Defects of splicing in antithrombin deficiency

Abstract

Summary Essentials Increasing evidences supports a role for splicing defects in multiple disorders.For antithrombin (AT) deficiency only 7% of mutations disturb intronic splicing sequences.Our study of 141 unrelated cases with AT deficiency found higher rate of splicing defects (>13%).A wide range of gene defects cause different types of AT deficiency through aberrant splicing. BackgroundThere is increasing evidence supporting the relevance of aberrant splicing in multiple disorders. In antithrombin deficiency only 22 intronic mutations affecting splicing sites (7% of SERPINC1 mutations) are considered as splicing mutations.Methods SERPINC1 was analyzed by Sanger sequencing and MLPA in 141 unrelated cases with antithrombin deficiency. Plasma antithrombin was studied by functional and western blot assays, purified by FPLC and characterized by proteomic analysis. In silico predictions on splicing was done with the Human Splicing Finder software.ResultsWe detected 89 different SERPINC1 defects, 13 with potential effect on splicing. Ten cases presented 9 mutations disturbing splicing sites, 5 new. Three gross or small gene defects also disturbed a correct splicing. Interestingly, the first duplication of a single exon ever described (c.1154‐13_1218+115dup), caused mild deficiency (75%). A deeper intronic mutation (c.1154‐14G>A), identified in three unrelated patients with traces of disulphide dimers of antithrombin in plasma, created a cryptic splicing site that might generate a variant with 4 additional in frame residues according to in silico predictions. This aberrant splicing was confirmed by proteomic analysis of the dimer purified from plasma.ConclusionsA high proportion of cases with antithrombin deficiency (up to 13%) may be explained by an aberrant splicing. Up to 15% of mutations in SERPINC1: splicing site variations, gross gene defects and deep intronic mutations, may affect a correct splicing with three potential consequences type I, type II, and even moderate antithrombin deficiency.

How I treat patients with hereditary antithrombin deficiency

“…18,[22][23][24] Type 1 AT deficiency is usually caused by nonsense or null mutations of SERPINC1 (which explains their phenotype) or defects in splicing. 25 Type 2 deficiency is almost exclusively caused by missense mutations. 1,26,27 The estimated prevalence of AT deficiency is ;0.02% to 0.2% in the general population and ;1% to 5% in patients with VTE.…”

Analysis of AlphaFold and molecular dynamics structure predictions of mutations in serpins

“…We also detected the presence of disulphide-linked dimers in plasma, which were slightly bigger than those identified in carriers of the p.Pro112Ser mutation. The purification and proteomic analysis of disulphide-linked dimers from carriers’ plasma confirmed the insertion of the predicted 4 additional amino acids in the variant antithrombin [48].…”

Utility of the SERPINC1 Gene Test in Ischemic Stroke Patients With Antithrombin Deficiency

“…Therefore, AT deficiency types 1 and 2 were not distinguished. However, SERPINC1 c.1154-14G>A variant is well-demonstrated, inducing type 1 deficiency (15). The missense mutations of amino acid 114 to 156, the region involved in heparin-binding, could lead to type 2 deficiency (16,17).…”