Clonogenic Assays to Detect Cell Fate in Mitotic Catastrophe

Methods Mol Biol. 2021:2267:227-239. doi: 10.1007/978-1-0716-1217-0_16.

Abstract

Mitotic catastrophe (MC) is a cell death modality induced by DNA damage that involves the activation of cell cycle checkpoints such as the "DNA structure checkpoint" and "spindle assembly checkpoint" (SAC) leading to aberrant mitosis. Depending on the signal, MC can drive the cell to death or to senescence. The suppression of MC favors aneuploidy. Several cancer therapies, included microtubular poisons and radiations, trigger MC. The clonogenic assay has been used to study the capacity of single cells to proliferate and to generate macroscopic colonies and to evaluate the efficacy of anticancer drugs. Nevertheless, this method cannot analyze MC events. Here, we report an improved technique based on the use of human colon cancer HCT116 stable expressing histone H2B-GFP and DsRed-centrin proteins, allowing to determine the capacity of cells to proliferate, and to determine changes in the nucleus and centrosomes.

Keywords: Centrin; Clonogenic assay; Histone; Mitotic catastrophe.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Antimitotic Agents / toxicity
  • Antineoplastic Agents / toxicity
  • Calcium-Binding Proteins / genetics
  • Calcium-Binding Proteins / metabolism
  • Cell Death*
  • Cell Proliferation*
  • Chromosomal Proteins, Non-Histone / genetics
  • Chromosomal Proteins, Non-Histone / metabolism
  • Genes, Reporter
  • Green Fluorescent Proteins / genetics
  • Green Fluorescent Proteins / metabolism
  • HCT116 Cells
  • Histones / genetics
  • Histones / metabolism
  • Humans
  • Mitosis*
  • Tumor Stem Cell Assay / methods*

Substances

  • Antimitotic Agents
  • Antineoplastic Agents
  • Calcium-Binding Proteins
  • Chromosomal Proteins, Non-Histone
  • Histones
  • caltractin
  • Green Fluorescent Proteins