Nestlé Research, Société des Produits Nestlé S.A., EPFL Innovation Park, Lausanne, 1015, Switzerland; School of Life Sciences, EPFL Innovation Park, Lausanne, 1015, Switzerland.
Centre for Metabolism, Obesity, and Diabetes Research, McMaster University, Hamilton, ON, L8N3Z5, Canada; Department of Medicine and Department of Biochemistry and Biomedical Sciences, McMaster University, Hamilton, ON, L8N3Z5, Canada.
Mol Metab. 2021 Sep;51:101228. doi: 10.1016/j.molmet.2021.101228. Epub 2021 Mar 30.
The metabolic master-switch AMP-activated protein kinase (AMPK) mediates insulin-independent glucose uptake in muscle and regulates the metabolic activity of brown and beige adipose tissue (BAT). The regulatory AMPKγ3 isoform is uniquely expressed in skeletal muscle and potentially in BAT. Herein, we investigated the role that AMPKγ3 plays in mediating skeletal muscle glucose uptake and whole-body glucose clearance in response to small-molecule activators that act on AMPK via distinct mechanisms. We also assessed whether γ3 plays a role in adipose thermogenesis and browning.
Global AMPKγ3 knockout (KO) mice were generated. A systematic whole-body, tissue, and molecular phenotyping linked to glucose homeostasis was performed in γ3 KO and wild-type (WT) mice. Glucose uptake in glycolytic and oxidative skeletal muscle ex vivo as well as blood glucose clearance in response to small molecule AMPK activators that target the nucleotide-binding domain of the γ subunit (AICAR) and allosteric drug and metabolite (ADaM) site located at the interface of the α and β subunit (991, MK-8722) were assessed. Oxygen consumption, thermography, and molecular phenotyping with a β3-adrenergic receptor agonist (CL-316,243) treatment were performed to assess BAT thermogenesis, characteristics, and function.
Genetic ablation of γ3 did not affect body weight, body composition, physical activity, and parameters associated with glucose homeostasis under chow or high-fat diet. γ3 deficiency had no effect on fiber-type composition, mitochondrial content and components, or insulin-stimulated glucose uptake in skeletal muscle. Glycolytic muscles in γ3 KO mice showed a partial loss of AMPKα2 activity, which was associated with reduced levels of AMPKα2 and β2 subunit isoforms. Notably, γ3 deficiency resulted in a selective loss of AICAR-, but not MK-8722-induced blood glucose-lowering in vivo and glucose uptake specifically in glycolytic muscle ex vivo. We detected γ3 in BAT and found that it preferentially interacts with α2 and β2. We observed no differences in oxygen consumption, thermogenesis, morphology of BAT and inguinal white adipose tissue (iWAT), or markers of BAT activity between WT and γ3 KO mice.
These results demonstrate that γ3 plays a key role in mediating AICAR- but not ADaM site binding drug-stimulated blood glucose clearance and glucose uptake specifically in glycolytic skeletal muscle. We also showed that γ3 is dispensable for β3-adrenergic receptor agonist-induced thermogenesis and browning of iWAT.
代谢主开关 AMP 激活的蛋白激酶（AMPK）介导肌肉中的胰岛素非依赖性葡萄糖摄取，并调节棕色和米色脂肪组织（BAT）的代谢活性。调节 AMPKγ3 同工型在骨骼肌中独特表达，并可能在 BAT 中表达。在此，我们研究了 AMPKγ3 在介导对通过不同机制作用于 AMPK 的小分子激活剂的反应中的骨骼肌葡萄糖摄取和全身葡萄糖清除中的作用。我们还评估了 γ3 是否在脂肪产热和棕色化中发挥作用。
生成了全球 AMPKγ3 敲除（KO）小鼠。对 γ3 KO 和野生型（WT）小鼠进行了系统的全身、组织和分子表型与葡萄糖稳态相关的研究。评估了对靶向 γ 亚基核苷酸结合结构域的小分子 AMPK 激活剂（AICAR）和位于 α 和 β 亚基界面的变构药物和代谢物（ADaM）位点的（991，MK-8722）的葡萄糖摄取以及对血糖清除率。进行了耗氧量、热成像和用β3-肾上腺素能受体激动剂（CL-316,243）处理的分子表型分析，以评估 BAT 产热、特征和功能。
γ3 的遗传缺失对在正常饮食或高脂肪饮食下的体重、身体成分、体力活动和与葡萄糖稳态相关的参数没有影响。γ3 缺乏对纤维类型组成、线粒体含量和组成或胰岛素刺激的骨骼肌葡萄糖摄取没有影响。在 γ3 KO 小鼠的糖酵解肌肉中，AMPKα2 活性部分丧失，这与 AMPKα2 和 β2 亚基同工型水平降低有关。值得注意的是，γ3 缺乏导致 AICAR 但不导致 MK-8722 诱导的体内血糖降低和糖酵解肌肉的体外葡萄糖摄取选择性缺失。我们在 BAT 中检测到 γ3，并发现它优先与 α2 和 β2 相互作用。我们在 WT 和 γ3 KO 小鼠之间未观察到耗氧量、产热、BAT 和腹股沟白色脂肪组织（iWAT）的形态或 BAT 活性的标志物之间的差异。
这些结果表明，γ3 在介导 AICAR 但不是 ADaM 结合药物刺激的血糖清除和糖酵解骨骼肌中的葡萄糖摄取中起关键作用。我们还表明，γ3 对于β3-肾上腺素能受体激动剂诱导的 iWAT 产热和棕色化是可有可无的。
