Towards an understanding of C9orf82 protein/CAAP1 function

PLoS One. 2019 Jan 10;14(1):e0210526. doi: 10.1371/journal.pone.0210526. eCollection 2019.

Abstract

C9orf82 protein, or conserved anti-apoptotic protein 1 or caspase activity and apoptosis inhibitor 1 (CAAP1) has been implicated as a negative regulator of the intrinsic apoptosis pathway by modulating caspase expression and activity. In contrast, an independent genome wide screen for factors capable of driving drug resistance to the topoisomerase II (Topo II) poisons doxorubicin and etoposide, implicated a role for the nuclear protein C9orf82 in delaying DSBs repair downstream of Topo II, hereby sensitizing cells to DSB induced apoptosis. To determine its function in a genetically defined setting in vivo and ex vivo, we here employed CRISPR/Cas9 technology in zygotes to generate a C9orf82 knockout mouse model. C9orf82ko/ko mice were born at a Mendelian ratio and did not display any overt macroscopic or histological abnormalities. DSBs repair dependent processes like lymphocyte development and class switch recombination (CSR) appeared normal, arguing against a link between the C9orf82 encoded protein and V(D)J recombination or CSR. Most relevant, primary pre-B cell cultures and Tp53 transformed mouse embryo fibroblasts (MEFs) derived from C9orf82ko/ko E14.5 and wild type embryos displayed comparable sensitivity to a number of DNA lesions, including DSBs breaks induced by the topoisomerase II inhibitors, etoposide and doxorubicin. Likewise, the kinetics of γH2AX formation and resolution in response to etoposide of C9orf82 protein proficient, deficient and overexpressing MEFs were indistinguishable. These data argue against a direct role of C9orf82 protein in delaying repair of Topo II generated DSBs and regulating apoptosis. The genetically defined systems generated in this study will be of value to determine the actual function of C9orf82 protein.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Apoptosis
  • Apoptosis Regulatory Proteins / genetics
  • Apoptosis Regulatory Proteins / physiology*
  • B-Lymphocytes / cytology
  • B-Lymphocytes / metabolism
  • CRISPR-Cas Systems
  • Caspase 3 / metabolism
  • Cell Proliferation
  • Cells, Cultured
  • DNA Breaks, Double-Stranded*
  • DNA Damage
  • DNA Repair
  • DNA Topoisomerases, Type II / metabolism*
  • Immunoglobulin Class Switching
  • Mice
  • Mice, Inbred C57BL
  • Mice, Knockout
  • T-Lymphocytes / cytology
  • T-Lymphocytes / metabolism

Substances

  • Apoptosis Regulatory Proteins
  • Caap1 protein, mouse
  • Caspase 3
  • DNA Topoisomerases, Type II

Grant support

This research was supported by a Top Grant from the Netherlands Organisation for Health Research and Development (ZonMw, project 91213018) to HJ. The funder had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.