Competition for calnexin binding regulates secretion and turnover of misfolded GPI-anchored proteins.
Competition for calnexin binding regulates secretion and turnover of misfolded GPI-anchored proteins.
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DOI：
10.1083/jcb.202108160
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发表时间：
2023-10-02
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错误折叠的GPI-AP从ER释放到高尔基体，通过组成型活性和应激诱导的GPI-AP途径在溶酶体中快速降解。使用多种互补方法，作者证明新生糖蛋白对钙连接蛋白结合的竞争促进了错误折叠的GPI-AP的释放。在哺乳动物细胞中，错误折叠的糖基磷脂酰肌醇（GPI）锚定蛋白（GPI-AP）通过组成性和应激诱导途径（称为内质网）从内质网清除到高尔基体。从高尔基体，错误折叠的GPI-AP短暂地进入细胞表面，然后快速内化以进行溶酶体降解。在稳态条件下，是什么调节了错误折叠的GPI-AP的释放，以及这种释放在ER应激期间如何加速尚不清楚。使用朊病毒蛋白或CD 59的突变体作为模型错误折叠的GPI-AP，我们证明，诱导钙连接蛋白降解或上调钙连接蛋白结合糖蛋白表达触发释放错误折叠的GPI-AP的pGE 2。相反，阻断蛋白质合成显著抑制错误折叠的GPI-AP从钙连接蛋白的解离和随后的周转。我们证明了新合成的钙连接蛋白底物和与钙连接蛋白共免疫沉淀的BMPs底物之间的负相关性。这些发现暗示了新合成的底物与钙连接蛋白的结合的竞争是调节钙连接蛋白释放错误折叠的GPI-AP以通过β途径进行周转的关键因素。
Misfolded GPI-APs are released from the ER to the Golgi for rapid degradation in lysosomes by the constitutively active and stress-inducible RESET pathway. Using multiple complementary approaches, the authors demonstrate that competition by nascent glycoproteins for calnexin-binding promotes the release of misfolded GPI-APs for RESET. In mammalian cells, misfolded glycosylphosphatidylinositol (GPI)-anchored proteins (GPI-APs) are cleared out of the ER to the Golgi via a constitutive and a stress-inducible pathway called RESET. From the Golgi, misfolded GPI-APs transiently access the cell surface prior to rapid internalization for lysosomal degradation. What regulates the release of misfolded GPI-APs for RESET during steady-state conditions and how this release is accelerated during ER stress is unknown. Using mutants of prion protein or CD59 as model misfolded GPI-APs, we demonstrate that inducing calnexin degradation or upregulating calnexin-binding glycoprotein expression triggers the release of misfolded GPI-APs for RESET. Conversely, blocking protein synthesis dramatically inhibits the dissociation of misfolded GPI-APs from calnexin and subsequent turnover. We demonstrate an inverse correlation between newly synthesized calnexin substrates and RESET substrates that coimmunoprecipitate with calnexin. These findings implicate competition by newly synthesized substrates for association with calnexin as a key factor in regulating the release of misfolded GPI-APs from calnexin for turnover via the RESET pathway.
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