Durocher Francine, Labrie Yvan, Soucy Penny, Sinilnikova Olga, Labuda Damian, Bessette Paul, Chiquette Jocelyne, Laframboise Rachel, Lépine Jean, Lespérance Bernard, Ouellette Geneviève, Pichette Roxane, Plante Marie, Tavtigian Sean V, Simard Jacques
Cancer Genomics Laboratory, Oncology and Molecular Endocrinology Research Centre, Centre Hospitalier Universitaire de Québec and Laval University, Québec, G1V 4G2, Canada.
BMC Cancer. 2006 Sep 29;6:230. doi: 10.1186/1471-2407-6-230.
Ataxia telangiectasia-mutated and Rad3-related (ATR) is a member of the PIK-related family which plays, along with ATM, a central role in cell-cycle regulation. ATR has been shown to phosphorylate several tumor suppressors like BRCA1, CHEK1 and TP53. ATR appears as a good candidate breast cancer susceptibility gene and the current study was designed to screen for ATR germline mutations potentially involved in breast cancer predisposition.
ATR direct sequencing was performed using a fluorescent method while widely available programs were used for linkage disequilibrium (LD), haplotype analyses, and tagging SNP (tSNP) identification. Expression analyses were carried out using real-time PCR.
The complete sequence of all exons and flanking intronic sequences were analyzed in DNA samples from 54 individuals affected with breast cancer from non-BRCA1/2 high-risk French Canadian breast/ovarian families. Although no germline mutation has been identified in the coding region, we identified 41 sequence variants, including 16 coding variants, 3 of which are not reported in public databases. SNP haplotypes were established and tSNPs were identified in 73 healthy unrelated French Canadians, providing a valuable tool for further association studies involving the ATR gene, using large cohorts. Our analyses led to the identification of two novel alternative splice transcripts. In contrast to the transcript generated by an alternative splicing site in the intron 41, the one resulting from a deletion of 121 nucleotides in exon 33 is widely expressed, at significant but relatively low levels, in both normal and tumoral cells including normal breast and ovarian tissue.
Although no deleterious mutations were identified in the ATR gene, the current study provides an haplotype analysis of the ATR gene polymorphisms, which allowed the identification of a set of SNPs that could be used as tSNPs for large-scale association studies. In addition, our study led to the characterization of a novel Delta33 splice form, which could generate a putative truncated protein lacking several functional domains. Additional studies in large cohorts and other populations will be needed to further evaluate if common and/or rare ATR sequence variants can be associated with a modest or intermediate breast cancer risk.
共济失调毛细血管扩张症突变和Rad3相关蛋白（ATR）是PIK相关家族的成员，它与ATM一起在细胞周期调控中发挥核心作用。ATR已被证明可磷酸化多种肿瘤抑制因子，如BRCA1、CHEK1和TP53。ATR似乎是一个很好的乳腺癌易感基因候选者，当前研究旨在筛查可能与乳腺癌易感性相关的ATR种系突变。
采用荧光法进行ATR直接测序，同时使用广泛可用的程序进行连锁不平衡（LD）、单倍型分析和标签单核苷酸多态性（tSNP）鉴定。使用实时PCR进行表达分析。
对来自非BRCA1/2高风险法裔加拿大乳腺癌/卵巢癌家族的54例乳腺癌患者的DNA样本进行了所有外显子及其侧翼内含子序列的完整测序。虽然在编码区未发现种系突变，但我们鉴定出41个序列变异，包括16个编码变异，其中3个未在公共数据库中报道。在73名健康的非亲属法裔加拿大人中建立了SNP单倍型并鉴定出tSNP，为使用大型队列进行涉及ATR基因的进一步关联研究提供了有价值的工具。我们的分析导致鉴定出两种新的可变剪接转录本。与由内含子41中的可变剪接位点产生的转录本不同，由外显子33中121个核苷酸缺失产生的转录本在包括正常乳腺和卵巢组织在内的正常和肿瘤细胞中均广泛表达，表达水平显著但相对较低。
虽然在ATR基因中未鉴定出有害突变，但当前研究提供了ATR基因多态性的单倍型分析，从而鉴定出一组可作为tSNP用于大规模关联研究的SNP。此外，我们的研究导致鉴定出一种新的Delta33剪接形式，它可能产生一种缺少几个功能域的推定截短蛋白。需要在大型队列和其他人群中进行进一步研究，以进一步评估常见和/或罕见的ATR序列变异是否与中度或中度乳腺癌风险相关。
