Positional mapping and candidate gene analysis of the mouse Ccs3 locus that regulates differential susceptibility to carcinogen-induced colorectal cancer

PLoS One. 2013;8(3):e58733. doi: 10.1371/journal.pone.0058733. Epub 2013 Mar 14.

Abstract

The Ccs3 locus on mouse chromosome 3 regulates differential susceptibility of A/J (A, susceptible) and C57BL/6J (B6, resistant) mouse strains to chemically-induced colorectal cancer (CRC). Here, we report the high-resolution positional mapping of the gene underlying the Ccs3 effect. Using phenotype/genotype correlation in a series of 33 AcB/BcA recombinant congenic mouse strains, as well as in groups of backcross populations bearing unique recombinant chromosomes for the interval, and in subcongenic strains, we have delineated the maximum size of the Ccs3 physical interval to a ∼2.15 Mb segment. This interval contains 12 annotated transcripts. Sequencing of positional candidates in A and B6 identified many either low-priority coding changes or non-protein coding variants. We found a unique copy number variant (CNV) in intron 15 of the Nfkb1 gene. The CNV consists of two copies of a 54 bp sequence immediately adjacent to the exon 15 splice site, while only one copy is found in CRC-susceptible A. The Nfkb1 protein (p105/p50) expression is much reduced in A tumors compared to normal A colonic epithelium as analyzed by immunohistochemistry. Studies in primary macrophages from A and B6 mice demonstrate a marked differential activation of the NfκB pathway by lipopolysaccharide (kinetics of stimulation and maximum levels of phosphorylated IκBα), with a more robust activation being associated with resistance to CRC. NfκB has been previously implicated in regulating homeostasis and inflammatory response in the intestinal mucosa. The interval contains another positional candidate Slc39a8 that is differentially expressed in A vs B6 colons, and that has recently been associated in CRC tumor aggressiveness in humans.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Base Sequence
  • Carcinogens / toxicity*
  • Chromosome Mapping*
  • Chromosomes, Mammalian / genetics*
  • Colorectal Neoplasms / chemically induced*
  • Colorectal Neoplasms / genetics*
  • Colorectal Neoplasms / pathology
  • Gene Expression Regulation, Neoplastic / drug effects
  • Gene Expression Regulation, Neoplastic / genetics
  • Genetic Loci / genetics*
  • Genetic Predisposition to Disease / genetics*
  • Humans
  • Hybridization, Genetic
  • Inbreeding
  • Intestinal Mucosa / drug effects
  • Intestinal Mucosa / metabolism
  • Intestinal Mucosa / pathology
  • Mice
  • Molecular Sequence Data
  • NF-kappa B p50 Subunit / metabolism
  • Sequence Analysis, DNA
  • Signal Transduction / drug effects
  • Signal Transduction / genetics
  • Species Specificity

Substances

  • Carcinogens
  • NF-kappa B p50 Subunit

Grants and funding

This work was supported by research grants to PG and NB from the Canadian Cancer Society Research Institute [www.cancer.ca/Research.aspx], the Cancer Research Society Inc. [www.src-crs.ca/en-CA] and the Canderel Initiative Program [www.deficanderel.com/3/donate.htm] of the Goodman Cancer Research Centre [cancercentre.mcgill.ca/research/]. PG is a James McGill Professor of Biochemistry. CM received studentship support and travel awards from the McGill Integrated Cancer Research Training Program (MICRTP) and the Peter Quinlan Foundation [www.mcgill.ca/gcc-research/funding/] through the McGill University Faculty of Medicine. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.