Calpain facilitates GLUT4 vesicle translocation during insulin-stimulated glucose uptake in adipocytes

Calpains are a family of non-lysosomal cysteine proteases. Recent studies have identified a member of the calpain family of proteases, calpain 10, as a putative diabetes-susceptibility gene that may be involved in the development of type 2 diabetes. Inhibition of calpain activity has been shown to reduce insulinstimulated glucose uptake in isolated rat-muscle strips and adipocytes. In this report, we examine the mechanism by which calpain affects insulin-stimulated glucose uptake in 3T3-L1 adipocytes. Inhibition of calpain activity resulted in approx. a 60% decrease in insulin-stimulated glucose uptake. Furthermore, inhibition of calpain activity prevented the translocation of insulin-responsive glucose transporter 4 (GLUT4) vesicles to the plasma membrane, as demonstrated by fluorescent microscopy of whole cells and isolated plasma membranes; it did not, however, alter the total GLUT4 protein content. While inhibition of calpain did not affect the insulin-mediated proximal steps of the phosphoinositide 3-kinase pathway, it did prevent the insulin-stimulated cortical actin reorganization required for GLUT4 translocation. Specific inhibition of calpain 10 by antisense expression reduced insulin-stimulated GLUT4 translocation and actin reorganization. Based on these findings, we propose a role for calpain in the actin reorganization required for insulin-stimulated GLUT4 translocation to the plasma membrane in 3T3-L1 adipocytes. These studies identify calpain as a novel factor involved in GLUT4 vesicle trafficking and suggest a link between calpain activity and the development of type 2 diabetes.

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Calpain-10 identified as susceptibility gene for type-2 diabetes

The Lancet ◽

10.1016/s0140-6736(05)72865-8 ◽

2000 ◽

Vol 356 (9236) ◽

pp. 1172

Author(s):

Kathryn Senior

Keyword(s):

Type 2 Diabetes ◽

Susceptibility Gene ◽

Calpain 10

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Loss of HIF-1α impairs GLUT4 translocation and glucose uptake by the skeletal muscle cells

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.00597.2012 ◽

2014 ◽

Vol 306 (9) ◽

pp. E1065-E1076 ◽

Cited By ~ 31

Author(s):

Hidemitsu Sakagami ◽

Yuichi Makino ◽

Katsutoshi Mizumoto ◽

Tsubasa Isoe ◽

Yasutaka Takeda ◽

...

Keyword(s):

Type 2 Diabetes ◽

Skeletal Muscle ◽

Protein Kinase ◽

Glucose Metabolism ◽

Glucose Uptake ◽

Intracellular Signaling ◽

Muscle Cells ◽

Skeletal Muscle Cells ◽

Glut4 Translocation

Defects in glucose uptake by the skeletal muscle cause diseases linked to metabolic disturbance such as type 2 diabetes. The molecular mechanism determining glucose disposal in the skeletal muscle in response to cellular stimuli including insulin, however, remains largely unknown. The hypoxia-inducible factor-1α (HIF-1α) is a transcription factor operating in the cellular adaptive response to hypoxic conditions. Recent studies have uncovered pleiotropic actions of HIF-1α in the homeostatic response to various cellular stimuli, including insulin under normoxic conditions. Thus we hypothesized HIF-1α is involved in the regulation of glucose metabolism stimulated by insulin in the skeletal muscle. To this end, we generated C2C12myocytes in which HIF-1α is knocked down by short-hairpin RNA and examined the intracellular signaling cascade and glucose uptake subsequent to insulin stimulation. Knockdown of HIF-1α expression in the skeletal muscle cells resulted in abrogation of insulin-stimulated glucose uptake associated with impaired mobilization of glucose transporter 4 (GLUT4) to the plasma membrane. Such defect seemed to be caused by reduced phosphorylation of the protein kinase B substrate of 160 kDa (AS160). AS160 phosphorylation and GLUT4 translocation by AMP-activated protein kinase activation were abrogated as well. In addition, expression of the constitutively active mutant of HIF-1α (CA-HIF-1α) or upregulation of endogenous HIF-1α in C2C12cells shows AS160 phosphorylation comparable to the insulin-stimulated level even in the absence of insulin. Accordingly GLUT4 translocation was increased in the cells expressing CA-HIF1α. Taken together, HIF-1α is a determinant for GLUT4-mediated glucose uptake in the skeletal muscle cells thus as a possible target to alleviate impaired glucose metabolism in, e.g., type 2 diabetes.

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Calpain-10 (NIDDM1) as a Susceptibility Gene for Common Type 2 Diabetes

Endocrine Journal ◽

10.1507/endocrj.kr-70 ◽

2006 ◽

Vol 53 (5) ◽

pp. 567-576 ◽

Cited By ~ 10

Author(s):

Yukio HORIKAWA

Keyword(s):

Type 2 Diabetes ◽

Susceptibility Gene ◽

Common Type ◽

Calpain 10

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Huntingtin-associated protein 1 plays an essential role in the pathogenesis of type 2 diabetes by regulating the translocation of GLUT4 in mouse adipocytes

BMJ Open Diabetes Research & Care ◽

10.1136/bmjdrc-2020-001199 ◽

2020 ◽

Vol 8 (1) ◽

pp. e001199

Author(s):

Yan-Ju Gong ◽

Ying Feng ◽

Yuan-Yuan Cao ◽

Jia Zhao ◽

Wei Wu ◽

...

