Subunit Interaction Differences Between the Replication Factor C Complexes in Arabidopsis and Rice.
Subunit Interaction Differences Between the Replication Factor C Complexes in Arabidopsis and Rice.
复制标题
拟南芥和水稻复制因子 C 复合物亚基相互作用的差异
DOI：
10.3389/fpls.2018.00779
复制
发表时间：
2018
影响因子：
5.6
通讯作者：
Zhao J
中科院分区：
文献类型：
--
作者：
Chen Y;Qian J;You L;Zhang X;Jiao J;Liu Y;Zhao J
复制因子C（RFC）是一种多亚基复合物，它以ATP依赖的方式打开滑动夹并将其加载到DNA链上，因此对高速DNA合成至关重要。在酵母（酿酒酵母）和人类，生化研究和结构分析揭示了亚基和架构的钳加载器之间的相互作用模式。ScRFC 1/2/3/4/5的突变导致细胞活力丧失和复制缺陷。然而，除AtRFC 1/3/4外，其他RFC亚基在高等植物中的功能尚不清楚，且其相互作用和排列方式也未得到研究。在此，我们鉴定了拟南芥中的rfc 2 -1/+、rfc 3 -2/+和rfc 5 -1/+突变体，发现胚和胚乳分别停滞在2/4细胞胚和6-8核期。亚细胞定位分析表明，AtRFC 1和OsRFC 1/4/5蛋白定位于细胞核，而AtRFC 2/3/4/5和OsRFC 2/3蛋白既定位于细胞核又定位于细胞质。利用酵母双杂交（Y2 H）和双分子荧光互补（BiFC）技术研究了拟南芥和水稻RFC亚基的相互作用，并提出了RFC复合体中5个亚基的排列顺序分别为AtRFC 5-AtRFC 4-AtRFC 3/2-AtRFC 2/3-AtRFC 1和OsRFC 5-OsRFC 2-OsRFC 3-OsRFC 4-OsRFC 1。此外，AtRFC 1与AtRFC 2/3/4/5在其他亚基存在时也能相互作用，而OsRFC 1则直接与其他4个亚基相互作用。为了进一步表征复合物形成所需的区域，产生了亚基的截短RFC蛋白。结果表明，RFC亚基的C-末端是复合物形成所必需的。我们的研究表明，拟南芥和水稻中的RFCs的定位和相互作用明显不同。
Replication factor C (RFC) is a multisubunit complex that opens the sliding clamp and loads it onto the DNA chain in an ATP-dependent manner and is thus critical for high-speed DNA synthesis. In yeast (Saccharomyces cerevisiae) and humans, biochemical studies and structural analysis revealed interaction patterns between the subunits and architectures of the clamp loaders. Mutations of ScRFC1/2/3/4/5 lead to loss of cell viability and defective replication. However, the functions of RFC subunits in higher plants are unclear, except for AtRFC1/3/4, and the interaction and arrangement of the subunits have not been studied. Here, we identified rfc2-1/+, rfc3-2/+, and rfc5-1/+ mutants in Arabidopsis, and found that embryos and endosperm arrested at the 2/4-celled embryo proper stage and 6-8 nuclei stages, respectively. Subcellular localization analysis revealed that AtRFC1 and OsRFC1/4/5 proteins were localized in the nucleus, while AtRFC2/3/4/5 and OsRFC2/3 proteins were present both in the nucleus and cytoplasm. By using yeast two-hybrid (Y2H) and bimolecular fluorescence complementation (BiFC) techniques, we demonstrated the interactions of Arabidopsis and rice (Oryza sativa) RFC subunits, and proposed arrangements of the five subunits within the RFC complex, which were AtRFC5-AtRFC4-AtRFC3/2-AtRFC2/3-AtRFC1 and OsRFC5-OsRFC2-OsRFC3-OsRFC4-OsRFC1, respectively. In addition, AtRFC1 could interact with AtRFC2/3/4/5 in the presence of other subunits, while OsRFC1 directly interacted with the other four subunits. To further characterize the regions required for complex formation, truncated RFC proteins of the subunits were created. The results showed that C-termini of the RFC subunits are required for complex formation. Our studies indicate that the localization and interactions of RFCs in Arabidopsis and rice are distinctly discrepant.
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影响因子：
4.8
作者：
Ellison, V;Stillman, B
通讯作者：
Stillman, B
DOI：
10.1073/pnas.93.23.12896
发表时间：
1996-11-12
影响因子：
11.1
作者：
Cai, JS;Uhlmann, F;Hurwitz, J
通讯作者：
Hurwitz, J
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影响因子：
6.9
作者：
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通讯作者：
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影响因子：
5.3
作者：
Krause, SA;Loupart, ML;Heck, MMS
通讯作者：
Heck, MMS
影响因子：
2.5
作者：
Gray, FC;MacNeill, SA
通讯作者：
MacNeill, SA