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Scalable platforms for understudied histone modifications and modifiers

用于未充分研究的组蛋白修饰和修饰剂的可扩展平台

基本信息

批准号：
10567849
负责人：
Albert Keung
金额：
$ 32.58万
依托单位：
NORTH CAROLINA STATE UNIVERSITY RALEIGH
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-09-06 至 2027-06-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10567849
关键词：
Acetylation Address Adopted Affinity Amino Acids Antibodies Arginine Binding Binding Proteins Biochemical Biochemistry Biological Biological Assay Biological Process Biology Biosensor Bypass Cell physiology ChIP-seq Chemicals Chromatin Chromatin Structure Complement Complex Cost Effectiveness Analysis DNA DNA Binding DNA Modification Process Disease Dopamine Engineering Enzymes Epigenetic Process Eukaryota Gene Expression Gene Expression Regulation Gene Silencing Generations Genetic Transcription Genome Genomic DNA Glutamine Goals Health Heart Heritability Histone Code Histones Human Lysine Measurement Mediating Methylation Modification Neurotransmitters Pattern Play Post-Translational Protein Processing Protein Engineering Proteins Quality Control RNA Reader Reagent Recombinant Proteins Regulation Research Research Personnel Role Serotonin Set protein Site Specificity Structure Surface System Transcriptional Regulation Work Writing Yeasts acyl group biochemical tools cell behavior cell growth regulation combinatorial cost efficient epigenetic silencing gene repression genome-wide histone acetyltransferase histone modification histone-binding proteins live cell imaging monoamine next generation open source preference tool virtual

项目摘要

PROJECT SUMMARY/ABSTRACT Eukaryotic genomic DNA is extensively associated with proteins and RNAs to form chromatin. Through its control of gene expression, changes in chromatin biochemistry and structure underlie nearly all cellular processes. Post-translational modifications of histone proteins that bind genomic DNA play an especially critical role in regulating chromatin structure and function. Modifications of certain histone residues influence the binding of histone proteins to DNA as well as the interactions of other proteins that specifically recognize these modifications. The specific pattern of histone modifications acts as a “histone code” to determine the set of proteins that interact with histones and histone-bound DNA, and consequently participate in diverse cellular processes. Therefore, identification of specific histone modifications, quantitative assessment of the interactions they mediate, and characterization of enzymes that modify histones is essential for understanding chromatin regulation of complex cellular behavior. However, unraveling the histone code is a daunting challenge due to its complexity – over eighty different amino acid residues on five histone proteins undergo over twenty distinct known post-translational modifications. Despite significant advances in research on some histone modifications such as acetylation and methylation, the vast majority of histone modifications remains understudied. Further, a vast majority of enzymes that write even extensively researched modifications like acetylation remain understudied. We propose to address the critical gap in essential biochemical tools and accessible experimental platforms that has hindered research on understudied histone modifications and modifiers. Specifically, we will harness next generation yeast surface display systems as scalable platforms for high throughput studies on understudied histone modifications and modifiers, as well as platforms for engineering biochemical reagents for chromatin research. In addition to understudied histone acetyltransferases, we will focus on three classes of histone modifications: (i) citrullination of arginine (ii) acyl modification of lysine by non-acetyl groups (propionylation, butyrylation, crotonylation), and (iii) monoamine modification of glutamine by serotonin and dopamine. In Aim 1, we will develop a platform for efficient generation of affinity reagents with high specificity that can serve as genetically encoded biosensors for live cell imaging, as well as in conventional analyses like ChIP-seq and CUT&Tag. In Aim 2, we will develop a platform for high throughput identification and quantification of binding interactions mediated by histone modifications. In Aim 3, we will develop a platform for high throughput interrogation of residue preferences of writers. Our work will develop “open source” platforms that are scalable, cost-efficient, and easily adopted by other investigators in chromatin biology. Such platforms will serve as strong complements to traditional biochemical assays by enabling research on both common and understudied histone modifications, and unlocking new high throughput measurements and research questions in chromatin biology.

项目总结/文摘