Department of Applied Biological Chemistry, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan.
Biochem Biophys Res Commun. 2011 Jan 28;404(4):946-51. doi: 10.1016/j.bbrc.2010.12.086. Epub 2010 Dec 23.
The polycystic kidney disease 1-like 3 (PKD1L3)-polycystic kidney disease 2-like 1 (PKD2L1) complex functions as a Ca(2+)-permeable, non-selective cation channel that is activated by acid and its subsequent removal; this is called an off-response. In this study, we identified a single aspartic residue in PKD2L1 that is responsible for the Ca(2+) permeation of the PKD1L3/PKD2L1 complex. Calcium imaging analysis using point mutants of negatively charged amino acids present in the putative pore regions of PKD1L3 and PKD2L1 revealed that neutralization of the aspartic residue in PKD2L1 (D523N), which is conserved among PKD2 family members, abolished Ca(2+) permeation, despite robust cell surface expression. In contrast, neutralization of the other negatively charged residues of PKD1L3 (D2049N and E2072Q) and PKD2L1 (D525N and D530N) as well as substitution of Asp(523) with a glutamate residue (D523E) had little effect on Ca(2+) permeation properties. These results demonstrate that Asp(523) in PKD2L1 is a key determinant of Ca(2+) permeation into the PKD1L3/PKD2L1 complex and that PKD2L1 contributes to forming the pore of the PKD1L3/PKD2L1 channel.
多囊肾病 1 样 3 (PKD1L3)-多囊肾病 2 样 1 (PKD2L1) 复合物作为一种 Ca(2+)可渗透、非选择性阳离子通道发挥作用，该通道可被酸激活并随后失活；这被称为关闭反应。在这项研究中，我们鉴定了 PKD2L1 中的一个单一天冬氨酸残基，该残基负责 PKD1L3/PKD2L1 复合物的 Ca(2+)渗透。使用 PKD1L3 和 PKD2L1 中假定的孔区域中带负电荷的氨基酸的点突变体进行钙成像分析表明，尽管 PKD2 家族成员中保守的 PKD2L1 中天冬氨酸残基（D523N）的中和作用，尽管细胞表面表达丰富，但 Ca(2+)渗透被完全消除。相比之下，中和 PKD1L3 的其他带负电荷的残基（D2049N 和 E2072Q）和 PKD2L1（D525N 和 D530N）以及用谷氨酸残基（D523E）取代 Asp(523)对 Ca(2+)渗透特性几乎没有影响。这些结果表明，PKD2L1 中的 Asp(523)是 Ca(2+)进入 PKD1L3/PKD2L1 复合物的关键决定因素，PKD2L1 有助于形成 PKD1L3/PKD2L1 通道的孔。
