The scaffold protein IRS-1 is an essential node in insulin/IGF signaling. It has long been recognized that the stability of IRS-1 is dependent on its endomembrane targeting. However, how IRS-1 targets the intracellular membrane, and what type of intracellular membrane is actually targeted, remains poorly understood. Here, we found that the phase separation-mediated IRS-1 puncta attached to endoplasmic reticulum (ER). VAPB, an ER-anchored protein that mediates tethers between ER and membranes of other organelles, was identified as a direct interacting partner of IRS-1. VAPB mainly binds active IRS-1 because IGF-1 enhanced the VAPB-IRS-1 association and replacing of the nine tyrosine residues of YXXM motifs disrupted the VAPB-IRS-1 association. We further delineated that the Y745 and Y746 residues in the FFAT-like motif of IRS-1 mediated the association with VAPB. Notably, VAPB targeted IRS-1 to the ER and subsequently maintained its stability. Consistently, ablation of VAPB in mice led to downregulation of IRS-1, suppression of insulin signaling, and glucose intolerance. The amyotrophic lateral sclerosis (ALS)-derived VAPB P56S mutant also impaired IRS-1 stability by interfering with the ER-tethering of IRS-1. Our findings thus revealed a previously unappreciated condensate-membrane contact (CMC), by which VAPB stabilizes the membraneless IRS-1 signalosome through targeting it to ER membrane.

支架蛋白 IRS-1 是胰岛素/IGF 信号传导的重要节点。人们早已认识到 IRS-1 的稳定性取决于其内膜靶向。然而，IRS-1 如何靶向细胞内膜，以及实际上靶向什么类型的细胞内膜，仍然知之甚少。在这里，我们发现相分离介导的 IRS-1 斑点附着在内质网 (ER) 上。VAPB 是一种 ER 锚定蛋白，介导 ER 和其他细胞器膜之间的连接，被确定为 IRS-1 的直接相互作用伙伴。VAPB 主要结合活性 IRS-1，因为 IGF-1 增强了 VAPB-IRS-1 关联，并且替换 YXXM 基序的九个酪氨酸残基破坏了 VAPB-IRS-1 关联。我们进一步描述了 IRS-1 的 FFAT 样基序中的 Y745 和 Y746 残基介导了与 VAPB 的关联。值得注意的是，VAPB 将 IRS-1 靶向 ER，并随后保持其稳定性。一致地，消除小鼠体内的 VAPB 会导致 IRS-1 下调、胰岛素信号传导抑制和葡萄糖耐受不良。肌萎缩侧索硬化症 (ALS) 衍生的 VAPB P56S 突变体也通过干扰 IRS-1 的 ER 束缚而损害 IRS-1 的稳定性。因此，我们的研究结果揭示了一种以前未被重视的凝结膜接触（CMC），通过这种接触，VAPB 通过将无膜 IRS-1 信号体靶向 ER 膜来稳定它。