Peracchia C, Wang X G, Peracchia L L
Department of Pharmacology and Physiology, University of Rochester, School of Medicine and Dentistry, 601 Elmwood Avenue, Rochester, NY 14642-8711, USA.
J Membr Biol. 2000 Nov 1;178(1):55-70. doi: 10.1007/s002320010015.
Certain COOH-terminus mutants of connexin32 (Cx32) were previously shown to form channels with unusual transjuctional voltage (V(j)) sensitivity when tested heterotypically in oocytes against Cx32 wild type. Junctional conductance (G(j)) slowly increased by severalfold or decreases to nearly zero with V(j) positive or negative, respectively, at mutant side, and V(j) positive at mutant side reversed CO(2)-induced uncoupling. This suggested that the CO(2)-sensitive gate might be a V(j)-sensitive slow gate. Based on previous data for calmodulin (CaM) involvement in gap junction function, we have hypothesized that the slow gate could be a CaM-like pore plugging molecule (cork gating model). This study describes a similar behavior in heterotypic channels between Cx32 and each of four new Cx32 mutants modified in cytoplasmic-loop and/or COOH-terminus residues. The mutants are: ML/NN+3R/N, 3R/N, ML/NN and ML/EE; in these mutants, N or E replace M105 and L106, and N replace R215, R219 and R220. This study also reports that inhibition of CaM expression strongly reduces V(j) and CO(2) sensitivities of two of the most effective mutants, suggesting a CaM role in slow and chemical gating.
先前的研究表明，连接蛋白32（Cx32）的某些羧基末端突变体在卵母细胞中与野生型Cx32进行异型测试时，会形成具有异常跨结电压（V(j)）敏感性的通道。在突变体一侧，当V(j)为正时，结电导（G(j)）缓慢增加几倍；当V(j)为负时，结电导则降至几乎为零，并且突变体一侧V(j)为正时可逆转CO(2)诱导的解偶联。这表明CO(2)敏感门可能是一个V(j)敏感的慢门。基于先前有关钙调蛋白（CaM）参与缝隙连接功能的数据，我们推测慢门可能是一种类似CaM的孔堵塞分子（软木塞门控模型）。本研究描述了Cx32与四个在胞质环和/或羧基末端残基上进行修饰的新Cx32突变体中的每一个之间在异型通道中的类似行为。这些突变体分别是：ML/NN+3R/N、3R/N、ML/NN和ML/EE；在这些突变体中，N或E取代M105和L106，N取代R215、R219和R220。本研究还报告称，抑制CaM表达会强烈降低两个最有效突变体的V(j)和CO(2)敏感性，这表明CaM在慢门控和化学门控中发挥作用。
