Development of a Novel Mouse Model to Evaluate HTLV Tax Transformation

开发新的小鼠模型来评估 HTLV 税收转型

• 批准号: 8637946
• 负责人: SUSAN J MARRIOTT
• 金额: $ 16.51万
• 依托单位: BAYLOR COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-04-01 至 2016-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8637946
• 关键词: Adult T-Cell Leukemia/Lymphoma; Apoptosis; Binding; Biological; Biological Models; Biological Process; Blood Cells; C57BL/6 Mouse; CREB1 gene; Cell Culture System; Cell physiology; Cells; Cellular biology; Communities; Development; Disease; Gene Dosage; Gene Expression; Gene Silencing; Genetic Recombination; Goals; Human; Human T-Cell Leukemia Viruses; Human T-lymphotropic virus 1; Infection; Knock-in Mouse; Knowledge; Link; Malignant Neoplasms; Mediating; Modeling; Mus; Mutation; NF-kappa B; Oncogene Proteins; Pathway interactions; Pharmaceutical Preparations; Play; Problem Solving; Protein Family; Proteins; Resources; Retroviridae; Role; Site; T-Lymphocyte; Taxes; Time; Transgenes; Transgenic Mice; Transgenic Model; Viral; Virus; Work; cancer type; cell transformation; improved; innovation; insight; leukemia; member; metaplastic cell transformation; mouse model; mutant; novel; novel therapeutics; promoter; protein expression; public health relevance; tax Gene Products; tax Genes; thymocyte; tool; transgene expression; tumor; tumorigenesis; tumorigenic; uncontrolled cell growth; vector

DESCRIPTION (provided by applicant): The goal of this proposal is to establish a novel mouse transformation model in which the human T cell leukemia virus type I (HTLV-1) Tax oncoprotein is conditionally expressed in T cells, and to use this model to determine the transforming activity of wild-type and mutant Tax proteins. HTLV-1 is a human retrovirus that currently infects approximately 15 million people in the world and is the cause of the aggressive malignancy, adult T cell leukemia/lymphoma. Although the Tax oncoprotein has been shown to transform cells in culture and to induce tumors in a variety of transgenic mouse models, the mechanism by which Tax transforms cells is not well understood. A large number of Tax mutants have been generated and their biological activities have been thoroughly characterized primarily in cell culture systems. A major problem currently facing the field is that the transforming activity of Tax mutants cannot be compared using available transgenic models due to random transgene integration sites, variable transgene copy number and inconsistent transgene expression levels. Thus, it is very difficult to link the biological activities of Tax mutants with their transforming potential. To solve this problem we will develop an innovative mouse model in which to study Tax transformation using vectors containing wild-type or mutant Tax genes that are silenced by a preceding floxed stop cassette. These vectors will be knocked in to the Rosa26 locus of recipient mice by recombination. By crossing these mice with Lck-CRE mice, the stop cassette will be specifically excised in developing thymocytes where the Lck promoter is active, allowing conditional expression of wild-type or mutant Tax proteins in T cells, the natural target of HTLV-1 infection. Insertion into the Rosa26 locus will eliminate inconsistent integration sites and standardize gene copy number resulting in consistent levels of wild-type and mutant Tax protein expression. The mouse model will be established in Aim 1 by creating targeting vectors containing silenced wild-type or mutant Tax genes. Targeting vectors will be knocked in to the Rosa26 locus of C57BL/6 mice, which will then be crossed with homozygous Lck-CRE mice, thereby excising the stop cassette to generate mice expressing wild-type or mutant Tax specifically in T cells. Aim 2 will examine the effect of mutations that disable specifi biological functions of Tax on Tax- mediated tumorigenesis. Tax can bind to and regulate the activity of members of the SRF, CREB, NF-kB and PBM protein families, each of which has been implicated in oncogenesis. Mice established in Aim 1 will be used to compare for the first time the tumorigenic potential of wild-type and mutant Tax proteins in an effort to identify pathways that are required for Tax tumorigenesis. The proposed studies will establish a new mouse model that will overcome current limitations and provide greater insight into the mechanism of HTLV-1 Tax transformation, knowledge that is currently lacking and that promises to yield novel insights into viral and cellular biology.

描述（由申请人提供）：该提案的目的是建立一个新型的小鼠转化模型，其中人类T细胞白血病I型（HTLV-1）税量癌蛋白在T细胞中有条件地表达，并使用该模型来确定野生型和突变税蛋白的转化活性。 HTLV-1是人类逆转录病毒，目前感染了世界上约1500万人，是侵略性恶性肿瘤，成人T细胞白血病/淋巴瘤的原因。尽管已显示税收癌蛋白可以转化培养物中的细胞并诱导各种转基因小鼠模型中的肿瘤，但税收转化细胞的机制尚不清楚。已经产生了大量的税收突变体，其生物活性主要在细胞培养系统中彻底表征。目前面临该领域的一个主要问题是，由于随机转基因整合位点，可变的转基因拷贝数和不一致的转基因表达水平，无法使用可用的转基因模型来比较税收突变体的转化活动。因此，很难将税收突变体的生物学活性与它们的转变潜力联系起来。为了解决这个问题，我们将开发一种创新的小鼠模型，在该模型中，使用含有野生型或突变税基因的向量研究税收转换，这些媒介被先前的floxed停止盒子沉默。这些向量将被重组撞到受体小鼠的Rosa26基因座。通过将这些小鼠与LCK-CRE小鼠跨越，将在发育中的LCK启动子处于活性的胸腺细胞中专门切开止盒，从而使T细胞中野生型或突变税蛋白的有条件表达 HTLV-1感染的自然靶标。插入ROSA26基因座将消除不一致的 整合位点并标准化基因拷贝数，导致野生型和突变税蛋白表达的水平一致。小鼠模型将通过创建含有沉默的野生型或突变税基因的靶向矢量来建立。靶向矢量将被撞到C57BL/6小鼠的Rosa26基因座，然后将其与纯合LCK-CRE小鼠交叉，从而使止动磁带分解，从而产生表达野生型或突变税的小鼠，这些小鼠在T细胞中专门在T细胞中特有。 AIM 2将检查禁用税收税对税收介导的肿瘤发生的突变的影响。税收可以结合并调节SRF，CREB，NF-KB和PBM蛋白质家族的活性，每个蛋白质都与肿瘤发生有关。在AIM 1中建立的小鼠将首次使用野生型和突变税蛋白的致瘤潜力进行比较，以识别税收肿瘤发生所需的途径。拟议的研究将建立一个新的小鼠模型，该模型将克服当前的局限性，并为HTLV-1税收转变的机制，当前缺乏的知识以及有望产生对病毒和细胞生物学的新见解。