1q21.1 deletion and a rare functional polymorphism in siblings with thrombocytopenia-absent radius-like phenotypes

Abstract

Thrombocytopenia-absent radii (TAR) syndrome, characterized by neonatal thrombocytopenia and bilateral radial aplasia with thumbs present, is typically caused by the inheritance of a 1q21.1 deletion and a single-nucelotide polymorphism in RBM8A on the nondeleted allele. We evaluated two siblings with TAR-like dysmorphology but lacking thrombocytopenia in infancy. Family NCI-107 participated in an IRB-approved cohort study and underwent comprehensive clinical and genomic evaluations, including aCGH, whole-exome, whole-genome, and targeted sequencing. Gene expression assays and electromobility shift assays (EMSAs) were performed to evaluate the variant of interest. The previously identified TAR-associated 1q21.1 deletion was present in the affected siblings and one healthy parent. Multiple sequencing approaches did not identify previously described TAR-associated SNPs or mutations in relevant genes. We discovered rs61746197 A > G heterozygosity in the parent without the deletion and apparent hemizygosity in both siblings. rs61746197 A > G overlaps a RelA-p65 binding motif, and EMSAs indicate the A allele has higher transcription factor binding efficiency than the G allele. Stimulation of K562 cells to induce megakaryocyte differentiation abrogated the shift of both reference and alternative probes. The 1q21.1 TAR-associated deletion in combination with the G variant of rs61746197 on the nondeleted allele is associated with a TAR-like phenotype. rs61746197 G could be a functional enhancer/repressor element, but more studies are required to identify the specific factor(s) responsible. Overall, our findings suggest a role of rs61746197 A > G and human disease in the setting of a 1q21.1 deletion on the other chromosome.

Trial registration: ClinicalTrials.gov NCT00027274.

Keywords: congenital thrombocytopenia; deformed radius.

Published by Cold Spring Harbor Laboratory Press.

Figures

Figure 1.

Skeletal features of study participants. (A–C) Proband, NCI-107-1. (A) Absent left radius and hypoplastic left ulna; (B) absent right radius and short right ulna; status post–multiple corrective surgeries for right thumb hypoplasia including tendon transfer and centralization; (C) normal right humerus. (D,E) Sister, NCI-107-2. (D) Wide left radius with mild hypoplasia and bony fusion of the proximal radial head to the distal humerus at the left elbow; (E) the right radius and ulna are normal; (D,E) both hands are normal with complete skeletal maturation.

Figure 2.

Schematic of 1q21.1 locus. The deleted region in family NCI-107 is shown in relation to the deletion previously reported in thrombocytopenia-absent radius (TAR) syndrome based on the hg38 reference genome. The transcription factor binding site (TFBS) affected by rs61746197 A > G is denoted as are the locations of the previously TAR-associated SNPs, rs139428292 and rs201779890.

Figure 3.

Electromobility shift assays (EMSAs) suggest that rs61746197 A > G is a functional SNP. (A) Ten micrograms of unstimulated nuclear extract from K562 cells was incubated with reference (Ref, R) or alternative (Alt, A) probe as indicated. Cold Ref probe titrates signal away from Alt at a lower stoichiometric ratio than the converse (arrow and compare lanes 11–13 with 6–8). (B) Unstimulated extract (left panel, U) exhibits a markedly stronger baseline shift than TPA stimulated extract (right panel, S). Unstimulated nuclear extract (U) when incubated with labeled alternative probe (A) produces a novel secondary shift not observed with either the reference probe (R) or the TPA stimulated extract (S). Addition of RelA–p65 antibody abrogates this secondary shift (arrow).