Browsing by Subject "GLUT4"

Now showing 1 - 5 of 5
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    Chromium Enhances Insulin Responsiveness via AMPK
    (Elsevier, 2014-05) Hoffman, Nolan J.; Penque, Brent A.; Habegger, Kirk M.; Sealls, Whitney; Tackett, Lixuan; Elmendorf, Jeffrey S.; Department of Cellular & Integrative Physiology, IU School of Medicine
    Trivalent chromium (Cr3+) is known to improve glucose homeostasis. Cr3+ has been shown to improve plasma membrane-based aspects of glucose transporter GLUT4 regulation and increase activity of the cellular energy sensor 5′ AMP-activated protein kinase (AMPK). However, the mechanism(s) by which Cr3+ improves insulin responsiveness and whether AMPK mediates this action is not known. In this study we tested if Cr3+ protected against physiological hyperinsulinemia-induced plasma membrane cholesterol accumulation, cortical filamentous actin (F-actin) loss and insulin resistance in L6 skeletal muscle myotubes. In addition, we performed mechanistic studies to test our hypothesis that AMPK mediates the effects of Cr3+ on GLUT4 and glucose transport regulation. Hyperinsulinemia-induced insulin-resistant L6 myotubes displayed excess membrane cholesterol and diminished cortical F-actin essential for effective glucose transport regulation. These membrane and cytoskeletal abnormalities were associated with defects in insulin-stimulated GLUT4 translocation and glucose transport. Supplementing the culture medium with pharmacologically relevant doses of Cr3+ in the picolinate form (CrPic) protected against membrane cholesterol accumulation, F-actin loss, GLUT4 dysregulation and glucose transport dysfunction. Insulin signaling was neither impaired by hyperinsulinemic conditions nor enhanced by CrPic, whereas CrPic increased AMPK signaling. Mechanistically, siRNA-mediated depletion of AMPK abolished the protective effects of CrPic against GLUT4 and glucose transport dysregulation. Together these findings suggest that the micronutrient Cr3+, via increasing AMPK activity, positively impacts skeletal muscle cell insulin sensitivity and glucose transport regulation.
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    DOC2B promotes insulin sensitivity in mice via a novel KLC1-dependent mechanism in skeletal muscle
    (Springer Verlag, 2019-05) Zhang, Jing; Oh, Eunjin; Merz, Karla E.; Aslamy, Arianne; Veluthakal, Rajakrishnan; Salunkhe, Vishal A.; Ahn, Miwon; Tunduguru, Ragadeepthi; Thurmond, Debbie C.; Cellular and Integrative Physiology, School of Medicine
    Aims/hypothesis: Skeletal muscle accounts for >80% of insulin-stimulated glucose uptake; dysfunction of this process underlies insulin resistance and type 2 diabetes. Insulin sensitivity is impaired in mice deficient in the double C2 domain β (DOC2B) protein, while whole-body overexpression of DOC2B enhances insulin sensitivity. Whether insulin sensitivity in the skeletal muscle is affected directly by DOC2B or is secondary to an effect on other tissues is unknown; the underlying molecular mechanisms also remain unclear. Methods: Human skeletal muscle samples from non-diabetic or type 2 diabetic donors were evaluated for loss of DOC2B during diabetes development. For in vivo analysis, new doxycycline-inducible skeletal-muscle-specific Doc2b-overexpressing mice fed standard or high-fat diets were evaluated for insulin and glucose tolerance, and insulin-stimulated GLUT4 accumulation at the plasma membrane (PM). For in vitro analyses, a DOC2B-overexpressing L6-GLUT4-myc myoblast/myotube culture system was coupled with an insulin resistance paradigm. Biochemical and molecular biology methods such as site-directed mutagenesis, co-immunoprecipitation and mass spectrometry were used to identify the molecular mechanisms linking insulin stimulation to DOC2B. Results: We identified loss of DOC2B (55% reduction in RNA and 40% reduction in protein) in the skeletal muscle of human donors with type 2 diabetes. Furthermore, inducible enrichment of DOC2B in skeletal muscle of transgenic mice enhanced whole-body glucose tolerance (AUC decreased by 25% for female mice) and peripheral insulin sensitivity (area over the curve increased by 20% and 26% for female and male mice, respectively) in vivo, underpinned by enhanced insulin-stimulated GLUT4 accumulation at the PM. Moreover, DOC2B enrichment in skeletal muscle protected mice from high-fat-diet-induced peripheral insulin resistance, despite the persistence of obesity. In L6-GLUT4-myc myoblasts, DOC2B enrichment was sufficient to preserve normal insulin-stimulated GLUT4 accumulation at the PM in cells exposed to diabetogenic stimuli. We further identified that DOC2B is phosphorylated on insulin stimulation, enhancing its interaction with a microtubule motor protein, kinesin light chain 1 (KLC1). Mutation of Y301 in DOC2B blocked the insulin-stimulated phosphorylation of DOC2B and interaction with KLC1, and it blunted the ability of DOC2B to enhance insulin-stimulated GLUT4 accumulation at the PM. Conclusions/interpretation: These results suggest that DOC2B collaborates with KLC1 to regulate insulin-stimulated GLUT4 accumulation at the PM and regulates insulin sensitivity. Our observation provides a basis for pursuing DOC2B as a novel drug target in the muscle to prevent/treat type 2 diabetes.
