Chianese Chiara, Gunning Adam C, Giachini Claudia, Daguin Fabrice, Balercia Giancarlo, Ars Elisabet, Lo Giacco Deborah, Ruiz-Castañé Eduard, Forti Gianni, Krausz Csilla
Department of Experimental and Clinical Biomedical Sciences, University of Florence and Centre of Excellence DeNothe, Florence, Italy; Molecular Biology Laboratory, Fundació Puigvert, Universitat Autonoma de Barcelona, Barcelona, Spain.
Department of Experimental and Clinical Biomedical Sciences, University of Florence and Centre of Excellence DeNothe, Florence, Italy.
PLoS One. 2014 Jun 10;9(6):e97746. doi: 10.1371/journal.pone.0097746. eCollection 2014.
Spermatogenesis is a highly complex process involving several thousand genes, only a minority of which have been studied in infertile men. In a previous study, we identified a number of Copy Number Variants (CNVs) by high-resolution array-Comparative Genomic Hybridization (a-CGH) analysis of the X chromosome, including 16 patient-specific X chromosome-linked gains. Of these, five gains (DUP1A, DUP5, DUP20, DUP26 and DUP40) were selected for further analysis to evaluate their clinical significance.
The copy number state of the five selected loci was analyzed by quantitative-PCR on a total of 276 idiopathic infertile patients and 327 controls in a conventional case-control setting (199 subjects belonged to the previous a-CGH study). For one interesting locus (intersecting DUP1A) additional 338 subjects were analyzed.
All gains were confirmed as patient-specific and the difference in duplication load between patients and controls is significant (p = 1.65 × 10(-4)). Two of the CNVs are private variants, whereas 3 are found recurrently in patients and none of the controls. These CNVs include, or are in close proximity to, genes with testis-specific expression. DUP1A, mapping to the PAR1, is found at the highest frequency (1.4%) that was significantly different from controls (0%) (p = 0.047 after Bonferroni correction). Two mechanisms are proposed by which DUP1A may cause spermatogenic failure: i) by affecting the correct regulation of a gene with potential role in spermatogenesis; ii) by disturbing recombination between PAR1 regions during meiosis. This study allowed the identification of novel spermatogenesis candidate genes linked to the 5 CNVs and the discovery of the first recurrent, X-linked gain with potential clinical relevance.
精子发生是一个高度复杂的过程，涉及数千个基因，其中只有少数基因在不育男性中得到研究。在先前的一项研究中，我们通过对X染色体进行高分辨率阵列比较基因组杂交（a-CGH）分析，鉴定出了一些拷贝数变异（CNV），包括16个患者特异性的X染色体连锁增益。其中，选择了五个增益（DUP1A、DUP5、DUP20、DUP26和DUP40）进行进一步分析，以评估它们的临床意义。
在传统的病例对照研究中，通过定量PCR分析了总共276例特发性不育患者和327例对照中五个选定基因座的拷贝数状态（199名受试者属于先前的a-CGH研究）。对于一个有趣的基因座（与DUP1A相交），又分析了另外338名受试者。
所有增益均被确认为患者特异性，患者和对照之间的重复负荷差异具有统计学意义（p = 1.65×10⁻⁴）。其中两个CNV是私人变异，而另外三个在患者中反复出现，在对照中均未发现。这些CNV包括睾丸特异性表达的基因，或与这些基因紧密相邻。定位于PAR1的DUP1A出现频率最高（1.4%），与对照（0%）有显著差异（经Bonferroni校正后p = 0.047）。提出了DUP1A可能导致生精失败的两种机制：i）通过影响在精子发生中可能起作用的基因的正确调控；ii）通过干扰减数分裂期间PAR1区域之间的重组。这项研究使得与5个CNV相关的新型精子发生候选基因得以鉴定，并发现了首个具有潜在临床相关性的反复出现的X连锁增益。
