Department of Basic and Applied Medical Sciences-Physiology Group, Ghent University, 9000 Ghent, Belgium.
Int J Mol Sci. 2023 Jul 18;24(14):11612. doi: 10.3390/ijms241411612.
Cx43 hemichannels (HCs) and Panx1 channels are two genetically distant protein families. Despite the lack of sequence homology, Cx43 and Panx1 channels have been the subject of debate due to their overlapping expression and the fact that both channels present similarities in terms of their membrane topology and electrical properties. Using the mimetic peptides Gap19 and Panx1, this study aimed to investigate the cross-effects of these peptides on Cx43 HCs and Panx1 channels. The single-channel current activity from stably expressing HeLa-Cx43 and C6-Panx1 cells was recorded using patch-clamp experiments in whole-cell voltage-clamp mode, demonstrating 214 pS and 68 pS average unitary conductances for the respective channels. Gap19 was applied intracellularly while Panx1 was applied extracellularly at different concentrations (100, 200 and 500 μM) and the average nominal open probability (NP) was determined for each testing condition. A concentration of 100 µM Gap19 more than halved the NP of Cx43 HCs, while 200 µM Panx1 was necessary to obtain a half-maximal NP reduction in the Panx1 channels. Gap19 started to significantly inhibit the Panx1 channels at 500 µM, reducing the NP by 26% while reducing the NP of the Cx43 HCs by 84%. In contrast Panx1 significantly reduced the NP of the Cx43 HCs by 37% at 100 µM and by 83% at 200 µM, a concentration that caused the half-maximal inhibition of the Panx1 channels. These results demonstrate that Panx1 inhibits Cx43 HCs over the 100-500 µM concentration range while 500 µM intracellular Gap19 is necessary to observe some inhibition of Panx1 channels.
间隙连接蛋白 43 半通道（HCs）和 Panx1 通道是两个遗传上相距较远的蛋白质家族。尽管缺乏序列同源性，但由于它们的重叠表达以及这两种通道在膜拓扑和电学特性方面具有相似性，因此 Cx43 和 Panx1 通道一直是争议的主题。本研究使用模拟肽 Gap19 和 Panx1 来研究这些肽对 Cx43 HCs 和 Panx1 通道的交叉影响。通过在全细胞膜片钳电压钳模式下记录稳定表达 HeLa-Cx43 和 C6-Panx1 细胞的单通道电流活动，分别证明了各自通道的平均单位电导为 214 pS 和 68 pS。Gap19 被应用于细胞内，而 Panx1 被应用于细胞外，浓度分别为 100、200 和 500 μM，并确定了每种测试条件下的平均名义开放概率（NP）。浓度为 100 μM 的 Gap19 使 Cx43 HCs 的 NP 减少了一半以上，而 200 μM 的 Panx1 则使 Panx1 通道的 NP 减少一半所需的浓度。Gap19 开始在 500 μM 时显著抑制 Panx1 通道，将 NP 降低 26%，同时将 Cx43 HCs 的 NP 降低 84%。相比之下，Panx1 在 100 μM 时使 Cx43 HCs 的 NP 降低 37%，在 200 μM 时降低 83%，这一浓度导致 Panx1 通道的半数最大抑制。这些结果表明，Panx1 在 100-500 μM 浓度范围内抑制 Cx43 HCs，而观察到 Panx1 通道的一些抑制作用则需要 500 μM 的细胞内 Gap19。
