Biochemical and cellular consequences of the antithrombin p.Met1? mutation identified in a severe thrombophilic family

Abstract

Nature is always the best inspiration for basic research. A family with severe thrombosis and antithrombin deficiency, the strongest anticoagulant, carried a new mutation affecting the translation-start codon of SERPINC1, the gene encoding antithrombin. Expression of this variant in a eukaryotic cell system produced three different antithrombins. Two downstream methionines were used as alternative initiation codons, generating highly expressed small aglycosylated antithrombins with cytoplasmic localization. Wild-type antithrombin was generated by the use of the mutated AUU as initiation codon. Actually, any codon except for the three stop codons might be used to initiate translation in this strong Kozak context.We show unexpected consequences of natural mutations affecting translation-start codons. Downstream alternative initiation AUG codons may be used when the start codon is mutated, generating smaller molecules with potential different cell localization, biochemical features and unexplored consequences. Additionally, our data further support the use of other codons apart from AUG for initiation of translation in eukaryotes.

Identification and characterization of two SERPINC1 mutations causing congenital antithrombin deficiency

“…Type I: Classical deficiency, characterized by impaired AT synthesis and a parallel decrease in plasma AT antigen and activity in patients. Its clinical manifestations are severe and usually associated with severe mutations in SER-PINC1, which lead to a significant decrease in the level of AT in plasma owing to mutations that destabilize mRNA, misfolded proteins, and intracellular retention, or lead to abnormal degradation [21][22][23]. Type II mainly affects the domain function bound to thrombin or heparin, with normal plasma AT antigen content, but its activity is weakened [24], its clinical manifestations are mild, and its incidence is relatively high [25].…”

“…SERPINC1 encodes 32 amino acid signal peptides and 432 amino acid maturation proteins [38,39]. The primary function of this signal peptide is to direct nascent polypeptide chains into the endoplasmic reticulum, followed by post-translational processing, including disulfide bond formation, N-glycosylation, signal peptide cleavage, and correct folding of the protein into a metastable conformation, which is required for the inhibitory activity of this serine protease inhibitor [22]. A portion of exons 2 and 3 encodes the heparin-binding site region [40].…”