期刊名称：Proceedings of the National Academy of Sciences
印刷版ISSN：0027-8424
电子版ISSN：1091-6490
出版年度：2017
卷号：114
期号：36
页码：9659-9664
DOI：10.1073/pnas.1705762114
语种：English
出版社：The National Academy of Sciences of the United States of America
摘要：Factor V Leiden ( F5 L ) is a common genetic risk factor for venous thromboembolism in humans. We conducted a sensitized N -ethyl- N -nitrosourea (ENU) mutagenesis screen for dominant thrombosuppressor genes based on perinatal lethal thrombosis in mice homozygous for F5 L ( F5 L/L ) and haploinsufficient for tissue factor pathway inhibitor ( Tfpi +/− ). F8 deficiency enhanced the survival of F5 L/L Tfpi +/− mice, demonstrating that F5 L/L Tfpi +/− lethality is genetically suppressible. ENU-mutagenized F5 L/L males and F5 L/+ Tfpi +/− females were crossed to generate 6,729 progeny, with 98 F5 L/L Tfpi +/− offspring surviving until weaning. Sixteen lines, referred to as “modifier of Factor 5 Leiden ( MF5L1–16 ),” exhibited transmission of a putative thrombosuppressor to subsequent generations. Linkage analysis in MF5L6 identified a chromosome 3 locus containing the tissue factor gene ( F3 ). Although no ENU-induced F3 mutation was identified, haploinsufficiency for F3 ( F3 +/− ) suppressed F5 L/L Tfpi +/− lethality. Whole-exome sequencing in MF5L12 identified an Actr2 gene point mutation (p.R258G) as the sole candidate. Inheritance of this variant is associated with suppression of F5 L/L Tfpi +/− lethality ( P = 1.7 × 10−6), suggesting that Actr2 p.R258G is thrombosuppressive. CRISPR/Cas9 experiments to generate an independent Actr2 knockin/knockout demonstrated that Actr2 haploinsufficiency is lethal, supporting a hypomorphic or gain-of-function mechanism of action for Actr2 p.R258G. Our findings identify F8 and the Tfpi/F3 axis as key regulators in determining thrombosis balance in the setting of F5 L and also suggest a role for Actr2 in this process.
