stimulation results in a rapid Ca2+-dependent threonine phosphor ylation of myosin heavy chain in rat pancreatic islets and RINm5F cells. J Biol Chem 273:22729-22737, 1998). Overall, our results demonstrate that in RINm5F cells and rat pancreatic islets, MHC phosphorylation correlates bett er with insulin secretion than phosphorylation of the RLC. We therefore pro pose that in beta-cells, in contrast to other secretory cells, phosphorylat ion of the MHC is more important than that of the RLC for regulation of the myosin II protein complex during insulin secretion.