Marion Bessin Liver Research Center, Albert Einstein College of Medicine and Montefiore Medical Center, Bronx, NY.
Department of Anatomy and Structural Biology, Albert Einstein College of Medicine and Montefiore Medical Center, Bronx, NY.
Hepatology. 2019 Dec;70(6):2156-2170. doi: 10.1002/hep.30772. Epub 2019 Jun 26.
Organic anion transport proteins (OATPs) on the basolateral surface of hepatocytes mediate uptake of a number of drugs and endogenous compounds. Previous studies showed that rat OATP1A1 (rOATP1A1) has a postsynaptic density protein, drosophila disc large tumor suppressor, zonula occludens-1 protein (PDZ) consensus binding motif at its C-terminus and binds to PDZ domain containing 1 (PDZK1), which is required for its cell-surface localization. PDZK1 associates with rOATP1A1-containing endocytic vesicles within cells, mediating recruitment of motor proteins required for microtubule-based trafficking to the plasma membrane. rOATP1A4 also traffics to the plasma membrane, although it lacks a PDZ binding consensus sequence. The current study was designed to test the hypothesis that trafficking of rOATP1A4 to the plasma membrane requires its direct interaction with rOATP1A1 resulting in a complex that traffics through the cell in common subcellular vesicles in which the cytosolic tail of rOATP1A1 is bound to PDZK1. We found that 74% of rOATP1A4-containing rat liver endocytic vesicles (n = 12,044) also contained rOATP1A1. Studies in transfected HEK293 cells showed surface localization of rOATP1A1 only when coexpressed with PDZK1 whereas rOATP1A4 required coexpression with rOATP1A1 and PDZK1. Studies in stably transfected HeLa cells that constitutively expressed PDZK1 showed that coexpression of rOATP1A4 with rOATP1A1 resulted in more rapid appearance of rOATP1A4 on the plasma membrane and faster maturation to its fully glycosylated form. Similar results were observed on immunofluorescence analysis of single cells. Immunoprecipitation of rat liver or transfected HeLa cell lysates with rOATP1A1 antibody specifically co-immunoprecipitated rOATP1A4 as determined by western blotting. Conclusion: These studies indicate that optimal rOATP1A4 trafficking to the cell surface is dependent upon coexpression and interaction with rOATP1A1. As rOATP1A1 binds to the chaperone protein, PDZK1, rOATP1A4 functionally hitchhikes through the cell with this complex.
有机阴离子转运蛋白（OATPs）位于肝细胞的基底外侧表面，介导多种药物和内源性化合物的摄取。先前的研究表明，大鼠 OATP1A1（rOATP1A1）在其 C 末端具有突触后密度蛋白、果蝇 disc large 肿瘤抑制因子、闭合蛋白-1（PDZ）的保守结合基序，与 PDZ 结构域包含蛋白 1（PDZK1）结合，这是其细胞表面定位所必需的。PDZK1 与细胞内含有 rOATP1A1 的内吞小泡结合，介导微管依赖性运输到质膜所需的运动蛋白的募集。rOATP1A4 也转运到质膜，尽管它缺乏 PDZ 结合保守序列。本研究旨在验证以下假设：rOATP1A4 向质膜的运输需要其与 rOATP1A1 的直接相互作用，从而形成一个复合物，通过共同的细胞内小泡运输，其中 rOATP1A1 的胞质尾部与 PDZK1 结合。我们发现，74%的 rOATP1A4 含有大鼠肝内吞小泡（n=12044）也含有 rOATP1A1。在转染的 HEK293 细胞中的研究表明，只有当与 PDZK1 共表达时，rOATP1A1 才能定位于质膜，而 rOATP1A4 则需要与 rOATP1A1 和 PDZK1 共表达。在稳定转染的 HeLa 细胞中的研究表明，rOATP1A4 与 rOATP1A1 共表达导致 rOATP1A4 更快地出现在质膜上，并更快地成熟为完全糖基化形式。在对单个细胞的免疫荧光分析中也观察到类似的结果。用 rOATP1A1 抗体免疫沉淀大鼠肝或转染的 HeLa 细胞裂解物，通过 Western blot 分析特异性共免疫沉淀 rOATP1A4。结论：这些研究表明，rOATP1A4 最佳的质膜转运依赖于与 rOATP1A1 的共表达和相互作用。由于 rOATP1A1 与伴侣蛋白 PDZK1 结合，rOATP1A4 可通过此复合物在细胞内功能性搭便车。
