Missense Variants Within GJB2 Gene Locus and the Risk of Hearing Defects in Nonsyndromic Cleft Lip and Palate

Abstract

Aim: The aim of the study was to investigate the role of variants in GJB2 gene in the etiology of hearing defects in nonsyndromic cleft lip/palate.

Method: Saliva samples were obtained from cases (subjects with orofacial clefts) and control (subjects without orofacial clefts) who consented to the study. Deoxyribonucleic acid (DNA) was extracted using standardized protocol at Butali Lab (Iowa, IA). Primers for the coding region of GJB2 was designed using Primer 3 (http://bioinfo.ut.ee/primer3-0.4.0/) and optimized in the Butali lab using a gradient polymerase chain reaction to determine the annealing temperature for each primer set (forward and reverse). We measured the DNA concentration using Qubit and XY genotyping done for quality control. A concentration of 5 ng/μL of DNA was used for Sanger sequencing.

Results: A total of 150 subjects were sequenced (66 cases; 84 controls). Mutations in GJB2 gene were detected in 2 individuals with cleft palate. We found p.Arg165Trp variant in 1 case and p.Leu81Val variant in the second case. Although p.Arg165Trp was predicted to be either benign or tolerated by SIFT/POLYPHEN, the single nucleotide change from C>T, that is, CGG>TGG leads to a premature stop codon preventing the protein formation. The p.Leu81Val variant was predicted to be probably damaging/ deleterious.

Conclusions: The present study implicates variants in the GJB2 gene in the etiology of hearing defects in nonsyndromic cleft lip and palate in the Nigerian population. Screening for variations in GJB2 gene is important for genetic counseling especially in high-risk families.