Tissue specific regulation of DNA replication during Drosophila development.

据报道，在果蝇的发育过程中观察到了细胞周期的几个变化。幼虫细胞大部分经历内复制周期，包括S期的重复周期。相反，假想的椎间盘细胞按照常规的细胞周期增殖，包括Gl期、S期、G2期和M期。我们已经确定了DNA复制因子140kD亚基的突变，Rf-C是由DREF的异位表达诱导的粗糙眼表型的显性抑制者，DREF是许多参与DNA复制的基因的共同转录因子。突变体在幼虫/蛹阶段死亡，因为在有丝分裂增殖的成像盘细胞中存在DNA复制缺陷，而在幼虫细胞中没有明显的内复制异常。考虑到参与复制的因子是作为母体产物提供的，表型表明，在没有干预有丝分裂的连续几轮S期的复制周期中，复制因子可能比有丝分裂DNA复制中的复制因子更稳定。此外，我们还分离了DNA聚合酶e的基因组克隆和cDNA克隆，并测定了整个一级结构。免疫组织化学分析表明，在果蝇发育过程中，DNA聚合酶e在所有的增殖组织中都有表达。此外，我们通过不精确地切除P元件分离到两个零突变体，从基因外部到TATA盒之前的调节区有1.8kb的长度缺失。这两个零突变体都在胚胎晚期死亡，表明DNA聚合酶e对果蝇DNA复制是必不可少的。缺失突变的杂合子在幼虫和蛹的生长方面也表现出显著的延迟。这些表型表明，DNA聚合酶e的缺失会影响有丝分裂和内复制组织中的DNA复制。

It has been reported that several variations of cell cycle are observed during Drosophila development. Most of larval cells undergoes endoreplication cycle which comprises repeated rounds of S phase. In contrast, the imaginal disc cells proliferate with conventional cell cycle consisted of Gl, S, G2 and M phases. We have identified a mutation for the 140 kD subunit of DNA replication factor, RF-C as a dominant suppressor of rough eye phenotypes induced by ectopic expression of DREF, a common transcription factor for many genes involved in DNAreplication. The mutant dies during larval/pupal stages because of DNA replication defects in imaginal disc cells that are mitotically proliferating, while any abnormalities in endoreplication was not evident in larval cells. Taking together that factors involved in DNA replication were supplied as maternal products, the phenotypes suggests an idea that the replication factor in endoreplication cycles consisting of consecutive rounds of S phases without intervening mitosis could be more stable than that in the mitotic DNA replication. Furthermore, we have isolated the genomic and cDNA clones for DNA polymerase e and determined the whole primary structure. Immunohistochemical analyses indicates that DNA polymerase e expresses in all proliferative tissues during Drosophila development. Also, we isolated two null mutants by imprecise excision of the P element, with a 1.8 kb long deletion in the regulatory region from outside of the gene to just before TATA box. Both null mutants dies at late embryonic stages, indicating the DNA polymerase e is indispensable for Drosophila DNA replication. The heterozygotes for the null mutations also show a significant delay in larval and pupal growth. These phenotypes suggest that a loss of DNA polymerase e affects on DNA replication in mitotic as well as endoreplication tissues.