SIPBS, University of Strathclyde, Glasgow, United Kingdom.
Institute of Molecular, Cell and Systems Biology, University of Glasgow, Glasgow, United Kingdom.
PeerJ. 2023 Jul 24;11:e15630. doi: 10.7717/peerj.15630. eCollection 2023.
The ability of insulin to stimulate glucose transport in muscle and fat cells is mediated by the regulated delivery of intracellular vesicles containing glucose transporter-4 (GLUT4) to the plasma membrane, a process known to be defective in disease such as Type 2 diabetes. In the absence of insulin, GLUT4 is sequestered in tubules and vesicles within the cytosol, collectively known as the GLUT4 storage compartment. A subset of these vesicles, known as the 'insulin responsive vesicles' are selectively delivered to the cell surface in response to insulin. We have previously identified Syntaxin16 (Sx16) and its cognate Sec1/Munc18 protein family member mVps45 as key regulatory proteins involved in the delivery of GLUT4 into insulin responsive vesicles. Here we show that mutation of a key residue within the Sx16 N-terminus involved in mVps45 binding, and the mutation of the Sx16 binding site in mVps45 both perturb GLUT4 sorting, consistent with an important role of the interaction of these two proteins in GLUT4 trafficking. We identify Threonine-7 (T7) as a site of phosphorylation of Sx16 . Mutation of T7 to D impairs Sx16 binding to mVps45 and overexpression of T7D significantly impaired insulin-stimulated glucose transport in adipocytes. We show that both AMP-activated protein kinase (AMPK) and its relative SIK2 phosphorylate this site. Our data suggest that Sx16 T7 is a potentially important regulatory site for GLUT4 trafficking in adipocytes.
胰岛素刺激肌肉和脂肪细胞葡萄糖转运的能力是通过含有葡萄糖转运蛋白-4（GLUT4）的细胞内囊泡的调节性递送至质膜来介导的，这个过程在 2 型糖尿病等疾病中被证明是有缺陷的。在没有胰岛素的情况下，GLUT4 被隔离在细胞质中的小管和囊泡中，统称为 GLUT4 储存室。这些囊泡中的一部分，称为“胰岛素反应性囊泡”，在胰岛素的作用下被选择性递送至细胞表面。我们之前已经确定 Syntaxin16（Sx16）及其同源的 Sec1/Munc18 蛋白家族成员 mVps45 是参与将 GLUT4 递送至胰岛素反应性囊泡的关键调节蛋白。在这里，我们表明，Sx16 N 端中一个关键残基的突变，该残基参与 mVps45 结合，以及 mVps45 中的 Sx16 结合位点的突变，都扰乱了 GLUT4 的分拣，这与这两种蛋白质在 GLUT4 运输中的相互作用的重要作用一致。我们确定苏氨酸-7（T7）是 Sx16 的磷酸化位点。将 T7 突变为 D 会损害 Sx16 与 mVps45 的结合，并且 T7D 的过表达显著损害了脂肪细胞中胰岛素刺激的葡萄糖转运。我们表明，AMP 激活的蛋白激酶（AMPK）及其相对 SIK2 都可以磷酸化这个位点。我们的数据表明，Sx16 T7 是脂肪细胞中 GLUT4 运输的一个潜在重要调节位点。
