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

Akiyo Hayashi, Nickolaos Nikiforos Giakoumakis, Tatjana Heidebrecht, Takashi Ishii, Andreas Panagopoulos, Christophe Caillat, Michiyo Takahara, Richard G. Hibbert, Naohiro Suenaga, Magda Stadnik-Spiewak, Tatsuro Takahashi, Yasushi Shiomi, Stavros Taraviras, Eleonore von Castelmur, Zoi Lygerou, Anastassis Perrakis, Hideo Nishitani

Research output: Contribution to journalArticlepeer-review

Abstract

The CRL4Cdt2 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 PCNA. 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, Cdt2PIP), which is necessary and sufficient for binding of Cdt2 to PCNA. Cdt2PIP 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 Cdt2PIP and Cdt1PIP show that the peptides occupy all three binding sites of the trimeric PCNA ring. Mutating Cdt2PIP weakens the interaction with PCNA, rendering CRL4Cdt2 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. Summary blurb The C-terminal end of Cdt2 contains a PIP-box for binding to PCNA to promote CRL4Cdt2 function, creating a new paradigm, where the substrate receptor and substrates bind to a common multivalent docking platform for ubiquitination.

Original languageEnglish
JournalUnknown Journal
DOIs
Publication statusPublished - Nov 12 2018

All Science Journal Classification (ASJC) codes

  • Biochemistry, Genetics and Molecular Biology(all)
  • Agricultural and Biological Sciences(all)
  • Immunology and Microbiology(all)
  • Neuroscience(all)
  • Pharmacology, Toxicology and Pharmaceutics(all)

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