Department of Animal Physiology, Cell Biology and Biosystems Technology, Institute of Biological Sciences, University of Rostock, Albert-Einstein-Str. 3, D-18059, Rostock, Germany.
Eur J Neurosci. 2013 Jul;38(2):2218-28. doi: 10.1111/ejn.12219. Epub 2013 Apr 22.
It was suggested that gap junctional intercellular communication (GJIC) and connexin (Cx) proteins play a crucial role in cell proliferation and differentiation. However, the mechanisms of cell coupling in regulating cell fate during embryonic development are poorly understood. To study the role of GJIC in proliferation and differentiation, we used a human neural progenitor cell line derived from the ventral mesencephalon. Fluorescence recovery after photobleaching (FRAP) showed that dye coupling was extensive in proliferating cells but diminished after the induction of differentiation, as indicated by a 2.5-fold increase of the half-time of fluorescence recovery. Notably, recovery half-time decreased strongly (five-fold) in the later stage of differentiation. Western blot analysis revealed a similar time-dependent expression profile of Cx43, acting as the main gap junction-forming protein. Interestingly, large amounts of cytoplasmic Cx43 were retained mainly in the Golgi network during proliferation but decreased when differentiation was induced. Furthermore, down-regulation of Cx43 by small interfering RNA reduced functional cell coupling, which in turn resulted in a 50% decrease of both the proliferation rate and neuronal differentiation. Our findings suggest a dual function of Cx43 and GJIC in the neural development of ReNcell VM197 human progenitor cells. GJIC accompanied by high Cx43 expression is necessary (1) to maintain cells in a proliferative state and (2) to complete neuronal differentiation, including the establishment of a neural network. However, uncoupling of cells is crucial in the early stage of differentiation during cell fate commitment.
有人提出，缝隙连接细胞间通讯（GJIC）和连接蛋白（Cx）在细胞增殖和分化中起着至关重要的作用。然而，细胞连接在胚胎发育过程中调节细胞命运的机制还知之甚少。为了研究 GJIC 在增殖和分化中的作用，我们使用了一种源自中脑腹侧的人神经祖细胞系。荧光漂白后恢复（FRAP）显示，在增殖细胞中染料偶联非常广泛，但在诱导分化后减少，荧光恢复的半衰期增加了 2.5 倍。值得注意的是，在分化的后期，恢复半衰期明显（五倍）下降。Western blot 分析显示 Cx43 的表达呈相似的时间依赖性，Cx43 作为主要的缝隙连接形成蛋白。有趣的是，大量的细胞质 Cx43 在增殖过程中主要保留在高尔基网络中，但在诱导分化时减少。此外，通过小干扰 RNA 下调 Cx43 减少了功能性细胞偶联，这反过来又导致增殖率和神经元分化降低了 50%。我们的研究结果表明，Cx43 和 GJIC 在 ReNcell VM197 人祖细胞的神经发育中具有双重功能。伴随着高 Cx43 表达的 GJIC 是必需的：（1）维持细胞的增殖状态，（2）完成神经元分化，包括神经网络的建立。然而，在细胞命运决定的早期分化阶段，解偶联对于细胞至关重要。
