Exome sequencing in mostly consanguineous Arab families with neurologic disease provides a high potential molecular diagnosis rate

Abstract

Background: Neurodevelopment is orchestrated by a wide range of genes, and the genetic causes of neurodevelopmental disorders are thus heterogeneous. We applied whole exome sequencing (WES) for molecular diagnosis and in silico analysis to identify novel disease gene candidates in a cohort from Saudi Arabia with primarily Mendelian neurologic diseases.

Methods: We performed WES in 31 mostly consanguineous Arab families and analyzed both single nucleotide and copy number variants (CNVs) from WES data. Interaction/expression network and pathway analyses, as well as paralog studies were utilized to investigate potential pathogenicity and disease association of novel candidate genes. Additional cases for candidate genes were identified through the clinical WES database at Baylor Miraca Genetics Laboratories and GeneMatcher.

Results: We found known pathogenic or novel variants in known disease genes with phenotypic expansion in 6 families, disease-associated CNVs in 2 families, and 12 novel disease gene candidates in 11 families, including KIF5B, GRM7, FOXP4, MLLT1, and KDM2B. Overall, a potential molecular diagnosis was provided by variants in known disease genes in 17 families (54.8 %) and by novel candidate disease genes in an additional 11 families, making the potential molecular diagnostic rate ~90 %.

Conclusions: Molecular diagnostic rate from WES is improved by exome-predicted CNVs. Novel candidate disease gene discovery is facilitated by paralog studies and through the use of informatics tools and available databases to identify additional evidence for pathogenicity.

Trial registration: Not applicable.

Keywords: Copy Number Variants (CNV); Developmental Delay/Intellectual Disability (DD/ID); GRM7; Neurodevelopment; Whole exome sequencing (WES).

Fig. 1

Clinical categories in the cohort and summary of WES findings. a Based on the brain structural findings and accompanying features, this cohort is grouped into nonsyndromic brain malformations (38 %), syndromic brain malformations (23 %), nonsyndromic DD/ID (16 %), and syndromic DD/ID (23 %). b Based on the brain structural defects, families can be further grouped into corpus callosum abnormalities, cortical dysgenesis, microcephaly, hindbrain malformations, and white matter changes; families may be counted in more than one of these groups. c Summary of WES findings (including CNVs): We identified a known variant in a known gene in 2 families (6 %), 10 novel variants in known genes in 9 families presenting reported clinical features (29 %), 7 novel variants in 6 known genes with phenotypic expansion in 6 families (19 %), and 12 novel disease candidates in 11 families (36 %). d The disease genes/candidates identified in this study can be grouped into several biological processes, including mitochondrial enzymes (metabolic or tRNA synthetase), lysosomal enzymes, motor proteins, trafficking proteins, DNA replication, DNA repair, transcription, mRNA splicing, synaptic transmission, signaling pathways, apoptosis, and transmembrane proteins

Fig. 2

A combination of SNV and CNV in RPS6KC1 in BAB6797. a CoNVex reveals a 42 kb heterozygous deletion (chr1:g.213403839_213445978del) in RPS6KC1 in BAB6797 (family 025). There is a SNV (NM_001287219.1:c.2074G > A:p.Gly692Ser) uncovered by this deletion. Gray dotted line: the read depth of other samples in the cohort; red line: the read depth for BAB6797; pink line: the predicted deletion region. b Droplet Digital PCR (ddPCR) detects heterozygous deletion of RPS6KC1 in proband BAB6797 and father BAB6799 but not in mother BAB6798; primer pair targeting RPS6KC1 around chr1:g.213415529[hg19] and three control primer pairs targeting copy-number neutral regions were used to perform ddPCR. Absolute positive droplet concentrations (copies/ul) are plotted from ddPCR results for each primer pair in each sample. Positive droplet concentrations of BAB6797 and BAB6799 for the RPS6KC1 primer pair (around 300 bp) are approximately half of the value in BAB6798 and the results from all control primers (around 600 bp), indicating heterozygous deletion of the RPS6KC1 gene in BAB6797 and BAB6799, but not in BAB6798. Corresponding raw data of ddPCR and primer sequences are shown in Table S5 . Ctrl, control. c This deletion is also confirmed by a customized array. The combination of SNV and CNV in RPS6KC1 segregates in the family. d Breakpoint junction is mapped to chr1:g.213396378-213592823[hg19] by long-range PCR and Sanger sequencing of the junction

Fig. 3

Segregation results and AOH plots of patients with variants in MLLT1, KDM2B, FOXP4, and KIF5B. In each panel, the segregation results are shown to the left and AOH plots on the corresponding chromosomes are shown to the right. a In family 058, the proband (BAB6950) has developmental delay, hypotonia, and infantile spasms with diffuse brain atrophy, delay and hypomyelination, as well as malformation of cortical development. A homozygous variant in MLLT1 (NM_005934.3:c.1418G > A:p.Arg473Gln) is identified and segregates with the phenotypes in the family. b In family 009, the proband (BAB6693) has developmental delay, hypotonia, and infantile spasms. A homozygous variant in KDM2B (NM_032590.4:c.3050G > A:p.Arg1017His) is identified and segregates with the phenotypes in the family. c In family 028, the proband (BAB6800) exhibits developmental delay, laryngeal hypoplasia, and ventricular septal defect (VSD). A homozygous variant in FOXP4 (NM_001012426.1:c.815delT:p.Lys272fs) is identified and segregates in the family. d In family 015, the proband (BAB6712) has developmental delay, seizures, increased reflexes in upper and lower extremities, short stature with mild diffuse brain atrophy, and thinning of the corpus callosum. The younger brother (BAB6713) is mildly affected with speech delay. A homozygous variant in KIF5B (NM_004521.2:c.2252A > G:p.His751Arg) is identified and segregates with the phenotypes in the family, present in a homozygous state in both brothers

Fig. 4

Comparison of the two families carrying GRM7 variants. a Homozygous variant in GRM7 (NM_000844.3:c.461 T > C:p.Ile154Thr) identified in family 014 with two affected children. b Compound heterozygous variants in GRM7 (NM_000844.3:c.1972C > T:p.Arg658Trp and NM_000844.3:c.2024C > A:p.Thr675Lys) segregates in the family with two affected children identified in the BMGL database. c MRI images of BAB6708 and BAB6709. d MRI images of BAB8506 and her brother. e Clinical features comparison among four patients