RG Adipocytes & Metabolism, Institute for Diabetes & Obesity, Helmholtz Diabetes Center, Helmholtz Zentrum München, German Research Center for Environmental Health GmbH, 85764, Neuherberg, Germany; German Center for Diabetes Research (DZD), 85764, Neuherberg, Germany.
Institute for Anatomy, University of Leipzig, 04103, Leipzig, Germany.
Mol Metab. 2022 Jul;61:101508. doi: 10.1016/j.molmet.2022.101508. Epub 2022 May 2.
Brown adipocytes play a key role in maintaining body temperature as well as glucose and lipid homeostasis. However, brown adipocytes need to adapt their thermogenic activity and substrate utilization to changes in nutrient availability. Amongst the multiple factors influencing brown adipocyte activity, autophagy is an important regulatory element of thermogenic capacity and activity. Nevertheless, a specific sensing mechanism of extracellular amino acid availability linking autophagy to nutrient availability in brown adipocytes is unknown.
To characterize the role of the amino acid transporter PAT2/SLC36A2 in brown adipocytes, loss or gain of function of PAT2 were studied with respect to differentiation, subcellular localization, lysosomal activity and autophagy. Activity of vATPase was evaluated by quenching of EGFP fused to LC3 or FITC-dextran loaded lysosomes in brown adipocytes upon amino acid starvation, whereas the effect of PAT2 on assembly of the vATPase was investigated by Native-PAGE.
We show that PAT2 translocates from the plasma membrane to the lysosome in response to amino acid withdrawal. Loss or overexpression of PAT2 impair lysosomal acidification and starvation-induced S6K re-phosphorylation, as PAT2 facilitates the assembly of the lysosomal vATPase, by recruitment of the cytoplasmic V1 subunit to the lysosome.
PAT2 is an important sensor of extracellular amino acids and regulator of lysosomal acidification in brown adipocytes.
棕色脂肪细胞在维持体温以及葡萄糖和脂质稳态方面发挥着关键作用。然而，棕色脂肪细胞需要适应其产热活性和底物利用，以应对营养物质可用性的变化。在影响棕色脂肪细胞活性的多种因素中，自噬是产热能力和活性的重要调节因素。然而，将自噬与棕色脂肪细胞中营养物质可用性联系起来的细胞外氨基酸可用性的特定感应机制尚不清楚。
为了研究氨基酸转运蛋白 PAT2/SLC36A2 在棕色脂肪细胞中的作用，研究了 PAT2 的缺失或功能获得对分化、亚细胞定位、溶酶体活性和自噬的影响。通过在氨基酸饥饿时猝灭与 LC3 融合的 EGFP 或 FITC-葡聚糖加载的溶酶体，评估 vATPase 的活性，而 PAT2 对 vATPase 组装的影响则通过 Native-PAGE 进行研究。
我们表明，PAT2 响应氨基酸耗竭从质膜转位到溶酶体。PAT2 的缺失或过表达会损害溶酶体酸化和饥饿诱导的 S6K 再磷酸化，因为 PAT2 通过将细胞质 V1 亚基募集到溶酶体来促进溶酶体 vATPase 的组装。
PAT2 是细胞外氨基酸的重要传感器，也是棕色脂肪细胞溶酶体酸化的调节剂。
