Inositol Phospholipid 3‐Kinase is Activated by Cellular Stress but is not Required for the Stress‐Induced Activation of Glucose Transport in L6 Rat Skeletal Muscle Cells

Abstract

A characteristic response of cells subjected to a stress stimulus is a rapid activation of cellular glucose transport. The mechanisms governing this increase in glucose transport are poorly understood, but it has been suggested that the response may involve the intracellular-signaling components that also participate in the hormonal activation of glucose transport. In skeletal muscle and fat tissue, inositol phospholipid 3-kinase plays an integral role in the regulation of both basal and insulin-stimulated glucose transport. In this study, we have investigated whether inositol phospholipid 3-kinase is activated by chemical stress and, if so, whether it has a role to play in the stress-induced increase in glucose transport in L6 muscle cells. Furthermore, we have attempted to assess the basis by which inositol phospholipid 3-kinase may participate in the regulation of basal glucose transport. Acute exposure (30 min) of L6 muscle cells to 0.5 mM arsenite induced an 80% stimulation in glucose transport. This activation was due to a rise in the number of cell-surface glucose transporters, based on an increase in the V,,,, of glucose transport and the observation that arsenite increases the plasma membrane content of GLUT1 and GLUT4 glucose transporters by 95 % and 60%, respectively, from an intracellular compartment. Arsenite induced rapid activation (< 2 min) of inositol phospholipid 3-kinase with an approximately fourfold increase in phosphatidylinositol 3,4,5-trisphosphate (PtdIns3,4,5P,). In contrast, phosphatidylinositol 3-phosphate (PtdIns3P) levels were unaffected. Prior treatment of L6 cells with 100 nM wortmannin suppressed the arsenite-induced increase in PtdIns3,4,5P3 and reduced the cellular content of PtdIns3P by 50%. Under these conditions however, wortmannin failed to prevent the stress-induced activation of glucose transport, but suppressed basal glucose transport by 60% with an IC,,, of about 10 nM. In the absence of arsenite, wortmannin caused a dose-dependent inhibition in the cellular levels of PtdIns3P and PtdIns3,4,5P3 with IC,, values of about 10 nM and 100 nM, respectively. In summary, the present results demonstrate that chemical stress activates inositol phospholipid 3-kinase and glucose transport in L6 muscle cells, but unlike the hormonal responses of these cells the activation of inositol phospholipid 3-kinase is not responsible for the stress-induced increase in glucose transport. This implies that stress-induced and hormonal stimulated increases in PtdIns3,4,5P, levels are functionally distinct. By contrast, the maintenance of PtdIns3P levels, presumably involving a PtdIns-specific, wortmannin-sensitive inositol phospholipid 3-kinase may be required to support basal glucose transport.

Evidence for a Role of p38 Kinase in Hypoxia-inducible Factor 1-independent Induction of Vascular Endothelial Growth Factor Expression by Sodium Arsenite

“…At least part of these effect are believed to be caused by the ability of trivalent arsenic to bind to SH groups of important cellular (signaling) proteins. Both sodium arsenite and arsenic trioxide can activate PI3K and/or the MAPK family members p44/p42 MAPK, SAPKs/ JNKs, and p38 (48,49). We demonstrated that these kinases were indeed activated by sodium arsenite in OVCAR-3 cells.…”

“…been suggested that sodium arsenite can activate PI3K in some cell types, we investigated the role of this pathway in sodium arsenite-induced HIF-1␣ protein accumulation and VEGF expression in OVCAR-3 cells (48). PI3K-mediated activation of Akt-1, the most frequently studied isoform of Akt, involves phosphorylation on its amino acid residues Ser 473 and Thr 308 .…”

Examination of the effects of arsenic on glucose homeostasis in cell culture and animal studies: Development of a mouse model for arsenic-induced diabetes

“…These studies used high concentrations of arsenicals that are cytotoxic for most cell types. Here, PAO (50 μM) or iAs III (200-1000 μM) have been shown to stimulate basal, insulin-independent, glucose uptake in baby hamster kidney (BHK) cells (Pasternak et al, 1991;Sviderskaya et al, 1996;Warren et al, 1986;Widnell et al, 1990), bovine chromaffin cells (Fladeby and Serck-Hanssen, 1999), 3T3-L1 adipocytes (Bazuine et al, 2004;Bazuine et al, 2003;Gould et al, 1989) and in L6 myotubes (McDowell et al, 1997). A modest 1-3 fold increase in basal glucose uptake was typically found with no effect on the translocation of GLUT4, an insulin-sensitive glucose transporter, to the plasma membrane.…”

“…Increased PI-3K-mediated PKB/Akt phosphorylation has been reported in cells exposed to toxic concentrations (200-500 μM) of iAs III (McDowell et al, 1997;Souza et al, 2001). Stress-induced phosphorylation of PKB/Akt is associated with the activation of pro-survival mechanisms aimed at preventing apoptosis and promoting cell proliferation (Dudek et al, 1997;Ibuki and Goto, 2000;Zhou et al, 2000).…”

Insulin Promotes the Cell Surface Recruitment of the SAT2/ATA2 System A Amino Acid Transporter from an Endosomal Compartment in Skeletal Muscle Cells

“…Subcellular Fractionation of L6 Myotubes-Subcellular membranes from L6 myotubes were isolated as described previously (21)(22)(23)(24). Briefly, 2 h prior to fractionation, cells were washed, and growth medium (containing 5 mM glucose) was supplemented with chloroquine (100 M).…”