A sensitized mutagenesis screen in Factor V Leiden mice identifies novel thrombosis suppressor loci

Factor V Leiden () is a common genetic risk factor for venous thromboembolism in humans. We conducted a sensitized ENU mutagenesis screen for dominant thrombosuppressor genes based on perinatal lethal thrombosis in mice homozygous for () and haploinsufficient for tissue factor pathway inhibitor (). deficiency enhanced survival of mice, demonstrating that lethality is genetically suppressible. ENU-mutagenized males and females were crossed to generate 6,729 progeny, with 98 offspring surviving until weaning. Sixteen lines exhibited transmission of a putative thrombosuppressor to subsequent generations, with these lines referred to as (odifier of actor eiden) 1-16. Linkage analysis in identified a chromosome 3 locus containing the tissue factor gene (). Though no ENU-induced mutation was identified, haploinsufficiency for () suppressed lethality. Whole exome sequencing in identified an gene point mutation (p.R258G) as the sole candidate. Inheritance of this variant is associated with suppression of lethality (p=1.7x10), suggesting that is thrombosuppressive. CRISPR/Cas9 experiments to generate an independent knockin/knockout demonstrated that haploinsufficiency is lethal, supporting a hypomorphic or gain of function mechanism of action for . Our findings identify and the axis as key regulators in determining thrombosis balance in the setting of and also suggest a novel role for in this process.