Evidence Coupling Increased Hexosamine Biosynthesis Pathway Activity to Membrane Cholesterol Toxicity and Cortical Filamentous Actin Derangement Contributing to Cellular Insulin Resistance†

Abstract

Hyperinsulinemia is known to promote the progression/worsening of insulin resistance. Evidence reveals a hidden cost of hyperinsulinemia on plasma membrane (PM) phosphatidylinositol 4,5-bisphosphate (PIP(2))-regulated filamentous actin (F-actin) structure, components critical to the normal operation of the insulin-regulated glucose transport system. Here we delineated whether increased glucose flux through the hexosamine biosynthesis pathway (HBP) causes PIP(2)/F-actin dysregulation and subsequent insulin resistance. Increased glycosylation events were detected in 3T3-L1 adipocytes cultured under conditions closely resembling physiological hyperinsulinemia (5 nm insulin; 12 h) and in cells in which HBP activity was amplified by 2 mm glucosamine (GlcN). Both the physiological hyperinsulinemia and experimental GlcN challenge induced comparable losses of PIP(2) and F-actin. In addition to protecting against the insulin-induced membrane/cytoskeletal abnormality and insulin-resistant state, exogenous PIP(2) corrected the GlcN-induced insult on these parameters. Moreover, in accordance with HBP flux directly weakening PIP(2)/F-actin structure, pharmacological inhibition of the rate-limiting HBP enzyme [glutamine-fructose-6-phosphate amidotransferase (GFAT)] restored PIP(2)-regulated F-actin structure and insulin responsiveness. Conversely, overexpression of GFAT was associated with a loss of detectable PM PIP(2) and insulin sensitivity. Even less invasive challenges with glucose, in the absence of insulin, also led to PIP(2)/F-actin dysregulation. Mechanistically we found that increased HBP activity increased PM cholesterol, the removal of which normalized PIP(2)/F-actin levels. Accordingly, these data suggest that glucose transporter-4 functionality, dependent on PIP(2) and/or F-actin status, can be critically compromised by inappropriate HBP activity. Furthermore, these data are consistent with the PM cholesterol accrual/toxicity as a mechanistic basis of the HBP-induced defects in PIP(2)/F-actin structure and impaired glucose transporter-4 regulation.

Exercise training prevents skeletal muscle plasma membrane cholesterol accumulation, cortical actin filament loss, and insulin resistance in C57BL/6J mice fed a western-style high-fat diet

“…In line with increased HBP activity transcriptionally provoking a membrane cholesterol‐based insulin‐resistant state, we found that HBP inhibition attenuated the transcription and expression of genes involved in cholesterol synthesis ( e.g., Hmgcr ) and prevented membrane cholesterol accumulation, F‐actin loss, and GLUT4/glucose transport dysfunction (Bhonagiri et al. 2011; Habegger et al. 2012a; Penque et al.…”

“…We have found that increased hexosamine biosynthesis pathway (HBP) activity in fat/muscle cells increases O‐linked N‐acetylglucosamine modification of the transcription factor Sp1, leading to transcriptional activation of HMG‐CoA reductase, the rate‐limiting enzyme in cholesterol biosynthesis (Bhonagiri et al. 2011; Habegger et al. 2012a; Penque et al.…”

“…Strikingly, inhibition of the HBP or Sp1 binding to DNA blocked both PM cholesterol accumulation, F‐actin loss, and GLUT4/glucose transport dysregulation (Bhonagiri et al. 2011; Habegger et al. 2012a; Penque et al.…”

“…2012a) and that normalization of PM cholesterol restores insulin responsivity (Bhonagiri et al. 2011; Habegger et al. 2012a,b).…”

New Aspects of Cellular Cholesterol Regulation on Blood Glucose Control—Review and Perspective on the Impact of Statin Medications on Metabolic Health

“…Aberrantly increased plasma membrane cholesterol is seen uniformly in insulin-resistant mice, rats, swine, and humans, and normalization restores insulin responsivity. [1][2][3][4] Mechanistic studies in clonal cells, as well as in fat and skeletal muscle tissue demonstrate that excess plasma membrane cholesterol reduces cortical filamentous actin (F-actin), which is essential for glucose transporter type 4 (GLUT4) regulation by insulin. In addition to this negative consequence of excess plasma membrane cholesterol on insulin action, Llanos et al found the ryanodine receptor calcium signals, which are important for GLUT4 regulation, are negatively affected by increased skeletal muscle membrane cholesterol.…”