Activation of the ERK Pathway and Atypical Protein Kinase C Isoforms in Exercise- and Aminoimidazole-4-carboxamide- 1-β-d-riboside (AICAR)-stimulated Glucose Transport

Abstract

Exercise increases glucose transport in muscle by activating 5-AMP-activated protein kinase (AMPK), but subsequent events are unclear. Presently, we examined the possibility that AMPK increases glucose transport through atypical protein kinase Cs (aPKCs) by activating proline-rich tyrosine kinase-2 (PYK2), ERK pathway components, and phospholipase D (PLD). In mice, treadmill exercise rapidly activated ERK and aPKCs in mouse vastus lateralis muscles. In rat extensor digitorum longus (EDL) muscles, (a) AMPK activator, 5-aminoimidazole-4-carboxamide-1-␤-D-riboside (AICAR), activated PYK2, ERK and aPKCs; (b) effects of AICAR on ERK and aPKCs were blocked by tyrosine kinase inhibitor, genistein, and MEK1 inhibitor, PD98059; and (c) effects of AICAR on aPKCs and 2-deoxyglucose (2-DOG) uptake were inhibited by genistein, PD98059, and PLD-inhibitor, 1-butanol. Similarly, in L6 myotubes, (a) AICAR activated PYK2, ERK, PLD, and aPKCs; (b) effects of AICAR on ERK were inhibited by genistein, PD98059, and expression of dominant-negative PYK2; (c) effects of AICAR on PLD were inhibited by MEK1 inhibitor UO126; (d) effects of AICAR on aPKCs were inhibited by genistein, PD98059, 1-butanol, and expression of dominant-negative forms of PYK2, GRB2, SOS, RAS, RAF, and ERK; and (e) effects of AICAR on 2DOG uptake/GLUT4 translocation were inhibited by genistein, PD98059, UO126, 1-butanol, cell-permeable myristoylated PKCpseudosubstrate, and expression of kinase-inactive RAF, ERK, and PKC-. AMPK activator dinitrophenol had effects on ERK, aPKCs, and 2-DOG uptake similar to those of AICAR. Our findings suggest that effects of exercise on glucose transport that are dependent on AMPK are mediated via PYK2, the ERK pathway, PLD, and aPKCs.

A pharmacological activator of AMP-activated protein kinase (AMPK) induces astrocyte stellation

“…Possible candidates would be atypical PKC isoforms. Incubation of isolated rat extensor digitorum longus muscles with two different pharmacological activators of AMPK, AICAR and dinitrophenol, resulted in PKC zeta activation [13]. In epithelial cells, polarity is regulated by activation of Par4, a LKB1 homolog [7].…”

Metformin improves atypical protein kinase C activation by insulin and phosphatidylinositol-3,4,5-(PO4)3 in muscle of diabetic subjects

“…Samples were homogenised (Polytron) in buffer containing 20 mmol/l Tris-HCl (pH 7.5), 50 mmol/l sucrose, 2 mmol/l EDTA, 2 mmol/l EGTA, 2 mmol/l Na 3 VO 4 , 2 mmol/l NaF, 2 mmol/l Na 4 P 2 O 7 , 1 mmol/l phenylmethylsulphonylfluoride (PMSF), 20 μg/ml leupeptin, 10 μg/ml aprotinin and 1 μmol/l LR-microcytstin, as described [15,16,19]. Muscle homogenates were supplemented with 150 mmol/l NaCl, 1% Triton X-100 and 0.5% Nonidet, and 500 μg of lysate were protein-immunoprecipitated at 4°C with antiserum for determination of total aPKC or IRS-1-dependent PI 3-kinase enzyme activity.…”

“…In muscle preparations of rodents and cultured L6 myotubes, substances that activate AMPK -e.g., aminoimidazole-4-carboxamide-1-β-D-riboside (AICAR), which enters cells and mimics 5'-AMP, and dinitrophenol (DNP), which uncouples mitochondrial oxidative phosphorylation, thereby increasing 5'-AMP levels -also activate atypical protein kinase C (aPKC) isoforms, PKC-ζ and PKC-λ. These are required for increases in GLUT4 translocation to the plasma membrane and glucose transport during AICAR and DNP action in L6 myotubes [15]. Importantly, this activation of aPKC by AMPK occurs in the absence of activation of phosphatidylinositol (PI) 3-kinase, which mediates increases in aPKC and protein kinase B (PKB/ Akt) activity during insulin-stimulated GLUT4 translocation and glucose transport.…”

“…Importantly, this activation of aPKC by AMPK occurs in the absence of activation of phosphatidylinositol (PI) 3-kinase, which mediates increases in aPKC and protein kinase B (PKB/ Akt) activity during insulin-stimulated GLUT4 translocation and glucose transport. Instead, increases in aPKC activity that accompany AMPK activation during AICAR and DNP action appear to depend upon the activation of proline-rich tyrosine kinase-2, extracellular receptor-activated kinase and phospholipase D, which generates the acidic phospholipid, phosphatidic acid (PA), a direct activator of aPKCs [15].…”

β2-Adrenergic activation increases glycogen synthesis in L6 skeletal muscle cells through a signalling pathway independent of cyclic AMP

“…AICAR increases glucose uptake in L6 skeletal muscle cells [29,44]. We have previously shown that only α 1 -adrenergic activation phosphorylates AMPK in these cells and AMPK activation contributes to α 1 -adrenergic-mediated increases in glucose uptake [29].…”