Solomon Obiri-Yeboah

Association Studies and Direct DNA Sequencing Implicate Genetic Susceptibility Loci in the Etiology of Nonsyndromic Orofacial Clefts in Sub-Saharan African Populations

Orofacial clefts (OFCs) are congenital dysmorphologies of the human face and oral cavity, with a global incidence of 1 per 700 live births. These anomalies exhibit a multifactorial pattern of inheritance, with genetic and environmental factors both playing crucial roles. Many loci have been implicated in the etiology of nonsyndromic cleft lip with or without cleft palate (NSCL/P) in populations of Asian and European ancestries, through genome-wide association studies and candidate gene studies. However, few populations of African descent have been studied to date. Here, the authors show evidence of an association of some loci with NSCL/P and nonsyndromic cleft palate only (NSCPO) in cohorts from Africa (Ghana, Ethiopia, and Nigeria). The authors genotyped 48 single-nucleotide polymorphisms that were selected from previous genome-wide association studies and candidate gene studies. These markers were successfully genotyped on 701 NSCL/P and 163 NSCPO cases, 1,070 unaffected relatives, and 1,078 unrelated controls. The authors also directly sequenced 7 genes in 184 nonsyndromic OFC (NSOFC) cases and 96 controls from Ghana. Population-specific associations were observed in the case-control analyses of the subpopulations, with West African subpopulations (Ghana and Nigeria) showing a similar pattern of associations. In meta-analyses of the case-control cohort, PAX7 (rs742071, P = 5.10 × 10−3), 8q24 (rs987525, P = 1.22 × 10−3), and VAX1 (rs7078160, P = 0.04) were nominally associated with NSCL/P, and MSX1 (rs115200552, P = 0.01), TULP4 (rs651333, P = 0.04), CRISPLD2 (rs4783099, P = 0.02), and NOG1 (rs17760296, P = 0.04) were nominally associated with NSCPO. Moreover, 7 loci exhibited evidence of threshold overtransmission in NSOFC cases through the transmission disequilibrium test and through analyses of the family-based association for disease traits. Through DNA sequencing, the authors also identified 2 novel, rare, potentially pathogenic variants (p.Asn323Asp and p.Lys426IlefsTer6) in ARHGAP29. In conclusion, the authors have shown evidence for the association of many loci with NSCL/P and NSCPO. To the best of this knowledge, this study is the first to demonstrate any of these association signals in any African population.

Loss-of-Function GRHL3 Variants Detected in African Patients with Isolated Cleft Palate

In contrast to the progress that has been made toward understanding the genetic etiology of cleft lip with or without cleft palate, relatively little is known about the genetic etiology for cleft palate only (CPO). A common coding variant of grainyhead like transcription factor 3 (GRHL3) was recently shown to be associated with risk for CPO in Europeans. Mutations in this gene were also reported in families with Van der Woude syndrome. To identify rare mutations in GRHL3 that might explain the missing heritability for CPO, we sequenced GRHL3 in cases of CPO from Africa. We recruited participants from Ghana, Ethiopia, and Nigeria. This cohort included case-parent trios, cases and other family members, as well as controls. We sequenced exons of this gene in DNA from a total of 134 nonsyndromic cases. When possible, we sequenced them in parents to identify de novo mutations. Five novel mutations were identified: 2 missense (c.497C>A; p.Pro166His and c.1229A>G; p.Asp410Gly), 1 splice site (c.1282A>C p.Ser428Arg), 1 frameshift (c.470delC; p.Gly158Alafster55), and 1 nonsense (c.1677C>A; p.Tyr559Ter). These mutations were absent from 270 sequenced controls and from all public exome and whole genome databases, including the 1000 Genomes database (which includes data from Africa). However, 4 of the 5 mutations were present in unaffected mothers, indicating that their penetrance is incomplete. Interestingly, 1 mutation damaged a predicted sumoylation site, and another disrupted a predicted CK1 phosphorylation site. Overexpression assays in zebrafish and reporter assays in vitro indicated that 4 variants were functionally null or hypomorphic, while 1 was dominant negative. This study provides evidence that, as in Caucasian populations, mutations in GRHL3 contribute to the risk of nonsyndromic CPO in the African population.

Novel GREM1 Variations in Sub-Saharan African Patients With Cleft Lip and/or Cleft Palate.

Objective: Cleft lip and/or cleft palate (CL/P) are congenital anomalies of the face and have multifactorial etiology, with both environmental and genetic risk factors playing crucial roles. Though at least 40 loci have attained genomewide significant association with nonsyndromic CL/P, these loci largely reside in noncoding regions of the human genome, and subsequent resequencing studies of neighboring candidate genes have revealed only a limited number of etiologic coding variants. The present study was conducted to identify etiologic coding variants in GREM1, a locus that has been shown to be largely associated with cleft of both lip and soft palate. Patients and Method: We resequenced DNA from 397 sub-Saharan Africans with CL/P and 192 controls using Sanger sequencing. Following analyses of the sequence data, we observed 2 novel coding variants in GREM1. These variants were not found in the 192 African controls and have never been previously reported in any public genetic variant database that includes more than 5000 combined African and African American controls or from the CL/P literature. Results: The novel variants include p.Pro164Ser in an individual with soft palate cleft only and p.Gly61Asp in an individual with bilateral cleft lip and palate. The proband with the p.Gly61Asp GREM1 variant is a van der Woude (VWS) case who also has an etiologic variant in IRF6 gene. Conclusion: Our study demonstrated that there is low number of etiologic coding variants in GREM1, confirming earlier suggestions that variants in regulatory elements may largely account for the association between this locus and CL/P.

Identification of paternal uniparental disomy on chromosome 22 and a de novo deletion on chromosome 18 in individuals with orofacial clefts

Orofacial clefts are the most common malformations of the head and neck region. Genetic and environmental factors have been implicated in the etiology of these traits.

The prevalence, penetrance, and expressivity of etiologicIRF6variants in orofacial clefts patients from sub-Saharan Africa

Orofacial clefts are congenital malformations of the orofacial region, with a global incidence of one per 700 live births. Interferon Regulatory Factor 6 (IRF6) (OMIM:607199) gene has been associated with the etiology of both syndromic and nonsyndromic orofacial clefts. The aim of this study was to show evidence of potentially pathogenic variants in IRF6 in orofacial clefts cohorts from Africa.