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    Excess membrane cholesterol is an early contributing reversible aspect of skeletal muscle insulin resistance in C57BL/6NJ mice fed a Western-style high-fat diet
    (American Physiological Society, 2019-08-06) Grice, Brian A.; Barton, Kelly J.; Covert, Jacob D.; Kreilach, Alec M.; Tackett, Lixuan; Brozinick, Joseph T.; Elmendorf, Jeffrey S.; Anatomy and Cell Biology, School of Medicine
    Skeletal muscle insulin resistance manifests shortly after high-fat feeding, yet mechanisms are not known. Here we set out to determine whether excess skeletal muscle membrane cholesterol and cytoskeletal derangement known to compromise glucose transporter (GLUT)4 regulation occurs early after high-fat feeding. We fed 6-wk-old male C57BL/6NJ mice either a low-fat (LF, 10% kcal) or a high-fat (HF, 45% kcal) diet for 1 wk. This HF feeding challenge was associated with an increase, albeit slight, in body mass, glucose intolerance, and hyperinsulinemia. Liver analyses did not reveal signs of hepatic insulin resistance; however, skeletal muscle immunoblots of triad-enriched regions containing transverse tubule membrane showed a marked loss of stimulated GLUT4 recruitment. An increase in cholesterol was also found in these fractions from HF-fed mice. These derangements were associated with a marked loss of cortical filamentous actin (F-actin) that is essential for GLUT4 regulation and known to be compromised by increases in membrane cholesterol. Both the withdrawal of the HF diet and two subcutaneous injections of the cholesterol-lowering agent methyl-β-cyclodextrin at 3 and 6 days during the 1-wk HF feeding intervention completely mitigated cholesterol accumulation, cortical F-actin loss, and GLUT4 dysregulation. Moreover, these beneficial membrane/cytoskeletal changes occurred concomitant with a full restoration of metabolic responses. These results identify skeletal muscle membrane cholesterol accumulation as an early, reversible, feature of insulin resistance and suggest cortical F-actin loss as an early derangement of skeletal muscle insulin resistance.
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    Regulation of skeletal muscle insulin sensitivity by PAK1
    (2016-012) Tunduguru, Ragadeepthi; Thurmond, Debbie C.; Elmendorf, Jeffrey S.; Atkinson, Simon J.; Brozinick, Joseph T.; Gunst, Susan J.
    Insulin-stimulated glucose uptake into skeletal muscle cells requires translocation of the glucose transporter-4 (GLUT4) from the cell interior to the plasma membrane. Insulin-stimulated GLUT4 vesicle translocation is dysregulated in Type 2 diabetes (T2D). The Group I p21–activated kinase (PAK1) is a required element in insulin-stimulated GLUT4 vesicle translocation in mouse skeletal muscle in vivo, although its placement and function(s) in the canonical insulin signaling cascade in skeletal muscle cells, remain undetermined. Therefore, the objective of my project is to determine the molecular mechanism(s) underlying the requirement for PAK1 in the process of insulin-stimulated GLUT4 vesicle translocation and subsequent glucose uptake by skeletal muscle cells. Toward this, my studies demonstrate that the pharmacological inhibition of PAK1 activation blunts insulin-stimulated GLUT4 translocation and subsequent glucose uptake into L6-GLUT4myc skeletal myotubes. Inhibition of PAK1 activation also ablates insulin-stimulated F-actin cytoskeletal remodeling, a process known to be required for mobilizing GLUT4 vesicles to the plasma membrane. Consistent with this mechanism, PAK1 activation was also required for the activation of cofilin, another protein implicated in F-actin remodeling. Interestingly, my studies reveal a novel molecular mechanism involving PAK1 signaling to p41-ARC, a regulatory subunit of the cytoskeletal Arp2/3 complex, and its interactions with another cytoskeletal factor, N-WASP, to elicit the insulin-stimulated F-actin remodeling in skeletal muscle cells. Pharmacological inactivation of N-WASP fully abrogated insulin-stimulated GLUT4 vesicle translocation to the cell surface, coordinate with blunted F-actin remodeling. Furthermore, my studies revealed new insulin-induced interactions amongst N WASP, actin, p41-ARC and PAK1; inactivation of PAK1 signaling blocked these dynamic interactions. Taken together, the above studies demonstrate the significance of PAK1 and its downstream signaling to F-actin remodeling in insulin-stimulated GLUT4 vesicle translocation and glucose uptake, revealing new signaling elements that may prove to be promising targets for future therapeutic design.
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    UBXN9 governs GLUT4-mediated spatial confinement of RIG-I-like receptors and signaling
    (Research Square, 2024-06-04) Wang, Penghua; Harrison, Andrew; Yang, Duomeng; Cahoon, Jason; Geng, Tingting; Cao, Ziming; Karginov, Timofey; Chiari, Conner; Li, Xin; Qyang, Yibing; Vella, Anthony; Fan, Zhichao; Vanaja, Sivapriya Kailasan; Rathinam, Vijay; Witczak, Carol; Bogan, Jonathan; Cellular and Integrative Physiology, School of Medicine
    The cytoplasmic RIG-I-like receptors (RLRs) recognize viral RNA and initiate innate antiviral immunity. RLR signaling also triggers glycolytic reprogramming through glucose transporters (GLUTs), whose role in antiviral immunity is elusive. Here, we unveil that insulin-responsive GLUT4 inhibits RLR signaling independently of glucose uptake in adipose and muscle tissues. At steady state, GLUT4 is docked at the Golgi matrix by ubiquitin regulatory X domain 9 (UBXN9, TUG). Following RNA virus infection, GLUT4 is released and translocated to the cell surface where it spatially segregates a significant pool of cytosolic RLRs, preventing them from activating IFN-β responses. UBXN9 deletion prompts constitutive GLUT4 trafficking, sequestration of RLRs, and attenuation of antiviral immunity, whereas GLUT4 deletion heightens RLR signaling. Notably, reduced GLUT4 expression is uniquely associated with human inflammatory myopathies characterized by hyperactive interferon responses. Overall, our results demonstrate a noncanonical UBXN9-GLUT4 axis that controls antiviral immunity via plasma membrane tethering of cytosolic RLRs.