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Chromatin Regulation of Tissue Regeneration and Stem Cell Function

组织再生和干细胞功能的染色质调节

基本信息

批准号：
10810081
负责人：
Elizabeth Marie Duncan
金额：
$ 1.05万
依托单位：
UNIVERSITY OF KENTUCKY
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-09-01 至 2026-06-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10810081
关键词：
Address Animal Model Animals Automobile Driving Binding Biochemical Biological Biomedical Research Cells Chromatin Cilia Cloning Competence Complex DNA Data Development Developmental Process Dissociation Enzymes Epitopes Essential Genes Family Functional Regeneration Gene Activation Gene Expression Genes Genetic Genetic Transcription Genome Genomics Goals Heterogeneity Histone H3 Histones Homeostasis Individual Injury Knowledge Laboratories Laboratory Study Link Lysine MLL gene MLL2 gene Mammalian Cell Methods Molecular Multienzyme Complexes Multipotent Stem Cells Mus Natural regeneration Neurons Organism Pattern Pharmaceutical Preparations Phenotype Planarians Platyhelminths Play Pluripotent Stem Cells Principal Investigator Process Proteins RNA Interference Recreation Regenerative capacity Regulation Research Research Personnel Retina Role Sequence Homology Signal Transduction Somatic Cell Specificity Testing Time Tissues Transcript Translating Width Work career cell fate specification cell type experience genomic locus histone methylation histone methyltransferase in vivo injured insight novel null mutation overexpression programs protein complex recruit regenerative replacement tissue response single cell analysis skills stem cell function stem cell population stem cells tissue regeneration tool transcription factor undergraduate student

项目摘要

Genomic regulators such as transcription factors and chromatin-modifying enzymes work together to instruct cell fate during developmental processes, including both the routine cell turnover that occurs during homeostasis and the major tissue replacement and repatterning that occurs during regeneration. One specific group of chromatin-modifying complexes whose activity is strongly associated with gene expression is the Set1/MLL family of histone methyltransferases. These enzyme complexes share a core of several essential subunits, yet each particular enzyme (Set1, MLL1, MLL2, etc.) is functionally non-redundant in multicellular animals i.e., individual null mutations are lethal. These data and others, including some from my laboratory, indicate that each Set1/MLL complex regulates its own distinct set of genes. However, many of the molecular and mechanistic details driving this biochemical and functional specificity remain unknown. To uncover the fundamental molecular mechanisms underlying this process, my laboratory studies the planarian model of animal regeneration. Planarians are free-living flatworms with incredible regenerative capacities. They are also amenable to genetic perturbation through RNAi, easily dissociated for single cell analyses, and encode chromatin modifying proteins with strong homology to those of other organisms. For example, planarian Set1 and MLL1/2 enzymes show strong sequence homology to their mammalian counterparts and my previous research has shown that loss of either enzyme leads to loss of their conserved histone modifying activity, trimethylation of histone H3 at lysine 4 (H3K4me3). Moreover, we find that each enzyme targets a specific set of genes that correlate with their phenotype after enzyme loss. Specifically, loss of MLL1/2 through RNAi of its transcript leads to loss of H3K4me3 at highly conserved cilia genes as well as loss of cilia and ciliated cells. However, we do not know how MLL1/2 targets these cilia genes specifically in vivo. To begin identifying the molecular and mechanistic basis of this targeting, we first need to characterize the planarian MLL1/2 complex in terms of its biochemical composition and function. Notably, the phenotype induced in planarians after RNAi of a core Set1/MLL complex subunit, dpy-30, may provide an opportunity to uncover the mechanism that distinguishes Set1 from MLL1/2 function in vivo. To test this idea, I have recruited a promising undergraduate student to identify planarian DPY-30 interacting proteins. He will begin by cloning the two planarian versions of DPY-30 with an epitope tag. He will then express these planarian proteins in mammalian cells and ask if they stably interact with mammalian MLL1/2 proteins and form a functional histone methyltransferase complex. These studies will inform whether planarian DPY-30 is required MLL1/2, but not Set1, assembly and/or function as a histone methyltransferase complex. This project will also allow Ethan to gain the experience and skills needed to build a career in biomedical research.

基因组调节因子如转录因子和染色质修饰酶共同作用