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PRODIGE: A high-throughput tool for joint profiling of protein-DNA interactions and gene expression in single cells

PRODIGE：一种高通量工具，用于联合分析单细胞中蛋白质-DNA 相互作用和基因表达

基本信息

批准号：
10303542
负责人：
Wei Wei
金额：
$ 27.45万
依托单位：
INSTITUTE FOR SYSTEMS BIOLOGY
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：
2021-09-30 至 2023-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10303542
关键词：
Address Antibodies Bar Codes Benchmarking Binding Biological Assay Cell Differentiation process Cell Line Cells Chromatin Clinical Complex DNA DNA-Binding Proteins DNA-Protein Interaction Data Data Set Development Disease Drops Epigenetic Process Gene Expression Genes Genetic Transcription Genome Genomics Heterogeneity Histones Hydrogels Individual Joints Libraries Maintenance Maps Measurement Measures Mediating Messenger RNA Methods Microfluidic Microchips Microfluidics Modality Modeling Molecular Output Pattern Performance Phenotype Play Poly A Population Post-Translational Protein Processing Process Protein Analysis Proteins Protocols documentation RNA Regulator Genes Reporting Reproducibility Research Personnel Resolution Role Sampling System Techniques Technology Testing Tissues cancer cell cell fixation cell type computational pipelines data analysis pipeline droplet sequencing epigenomics genetic manipulation genome-wide histone modification insight interest methylome multiple omics next generation next generation sequencing novel programs protein profiling single-cell RNA sequencing tool transcription factor transcriptome

项目摘要

PROJECT SUMMARY/ABSTRACT Protein-DNA interactions control access to genes in chromatins and regulate the spatial- and temporal-specific gene expression. These interactions are crucial for cell differentiation and the maintenance of cellular diversity. Dysregulated patterns of protein-DNA interactions are often associated with disease states. Applications of next- generation epigenomic sequencing techniques have enabled deconvolution of protein-DNA interactions at single-cell level. However, it remains challenging to define how cell-to-cell heterogeneity in protein-DNA interactions impacts gene expression variability. Currently available methods for joint profiling protein-DNA interactions and gene expression from single cells require laborious genetic manipulation, suffer relatively low throughput, and are impossible to specifically map proteins with post-translational modifications, such as histone marks. To address these unmet needs, we propose to develop PRODIGE – a high-throughput tool for joint profiling of protein-DNA interactions and gene expression in the same single cells without the need of genetic manipulations. It is enabled by a strategic integration of reversible cell fixation, antibody-guided chromatin tagmentation, novel deformable and degradable barcoded hydrogel beads, droplet microfluidics, and next- generation sequencing technology. In Aim 1, we will develop technical components and analytical pipelines of PRODIGE. We will prepare protein A-Tn5 transposome for antibody-guided Tn5 tagmentation, fabricate microfluidic devices for single-cell PRODIGE assay, and synthesize a barcoded library of deformable and degradable barcoded hydrogel beads for single-cell barcoding. In Aim 2, we will optimize the method in bulk and single-cell levels, and assess the performance of PRODIGE (joint analysis) in comparison to other stand-alone counterparts. In Aim 3, we will benchmark PRODIGE to established tools for joint single-cell profiling of protein- DNA interactions and transcriptome. Successful implementation of the proposed program will deliver a high- throughput tool to quantify how cell-to-cell heterogeneity in protein-DNA binding influences gene expression variability, decipher cell-type-specific patterns of upstream protein-DNA interactions and their transcriptional outputs, and identify protein-mediated mechanisms that regulate cell-type-associated transcriptional programs in heterogeneous cell populations and under different developmental and/or disease conditions.

项目总结/文摘