Abstract Aims Proprotein convertase subtilisin/kexin type-9 (PCSK9), a molecular determinant of low-density lipoprotein (LDL) receptor (LDLR) fate, has emerged as a promising therapeutic target for atherosclerotic cardiovascular diseases. However, the precise mechanism by which PCSK9 regulates the internalization and lysosomal degradation of LDLR is unknown. Recently, we identified adenylyl cyclase-associated protein 1 (CAP1) as a receptor for human resistin whose globular C-terminus is structurally similar to the C-terminal cysteine-rich domain (CRD) of PCSK9. Herein, we investigated the role of CAP1 in PCSK9-mediated lysosomal degradation of LDLR and plasma LDL cholesterol (LDL-C) levels. Methods and results The direct binding between PCSK9 and CAP1 was confirmed by immunoprecipitation assay, far-western blot, biomolecular fluorescence complementation, and surface plasmon resonance assay. Fine mapping revealed that the CRD of PCSK9 binds with the Src homology 3 binding domain (SH3BD) of CAP1. Two loss-of-function polymorphisms found in human PCSK9 (S668R and G670E in CRD) were attributed to a defective interaction with CAP1. siRNA against CAP1 reduced the PCSK9-mediated degradation of LDLR in vitro. We generated CAP1 knock-out mice and found that the viable heterozygous CAP1 knock-out mice had higher protein levels of LDLR and lower LDL-C levels in the liver and plasma, respectively, than the control mice. Mechanistic analysis revealed that PCSK9-induced endocytosis and lysosomal degradation of LDLR were mediated by caveolin but not by clathrin, and they were dependent on binding between CAP1 and caveolin-1. Conclusion We identified CAP1 as a new binding partner of PCSK9 and a key mediator of caveolae-dependent endocytosis and lysosomal degradation of LDLR.

中文翻译：

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环化酶相关蛋白 1 是前蛋白转化酶枯草杆菌蛋白酶/kexin type-9 的结合伴侣，是前蛋白转化酶枯草杆菌蛋白酶/kexin type-9 降解低密度脂蛋白受体所必需的

摘要 目的 Proprotein convertase subtilisin/kexin type-9 (PCSK9) 是低密度脂蛋白 (LDL) 受体 (LDLR) 命运的分子决定因素，已成为动脉粥样硬化心血管疾病的有希望的治疗靶点。然而，PCSK9 调节 LDLR 内化和溶酶体降解的确切机制尚不清楚。最近，我们将腺苷酸环化酶相关蛋白 1 (CAP1) 鉴定为人类抵抗素的受体，其球状 C 末端在结构上类似于 PCSK9 的 C 末端富含半胱氨酸结构域 (CRD)。在此，我们研究了 CAP1 在 PCSK9 介导的 LDLR 溶酶体降解和血浆 LDL 胆固醇 (LDL-C) 水平中的作用。方法和结果 PCSK9 与 CAP1 之间的直接结合通过免疫沉淀试验、远蛋白质印迹、生物分子荧光互补和表面等离子体共振测定。精细定位显示 PCSK9 的 CRD 与 CAP1 的 Src 同源 3 结合域 (SH3BD) 结合。在人类 PCSK9（CRD 中的 S668R 和 G670E）中发现的两种功能缺失多态性归因于与 CAP1 的相互作用缺陷。针对 CAP1 的 siRNA 在体外减少了 PCSK9 介导的 LDLR 降解。我们生成了 CAP1 敲除小鼠，发现与对照小鼠相比，活的杂合 CAP1 敲除小鼠在肝脏和血浆中具有更高的 LDLR 蛋白质水平和更低的 LDL-C 水平。机制分析表明，PCSK9 诱导的 LDLR 内吞作用和溶酶体降解是由小窝蛋白介导的，而不是由网格蛋白介导的，并且它们依赖于 CAP1 和小窝蛋白-1 之间的结合。