title: Detecting differential allelic expression using high-resolution melting curve analysis: application to the breast cancer susceptibility gene CHEK2.
abstract: BACKGROUND
  The gene CHEK2 encodes a checkpoint kinase playing a key role in the DNA damage pathway. Though CHEK2 has been identified as an intermediate breast cancer susceptibility gene, only a small proportion of high-risk families have been explained by genetic variants located in its coding region. Alteration in gene expression regulation provides a potential mechanism for generating disease susceptibility. The detection of differential allelic expression (DAE) represents a sensitive assay to direct the search for a functional sequence variant within the transcriptional regulatory elements of a candidate gene. We aimed to assess whether CHEK2 was subject to DAE in lymphoblastoid cell lines (LCLs) from high-risk breast cancer patients for whom no mutation in BRCA1 or BRCA2 had been identified.
  
  
  METHODS
  We implemented an assay based on high-resolution melting (HRM) curve analysis and developed an analysis tool for DAE assessment.
  
  
  RESULTS
  We observed allelic expression imbalance in 4 of the 41 LCLs examined. All four were carriers of the truncating mutation 1100delC. We confirmed previous findings that this mutation induces non-sense mediated mRNA decay. In our series, we ruled out the possibility of a functional sequence variant located in the promoter region or in a regulatory element of CHEK2 that would lead to DAE in the transcriptional regulatory milieu of freely proliferating LCLs.
  
  
  CONCLUSIONS
  Our results support that HRM is a sensitive and accurate method for DAE assessment. This approach would be of great interest for high-throughput mutation screening projects aiming to identify genes carrying functional regulatory polymorphisms.
authors.affiliation: Genetic Cancer Susceptibility Group, IARC, 69372 Lyon, France.
authors.forename: Tú
authors.initials: T
authors.lastname: Nguyen-Dumont
authors.forename: Lars P
authors.initials: LP
authors.lastname: Jordheim
authors.forename: Jocelyne
authors.initials: J
authors.lastname: Michelon
authors.forename: Nathalie
authors.initials: N
authors.lastname: Forey
authors.forename: Sandrine
authors.initials: S
authors.lastname: McKay-Chopin
authors.forename: Olga
authors.initials: O
authors.lastname: Sinilnikova
authors.forename: Florence
authors.initials: F
authors.lastname: Le Calvez-Kelm
authors.forename: Melissa C
authors.initials: MC
authors.lastname: Southey
authors.forename: Sean V
authors.initials: SV
authors.lastname: Tavtigian
authors.forename: Fabienne
authors.initials: F
authors.lastname: Lesueur
mesh_terms: D000483:Alleles; D022981:Allelic Imbalance / Q000235:genetics; D001943:Breast Neoplasms / Q000201:enzymology* / Q000235:genetics*; D045744:Cell Line, Tumor; D064447:Checkpoint Kinase 2; D018389:Codon, Nonsense / Q000235:genetics; D004252:DNA Mutational Analysis; D005091:Exons / Q000235:genetics; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D052138:Genes, Neoplasm; D020022:Genetic Predisposition to Disease; D005838:Genotype; D006579:Heterozygote; D006801:Humans; D009691:Nucleic Acid Denaturation / Q000235:genetics*; D020641:Polymorphism, Single Nucleotide / Q000235:genetics; D017346:Protein Serine-Threonine Kinases / Q000235:genetics*; D020871:RNA Stability / Q000235:genetics
chemical_list: D018389:Codon, Nonsense; D064447:Checkpoint Kinase 2; C578634:CHEK2 protein, human; D017346:Protein Serine-Threonine Kinases