Reynolds N, Warbrick E, Fantes P A, MacNeill S A
Institute of Cell and Molecular Biology, University of Edinburgh, King's Buildings, Mayfield Road, Edinburgh EH9 3JR.
EMBO J. 2000 Mar 1;19(5):1108-18. doi: 10.1093/emboj/19.5.1108.
Direct interaction between DNA polymerase delta and its processivity factor proliferating cell nuclear antigen (PCNA) is essential for effective replication of the eukaryotic genome, yet the precise manner by which this occurs is unclear. We show that the 54 kDa subunit of DNA polymerase delta from Schizosaccharomyces pombe interacts directly with Pcn1 (PCNA) both in vivo and in vitro. Binding is effected via a short sequence at the C-terminus of Cdc27 with significant similarity to the canonical PCNA binding motif first identified in the mammalian p21(Cip1) protein. This motif is both necessary and sufficient for binding of Pcn1 by Cdc27 in vitro and is essential for Cdc27 function in vivo. We also show that the Pcn1 binding motif in Cdc27 is distinct from its binding site for Cdc1, the 55 kDa B-subunit of polymerase delta, and present evidence that Cdc27 can bind to Pcn1 and Cdc1 simultaneously. Finally, we show that Cdc27 performs at least two distinct essential functions, one of which is independent of Pcn1 binding.
DNA聚合酶δ与其持续性因子增殖细胞核抗原（PCNA）之间的直接相互作用对于真核基因组的有效复制至关重要，然而其发生的确切方式尚不清楚。我们发现，粟酒裂殖酵母DNA聚合酶δ的54 kDa亚基在体内和体外均能与Pcn1（PCNA）直接相互作用。结合是通过Cdc27 C末端的一段短序列实现的，该序列与在哺乳动物p21（Cip1）蛋白中首次鉴定出的典型PCNA结合基序具有显著相似性。该基序在体外对于Cdc27结合Pcn1既必要又充分，且在体内对于Cdc27的功能至关重要。我们还表明，Cdc27中的Pcn1结合基序与其与DNA聚合酶δ的55 kDa B亚基Cdc1的结合位点不同，并提供证据表明Cdc27可以同时结合Pcn1和Cdc1。最后，我们表明Cdc27至少执行两种不同的基本功能，其中一种功能独立于Pcn1结合。
