Distinct and opposing roles for the phosphatidylinositol 3-OH kinase catalytic subunits p110α and p110β in the regulation of insulin secretion from rodent and human beta cells

Abstract

Aims/hypothesis Phosphatidylinositol 3-OH kinases (PI3Ks) regulate beta cell mass, gene transcription, and function, although the contribution of the specific isoforms is unknown. As reduced type 1A PI3K signalling is thought to contribute to impaired insulin secretion, we investigated the role of the type 1A PI3K catalytic subunits α and β (p110α and -β) in insulin granule recruitment and exocytosis in rodent and human islets. Methods The p110α and p110β subunits were inhibited pharmacologically or by small hairpin (sh)RNA-mediated knockdown, and were directly infused or overexpressed in mouse and human islets, beta cells and INS-1 832/13 cells. Glucose-stimulated insulin secretion (GSIS), single-cell exocytosis, Ca 2+ signalling, plasma membrane granule localisation, and actin density were monitored. Results Inhibition or knockdown of p110α increased GSIS. This was not due to altered Ca 2+ responses, depolymerisation of cortical actin or increased cortical granule density, but to enhanced Ca 2+ -dependent exocytosis. Intracellular infusion of recombinant PI3Kα (p110α/p85β) blocked exocytosis. Conversely, knockdown (but not pharmacological inhibition) of p110β blunted GSIS, reduced cortical granule density and impaired exocytosis. Exocytosis was rescued by direct intracellular infusion of recombinant PI3Kβ (p110β/p85β) even when p110β catalytic activity was inhibited. Conversely, both the wild-type p110β and a catalytically inactive mutant directly facilitated exocytosis. Conclusions/interpretation Type 1A PI3K isoforms have distinct and opposing roles in the acute regulation of insulin secretion. While p110α acts as a negative regulator of beta cell exocytosis and insulin secretion, p110β is a positive regulator of insulin secretion through a mechanism separate from its catalytic activity.

Ubiquitous expression of the Pik3ca H1047R mutation promotes hypoglycemia, hypoinsulinemia, and organomegaly

“…The mechanism of the hypoinsulinemia observed in this model remains unclear. Our findings of normal pancreatic insulin staining but reduced glucose stimulated insulin release in the Pik3ca H1047R mice are consistent with the critical role that p110α plays in inhibiting Ca 2+ ‐dependent exocytosis in pancreatic β cells (43, 44). Additionally, the inability to significantly raise insulin serum levels following insulin infusion in the glucose turnover studies suggests that increased clearance is a contributing factor to the hypoinsulinemia.…”

Insulin Secretion Induced by Glucose-dependent Insulinotropic Polypeptide Requires Phosphatidylinositol 3-Kinase γ in Rodent and Human β-Cells

“…Although it is possible, because PI3K␣ and PI3K␤ (class 1A PI3Ks) also regulate insulin secretion (27,30), that these may link the GLP-1-R to actin depolymerization, it should be noted that selective inhibition of PI3K␣ potentiates glucose-stimulated insulin secretion (30). This would be inconsistent with a positive role in GLP-1-dependent insulin secretion.…”

“…INS-1 832/13 cells were a gift of Prof. Christopher Newgard (Duke University). Islet and cell culture was as described previously (29,30). All studies were approved by the Animal Care and Use Committee and the Human Research Ethics Board at the University of Alberta.…”

“…Insulin Secretion Measurements-Islets (either mouse or human) were treated overnight with 1 mol/liter AS604850, (or vehicle) or infected with a p110␥ shRNA adenovirus (or scrambled control) for 72 h. Static insulin secretion measurements were performed at 37°C in KRB, as described previously (29,30). GIP (100 nmol/liter), Ex-4 (100 nmol/liter), GLP-1 (10 nmol/liter), or latrunculin B (10 mol/liter) was present during the 60-min 16.7-mmol/liter glucose KRB stimulation as indicated.…”

“…Experiments were performed at 32-35°C. Solutions used for capacitance measurements are previously described (29,30). For GIP (100 nmol/liter) or Ex-4 (100 nmol/ liter) stimulation experiments, the peptides were added to the bath solution prior to patch-clamping.…”