Lens ion homeostasis relies on the assembly and/or stability of large connexin 46 gap junction plaques on the broad sides of differentiating fiber cells
Lens ion homeostasis relies on the assembly and/or stability of large connexin 46 gap junction plaques on the broad sides of differentiating fiber cells
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DOI：
10.1152/ajpcell.00372.2014
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发表时间：
2015-05-15
影响因子：
5.5
通讯作者：
Fowler, Velia M.
中科院分区：
文献类型：
--
作者：
Cheng, Catherine;Nowak, Roberta B.;Fowler, Velia M.
眼睛晶状体由多层紧密堆积的纤维细胞组成，形成透明且无血管的器官，对于将光线聚焦到视网膜上非常重要。假设微循环系统由连接蛋白 46 和 50（Cx46 和 Cx50）组成的间隙连接通道网络促进，以维持和滋养晶状体纤维细胞。我们测量了缺乏原调节蛋白 1（Tmod1，一种肌动蛋白尖端加帽蛋白）、CP49（一种晶状体特异性中间丝蛋白）或同时缺乏 Tmod1 和 CP49 的小鼠的晶状体阻抗。我们惊讶地发现，Tmod1 和 CP49 的同时丢失会破坏晶状体纤维细胞中的细胞骨架网络，导致间隙连接耦合电阻、静水压和钠浓度增加。 Tmod1(-/-)中Cx46和Cx50的蛋白水平； CP49(-/-) 双敲除 (DKO) 镜片未发生变化，电子显微镜显示间隙连接正常。然而，免疫染色和三维共焦图像的定量分析表明，Cx46间隙连接斑块在DKO分化纤维细胞中更小且更分散。 DKO 纤维中 Cx50 间隙连接斑块的定位和大小未受影响，表明 Cx46 和 Cx50 形成同聚通道。我们还证明间隙连接斑块位于膜相关肌动蛋白-血影蛋白网络的空隙中，这表明DKO纤维中肌动蛋白-血影蛋白网络的破坏可能会干扰间隙连接斑块积聚成微米大小的区域或改变大斑块的稳定性。这是第一项揭示正常间隙连接斑块定位和大小与正常晶状体耦合电导相关的工作。
The eye lens consists of layers of tightly packed fiber cells, forming a transparent and avascular organ that is important for focusing light onto the retina. A microcirculation system, facilitated by a network of gap junction channels composed of connexins 46 and 50 (Cx46 and Cx50), is hypothesized to maintain and nourish lens fiber cells. We measured lens impedance in mice lacking tropomodulin 1 (Tmod1, an actin pointed-end capping protein), CP49 (a lens-specific intermediate filament protein), or both Tmod1 and CP49. We were surprised to find that simultaneous loss of Tmod1 and CP49, which disrupts cytoskeletal networks in lens fiber cells, results in increased gap junction coupling resistance, hydrostatic pressure, and sodium concentration. Protein levels of Cx46 and Cx50 in Tmod1(-/-); CP49(-/-) doubleknockout (DKO) lenses were unchanged, and electron microscopy revealed normal gap junctions. However, immunostaining and quantitative analysis of three-dimensional confocal images showed that Cx46 gap junction plaques are smaller and more dispersed in DKO differentiating fiber cells. The localization and sizes of Cx50 gap junction plaques in DKO fibers were unaffected, suggesting that Cx46 and Cx50 form homomeric channels. We also demonstrate that gap junction plaques rest in lacunae of the membrane-associated actin-spectrin network, suggesting that disruption of the actin-spectrin network in DKO fibers may interfere with gap junction plaque accretion into micrometer-sized domains or alter the stability of large plaques. This is the first work to reveal that normal gap junction plaque localization and size are associated with normal lens coupling conductance.