Pan Z Q, Reardon J T, Li L, Flores-Rozas H, Legerski R, Sancar A, Hurwitz J
Graduate Program in Molecular Biology, Memorial Sloan-Kettering Cancer Center, New York, New York 10021, USA.
J Biol Chem. 1995 Sep 15;270(37):22008-16. doi: 10.1074/jbc.270.37.22008.
p21, a p53-induced gene product that blocks cell cycle progression at the G1 phase, interacts with both cyclin-dependent kinases and proliferating cell nuclear antigen (PCNA). PCNA functions as a processivity factor for DNA polymerases delta and epsilon and is required for both DNA replication and nucleotide excision repair. Previous studies have shown that p21 inhibits simian virus 40 (SV40) DNA replication in HeLa cell extracts by interacting with PCNA. In this report we show that p21 blocks nucleotide excision repair of DNA that has been damaged by either ultraviolet radiation or alkylating agents, and that this inhibition can be reversed following addition of PCNA. We have determined that p21 is more effective in blocking DNA resynthesis than in inhibiting the excision step. We further show that a peptide derived from the carboxyl terminus of p21, which specifically interacts with PCNA, inhibits polymerase delta-catalyzed elongation of DNA chains almost stoichiometrically relative to the concentration of PCNA. When added at higher levels, this peptide also blocks both SV40 DNA replication and nucleotide excision repair in HeLa cell extracts. These results indicate that p21 interferes with the function of PCNA in both in vitro DNA replication and nucleotide excision repair.
p21是一种由p53诱导产生的基因产物，它在G1期阻断细胞周期进程，能与细胞周期蛋白依赖性激酶和增殖细胞核抗原（PCNA）相互作用。PCNA作为DNA聚合酶δ和ε的持续合成因子，在DNA复制和核苷酸切除修复过程中均发挥作用。先前的研究表明，p21通过与PCNA相互作用，抑制HeLa细胞提取物中的猿猴病毒40（SV40）DNA复制。在本报告中，我们发现p21可阻断因紫外线辐射或烷化剂而受损的DNA的核苷酸切除修复，且加入PCNA后这种抑制作用可被逆转。我们还确定，p21在阻断DNA再合成方面比抑制切除步骤更有效。我们进一步表明，源自p21羧基末端的一种肽，其能与PCNA特异性相互作用，相对于PCNA的浓度，几乎以化学计量的方式抑制聚合酶δ催化的DNA链延伸。当以更高水平添加时，这种肽还会阻断HeLa细胞提取物中的SV40 DNA复制和核苷酸切除修复。这些结果表明，p21在体外DNA复制和核苷酸切除修复过程中均干扰了PCNA的功能。
