BACKGROUND The proliferating cell nuclear antigen (PCNA or PCN-1 in C. elegans), an essential processivity factor for DNA polymerase δ, has been widely used as a marker of S-phase. In C. elegans early embryos, PCN-1 accumulation is cyclic, localizing to the nucleus during S-phase and the cytoplasm during the rest of the cell cycle. The C. elegans larval and adult germline is an important model systems for studying cell cycle regulation, and it was observed that the cell cycle regulator cyclin E (CYE-1 in C. elegans) displays a non-cyclic, continuous accumulation pattern in this tissue. The accumulation pattern of PCN-1 has not been well defined in the larval and adult germline, and the objective of this study was to determine if the accumulation pattern is cyclic, as in other cells and organisms, or continuous, similar to cyclin E. RESULTS To study the larval and adult germline accumulation of PCN-1 expressed from its native locus, we used CRISPR/Cas9 technology to engineer a novel allele of pcn-1 that encodes an epitope-tagged protein. S-phase nuclei were labeled using EdU nucleotide incorporation, and FLAG::PCN-1 was detected by antibody staining. All progenitor zone nuclei, including those that were not in S-phase (as they were negative for EdU staining) showed PCN-1 accumulation, indicating that PCN-1 accumulated during all cell cycle phases in the germline progenitor zone. The same result was observed with a GFP::PCN-1 fusion protein expressed from a transgene. pcn-1 loss-of-function mutations were analyzed, and pcn-1 was necessary for robust fertility and embryonic development. CONCLUSIONS In the C. elegans early embryo as well as other organisms, PCN-1 accumulates in nuclei only during S-phase. By contrast, in the progenitor zone of the germline of C. elegans, PCN-1 accumulated in nuclei during all cell cycle stages. This pattern is similar to accumulation pattern of cyclin E. These observations support the model that mitotic cell cycle regulation in the germline stem and progenitor cells is distinct from somatic cells, as it does not heavily rely on cyclic accumulation of classic cell cycle proteins.

中文翻译：

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增殖细胞核抗原 PCN-1 的细胞周期积累从成年种系中的连续积累转变为秀丽隐杆线虫早期胚胎中的间歇性积累。

背景增殖细胞核抗原（线虫中的PCNA或PCN-1）是DNA聚合酶δ的必需持续合成因子，已被广泛用作S期的标志物。在秀丽隐杆线虫早期胚胎中，PCN-1 积累是周期性的，在 S 期定位于细胞核，在细胞周期的其余时间定位于细胞质。线虫幼虫和成虫种系是研究细胞周期调控的重要模型系统，观察到细胞周期调节因子cyclin E（线虫中的CYE-1）在该系统中表现出非循环、连续积累模式。组织。PCN-1 在幼虫和成虫种系中的积累模式尚未明确，本研究的目的是确定积累模式是否是循环的（如其他细胞和生物体中那样），还是连续的（类似于细胞周期蛋白 E）。结果 为了研究从其天然位点表达的 PCN-1 的幼虫和成虫种系积累，我们使用 CRISPR/Cas9 技术设计了编码表位标记蛋白的新型 PCN-1 等位基因。使用 EdU 核苷酸掺入标记 S 期细胞核，并通过抗体染色检测 FLAG::PCN-1。所有祖细胞区细胞核，包括那些不处于 S 期的细胞核（因为它们的 EdU 染色呈阴性）均显示出 PCN-1 积累，表明 PCN-1 在种系祖细胞区的所有细胞周期阶段均积累。用转基因表达的 GFP::PCN-1 融合蛋白观察到相同的结果。对 pcn-1 功能丧失突变进行了分析，发现 pcn-1 对于强大的生育能力和胚胎发育是必需的。结论 在线虫早期胚胎以及其他生物体中，PCN-1 仅在 S 期在细胞核中积累。相比之下，在秀丽隐杆线虫种系的祖细胞区，PCN-1 在所有细胞周期阶段都在细胞核中积累。这种模式类似于细胞周期蛋白 E 的积累模式。这些观察结果支持这样的模型：生殖干细胞和祖细胞中的有丝分裂细胞周期调节与体细胞不同，因为它并不严重依赖于经典细胞周期蛋白的循环积累。