Existence of two populations of cyclin/proliferating cell nuclear antigen during the cell cycle: association with DNA replication sites.
Existence of two populations of cyclin/proliferating cell nuclear antigen during the cell cycle: association with DNA replication sites.
复制标题
在细胞周期中存在两个细胞周期蛋白/增殖细胞核抗原的种群：与DNA复制位点的关联。
DOI：
10.1083/jcb.105.4.1549
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发表时间：
1987-10
影响因子：
7.8
通讯作者：
Macdonald-Bravo, H
中科院分区：
文献类型：
--
作者：
Bravo, R;Macdonald-Bravo, H
脉冲追逐实验表明，在3T3细胞从生长到静止的过程中，DNA聚合酶Delta的辅助蛋白Cyclin是稳定的。免疫印迹结合免疫荧光分析表明，静息24小时后细胞周期蛋白的含量是生长细胞的30-40%，呈核质染色。免疫荧光研究表明，在S期存在两种细胞周期蛋白，一种是像静止细胞一样的核质，很容易被洗涤剂提取，另一种与特定的核结构有关。用抗溴脱氧尿核苷免疫荧光检测DNA合成的部位，结果表明，在整个S期，复制子簇的染色模式和它们的出现顺序与细胞周期蛋白的相同。对Triton提取的细胞进行的二维凝胶分析表明，20-30%的细胞周期蛋白仍然与复制子簇有关。这种细胞周期蛋白不能用高盐抽提物从细胞核中释放出来。这表明细胞周期蛋白与DNA复制位点密切相关，它在真核细胞的DNA合成中一定起着重要作用。
Pulse-chase experiments have revealed that cyclin, the auxiliary protein of DNA polymerase-delta, is stable during the transition from growth to quiescence in 3T3 cells. Immunoblotting together with immunofluorescence analysis has shown that the amount of cyclin after 24 h of quiescence is 30-40% of that of growing cells and that it presents a nucleoplasmic staining. Immunofluorescence studies show the existence of two populations of cyclin during the S phase, one that is nucleoplasmic as in quiescent cells and is easily extracted by detergent, and another that is associated to specific nuclear structures. By using antibromodeoxyuridine immunofluorescence to detect the sites of DNA synthesis, it was shown that the staining patterns of the replicon clusters and their order of appearance throughout the S phase are identical to those observed for cyclin. Two-dimensional gel analysis of Triton-extracted cells show that 20-30% of cyclin remains associated with the replicon clusters. This population of cyclin could not be released from the nucleus using high-salt extractions. This demonstrates that cyclin is tightly associated to the sites of DNA replication and that it must have a fundamental role in DNA synthesis in eukaryotic cells.