Identifying Thrombosis Modifier Genes in the Mouse

鉴定小鼠血栓形成修饰基因

• 批准号: 7657076
• 负责人: David Ginsburg
• 金额: $ 37.62万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-04-01 至 2014-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7657076
• 关键词: Alleles; Animals; Behavior Therapy; Blood; Blood Clot; Blood Coagulation Disorders; Blood coagulation; Breeding; Chromosome Mapping; Clinical; Development; Diagnostic; Disease; Disease susceptibility; Ethylnitrosourea; Factor V Leiden mutation; Female; Funding; Genes; Genetic; Genetic Crosses; Genetic Variation; Genome; Genotype; Hemorrhage; Heterozygote; Human; Inbred Strains Mice; Infertility; Inherited; Injection of therapeutic agent; Knock-out; Knockout Mice; Maps; Metalloproteases; Molecular Genetics; Morbidity - disease rate; Mouse Strains; Mus; Mutagenesis; Mutate; Mutation; Pathogenesis; Pathologic; Patients; Perinatal; Phenotype; Predisposition; Prognostic Marker; Regulation; Reporting; Research Personnel; Severities; Source; Suppressor Mutations; Testing; Thrombocytopenic Purpura; Thrombophilia; Thrombosis; Thrombotic Thrombocytopenic Purpura; Variant; Work; base; factor V Leiden; genetic linkage analysis; ilium; insight; male; mammalian genome; mortality; mouse genome; mouse model; mutant; novel strategies; offspring; positional cloning; programs; resistant strain

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.

小鼠血栓形成修饰基因的鉴定 导致凝血因子V Leiden患者高度可变临床病程的遗传因素， 其他形式的血栓形成倾向是未知的。该项目的目标1将确定一个大的潜在子集 使用全基因组ENU诱变策略在哺乳动物基因组内的血栓形成修饰基因 在老鼠。我们先前报道了一个均匀致死，围产期血栓形成的小鼠纯合子为 因子V Leiden突变（FvQ/Q）和Tfpi杂合（Tfpi+/-）。在初步研究中，我们用这个 表型作为大规模遗传抑制筛选的基础。将突变的FvQ/Q雄性交配， 用双杂合子（FvQ/+Tfpi+/-）雌性和> 6300个存活的G1后代进行基因分型以鉴定 罕见的FvQ/QTfpi+/-动物存活，否则围产期死亡率一致，可能是由于 突变（单倍不足）修饰基因的抑制作用。作为原理的证明， 用TF敲除小鼠的研究表明，TF基因座处的突变将拯救致死性FvQ/Q Tfpi+/-突变。 基因型到目前为止，已经鉴定出82个候选抑制突变体。虽然大多数变种人 由于存活率下降和/或不育，这些品系中有7个的遗传图谱正在进行中 另外4个品系正在进行后代测试。我们会把这块屏幕扩大到4-6倍 基因组覆盖率，并通过定位克隆鉴定相应的基因。目标2将进一步描述 在DBA/2 J菌株中鉴定的FvQ/QTfpi+/-致死表型的菌株特异性修饰物，通过类似的 定位克隆策略。Aim 3将利用小鼠的自然品系变异来识别 血栓性血小板减少性紫癜（TTP）易感性的关键遗传因素。我们以前的工作 其他人认为金属蛋白酶ADAMTS 13的缺乏是必要的，但不足以 TTP在人类和小鼠中的发展。在初步研究中，我们已经表明， 在ADAMTS 13缺失小鼠中TTP的发展取决于不同小鼠品系之间的遗传因素。 将进行敏感和耐药菌株之间的遗传杂交，以确定潜在的 基因，这应该提供关键的洞察TTP的发病机制，以及提供候选人， 在人类中有用的诊断和预后标志物。 相关性：这些研究应该为凝血的调节提供新的见解，并确定一个新的机制。 许多基因可能是许多常见遗传性凝血严重程度的重要预测因子 疾病这些信息也可能为这些疾病的治疗提供新的方法。