Genetic Analysis in Fetal Skeletal Dysplasias by Trio Whole-Exome Sequencing

Abstract

Skeletal dysplasias (SDs) comprise a series of severe congenital disorders that have strong clinical heterogeneity and usually attribute to diverse genetic variations. The pathogenesis of more than half of SDs remains unclear. Additionally, the clinical manifestations of fetal SDs are ambiguous, which poses a big challenge for accurate diagnosis. In this study, eight unrelated families with fetal SD were recruited and subjected to sequential tests including chromosomal karyotyping, chromosomal microarray analysis (CMA), and trio whole-exome sequencing (WES). Sanger sequencing and quantitative fluorescence PCR (QF-PCR) were performed as affirmative experiments. In six families, a total of six pathogenic/likely pathogenic variations were identified in four genes including SLC26A2, FGFR3, FLNB, and TMEM38B. These variations caused disorders following autosomal dominant or autosomal recessive inheritance patterns, respectively. The results provided reliable evidence for the subsequent genetic counseling and reproductive options to these families. With its advantage in variation calling and interpreting, trio WES is a promising strategy for the investigation of fetal SDs in cases with normal karyotyping and CMA results. It has considerable prospects to be utilized in prenatal diagnosis.

Figure 1

Results of Sanger sequencing and QF-PCR: (a) a single-base substitution in SLC26A2 (c.292T>C); (b) a three-base deletion in SLC26A2 (c.1018_1020del); (c) a single-base substitution in FGFR3 (c.742C>T); (d) a single-base substitution in FLNB (c.601G>A); (e) a single-base substitution in FGFR3 (c.1138G>A); (f) a single-base substitution in FLNB (c.685T>C); (g) a single-base substitution in exon 3 of TMEM38B (c.344C>A); (h) one copy loss of exon 3 in TMEM38B; (i) one copy loss of exon 4 in TMEM38B.

Figure 2

Biophysical analyses of the FLNB: c.601G>A(p.Ala201Thr) variation: (a) the conservation of FLNB Ala201 between multiple species; (b) part of the three-dimensional structure of the wild type FLNB, with the CH2 subdomain in blue and the actin-binding site 3 in yellow; (c) part of the three-dimensional structure of the mutant FLNB, with the CH2 subdomain in blue and the actin-binding site 3 in yellow; (d) the detailed red block in (b) showing Ala201; (e) the detailed red block in (c) showing Thr201 and the extra hydrogen bonds among T201 and L181, K152, and S177.