Bertolesi G E, Walia Da Silva R, Jollimore C A B, Shi C, Barnes S, Kelly M E M
Department of Ophthalmology and Visual Science, Laboratory of Retina and Optic Nerve Research, Dalhousie University, Halifax, Nova Scotia, Canada.
Neuroscience. 2006 Aug 11;141(1):259-68. doi: 10.1016/j.neuroscience.2006.03.067. Epub 2006 May 6.
A decrease in transient-type calcium channel current, Ca(v)3.1 protein and the mRNA encoding these channels has been reported during differentiation of human retinoblastoma cells. In this study, we examined splice variants of Ca(v)3.1 before and after neuronal differentiation of the Y-79 retinoblastoma cell line to investigate the potential contribution of Ca(v)3.1 to Y-79 differentiation. In Ca(v)3.1, alternative splicing induces variations in three cytoplasmic regions, e.g. the link between domains II and III (producing isoforms e+ and e-), the link between domains III and IV (producing isoforms a, b, ac and bc) and the carboxy terminal region (producing isoforms f and d). Our results demonstrate that Ca(v)3.1e was not expressed in either undifferentiated or differentiated retinoblastoma cells. Splice variants Ca(v)3.1ac; Ca(v)3.1bc and Ca(v)3.1b were all identified in undifferentiated retinoblastoma cells, while expression of these variants in differentiated cells was restricted to the Ca(v)3.1bc isoform. The carboxy terminal variant Ca(v)3.1f is expressed independently of the differentiation status of retinoblastoma cells with or without Ca(v)3.1d. Examination of the functional contribution of Ca(v)3.1 protein to Y-79 cell differentiation revealed that in Y-79 cells transfected with Ca(v)3.1 antisense oligodeoxynucleotides, knockdown of Ca(v)3.1 did not alter the time-course of differentiation or neuritogenesis. The changes in Ca(v)3.1 splice variants were not required for the initiation of differentiation but may be associated with tissue-specific expression or localized alterations in Ca(2+) signaling that are essential for establishment of the mature differentiated phenotype.
据报道，在人视网膜母细胞瘤细胞分化过程中，瞬时型钙通道电流、Ca(v)3.1蛋白以及编码这些通道的mRNA均有所减少。在本研究中，我们检测了Y-79视网膜母细胞瘤细胞系在神经元分化前后Ca(v)3.1的剪接变体，以研究Ca(v)3.1对Y-79分化的潜在作用。在Ca(v)3.1中，可变剪接在三个细胞质区域诱导变异，例如结构域II和III之间的连接（产生异构体e+和e-）、结构域III和IV之间的连接（产生异构体a、b、ac和bc）以及羧基末端区域（产生异构体f和d）。我们的结果表明，Ca(v)3.1e在未分化或分化的视网膜母细胞瘤细胞中均未表达。剪接变体Ca(v)3.1ac、Ca(v)3.1bc和Ca(v)3.1b在未分化的视网膜母细胞瘤细胞中均被鉴定出来，而这些变体在分化细胞中的表达仅限于Ca(v)3.1bc异构体。羧基末端变体Ca(v)3.1f的表达与视网膜母细胞瘤细胞的分化状态无关，无论有无Ca(v)3.1d。对Ca(v)3.1蛋白对Y-79细胞分化的功能贡献的研究表明，在转染了Ca(v)3.1反义寡脱氧核苷酸的Y-79细胞中，Ca(v)3.1的敲低并未改变分化或神经突发生的时间进程。Ca(v)3.1剪接变体的变化对于分化的启动并非必需，但可能与组织特异性表达或Ca(2+)信号传导的局部改变有关，而这些对于成熟分化表型的建立至关重要。
