Abstract
In silico predictive software allows assessing the effect of amino acid substitutions on the structure or function of a protein without conducting functional studies. The accuracy of in silico pathogenicity prediction tools has not been previously assessed for variants associated with autosomal recessive deafness 1A (DFNB1A). Here, we identify in silico tools with the most accurate clinical significance predictions for missense variants of the GJB2 (Cx26), GJB6 (Cx30), and GJB3 (Cx31) connexin genes associated with DFNB1A. To evaluate accuracy of selected in silico tools (SIFT, FATHMM, MutationAssessor, PolyPhen-2, CONDEL, MutationTaster, MutPred, Align GVGD, and PROVEAN), we tested nine missense variants with previously confirmed clinical significance in a large cohort of deaf patients and control groups from the Sakha Republic (Eastern Siberia, Russia): Сх26: p.Val27Ile, p.Met34Thr, p.Val37Ile, p.Leu90Pro, p.Glu114Gly, p.Thr123Asn, and p.Val153Ile; Cx30: p.Glu101Lys; Cx31: p.Ala194Thr. We compared the performance of the in silico tools (accuracy, sensitivity, and specificity) by using the missense variants in GJB2 (Cx26), GJB6 (Cx30), and GJB3 (Cx31) genes associated with DFNB1A. The correlation coefficient (r) and coefficient of the area under the Receiver Operating Characteristic (ROC) curve as alternative quality indicators of the tested programs were used. The resulting ROC curves demonstrated that the largest coefficient of the area under the curve was provided by three programs: SIFT (AUC = 0.833, p = 0.046), PROVEAN (AUC = 0.833, p = 0.046), and MutationAssessor (AUC = 0.833, p = 0.002). The most accurate predictions were given by two tested programs: SIFT and PROVEAN (Ac = 89%, Se = 67%, Sp = 100%, r = 0.75, AUC = 0.833). The results of this study may be applicable for analysis of novel missense variants of the GJB2 (Cx26), GJB6 (Cx30), and GJB3 (Cx31) connexin genes.
Highlights
The most common form of hereditary nonsyndromic hearing loss is autosomal recessive deafness 1A (DFNB1A, MIM#220290) caused by pathogenic variants in the GJB2, GJB6, and GJB3 genes encoding connexin 26 (Cx26), connexin (Cx30), and connexin (Cx31) proteins, respectively
High rates of predictability of positive and negative results were provided by the SIFT and PROVEAN programs (PPV = 100% and Negative predictive values (NPV) = 86% for both programs) while the FATHMM and Align GVGD programs were the most inaccurate, which resulted in a decrease in almost all of the analyzed parameters
The resulting Receiver Operating Characteristic (ROC) curves demonstrated that the largest coefficient of the area under the curve was shown by three programs: SIFT (AUC = 0.833, p = 0.046, 95% CI: 0.45-0.98), PROVEAN (AUC = 0.833, p = 0.046, 95% CI: 0.45-0.98), and MutationAssessor (AUC = 0.833, p = 0.002, 95% CI: 0.450.98)
Summary
The most common form of hereditary nonsyndromic hearing loss is autosomal recessive deafness 1A (DFNB1A, MIM#220290) caused by pathogenic variants in the GJB2, GJB6, and GJB3 genes encoding connexin 26 (Cx26), connexin (Cx30), and connexin (Cx31) proteins, respectively. The estimated prevalence of DFNB1A among general human population is 14:100 000, and the main cause of DFNB1A is biallelic recessive pathogenic variants in the GJB2 gene (MIM#121011) (http://www.ncbi.nlm.nih.gov/ books/NBK1272/, 2018). About 400 different pathogenic variations of GJB2 sequence (more than 70% are missense or nonsense amino acid substitutions) are presented in the Human Gene Mutation Database (HGMD, http://www.hgmd.cf.ac.uk/ac/ all.php), and this list is regularly updated by novel yet unclassified variants. The majority of nonsense variants are pathogenic since they lead to a premature termination of translation and protein synthesis, while missense variants depending on their location in the amino acid sequence can be neutral, damaging, or partially damaging to the structure and function of protein. The impact of a missense substitution depends on criteria such as the evolutionary conservatism of an amino acid/nucleotide, location, and context within the protein sequence and the biochemical consequence of the amino acid substitution [4]
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