Indentification of Hox target genes controlling tissue growth

描述(申请人提供)：破坏组织生长和模式是癌症发展的常见步骤，适当的组织形态依赖于细胞间信号通路和区域特异性转录因子的组合输入。在果蝇中，超双胸蛋白(Ubx)是一种区域特异性转录因子，是Hox同源结构域蛋白家族的成员，它通过调节有丝分裂十五肢瘫痪(DPP)途径的几个组成部分的表达来限制后翅(Hatere)组织的生长。该项目的目标是确定果蝇基因组中受空腹Ubx直接调控的靶基因和转录调控元件；将使用三种实验方法来实现这一目标。首先，一种新的基因表达谱方法将被用来分析序列同源Ubx表达(Haltere)和非表达(WINE)组织中的区域基因表达差异。第二步将使用染色质免疫沉淀和DamID结合果蝇基因组拼接阵列来确定与Hatere中的Ubx直接结合的DNA区域。总而言之，这些实验将允许在全基因组范围内识别龟甲中的直接Ubx靶基因。这项分析的第三步将是使用Cognate Site Identity(CSI)微阵列技术来表征Ubx的DNA结合特异性，无论是作为单体还是与Hox辅因子牙外环(EXD)和同胸(HTH)结合。这将允许探索Ubx可以和不能结合的DNA序列，结合前两个步骤，应该可以更好地理解HOX调控网络。对Ubx靶基因子集的后续分析，特别是那些可能参与生长调节的基因，将包括验证它们在翅膀和龟头中的表达模式，并使用标准的果蝇遗传方法进行功能分析。公共卫生相关性：HOX蛋白和许多控制生长、增殖和形态发生的信号分子在果蝇和人类之间是保守的；本研究的目标是深入了解HOX转录因子调节组织生长的机制。最终，详细描述Hox蛋白在正常发育过程中如何调节组织生长，对于理解它们在癌症等疾病状态下生长受到错误调控时的作用至关重要。

DESCRIPTION (provided by applicant): Disruption of tissue growth and patterning is a common step in the development of cancer, and proper tissue morphology depends upon combined input from intercellular signaling pathways and region-specific transcription factors. In Drosophila melanogaster the protein Ultrabithorax (Ubx), a region-specific transcription factor and member of Hox family of homeodomain proteins, restricts growth of hindwing (haltere) tissue in part by modulating the expression of several components of the mitogenic Decapentaplegic (Dpp) pathway. The goal of this project is to identify target genes and transcriptional regulatory elements in the Drosophila genome that are directly regulated by Ubx in the haltere; three experimental approaches will be used to accomplish this goal. First, a novel gene expression profiling .method will be used to analyze regional gene expression differences in serially homologous Ubx-expressing (haltere) and non-expressing (wing) tissues. The second step will use chromatin immunoprecipitation and DamID in combination with Drosophila genome tiling arrays to determine the DNA regions that are directly bound by Ubx in the haltere. Together, these experiments will allow the identification of direct Ubx target genes in the haltere on a genome-wide scale. The third step in this analysis will be to use Cognate Site Identity (CSI) microarray technology to characterize the DNA-binding specificity of Ubx, both as a monomer and in conjunction with the Hox cofactors Extradenticle (Exd) and Homothorax (Hth). This will permit exploration of the DNA sequences to which Ubx can and cannot bind, and, in combination with the first two steps, should allow a better understanding of a Hox regulatory network. Follow-up analysis on a subset of Ubx target genes, particularly those likely to be involved in growth regulation, will include the validation of their expression patterns in both the wing and haltere and functional analysis using standard Drosophila genetic approaches. Public Health Relevance: The Hox proteins and many of the signaling molecules controlling growth, proliferation and morphogenesis are conserved between Drosophila and humans; the goal of this research is to provide insight into the mechanisms by which a Hox transcription factor regulates tissue growth. Ultimately, a detailed description of how Hox proteins regulate tissue growth during normal development will be crucial to understanding their role when growth is misregulated in diseases states such as cancer.