Data_Sheet_5_Identification of Novel Variants in Cleft Palate-Associated Genes in Brazilian Patients With Non-syndromic Cleft Palate Only.docx

The identification of genetic risk factors for non-syndromic oral clefts is of great importance for better understanding the biological processes related to this heterogeneous and complex group of diseases. Herein we applied whole-exome sequencing to identify potential variants related to non-syndromic cleft palate only (NSCPO) in the multiethnic Brazilian population. Thirty NSCPO samples and 30 sex- and genetic ancestry-matched healthy controls were pooled (3 pools with 10 samples for each group) and subjected to whole-exome sequencing. After filtering, the functional affects, individually and through interactions, of the selected variants and genes were assessed by bioinformatic analyses. As a group, 399 variants in 216 genes related to palatogenesis/cleft palate, corresponding to 6.43%, were exclusively identified in the NSCPO pools. Among those genes are 99 associated with syndromes displaying cleft palate in their clinical spectrum and 92 previously related to cleft lip palate. The most significantly biological processes and pathways overrepresented in the NSCPO-identified genes were associated with the folic acid metabolism, highlighting the interaction between LDL receptor-related protein 6 (LRP6) and 5-methyltetrahydrofolate-homocysteine methyltransferase (MTR) that interconnect two large networks. This study yields novel data on characterization of specific variants and complex processes and pathways related to NSCPO, including many variants in genes of the folate/homocysteine pathway, and confirms that variants in genes related to syndromic cleft palate and cleft lip-palate may cause NSCPO.

鉴定非综合征性口腔裂的遗传风险因子，对于深入理解这类异质性复杂疾病相关的生物学过程具有重要意义。本研究针对多族裔巴西人群，采用全外显子组测序（whole-exome sequencing）技术筛选与非综合征性单纯腭裂（non-syndromic cleft palate only, NSCPO）相关的潜在变异。本研究收集30例非综合征性单纯腭裂患者样本与30例性别、遗传祖先匹配的健康对照样本，将每组样本分为3个混合池（每个混合池含10例样本）后进行全外显子组测序。过滤完成后，通过生物信息学分析评估筛选出的变异与基因的单独功能效应及互作功能效应。整体而言，仅在非综合征性单纯腭裂混合池中检出的、与腭发育/腭裂相关的216个基因中共包含399个变异，占总变异的6.43%。其中99个基因对应临床表型包含腭裂的综合征，另有92个基因此前已被报道与唇腭裂相关。在非综合征性单纯腭裂患者样本中检出的基因里，富集度最高的生物学过程与通路均与叶酸代谢相关，其中尤为值得关注的是低密度脂蛋白受体相关蛋白6（LDL receptor-related protein 6, LRP6）与5-甲基四氢叶酸-同型半胱氨酸甲基转移酶（5-methyltetrahydrofolate-homocysteine methyltransferase, MTR）之间的互作，该互作连接了两个大型基因调控网络。本研究为非综合征性单纯腭裂相关的特异性变异、复杂生物学过程及通路的特征解析提供了全新数据，其中包含叶酸/同型半胱氨酸通路中多个基因的变异，并证实与综合征性腭裂及唇腭裂相关的基因变异可能导致非综合征性单纯腭裂的发生。