Keyword(s):

Type 2 Diabetes ◽

Body Weight ◽

Adipose Tissue ◽

Glucose Uptake ◽

Hek293 Cells ◽

Diabetic Mice ◽

Glut4 Translocation ◽

Mouse Adipocytes

ObjectiveGlucose disposal by insulin-responsive tissues maintains the body glucose homeostasis and insulin resistance leads to a risk of developing type 2 diabetes (T2DM). Insulin stimulates the translocation of glucose transporter isoform 4 (GLUT4) vesicles from intracellular compartments to the plasma membrane to facilitate glucose uptake. However, the underlying mechanisms of GLUT4 vesicle translocation are not well defined. Here we show the role of huntingtin-associated protein 1 (HAP1) in GLUT4 translocation in adipocytes and the pathogenesis of T2DM.Research design and methodsThe parameters for glucose metabolism including body weight, glucose tolerance and insulin tolerance were assessed in wild-type (WT) and Hap1+/- mice. HAP1 protein expression was verified in adipose tissue. Hap1 mRNA and protein expression was monitored in adipose tissue of high-fat diet (HFD)-induced diabetic mice. Insulin-stimulated GLUT4 vesicle translocation and glucose uptake were detected using immunofluorescence techniques and quantified in primary adipocytes from Hap1-/- mice. The interaction between HAP1 and GLUT4 was assessed by immunofluorescence colocalization and co-immunoprecipitation in HEK293 cells and adipose tissue. The role of sortilin in HAP1 and GLUT4 interaction was approved by co-immunoprecipitation and RNA interference.ResultsThe expression of Hap1 mRNA and protein was detected in WT mouse adipose tissue and downregulated in adipose tissue of HFD-induced diabetic mice. Hap1+/- mice exhibited increased body weight, pronounced glucose tolerance and significant insulin intolerance compared with the WT mice. HAP1 colocalized with GLUT4 in mouse adipocytes and cotransfected HEK293 cells. Furthermore, the insulin-stimulated GLUT4 vesicle translocation and glucose uptake were defective in Hap1-/- adipocytes. Finally, sortilin mediated the interaction of HAP1 and GLUT4.ConclusionsOur study showed that HAP1 formed a protein complex with GLUT4 and sortilin, and played a critical role in insulin-stimulated GLUT4 translocation in adipocytes. Its downregulation may contribute to the pathogenesis of diabetes.

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Impairment of insulin-stimulated GLUT4 translocation in skeletal muscle and adipose tissue in the Tsumura Suzuki obese diabetic mouse: a new genetic animal model of type 2 diabetes

Acta Endocrinologica ◽

10.1530/eje.0.1450785 ◽

2001 ◽

pp. 785-790 ◽

Cited By ~ 45

Author(s):

T Miura ◽

W Suzuki ◽

E Ishihara ◽

I Arai ◽

H Ishida ◽

...

Keyword(s):

Type 2 Diabetes ◽

Skeletal Muscle ◽

Animal Model ◽

Adipose Tissue ◽

Plasma Membrane ◽

Blood Glucose ◽

Low Density ◽

Glut4 Translocation ◽

Genetic Animal Model

BACKGROUND: In skeletal muscle and adipocytes, insulin-stimulated glucose transport has been known to occur through the translocation of glucose transporter (GLUT) 4 from the intracellular pool to the plasma membrane. The Tsumura Suzuki obese diabetic (TSOD) mouse, a new genetic animal model of type 2 diabetes, develops moderate degrees of obesity and diabetes that are especially apparent in animals more than 11 weeks old. A defect in insulin stimulation of GLUT4 translocation also contributes to the characteristics of type 2 diabetes. OBJECTIVE: To characterize this mouse further, we examined the alteration in insulin-stimulated GLUT4 translocation in the skeletal muscle and adipose tissue. METHODS: For glucose and insulin tolerance tests, the mice were given glucose or insulin and blood samples were collected. After isolation of low-density microsomal membrane and plasma membrane from skeletal muscle and adipose tissue, insulin-stimulated translocation of GLUT4 in these TSOD mice was examined by Western blot. RESULTS AND CONCLUSIONS: TSOD mice showed a significant increase in blood glucose after the glucose load, and exhibited a significantly attenuated decrease in blood glucose concentrations after administration of insulin, compared with that in control Tsumura Suzuki non-obese (TSNO) mice. The insulin-stimulated translocation of GLUT4 from low-density microsomal membranes to plasma membrane was significantly reduced in both skeletal muscle and adipose tissue of TSOD mice. These results indicate that the reduced insulin sensitivity in diabetic TSOD mice is presumably due, at least in part, to the impaired GLUT4 translocation by insulin in both skeletal muscle and adipocytes.

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