[Exploring parent-of-origin effects for non-syndromic cleft lip with or without cleft palate on PTCH1, PTCH2, SHH, SMO genes in Chinese case-parent trios]

Beijing Da Xue Xue Bao Yi Xue Ban. 2020 Oct 18;52(5):809-814. doi: 10.19723/j.issn.1671-167X.2020.05.003.
[Article in Chinese]


Objective: Non-syndromic cleft lip with or without cleft palate (NSCL/P) is a common birth defect, affecting 1.4 per 1 000 live births, and multiple genetic and environmental risk factors influencing its risk. All the known genetic risk factors accounted for a small proportion of the heritability. Several authors have suggested parent-of-origin effects (PoO) may play an important role in the etiology of this complex and heterogeneous malformation. To clarify the genetic association between PTCH1, PTCH2, SHH and SMO in hedgehog (HH) pathway and NSCL/P, as well as testing for potential PoO effects in Chinese case-parent trios.

Methods: We tested for transmission disequilibrium tests (TDT) and PoO effects using 83 common single nucleotide polymorphic (SNP) markers of HH pathway genes from 806 NSCL/P case-parent trios. These trios were drawn from an international consortium established for a genome-wide association studies (GWAS) of non-syndromic oral clefts of multiple ethnicities. DNA samples were collected from each trio. Single marker and haplotype based analysis were performed both in TDT tests and PoO effects. SNPs were excluded if they (ⅰ) had a call rate of &#60; 95%, (ⅱ) had a minor allele frequency (MAF) of &#60; 0.05, (ⅲ) had Mendelian errors over all trios of >5%, (ⅳ) had a genotype distribution in the parents that deviated from the Hardy-Weinberg equilibrium (HWE) (<i>P</i> &#60; 0.000 1). The process was done using Plink (version 1.07, <a href="http://pngu.mgh.harvard.edu/~purcell/plink/data.shtml" target="_blank">http://pngu.mgh.harvard.edu/~purcell/plink/data.shtml</a>). TDT test was performed in Plink v1.07. A log-linear model was used to explore PoO effects using Haplin v6.2.1 as implemented in R package v3.4.2. Significance level was assessed using the Bonferroni correction.

Results: A total of 18 SNPs were dropped due to low MAF, thus leaving 65 SNPs available for the analysis. Thus the Bonferroni threshold was 7.7×10-4 (0.05/65). Nominal significant association with NSCL/P was found at a SNP (rs4448343 in PTCH1, P=0.023) and six haplotypes (rs10512249-rs4448343, rs1461208-rs7786445, rs10512249-rs4448343, rs16909865-rs10512249-rs4448343, rs1461208-rs7786445-rs12698335, and rs288756-rs288758-rs1151790, P < 0.05). A total of six haplotypes (rs288765-rs1233563, rs12537550-rs11765352, rs872723-rs288765-rs1233563, rs288765-rs1233563-rs288756, rs6459952-rs12537550-rs11765352, and rs12537550-rs11765352-rs6971211) showed PoO effect (P < 0.05). None of the results remained significant after the Bonferroni correction (P>7.7×10-4).

Conclusion: Neither significant association between SNPs within HH pathway and the risk of NSCL/P nor PoO effects was seen in this study.

目的: 探索非综合征型唇腭裂(non-syndromic cleft lip with or without cleft palate,NSCL/P)这一类常见出生缺陷的可能致病机制,在Hedgehog(HH)通路基因中(PTCH1PTCH2SHHSMO)探索基因多态性对NSCL/P的关联关系以及亲源效应(parent-of-origin effects,PoO)对NSCL/P发病风险的影响。

方法: 纳入806个中国非综合征型唇腭裂核心家系,对HH通路基因(PTCH1PTCH2SHHSMO)的83个单核苷酸多态性位点(single nucleotide polymorphisms,SNPs)进行传递不平衡检验(transmission disequilibrium test,TDT),并采用对数线性模型进行亲源效应分析。家系样本来自“唇腭裂基因和交互作用的国际合作研究”项目。采用Plink进行TDT检验;通过R软件中的Haplin v6.2.1软件包开展亲源效应分析。采用Bonferroni法进行多重检验校正。

结果: 经过质量控制,共纳入65个SNPs进行分析,Bonferroni显著性水平为7.7×10-4(0.05/65)。未校正P值前,关联分析发现rs4448343与NSCL/P存在关联(P=0.023),6个单体型(rs10512249-rs4448343、rs1461208-rs7786445、rs10512249-rs4448343、rs16909865-rs10512249-rs4448343、rs1461208-rs7786445-rs12698335、rs288756-rs288758-rs1151790)与NSCL/P存在关联(P < 0.05);6个单体型(rs288765-rs1233563、rs12537550-rs11765352、rs872723-rs288765-rs1233563、rs288765-rs1233563-rs288756、rs6459952-rs12537550-rs11765352、rs12537550-rs11765352-rs6971211)具有潜在的PoO效应(P < 0.05)。以上结果经过多重检验校正,均无统计学意义(P>7.7×10-4)。

结论: 未发现HH通路基因多态性与NSCL/P的关联,未发现HH通路基因通过PoO效应影响NSCL/P发病风险。

Keywords: Hedgehog pathway; Non-syndromic cleft lip with or without cleft palate; Parent-of-origin effects.

MeSH terms

  • Asian Continental Ancestry Group
  • Cleft Lip* / genetics
  • Cleft Palate* / genetics
  • Genome-Wide Association Study
  • Hedgehog Proteins / genetics
  • Humans
  • Patched-2 Receptor
  • Smoothened Receptor


  • Hedgehog Proteins
  • PTCH2 protein, human
  • Patched-2 Receptor
  • SHH protein, human
  • SMO protein, human
  • Smoothened Receptor

Grant support

国家自然科学基金(81102178, 81573225)、北京市自然科学基金(7172115)和北京大学医学交叉研究基金(BMU2017MX018)