Role of FIR in limb morphogenesis

FIR 在肢体形态发生中的作用

基本信息

批准号：
8553102
负责人：
Susan Mackem
金额：
$ 6.2万
依托单位：
DIVISION OF BASIC SCIENCES - NCI
依托单位国家：
美国
项目类别：
财政年份：
资助国家：
美国
起止时间：
至
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/8553102
关键词：
Alleles Apical Ectodermal Ridge Apoptosis Biological CCR Cancer Biology Cell Death Cell Proliferation Cell Survival Cell physiology Cells Cessation of life Collaborations Colorectal Cancer Complex Development Developmental Biology Differentiation and Growth Digit structure Drosophila genus Embryo Ensure Erinaceidae FBP interacting repressor Factor Analysis Fibroblast Growth Factor Fingers Fluorescence Gene Expression Regulation Gene Targeting Genes Genetic Genetic Transcription Genomics Goals Growth Homologous Gene Human Intercept Joints Knock-out Lead Learning Length Light Limb Bud Limb Development Limb structure Link Malignant Neoplasms Mediator of activation protein Mesoderm Mission Morphogenesis Mus Neoplasm Metastasis Neoplasms Normal tissue morphology Organ Pathology Pathway interactions Pattern Phalanx Phenotype Physiological Processes Play Process Proteomics RNA Splicing Regulation Reporter Research Role Shapes Signal Transduction Skeleton Structure Study models System Systems Biology Tamoxifen Testing Tissues Tumor Biology Vertebrates Wing base beta catenin c-myc Genes cancer therapy cell behavior cell motility cellular targeting design gene function genome wide association study implantation in vivo mutant neoplastic cell null mutation programs promoter recombinase skeletal smoothened signaling pathway transcription factor tumorigenesis

项目摘要

The developing limb is a well studied model for morphogenesis in which many of the regulatory components governing skeletal pattern and growth have been elucidated, providing a strong biological framework for interpreting gene function based on mutant phenotypes. FIR (FBP interacting repressor) is a highly evolutionarily conserved negative regulator of myc transcription present in both vertebrates and Drosophila. The Drosophila homolog (half pint) controls the zone of non-proliferation in the developing embryonic wing disc. The human homolog has been implicated in colorectal cancer; alternatively spliced dominant-interfering forms of FIR are associated with these cancers. Elucidation of the normal function of FIR in mammalian tissues has been hampered by the very early embryonic lethal (prior to implantation) of FIR null mutant embryos, limiting the ability to assess the role of this gene in normal tissue morphogenesis. In collaboration with Dr. Levens (CCR, NCI), who has generated a conditional (floxed) allele of mouse FIR, we are using several Cre drivers to selectively remove FIR from the early limb bud. We want to test whether FIR plays a role in regulating limb outgrowth, and if it does, learn how it acts to regulate growth. In preliminary results, we have found that, surprisingly, loss of FIR function in early limb bud mesoderm results in considerable apoptosis leading to truncations of the limb skeleton. In Drosophila, half pint is a target of Wg (wingless, Wnt) pathway. Interestingly, loss of the Wnt signaling effector, beta-catenin, from limb bud mesoderm also causes limb truncation phenotypes in mouse embryos, related to abnormalities in formation of an early Fgf signaling center in the limb bud (AER, apical ectodermal ridge) that is necessary for limb outgrowth. We are analyzing components of the Wnt pathway to determine whether they are altered in the FIR mutant limbs, along with other analyses of factors and signaling centers critical for cell survival in the early limb. In parallel studies we are analyzing the role of FIR in cell survival and proliferation at the cellular level using MEF cells derived from the conditional knock-out line that also carry a conditional tamoxifen-activated Cre recombinase and a dual fluorescence reporter to allow FACS isolation of recombined (FIR knock-out) cells. These studies will shed new light on the varied roles of FIR in regulating growth, and will have implications for developing cancer therapies that may target function of this Myc-regulator.

发育中的肢体是一个经过充分研究的形态发生模型，其中许多控制骨骼模式和生长的调节成分已被阐明，为解释基于突变表型的基因功能提供了强大的生物学框架。 FIR（FBP 相互作用阻遏蛋白）是一种高度进化保守的 myc 转录负调节因子，存在于脊椎动物和果蝇中。果蝇同源物（半品脱）控制发育中的胚胎翼盘的非增殖区。人类同源物与结直肠癌有关； FIR 的选择性剪接显性干扰形式与这些癌症相关。 FIR 在哺乳动物组织中的正常功能的阐明受到 FIR 无效突变胚胎的极早期胚胎致死（在植入之前）的阻碍，限制了评估该基因在正常组织形态发生中的作用的能力。 Levens 博士（CCR、NCI）已生成了小鼠 FIR 的条件（floxed）等位基因，我们与该博士合作，使用多个 Cre 驱动程序选择性地从早期肢芽中去除 FIR。我们想要测试 FIR 是否在调节肢体生长方面发挥作用，如果有的话，了解它如何发挥调节生长的作用。在初步结果中，我们令人惊讶地发现，早期肢芽中胚层中 FIR 功能的丧失会导致大量细胞凋亡，从而导致肢体骨骼截断。在果蝇中，半品脱是 Wg（wingless，Wnt）途径的目标。有趣的是，肢芽中胚层中 Wnt 信号效应子 β-catenin 的缺失也会导致小鼠胚胎出现肢体截断表型，这与肢芽中早期 Fgf 信号中心（AER，顶端外胚层脊）形成的异常有关。肢体生长所必需的。我们正在分析 Wnt 通路的组成部分，以确定它们是否在 FIR 突变肢体中发生改变，同时对早期肢体中细胞存活至关重要的因素和信号中心进行其他分析。在平行研究中，我们正在使用源自条件敲除系的 MEF 细胞在细胞水平上分析 FIR 在细胞存活和增殖中的作用，该细胞系还携带条件他莫昔芬激活的 Cre 重组酶和双荧光报告基因，以允许 FACS 分离重组（FIR 敲除）细胞。这些研究将为 FIR 在调节生长中的不同作用提供新的线索，并将对开发可能针对这种 Myc 调节器功能的癌症疗法产生影响。