An intron-retaining splice variant of human cyclin A2, expressed in adult differentiated tissues, induces a G1/S cell cycle arrest in vitro.
An intron-retaining splice variant of human cyclin A2, expressed in adult differentiated tissues, induces a G1/S cell cycle arrest in vitro.
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DOI：
10.1371/journal.pone.0039249
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发表时间：
2012
期刊：
影响因子：
3.7
通讯作者：
Faivre J
中科院分区：
文献类型：
--
作者：
Honda A;Valogne Y;Bou Nader M;Bréchot C;Faivre J
人细胞周期蛋白 A2 是 S 期进展和进入有丝分裂的关键调节因子。 G1 和有丝分裂细胞周期蛋白的选择性剪接变体已被证明会干扰全长细胞周期蛋白功能以调节细胞周期进展，因此可能在分化或肿瘤发生中发挥作用。人类细胞周期蛋白 A2 的选择性剪接尚未被研究。序列特异性引物被设计用于扩增细胞系和人体组织中细胞周期蛋白 A2 mRNA 的各个外显子-内含子区域。对保留内含子的 PCR 产物进行克隆和测序，然后在 HeLa 细胞中过表达。使用共聚焦和延时显微镜研究了剪接变体的亚细胞定位，并通过流式细胞术、免疫印迹和组蛋白 H1 激酶活性研究了它们对细胞周期的影响。我们发现了一种名为A2V6的细胞周期蛋白A2 mRNA剪接变体，它部分保留了内含子6。A2V6 mRNA在人体组织中的基因表达模式与野生型细胞周期蛋白A2 (A2WT) mRNA明显不同。它在增殖的胎儿组织中较低，而在某些分化的成人组织中较强，尤其是心脏。在转染的 HeLa 细胞中，A2V6 仅位于细胞质中，而 A2WT 则积累在细胞核中。我们发现 A2V6 诱导明显的 G1/S 细胞周期停滞，与 p21 和 p27 上调以及视网膜母细胞瘤蛋白磷酸化抑制相关。与 A2WT 一样，A2V6 结合 CDK2，但 A2V6/CDK2 复合物不会磷酸化组蛋白 H1。这项研究表明，一些高度分化的人体组织表达保留内含子的细胞周期蛋白 A2 mRNA，可在体外诱导 G1/S 阻滞。与调节细胞增殖的全长细胞周期蛋白 A2 相反，A2V6 剪接变体可能在调节非分裂细胞状态（例如终末分化或衰老）中发挥作用。
Human cyclin A2 is a key regulator of S phase progression and entry into mitosis. Alternative splice variants of the G1 and mitotic cyclins have been shown to interfere with full-length cyclin functions to modulate cell cycle progression and are therefore likely to play a role in differentiation or oncogenesis. The alternative splicing of human cyclin A2 has not yet been studied. Sequence-specific primers were designed to amplify various exon–intron regions of cyclin A2 mRNA in cell lines and human tissues. Intron retaining PCR products were cloned and sequenced and then overexpressed in HeLa cells. The subcellular localization of the splice variants was studied using confocal and time-lapse microscopy, and their impact on the cell cycle by flow cytometry, immunoblotting and histone H1 kinase activity. We found a splice variant of cyclin A2 mRNA called A2V6 that partly retains Intron 6. The gene expression pattern of A2V6 mRNA in human tissues was noticeably different from that of wild-type cyclin A2 (A2WT) mRNA. It was lower in proliferating fetal tissues and stronger in some differentiated adult tissues, especially, heart. In transfected HeLa cells, A2V6 localized exclusively in the cytoplasm whereas A2WT accumulated in the nucleus. We show that A2V6 induced a clear G1/S cell cycle arrest associated with a p21 and p27 upregulation and an inhibition of retinoblastoma protein phosphorylation. Like A2WT, A2V6 bound CDK2, but the A2V6/CDK2 complex did not phosphorylate histone H1. This study has revealed that some highly differentiated human tissues express an intron-retaining cyclin A2 mRNA that induced a G1/S block in vitro. Contrary to full-length cyclin A2, which regulates cell proliferation, the A2V6 splice variant might play a role in regulating nondividing cell states such as terminal differentiation or senescence.
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影响因子：
8
作者：
Mateo, F.;Vidal-Laliena, M.;Canela, N.;Busino, L.;Martinez-Balbas, M. A.;Pagano, M.;Agell, N.;Bachs, O.
通讯作者：
Bachs, O.
影响因子：
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LEES, E;FAHA, B;HARLOW, E
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HARLOW, E