Abstract

Transmembrane protein 168 (TMEM168) was found to be localized on the nuclear membrane. A heterozygous mutation (c.1616G>A, p. R539Q) in TMEM168 was identified in patients with Brugada syndrome. This mutation reduced expression of cardiomyocyte sodium channel Na_v1.5 via Nedd4-2 E3 ubiquitin ligase-induced ubiquitination and degradation. However, the detailed molecular mechanism provoked by the TMEM168 mutant remains unclear. Here, we demonstrated that small heat shock protein αB-crystallin, which can bind to Na_v1.5 and Nedd4-2 and interfere with the association of both proteins, was strongly recruited from the cell surface to the perinuclear region because of the much higher interaction of αB-crystallin with the TMEM168 mutant than with wild-type TMEM168. Following knockdown of αB-crystallin in HL-1 cardiomyocytes, the interaction of Na_v1.5 with Nedd4-2 was increased, despite a reduction of the expression level of Na_v1.5. Moreover, αB-crystallin-mediated reduction of Na_v1.5 expression was rescued in the presence of a proteasome inhibitor MG-132, suggesting the importance of the αB-crystallin-modulated ubiquitin–proteasome system for the stability of Na_v1.5 expression. Collectively, the balance of molecular interactions among Na_v1.5, Nedd4-2, and αB-crystallin plays a role in the regulation of cardiomyocyte cell surface expression of Na_v1.5, and the TMEM168 mutant disturbs this balance, resulting in a decrease in Na_v1.5 expression.

中文翻译：

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跨膜蛋白168突变通过αB-晶状体蛋白细胞内动力学降低Nav1.5的心肌细胞表面表达

摘要

发现跨膜蛋白 168 (TMEM168) 位于核膜上。在 Brugada 综合征患者中发现了 TMEM168 中的杂合突变 (c.1616G>A, p. R539Q)。这种突变通过 Nedd4-2 E3 泛素连接酶诱导的泛素化和降解降低了心肌细胞钠通道 Na _{v 1.5 的表达。}然而，TMEM168 突变体引发的详细分子机制仍不清楚。在这里，我们证明了可以与 Na _{v结合的小热休克蛋白 αB-晶状体蛋白}1.5 和 Nedd4-2 并干扰两种蛋白质的结合，由于 αB-晶状体蛋白与 TMEM168 突变体的相互作用比野生型 TMEM168 高得多，因此从细胞表面强烈募集到核周区域。在 HL-1 心肌细胞中敲低 αB-晶状体蛋白后，尽管 Na v 1.5 的表达水平降低，但 Na _v 1.5 与 Nedd4-2_的相互作用增加。此外，在蛋白酶体抑制剂 MG-132 存在的情况下，αB-晶状体蛋白介导的 Na _v 1.5 表达降低得以挽救，这表明 αB-晶状体蛋白调节的泛素-蛋白酶体系统对于 Na _v 1.5 表达稳定性的重要性。总的来说，Na 之间的分子相互作用的平衡_v 1.5、Nedd4-2 和 αB-晶状体蛋白在 Na _v 1.5 的心肌细胞表面表达的调节中起作用，而 TMEM168 突变体扰乱了这种平衡，导致 Na _v 1.5 表达下降。