Division of Biochemistry, Chungbuk National University, Cheongju, 361-763, Korea.
Mol Cells. 2013 Jan;35(1):41-6. doi: 10.1007/s10059-013-2268-7. Epub 2013 Jan 9.
Store-operated calcium entry (SOCE) channels composed of Stim and Orai proteins play a critical role in diverse biological processes. Upon endoplasmic reticulum (ER)-mediated calcium (Ca(2+)) depletion, Stim proteins oligomerize with Orai to initiate Ca(2+) influx across the plasma membrane. The ubiquitin-like (UBL) and ubiquitin-associated (UBA) domains of ubiquilin 1 are involved in the degradation of presenilin and polyglutamine proteins. Through screening of Orai1 interaction partner(s) that might have an effect on SOCE, ubiquilin 1 was identified as a target of Orai1. However, the UBL and UBA domains of ubiquilin 1 were dispensable for this interaction. Additionally, ubiquilin 1 and Orai1 colocalized in the cytosolic compartment. Ubiquilin 1 increased the ubiquitination of Orai1, resulting in the formation of a high-molecular-weight form. MG132, a proteasome inhibitor, failed to block the degradation of Orai1, whereas bafilomycin A, a lysosome inhibitor, prevented Orai1 degradation. Confocal microscopy studies demonstrated that a fraction of Orai1 colocalized with ubiquilin 1 and the autophagosomal marker LC3. Because Orai1 is a constituent of SOCE, we determined the effect of ubiquilin 1 on Orai1-mediated Ca(2+) influx. As we expected, intracellular Ca(2+) mobilization, a process normally potentiated by Orai1, was downregulated by ubiquilin 1. Taken together, these findings suggest that ubiquilin 1 downregulates intracellular Ca(2+) mobilization and its downstream signaling by promoting the ubiquitination and lysosomal degradation of Orai1.
钙库操纵性钙内流（SOCE）通道由 Stim 和 Orai 蛋白组成，在多种生物过程中发挥着关键作用。在内质网（ER）介导的钙离子（Ca(2+)）耗竭后，Stim 蛋白寡聚化与 Orai 结合，启动质膜 Ca(2+)内流。泛素样（UBL）和泛素相关（UBA）结构域的泛素结合蛋白 1 参与早老素和多聚谷氨酰胺蛋白的降解。通过筛选可能影响 SOCE 的 Orai1 相互作用伙伴，发现泛素结合蛋白 1 是 Orai1 的靶标。然而，泛素结合蛋白 1 的 UBL 和 UBA 结构域对于这种相互作用是可有可无的。此外，泛素结合蛋白 1 和 Orai1 在细胞质区室中共定位。泛素结合蛋白 1 增加了 Orai1 的泛素化，导致形成高分子量形式。蛋白酶体抑制剂 MG132 未能阻止 Orai1 的降解，而溶酶体抑制剂巴弗洛霉素 A 则阻止了 Orai1 的降解。共聚焦显微镜研究表明，一部分 Orai1 与泛素结合蛋白 1 和自噬体标记物 LC3 共定位。由于 Orai1 是 SOCE 的组成部分，我们确定了泛素结合蛋白 1 对 Orai1 介导的 Ca(2+)内流的影响。正如我们所预期的，通常由 Orai1 增强的细胞内 Ca(2+)动员过程被泛素结合蛋白 1 下调。总之，这些发现表明，泛素结合蛋白 1 通过促进 Orai1 的泛素化和溶酶体降解来下调细胞内 Ca(2+)动员及其下游信号转导。
