Modulation of ADAMTS13 secretion and specific activity by a combination of common amino acid polymorphisms and a missense mutation

Abstract

Sequence analysis of the ADAMTS13 locus of 2 patients with hereditary thrombotic thrombocytopenic purpura (TTP) revealed the homozygous presence of 4 single nucleotide polymorphisms (SNPs) (R7W, Q448E, P618A, A732V) and a rare missense mutation (R1336W). Analysis of the individual effect of any amino acid exchanges showed that several sequence variations can interact with each other, thereby altering the phenotype of ADAMTS13 deficiency. Introduction of polymorphisms R7W, Q448E, and A732V had no or only minor effects on ADAMTS13 secretion. In contrast, P618A, R1336W, and the A732V-P618A combination strongly reduced ADAMTS13-specific activity and antigen levels. Surprisingly, R7W and Q448E were positive modifiers of ADAMTS13 secretion in the context of P618A and A732V but neither could rescue the severely reduced specific activity conferred by P618A. However, in the context of R1336W, polymorphisms R7W and Q448E enhanced the detrimental effect of the missense mutation and led to undetectable enzyme activity. We show that dependent on the sequence context, the same polymorphisms might be either positive or negative modifiers of gene expression. Our results might therefore be widely relevant to understanding the influence of polymorphisms on the phenotypic expression of complex diseases. Introductionvon Willebrand factor (VWF) is crucial for primary hemostasis by mediating extracellular matrix-platelet interactions and delivering factor VIII to the site of vascular injury. The large and heterogeneous glycoprotein circulates as a multimer composed of identical units, ranging in molecular weight from 500 kDa to more than 20 000 kDa. 1 Plasma VWF is predominately secreted as high molecular weight, "unusually large," VWF (ULVWF) multimers. Because ULVWF binds more avidly to components of the extracellular matrix 2 and to various platelet receptors 3,4 than low molecular weight VWF, the hemostatic activity of VWF is size dependent and much more pronounced for high molecular weight VWF than for smaller multimers. This intrinsic property of ULVWF allows for platelet attachment to sites of vascular injury under high shear stress conditions of fastflowing blood. 5 However, immediately downstream from the site of injury, the multimeric size of VWF needs to be physiologically regulated to prevent the formation of platelet and VWF-rich thrombi.A VWF-cleaving protease, now called ADAMTS13 (a disintegrin and metalloprotease with thrombospondin type-1 motifs) has been identified. [6][7][8][9][10][11][12][13] ADAMTS13 cleaves VWF between Tyr1605 and Met1606 in the A2 domain of the VWF monomer, 6,7,14,15 yielding typical fragments of 176 kDa and 140 kDa and smaller VWF multimers. Severe deficiency of ADAMTS13 may result in ULVWF spontaneously interacting with platelet receptors 5 and cause the life-threatening disorder thrombotic thrombocytopenic purpura (TTP). 16,17 Acquired TTP has been associated with the inhibition of ADAMTS13 activity by autoimmune antibodies, [18][19][20][21] whereas mutations in the ADAMTS13 gene have been ...

Thrombotic Thrombocytopenic Purpura: A Thrombotic Disorder Caused by ADAMTS13 Deficiency

“…More than 65 mutations, including nonsense, missense, frame-shifting insertion or deletion and splicing mutations, have been detected in patients with hereditary TTP [99][100][101][102][103]. The mutations affect the process of protease synthesis, activity, or more commonly, secretion.…”

“…More than 25 polymorphisms have also been detected in the coding sequence of ADAMTS13. Certain combinations of polymorphisms or mutation-polymorphism may result in severe ADAMTS13 deficiency [100].…”

The D173G mutation in ADAMTS-13 causes a severe form of congenital thrombotic thrombocytopenic purpura

“…Inherited TTP is a rare autosomal recessive disorder due to homozygous or double heterozygous mutations in the ADAMTS-13 gene, causing a severe decrease of ADAMTS-13 level and activity (10)(11)(12)(13)(14)(15)(16). About 100 mutations causing inherited ADAMTS-13 deficiency have been identified so far in regions of the gene encoding different domains (10)(11)(12)(13)(14)(15)(16)(17) with only a few of them characterized by in vitro expression studies (12,15,(18)(19)(20)(21)(22). In this manuscript, we report cases of congenital TTP, due to the homozygous mutation in ADAMTS-13 gene in two young brothers born from two consanguineous parents of Romanian origin.…”

Linkage analysis and mutation screening of the rhodopsin gene in a Chinese Bai family with autosomal dominant retinitis pigmentosa

“…Previous studies on thrombotic thrombocytopenic purpura have shown that, in addition to ADAMTS13 mutations, SNPs located in the ADAMTS13 coding region can influence plasmatic ADAMTS13 activity levels, 22 and a combination of SNPs and a missense mutation can also modulate ADAMTS13 secretion. 23 Therefore, in view of the implication of SNPs as genetic risk factors contributing to disease development and disease susceptibility, detailed investigations of a potential association between the two SNPs and adRP in the Chinese Bai population, and of the effect of combinations of the Pro347leu mutation with either or both SNPs, are needed.…”