Direct binding of Cdt2 to PCNA is important for targeting the CRL4Cdt2 E3 ligase activity to Cdt1

被引:16
|
作者
Hayashi, Akiyo [1 ]
Giakoumakis, Nickolaos Nikiforos [2 ]
Heidebrecht, Tatjana [3 ]
Ishii, Takashi [1 ,6 ]
Panagopoulos, Andreas [2 ]
Caillat, Christophe [3 ,7 ]
Takahara, Michiyo [1 ]
Hibbert, Richard G. [3 ,8 ]
Suenaga, Naohiro [1 ]
Stadnik-Spiewak, Magda [3 ]
Takahashi, Tatsuro [5 ]
Shiomi, Yasushi [1 ]
Taraviras, Stavros [4 ]
von Castelmur, Eleonore [3 ]
Lygerou, Zoi [2 ]
Perrakis, Anastassis [3 ]
Nishitani, Hideo [1 ]
机构
[1] Univ Hyogo, Grad Sch Life Sci, Kamigori, Japan
[2] Univ Patras, Sch Med, Dept Biol, Patras, Greece
[3] Netherlands Canc Inst, Dept Biochem, Amsterdam, Netherlands
[4] Univ Patras, Sch Med, Dept Physiol, Patras, Greece
[5] Kyushu Univ, Fac Sci, Fukuoka, Fukuoka, Japan
[6] Fukuoka Dent Coll, Dept Biochem, Fukuoka, Fukuoka, Japan
[7] UVHCI, CNRS, 71 Ave Martyrs, Grenoble, France
[8] Genmab BV, Utrecht, Netherlands
基金
欧洲研究理事会;
关键词
CELL NUCLEAR ANTIGEN; THYMINE DNA GLYCOSYLASE; LICENSING FACTOR CDT1; UBIQUITIN LIGASE; S-PHASE; REPLICATION FACTOR; PIP-BOX; FLUORESCENCE RECOVERY; PROTEIN KINETICS; DAMAGED SITES;
D O I
10.26508/lsa.201800238
中图分类号
Q [生物科学];
学科分类号
07 ; 0710 ; 09 ;
摘要
The CRL4(Cdt2) ubiquitin ligase complex is an essential regulator of cell-cycle progression and genome stability, ubiquitinating substrates such as p21, Set8, and Cdt1, via a display of substrate degrons on proliferating cell nuclear antigens (PCNAs). Here, we examine the hierarchy of the ligase and substrate recruitment kinetics onto PCNA at sites of DNA replication. We demonstrate that the C-terminal end of Cdt2 bears a PCNA interaction protein motif (PIP box, Cdt2(PIP)), which is necessary and sufficient for the binding of Cdt2 to PCNA. Cdt2(PIP) binds PCNA directly with high affinity, two orders of magnitude tighter than the PIP box of Cdt1. X-ray crystallographic structures of PCNA bound to Cdt2(PIP) and Cdt1(PIP) show that the peptides occupy all three binding sites of the trimeric PCNA ring. Mutating Cdt2(PIP) weakens the interaction with PCNA, rendering CRL4(Cdt2) less effective in Cdt1 ubiquitination and leading to defects in Cdt1 degradation. The molecular mechanism we present suggests a new paradigm for bringing substrates to the CRL4-type ligase, where the substrate receptor and substrates bind to a common multivalent docking platform to enable subsequent ubiquitination.
引用
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页数:17
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