A novel assay to screen siRNA libraries identifies protein kinases required for chromosome transmission

Genome Res. 2019 Oct;29(10):1719-1732. doi: 10.1101/gr.254276.119. Epub 2019 Sep 12.

Abstract

One of the hallmarks of cancer is chromosome instability (CIN), which leads to aneuploidy, translocations, and other chromosome aberrations. However, in the vast majority of human tumors the molecular basis of CIN remains unknown, partly because not all genes controlling chromosome transmission have yet been identified. To address this question, we developed an experimental high-throughput imaging (HTI) siRNA assay that allows the identification of novel CIN genes. Our method uses a human artificial chromosome (HAC) expressing the GFP transgene. When this assay was applied to screen an siRNA library of protein kinases, we identified PINK1, TRIO, IRAK1, PNCK, and TAOK1 as potential novel genes whose knockdown induces various mitotic abnormalities and results in chromosome loss. The HAC-based assay can be applied for screening different siRNA libraries (cell cycle regulation, DNA damage response, epigenetics, and transcription factors) to identify additional genes involved in CIN. Identification of the complete spectrum of CIN genes will reveal new insights into mechanisms of chromosome segregation and may expedite the development of novel therapeutic strategies to target the CIN phenotype in cancer cells.

Publication types

  • Research Support, N.I.H., Intramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Aneuploidy
  • Calcium-Calmodulin-Dependent Protein Kinase Type 1 / genetics
  • Cell Line, Tumor
  • Chromosomal Instability / genetics*
  • Chromosomes, Artificial, Human / genetics
  • Chromosomes, Human / genetics*
  • Guanine Nucleotide Exchange Factors / genetics
  • Humans
  • Interleukin-1 Receptor-Associated Kinases / genetics
  • Mitosis / genetics
  • Protein Kinases / genetics*
  • Protein Kinases / isolation & purification
  • Protein-Serine-Threonine Kinases / genetics
  • RNA, Double-Stranded / genetics
  • RNA, Small Interfering / genetics*
  • Transgenes
  • Translocation, Genetic / genetics

Substances

  • Guanine Nucleotide Exchange Factors
  • RNA, Double-Stranded
  • RNA, Small Interfering
  • Protein Kinases
  • IRAK1 protein, human
  • Interleukin-1 Receptor-Associated Kinases
  • PTEN-induced putative kinase
  • Protein-Serine-Threonine Kinases
  • TRIO protein, human
  • Calcium-Calmodulin-Dependent Protein Kinase Type 1
  • PNCK protein, human