1. Chromosomes and Gene Expression
Download icon

Cdc7 activates replication checkpoint by phosphorylating the Chk1 binding domain of Claspin in human cells

  1. Chi-Chun Yang
  2. Hiroyuki Kato
  3. Mayumi Shindo
  4. Hisao Masai  Is a corresponding author
  1. Tokyo Metropolitan Institute of Medical Science, Japan
Research Article
  • Cited 5
  • Views 1,835
  • Annotations
Cite this article as: eLife 2019;8:e50796 doi: 10.7554/eLife.50796

Abstract

Replication checkpoint is essential for maintaining genome integrity in response to various replication stresses as well as during the normal growth. The evolutionally conserved ATR-Claspin-Chk1 pathway is induced during replication checkpoint activation. Cdc7 kinase, required for initiation of DNA replication at replication origins, has been implicated in checkpoint activation but how it is involved in this pathway has not been known. Here, we show that Cdc7 is required for Claspin-Chk1 interaction in human cancer cells by phosphorylating CKBD (Chk1-binding-domain) of Claspin. The residual Chk1 activation in Cdc7-depleted cells is lost upon further depletion of casein kinase1 (CK1g1), previously reported to phosphorylate CKBD. Thus, Cdc7, in conjunction with CK1g1, facilitates the interaction between Claspin and Chk1 through phosphorylating CKBD. We also show that, whereas Cdc7 is predominantly responsible for CKBD phosphorylation in cancer cells, CK1g1plays a major role in non-cancer cells, providing rationale for targeting Cdc7 for cancer cell-specific cell killing.

Data availability

All data generated or analyzed during this study are included in the manuscript and supporting files.Figure 1-source data 1 has been provided for Figure 1AFigure 4 -source data 1-3 have been provided for Figure 4Figure 5-figure supplement 2-source data 1 has been provided for Figure 5-figure supplement 2Figure 5-figure supplement 3-source data 1 has been provided for Figure 5-figure supplement 3BFigure 6-source data 1has been provided for Figure 6BFigure 7-source data 1 has been provided for Figure 7BFigure 7-source data 2 has been provided for Figure 7D

Article and author information

Author details

  1. Chi-Chun Yang

    Department of Genome Medicine, Tokyo Metropolitan Institute of Medical Science, Tokyo, Japan
    Competing interests
    The authors declare that no competing interests exist.
  2. Hiroyuki Kato

    Department of Genome Medicine, Tokyo Metropolitan Institute of Medical Science, Tokyo, Japan
    Competing interests
    The authors declare that no competing interests exist.
  3. Mayumi Shindo

    Protein Analyses Laboratory, Tokyo Metropolitan Institute of Medical Science, Tokyo, Japan
    Competing interests
    The authors declare that no competing interests exist.
  4. Hisao Masai

    Department of Genome Medicine, Tokyo Metropolitan Institute of Medical Science, Tokyo, Japan
    For correspondence
    masai-hs@igakuken.or.jp
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-1268-5302

Funding

Japan Society for the Promotion of Science (23247031)

  • Hisao Masai

Japan Society for the Promotion of Science (26251004)

  • Hisao Masai

Japan Society for the Promotion of Science (24114520)

  • Hisao Masai

Japan Society for the Promotion of Science (25125724)

  • Hisao Masai

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Reviewing Editor

  1. Bruce Stillman, Cold Spring Harbor Laboratory, United States

Publication history

  1. Received: August 2, 2019
  2. Accepted: December 30, 2019
  3. Accepted Manuscript published: December 31, 2019 (version 1)
  4. Version of Record published: February 3, 2020 (version 2)
  5. Version of Record updated: February 4, 2020 (version 3)

Copyright

© 2019, Yang et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

Metrics

  • 1,835
    Page views
  • 289
    Downloads
  • 5
    Citations

Article citation count generated by polling the highest count across the following sources: Crossref, PubMed Central, Scopus.

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Download citations (links to download the citations from this article in formats compatible with various reference manager tools)

Open citations (links to open the citations from this article in various online reference manager services)

Further reading

    1. Chromosomes and Gene Expression
    Alessandro Stirpe et al.
    Research Article

    The SUV39 class of methyltransferase enzymes deposits histone H3 lysine 9 di- and trimethylation (H3K9me2/3), the hallmark of constitutive heterochromatin. How these enzymes are regulated to mark specific genomic regions as heterochromatic is poorly understood. Clr4 is the sole H3K9me2/3 methyltransferase in the fission yeast Schizosaccharomyces pombe, and recent evidence suggests that ubiquitination of lysine 14 on histone H3 (H3K14ub) plays a key role in H3K9 methylation. However, the molecular mechanism of this regulation and its role in heterochromatin formation remain to be determined. Our structure-function approach shows that the H3K14ub substrate binds specifically and tightly to the catalytic domain of Clr4, and thereby stimulates the enzyme by over 250-fold. Mutations that disrupt this mechanism lead to a loss of H3K9me2/3 and abolish heterochromatin silencing similar to clr4 deletion. Comparison with mammalian SET domain proteins suggests that the Clr4 SET domain harbors a conserved sensor for H3K14ub, which mediates licensing of heterochromatin formation.

    1. Chromosomes and Gene Expression
    2. Microbiology and Infectious Disease
    Michele Felletti et al.
    Research Article

    The ability to regulate DNA replication initiation in response to changing nutrient conditions is an important feature of most cell types. In bacteria, DNA replication is triggered by the initiator protein DnaA, which has long been suggested to respond to nutritional changes; nevertheless, the underlying mechanisms remain poorly understood. Here, we report a novel mechanism that adjusts DnaA synthesis in response to nutrient availability in Caulobacter crescentus. By performing a detailed biochemical and genetic analysis of the dnaA mRNA, we identified a sequence downstream of the dnaA start codon that inhibits DnaA translation elongation upon carbon exhaustion. Our data show that the corresponding peptide sequence, but not the mRNA secondary structure or the codon choice, is critical for this response, suggesting that specific amino acids in the growing DnaA nascent chain tune translational efficiency. Our study provides new insights into DnaA regulation and highlights the importance of translation elongation as a regulatory target. We propose that translation regulation by nascent chain sequences, like the one described, might constitute a general strategy for modulating the synthesis rate of specific proteins under changing conditions.