Madine M A, Khoo C Y, Mills A D, Laskey R A
Wellcome/CRC Institute, Cambridge, UK.
Nature. 1995 Jun 1;375(6530):421-4. doi: 10.1038/375421a0.
An intact nuclear membrane restricts DNA replication to only one round in each cell cycle, apparently by excluding an essential replication-licensing factor throughout interphase. A family of related yeast replication proteins, MCM2, 3 and 5 (also called, after cell-division cycle, CDC46), resemble licensing factor, entering the nucleus only during mitosis. We have cloned a Xenopus homologue of MCM3 (XMCM3) and raised antibodies against expressed protein. Immunodepletion of Xenopus egg extracts removes a complex of MCM2, 3 and 5 homologues and inhibits replication of Xenopus sperm nuclei or permeable G2 HeLa nuclei. However, G1 HeLa nuclei still replicate efficiently. Mock-depleted extracts replicate all three templates. XMCM3 accumulates in nuclei before replication but anti-XMCM3 staining decreases during replication. These results can explain why replicated nuclei are unable to reinitiate replication in a single cell cycle.
完整的核膜将DNA复制限制在每个细胞周期仅一轮，显然是通过在整个间期排除一种必需的复制许可因子来实现的。一族相关的酵母复制蛋白，MCM2、3和5（在细胞分裂周期后也称为CDC46），类似于许可因子，仅在有丝分裂期间进入细胞核。我们克隆了非洲爪蟾MCM3的同源物（XMCM3），并制备了针对表达蛋白的抗体。用非洲爪蟾卵提取物进行免疫去除可去除MCM2、3和5同源物的复合物，并抑制非洲爪蟾精子细胞核或可渗透的G2期HeLa细胞核的复制。然而，G1期HeLa细胞核仍能高效复制。模拟去除提取物能复制所有三种模板。XMCM3在复制前在细胞核中积累，但在复制期间抗XMCM3染色减少。这些结果可以解释为什么复制后的细胞核在单个细胞周期中无法重新启动复制。
