Rac-mediated actin remodeling and myosin II are involved in KATP channel trafficking in pancreatic β-cells

Abstract

AMP-activated protein kinase (AMPK) is a metabolic sensor activated during metabolic stress and it regulates various enzymes and cellular processes to maintain metabolic homeostasis. We previously reported that activation of AMPK by glucose deprivation (GD) and leptin increases KATP currents by increasing the surface levels of KATP channel proteins in pancreatic β-cells. Here, we show that the signaling mechanisms that mediate actin cytoskeleton remodeling are closely associated with AMPK-induced KATP channel trafficking. Using F-actin staining with Alexa 633-conjugated phalloidin, we observed that dense cortical actin filaments present in INS-1 cells cultured in 11 mM glucose were disrupted by GD or leptin treatment. These changes were blocked by inhibiting AMPK using compound C or siAMPK and mimicked by activating AMPK using AICAR, indicating that cytoskeletal remodeling induced by GD or leptin was mediated by AMPK signaling. AMPK activation led to the activation of Rac GTPase and the phosphorylation of myosin regulatory light chain (MRLC). AMPK-dependent actin remodeling induced by GD or leptin was abolished by the inhibition of Rac with a Rac inhibitor (NSC23766), siRac1 or siRac2, and by inhibition of myosin II with a myosin ATPase inhibitor (blebbistatin). Immunocytochemistry, surface biotinylation and electrophysiological analyses of KATP channel activity and membrane potentials revealed that AMPK-dependent KATP channel trafficking to the plasma membrane was also inhibited by NSC23766 or blebbistatin. Taken together, these results indicate that AMPK/Rac-dependent cytoskeletal remodeling associated with myosin II motor function promotes the translocation of KATP channels to the plasma membrane in pancreatic β-cells.

Leptin-induced Trafficking of KATP Channels: A Mechanism to Regulate Pancreatic β-cell Excitability and Insulin Secretion

“…Soon after leptin was found to recruit K ATP channels to the β-cell surface the role of PTEN, which was previously implicated in mediating the effect of leptin on K ATP conductance, was re-examined. In agreement with early studies, inhibition of PTEN’s lipid and phosphatase activities are required for actin remodeling and leptin-induced K ATP channel trafficking [55,56]. However, in contrast to findings by the Ashford lab [23], PTEN inactivation was shown to require phosphorylation by Glycogen Synthase Kinase 3 β (GSK3β) but not Casein Kinase 2 (CK2) [55].…”

“…In an effort to elucidate how PTEN inactivation following leptin stimulation leads to actin remodeling, Han et al examined the role of Rac [56], a Rho family small GTPase molecular switch that is known to regulate actin dynamics for vesicle transport [57]. They showed that Rac activity is increased and that this is necessary for the recruitment of K ATP channels to the β-cell surface [56].…”

Modulation of Ionic Channels and Insulin Secretion by Drugs and Hormones in Pancreatic Beta Cells

“…The trafficking of K1 channels to cell surface could also be considered as a novel autocrine negative feedback mechanism for insulin secretion (Han et al, 2015). In this way in MIN6 cells, high glucose and exogenous insulin treatment increase K ATP channels trafficking to the surface, stabilizing the membrane potential and avoiding insulin secretion.…”

Effect of Combined Treatment With Insulin and Other Hypoglycemic Drugs on 5'amp-Activated Protein Kinase Activity in Leucocytes of Patients With Diabetes Mellitus

“…In β-cells, ATP-sensitive potassium channels (KATP) associate glucose stimulation with membrane depolarization. It was shown that AMPK affects the activity of KATP in β-cells, enhancing their migration to the cell membrane by remodeling the cytoskeleton [19]. The effect of leptin, which inhibits the secretion of insulin, is mediated by AMPK and also associated with increased movement of KATP to the cell membrane [20].…”