Corradi Gerardo R, Mazzitelli Luciana R, Petrovich Guido D, de Tezanos Pinto Felicitas, Rochi Lucia, Adamo Hugo P
Departamento de Química Biológica, Instituto de Química y Fisicoquímica Biológicas, Facultad de Farmacia y Bioquímica, Consejo Nacional de Investigaciones Científicas y Técnicas-Universidad de Buenos Aires, Buenos Aires, Argentina.
Front Cell Neurosci. 2021 Aug 26;15:668371. doi: 10.3389/fncel.2021.668371. eCollection 2021.
The plasma membrane Ca pumps (PMCA) are P-ATPases that control Ca signaling and homeostasis by transporting Ca out of the eukaryotic cell. Humans have four genes that code for PMCA isoforms (PMCA1-4). A large diversity of PMCA isoforms is generated by alternative mRNA splicing at sites A and C. The different PMCA isoforms are expressed in a cell-type and developmental-specific manner and exhibit differential sensitivity to a great number of regulatory mechanisms. PMCA4 has two A splice variants, the forms "x" and "z". While PMCA4x is ubiquitously expressed and relatively well-studied, PMCA4z is less characterized and its expression is restricted to some tissues such as the brain and heart muscle. PMCA4z lacks a stretch of 12 amino acids in the so-called A-M3 linker, a conformation-sensitive region of the molecule connecting the actuator domain (A) with the third transmembrane segment (M3). We expressed in yeast PMCA4 variants "x" and "z", maintaining constant the most frequent splice variant "b" at the C-terminal end, and obtained purified preparations of both proteins. In the basal autoinhibited state, PMCA4zb showed a higher ATPase activity and a higher apparent Ca affinity than PMCA4xb. Both isoforms were stimulated by calmodulin but PMCA4zb was more strongly activated by acidic lipids than PMCA4xb. The results indicate that a PMCA4 intrinsically more active and more responsive to acidic lipids is produced by the variant "z" of the splicing site A.
质膜钙泵（PMCA）是一种P型ATP酶，通过将钙离子转运出真核细胞来控制钙信号传导和体内平衡。人类有四个编码PMCA亚型（PMCA1 - 4）的基因。通过A和C位点的可变mRNA剪接产生了多种PMCA亚型。不同的PMCA亚型以细胞类型和发育特异性的方式表达，并且对大量调节机制表现出不同的敏感性。PMCA4有两种A剪接变体，即“x”和“z”形式。虽然PMCA4x广泛表达且研究相对充分，但PMCA4z的特征较少，其表达仅限于某些组织，如脑和心肌。PMCA4z在所谓的A - M3连接区缺少一段12个氨基酸的序列 , 该区域是连接驱动域（A）和第三个跨膜片段（M3）的分子构象敏感区。我们在酵母中表达了PMCA4变体“x”和“z”，在C末端保持最常见的剪接变体“b”不变，并获得了两种蛋白质的纯化制剂。在基础自抑制状态下，PMCA4zb比PMCA4xb表现出更高的ATP酶活性和更高的表观钙亲和力。两种亚型都受到钙调蛋白的刺激，但PMCA4zb比PMCA4xb更强烈地被酸性脂质激活。结果表明，剪接位点A的变体“z”产生了一种本质上更活跃且对酸性脂质更敏感的PMCA4。
