Identifying Thrombosis Modifier Genes in the Mouse

鉴定小鼠血栓形成修饰物基因 F V Leiden和FV患者临床病程高度可变的遗传因素 其他形式的血栓形成尚不清楚。该项目的目标1将确定潜在的一个大的子集 利用全基因组ENU突变策略在哺乳动物基因组中检测血栓形成因子基因 在老鼠身上。我们之前报告了一种一致致命的围产期血栓形成小鼠的纯合子 因子V Leiden突变(FvQ/Q)和TFPI杂合子(TFPI+/-)。在初步研究中，我们使用了这个 表型作为大规模基因抑制基因筛选的基础。培育出诱变的FvQ/Q雄性 对双杂合子(FvQ/+TfpI+/-)雌性和&gt；6300存活的G1后代进行基因分型以识别 罕见的Fvq/QTfpi+/-动物在其他方面一致的围产期死亡中幸存下来，推测是由于 突变(单倍体不足)修饰基因的抑制作用。作为原则的证明，基因杂交 Tf基因敲除小鼠证明Tf基因突变将挽救致死性FvQ/Q TFPI+/- 基因分型。到目前为止，已经确定了82个候选抑制突变体。尽管这些突变体中的大多数 由于存活率下降和/或不育性而丢失，其中7个品系的遗传图谱正在进行中 另有4个品系正在进行子代测试。我们将扩展此屏幕以实现4-6倍 基因组覆盖，并通过定位克隆识别相应的基因。目标2将进一步描述 DBA/2J株中Fvq/QTfpi+/-致死表型的株特异性修饰物，由类似的 位置克隆策略。目标3将利用小鼠体内的自然菌株变异来识别 遗传因素对血栓性血小板减少性紫癜(TTP)的易感性至关重要。我们之前的工作 另一些人认为，金属蛋白酶ADAMTS13的缺乏是必要的，但不是足够的 TTP在人和小鼠体内的发展。在初步研究中，我们已经表明 ADAMTS13基因缺失小鼠TTP的发生依赖于不同品系小鼠不同的遗传因素。 将进行敏感和抗性品系之间的遗传杂交，以确定潜在的 基因(S)，这将为TTP的发病机制提供关键的洞察力，并为 对人类有用的诊断和预后标记物。 相关性：这些研究应该为血液凝结的调节提供新的见解，并确定 可能是许多常见遗传性凝血严重程度的重要预测因子的基因数量 疾病。这一信息也可能为这些疾病的治疗提供新的方法。

Identifying Thrombosis Modifier Genes in the Mouse The genetic factors responsible for the highly variable clinical course of patients with factor V Leiden and other forms of thrombophilia are unknown. Aim 1 of this project will identify a large subset of potential thrombosis modifier genes within the mammalian genome using a whole genome ENU mutagenesis strategy in the mouse. We previously reported a uniformly lethal, perinatal thrombosis in mice homozygous for the factor V Leiden mutation (FvQ/Q) and heterozygous for Tfpi (Tfpi+/-). In preliminary studies, we used this phenotype as the basis for a large scale genetic suppressor screen. Mutagenized FvQ/Q males were bred with doubly heterozygote (FvQ/+Tfpi+/-) females and > 6300 surviving G1 offspring genotyped to identify rare FvQ/QTfpi+/- animals who have survived the otherwise uniform perinatal lethality, presumably due to the suppressing effect of a mutated (haploinsufficient) modifier gene. As proof of principal, a genetic cross with TF knockout mice demonstrates that mutation at the TF locus will rescue the lethal FvQ/Q Tfpi+/- genotype. To date, 82 candidate suppressor mutants have been identified. Though most of these mutants have been lost due to decreased survival and/or infertility, genetic mapping for 7 of these lines is in progress and an additional 4 lines are undergoing progeny testing. We will expand this screen to achieve 4-6 fold genome coverage and identify the corresponding genes by positional cloning. Aim 2 will further characterize a strain specific modifier of the FvQ/QTfpi+/- lethal phenotype identified in the DBA/2J strain, by a similar positional cloning strategy. Aim 3 will take advantage of natural strain variation in the mouse to identify genetic factors critical for susceptibility to thrombotic thrombocytopenic purpura (TTP). Previous work by us and others suggests that deficiency of the metalloprotease ADAMTS13 is necessary, but not sufficient for the development of TTP in humans and in mice. In preliminary studies, we have shown that the development of TTP in ADAMTS13 null mice is dependent on genetic factors differing among mouse strains. Genetic crosses between susceptible and resistant strains will be performed to identify the underlying gene(s), which should provide critical insight into the pathogenesis of TTP as well as offering candidates for useful diagnostic and prognostic markers in humans. Relevance: These studies should provide new insight into the regulation of blood clotting, and identify a number of genes that may be important predictors of severity for many common inherited blood clotting diseases. This information may also suggest new approaches to therapy for these conditions.