Identification and Classification of Novel Genetic Variants: En Route to the Diagnosis of Primary Ciliary Dyskinesia.

Nina Stevanovic,Anita Skakic,Aleksandar Sovtic,Marina Andjelkovic,Maja Stojiljkovic,Sonja Pavlovic,Predrag Minic

doi:10.3390/ijms22168821

Abstract

Primary ciliary dyskinesia (PCD) is a disease caused by impaired function of motile cilia. PCD mainly affects the lungs and reproductive organs. Inheritance is autosomal recessive and X-linked. PCD patients have diverse clinical manifestations, thus making the establishment of proper diagnosis challenging. The utility of next-generation sequencing (NGS) technology for diagnostic purposes allows for better understanding of the PCD genetic background. However, identification of specific disease-causing variants is difficult. The main aim of this study was to create a unique guideline that will enable the standardization of the assessment of novel genetic variants within PCD-associated genes. The designed pipeline consists of three main steps: (1) sequencing, detection, and identification of genes/variants; (2) classification of variants according to their effect; and (3) variant characterization using in silico structural and functional analysis. The pipeline was validated through the analysis of the variants detected in a well-known PCD disease-causing gene (DNAI1) and the novel candidate gene (SPAG16). The application of this pipeline resulted in identification of potential disease-causing variants, as well as validation of the variants pathogenicity, through their analysis on transcriptional, translational, and posttranslational levels. The application of this pipeline leads to the confirmation of PCD diagnosis and enables a shift from candidate to PCD disease-causing gene.

Highlights

Primary ciliary dyskinesia (PCD(OMIM #244400)) is a rare genetic disorder most often inherited in an autosomal recessive and X-linked manner; recent studies have shown that autosomal dominant variants in the FOXJ1 gene cause PCD [1]
Classify, and characterize novel genetic variants and novel candidate genes using in silico and functional analysis as tools, we designed the guideline in a form of a pipeline (Figure 1)
We report a very comprehensive genetic pipeline designed for identification, classification, and structural and functional analysis of novel genetic variants and candidate genes

Summary

Introduction

Primary ciliary dyskinesia (PCD(OMIM #244400)) is a rare genetic disorder most often inherited in an autosomal recessive and X-linked manner; recent studies have shown that autosomal dominant variants in the FOXJ1 gene cause PCD [1]. The main features of PCD are structural defects of motile cilia, leading to abnormal ciliary motility, ciliary immotility, or absence of cilia [2]. Because motile cilia are present in humans in the respiratory tract, middle ear, paranasal sinuses, female reproductive tract, ependyma of the brain, and on the embryonic node, this disorder predominantly affects the respiratory system, reproductive organs, and visceral organ laterality (about 50% of the PCD patients have situs inversus) [3]. Manifestations in other systems include subfertility in both males, due to immotile spermatozoa [8], and females, due to altered motility of Fallopian tube cilia [9]. The estimated prevalence is between 1:4000 and 1:40,000, with the true prevalence of probably about 1:10,000 [5,10] and higher rates in certain ethnic groups due to consanguinity [11,12]

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