Department of Biochemistry and Cell Biology, School of Medicine, Kyungpook National University, Taegu, Korea.
BK21 FOUR KNU Convergence Educational Program, Department of Biomedical Science, School of Medicine, Kyungpook National University, Taegu, Korea.
Am J Physiol Renal Physiol. 2024 Jan 1;326(1):F69-F85. doi: 10.1152/ajprenal.00144.2023. Epub 2023 Oct 19.
Poly(ADP-ribosyl)ation (PARylation), as a posttranslational modification mediated by poly(ADP-ribose) polymerases (PARPs) catalyzing the transfer of ADP-ribose from NAD molecules to acceptor proteins, involves a number of cellular processes. As mice lacking the PARP-1 gene () produce more urine, we investigated the role of PARP-1, the most prevalent member of the PARP family, in the vasopressin-responsive expression of aquaporin-2 (AQP2). In biotin-conjugated nicotinamide adenine dinucleotide (biotin-NAD) pulldown and immunoprecipitation assays of poly(ADP)-ribose in mpkCCDc14 cells, immunoblots demonstrated that 1-deamino-8-D-arginine vasopressin (dDAVP) induced the PARylation of total proteins, associated with an increase in the cleavage of PARP-1 and cleaved caspase-3 expression. By inhibiting PARP-1 with siRNA, the abundance of dDAVP-induced AQP2 mRNA and protein was significantly diminished. In contrast, despite a substantial decrease in PARylation, the PARP-1 inhibitor (PJ34) had no effect on the dDAVP-induced regulation of AQP2 expression. The findings suggest that PARP-1 protein expression itself, and not PARP-1-mediated PARylation, is necessary for dDAVP-regulated AQP2 expression. Bioinformatic analysis revealed that 408 proteins interact with PARP-1 in the collecting duct (CD) cells of the kidney. Among them, the signaling pathway of the vasopressin V2 receptor was identified for 49 proteins. In particular, β-catenin, which is phosphorylated at Ser by dDAVP, was identified as the PARP-1-interacting protein. A significant decrease of β-catenin phosphorylation (Ser) in response to dDAVP was associated with siRNA-mediated PARP-1 knockdown. Taken together, PARP-1 is likely to play a role in vasopressin-induced AQP2 expression by interacting with β-catenin in renal CD cells. The poly(ADP-ribose) polymerase (PARP) family catalyzes poly(ADP-ribosylation) (PARylation), which is one of the posttranslational modifications of largely undetermined physiological significance. This study investigated the role of PARP-1, the most prevalent member of the PARP family, in the vasopressin-responsive expression of aquaporin-2 (AQP2). The results demonstrated that PARP-1 protein expression itself, and not PARP-1-mediated PARylation, is necessary for dDAVP-regulated AQP2 expression. β-Catenin, which is phosphorylated at Ser by dDAVP, was identified as the PARP-1-interacting protein.
聚（ADP-核糖）化（PAR 化）是一种由聚（ADP-核糖）聚合酶（PARPs）催化的将 ADP-核糖从 NAD 分子转移到受体蛋白上的翻译后修饰，涉及许多细胞过程。由于缺乏 PARP-1 基因的小鼠（）会产生更多的尿液，因此我们研究了 PARP-1 在血管加压素反应性表达水通道蛋白-2（AQP2）中的作用。在 biotin-conjugated nicotinamide adenine dinucleotide（biotin-NAD）pulldown 和 mpkCCDc14 细胞中的多聚（ADP-核糖）的免疫沉淀测定中，免疫印迹表明 1-脱氨基-8-D-精氨酸血管加压素（dDAVP）诱导总蛋白的 PAR 化，与 PARP-1 的切割增加和切割 caspase-3 的表达增加有关。通过 siRNA 抑制 PARP-1，dDAVP 诱导的 AQP2 mRNA 和蛋白的丰度显著减少。相比之下，尽管 PAR 化有很大减少，但 PARP-1 抑制剂（PJ34）对 dDAVP 诱导的 AQP2 表达调节没有影响。这些发现表明，PARP-1 蛋白表达本身，而不是 PARP-1 介导的 PAR 化，是 dDAVP 调节的 AQP2 表达所必需的。生物信息学分析显示，肾脏集合管（CD）细胞中 408 种蛋白与 PARP-1 相互作用。其中，血管加压素 V2 受体的信号通路被确定为 49 种蛋白。特别是，β-连环蛋白，其在 Ser 处被 dDAVP 磷酸化，被鉴定为与 PARP-1 相互作用的蛋白。在对 dDAVP 的反应中，β-连环蛋白磷酸化（Ser）的显著减少与 siRNA 介导的 PARP-1 敲低有关。总之，PARP-1 可能通过与肾脏 CD 细胞中的β-连环蛋白相互作用在血管加压素诱导的 AQP2 表达中发挥作用。多聚（ADP-核糖）聚合酶（PARP）家族催化多聚（ADP-核糖）化（PAR 化），这是一种具有很大未确定生理意义的翻译后修饰之一。本研究调查了 PARP-1，即 PARP 家族中最常见的成员，在血管加压素反应性表达水通道蛋白-2（AQP2）中的作用。结果表明，PARP-1 蛋白表达本身，而不是 PARP-1 介导的 PAR 化，是 dDAVP 调节的 AQP2 表达所必需的。dDAVP 磷酸化的 Ser 处的β-连环蛋白被鉴定为与 PARP-1 相互作用的蛋白。
