A genome-scale CRISPR-Cas9 screening method for protein stability reveals novel regulators of Cdc25A

Yuanzhong Wu, Liwen Zhou, Xin Wang, Jinping Lu, Ruhua Zhang, Xiaoting Liang, Li Wang, Wuguo Deng, Yi Xin Zeng, Haojie Huang, Tiebang Kang

Research output: Contribution to journalArticle

14 Scopus citations

Abstract

The regulation of stability is particularly crucial for unstable proteins in cells. However, a convenient and unbiased method of identifying regulators of protein stability remains to be developed. Recently, a genome-scale CRISPR-Cas9 library has been established as a genetic tool to mediate loss-of-function screening. Here, we developed a protein stability regulators screening assay (Pro-SRSA) by combining the whole-genome CRISPR-Cas9 library with a dual-fluorescence-based protein stability reporter and high-throughput sequencing to screen for regulators of protein stability. Using Cdc25A as an example, Cul4B-DDB1DCAF8 was identified as a new E3 ligase for Cdc25A. Moreover, the acetylation of Cdc25A at lysine 150, which was acetylated by p300/CBP and deacetylated by HDAC3, prevented the ubiquitin-mediated degradation of Cdc25A by the proteasome. This is the first study to report that acetylation, as a novel posttranslational modification, modulates Cdc25A stability, and we suggest that this unbiased CRISPR-Cas9 screening method at the genome scale may be widely used to globally identify regulators of protein stability.

Original languageEnglish (US)
Article number16014
JournalCell Discovery
Volume2
DOIs
StatePublished - May 24 2016
Externally publishedYes

Keywords

  • acetylation
  • Cdc25A
  • CRISPR-Cas9 screening
  • protein stability
  • ubiquitination

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Genetics
  • Cell Biology

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  • Cite this

    Wu, Y., Zhou, L., Wang, X., Lu, J., Zhang, R., Liang, X., Wang, L., Deng, W., Zeng, Y. X., Huang, H., & Kang, T. (2016). A genome-scale CRISPR-Cas9 screening method for protein stability reveals novel regulators of Cdc25A. Cell Discovery, 2, [16014]. https://doi.org/10.1038/celldisc.2016.14