权益分类	功能权益	普通用户	{{item.name}}会员
{{category.name}}	{{benefitItem.name}}

Mechanisms of Homeodomain Transcriptional Specificity

同源域转录特异性的机制

基本信息

批准号：
10673333
负责人：
BRIAN GEBELEIN
金额：
$ 5.96万
依托单位：
CINCINNATI CHILDRENS HOSP MED CTR
依托单位国家：
美国
项目类别：
财政年份：
2008
资助国家：
美国
起止时间：
2008-04-01 至 2024-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10673333
关键词：
Abdomen Address Animals Anterior Architecture Binding Binding Sites Biochemical Bioinformatics Biological Assay Biological Process Cell Culture Techniques Cells Complex DNA DNA Binding DNA Sequence Data Development Drosophila genus Drosophila melanogaster Embryo Ensure Event Family Family member Gene Activation Gene Expression Genes Genetic Genetic Transcription Genomics Goals Health Heterodimerization Homeodomain Proteins Homo Human Human Development In Vitro Individual Logic Luciferases Mammalian Cell Mammals Mediating Neurons Organism Organogenesis Outcome Output Pattern Physiological Process Proteins Publishing Regulation Regulator Genes Reporter Repression Research Proposals Role Site Specific qualifier value Specificity System Testing To specify Transcription Repressor Transgenic Organisms body system cell type comparative dimer fly gene interaction gene repression homeodomain in vivo insight leukemia member molecular domain monomer neuroblast novel programs structural biology transcription factor transcriptome sequencing

项目摘要

PROJECT ABSTRACT Homeodomain (HD) proteins comprise a large family of transcription factors (TFs) that regulate numerous aspects of animal development. For example, members of the Hox-like (HoxL) and Nkx-like (NKL) HD proteins regulate processes ranging from patterning of the anterior-posterior axis (A-P) of the embryo to specifying individual cell fates within different organ systems. Intriguingly, the HoxL and NKL proteins have highly similar HDs that bind largely overlapping AT-rich DNA sequences in vitro. These findings provide a classic TF specificity paradox: How do TFs with highly similar in vitro DNA binding activities achieve sufficient in vivo specificity to ensure the accurate regulation of genetic programs in different cell types? To address this paradox, my lab is focused on defining how HD TFs achieve in vivo specificity by forming cooperative TF complexes on cis- regulatory modules. Our preliminary and published data reveal that members of the HoxL and NKL TFs differ in their ability to form homo- and heterodimer TF complexes on DNA. For instance, we unexpectedly found that the Gsx/Ind TFs, which specify neuronal cell fates in animals from flies to mammals, differentially regulate gene expression when bound to DNA as monomers versus homodimers. In contrast, the Abdominal-A (Abd-A) Hox TF, which specifies distinct cell fates in the Drosophila abdomen, does not bind DNA as a homodimer, but instead cooperatively binds DNA with three other HD proteins: Extradenticle (Exd), Homothorax (Hth), and Engrailed (En). These data support the hypothesis that HD TFs achieve target and regulatory specificity by binding distinct combinations of AT-rich DNA sites as monomers, cooperative homodimers, or cooperative heterodimers. To test this hypothesis, we propose two aims: In Aim1, we propose to determine how HD monomer versus homodimer binding impacts target gene binding and regulation. To achieve this goal, we will (1) systematically define which HoxL and NKL HDs cooperatively bind DNA as homodimers; (2) assess the regulatory potential of each HD on monomer vs dimer sites in cell culture assays; and (3) define the mechanism and function of Ind homodimer formation on Drosophila neuroblast gene expression using structural biology and transgenic reporter, CUT&RUN, and RNA-seq assays. In Aim2, we propose to define how the choice of Hox heterodimer partner impacts the DNA binding and regulatory specificity of the Abd-A Hox TF. To achieve this goal, we will (1) define the DNA motifs and molecular domains required for cooperative Abd-A/Hth and Abd-A/En complexes; (2) test the role of Abd-A heterodimerization domains in gene activation and repression assays in the Drosophila embryo; (3) define the in vivo binding motifs and target genes regulated by Abd-A with a focus on identifying heterodimer binding events using CUT&RUN and RNA-seq assays. Since the TFs and biological processes studied are highly conserved between flies and mammals, we are optimistic our studies will uncover gene regulatory mechanisms relevant to human health and development.

项目摘要