SBIR-STTR Award

Epigenome analysis can help dissect the transcriptional regulatory sequences that control spatiotemporal patterns of gene expression during anima development and disease pathogenesis, but a major challenge to epigenome analysis is the heterogeneity of primary tissues. Conventional epigenome assays that take bulk tissues as input only produce population average signals. To address this major bottleneck, we propose to develop an ultra-high throughput single-cell multi-omics method, Paired-Tag, for joint profiling of histone modifications and transcriptome. In preliminary experiments, we have demonstrated the feasibility and utility of this method through analysis of the nuclear transcriptome and multiple histone modifications at single cell resolution in the adult mouse frontal cortex and hippocampus. In the proposed study, we will further optimize the current Paired-Tag protocol and demonstrate its utility in cancer epigenome analysis. If successful, the research would add a major toolkit for the production of cell-type-resolved maps of chromatin state and transcriptome in complex tissues and enable next generation epigenome analysis of tumor samples. Public Health Relevance Statement We proposed to develop a powerful high throughput method to investigate the inner workings of human cells at single cell resolution. The resulting tool could have broad use in biomedical research.

Project Terms:
Adoption ; Adult ; 21+ years old ; Adult Human ; adulthood ; Antibodies ; Architecture ; Engineering / Architecture ; Bar Codes ; barcode ; Biological Assay ; Assay ; Bioassay ; Biologic Assays ; Biomedical Research ; Biopsy ; Complementary DNA ; cDNA ; Malignant Neoplasms ; Cancers ; Malignant Tumor ; malignancy ; neoplasm/cancer ; Cell Nucleus ; Nucleus ; Cells ; Cell Body ; Chromatin ; Chromosomes ; Communities ; Disease ; Disorder ; DNA ; Deoxyribonucleic Acid ; frontal lobe ; frontal cortex ; Gene Expression ; Genes ; Regulator Genes ; Transcriptional Regulatory Elements ; regulatory gene ; trans acting element ; Goals ; Heterogeneity ; Hippocampus (Brain) ; Ammon Horn ; Cornu Ammonis ; Hippocampus ; hippocampal ; Histones ; Homeostasis ; Autoregulation ; Physiological Homeostasis ; Human ; Modern Man ; Joints ; Laboratories ; Lead ; Pb element ; heavy metal Pb ; heavy metal lead ; Libraries ; Ligation ; Closure by Ligation ; Maps ; Memory ; Methods ; Mus ; Mice ; Mice Mammals ; Murine ; Organism ; living system ; Production ; Proteins ; Reagent ; Research ; Reverse Transcription ; Nuclear RNA ; Signal Transduction ; Cell Communication and Signaling ; Cell Signaling ; Intracellular Communication and Signaling ; Signal Transduction Systems ; Signaling ; biological signal transduction ; Technology ; Time ; Tissues ; Body Tissues ; transcription factor ; Basal Transcription Factor ; Basal transcription factor genes ; General Transcription Factor Gene ; General Transcription Factors ; Transcription Factor Proto-Oncogene ; Transcription factor genes ; Genetic Transcription ; Gene Transcription ; RNA Expression ; Transcription ; Work ; Transposase ; Generations ; base ; Label ; improved ; Procedures ; Clinical ; Phase ; Series ; Individual ; Malignant Cell ; cancer cell ; tool ; Brain Glioblastoma Multiforme ; Grade IV Brain Astrocytic Neoplasm ; Grade IV Brain Astrocytic Tumor ; Grade IV Brain Astrocytoma ; Brain Glioblastoma ; Nature ; DNA Methylation ; programs ; Complex ; Genetic Materials ; Protocol ; Protocols documentation ; Reaction ; cell type ; Techniques ; brain tissue ; Nuclear ; Best Practice Analysis ; Benchmarking ; brain cell ; Tumor Cell ; neoplastic cell ; Performance ; Biopsy Sample ; Biopsy Specimen ; novel ; Modality ; Pathogenesis ; Gene Expression Monitoring ; Gene Expression Pattern Analysis ; Transcript Expression Analyses ; Transcript Expression Analysis ; gene expression analysis ; gene expression assay ; transcriptional profiling ; Gene Expression Profiling ; Sampling ; histone modification ; Oncogenesis ; tumorigenesis ; Address ; Data ; Molecular Fingerprinting ; molecular profile ; molecular signature ; Molecular Profiling ; Resolution ; Collection ; Small Business Technology Transfer Research ; STTR ; Pathologic ; Development ; developmental ; Gene Expression Profile ; Expression Signature ; gene expression pattern ; gene expression signature ; transcriptional profile ; transcriptional signature ; epigenomics ; virtual ; next generation ; Population ; combinatorial ; tumor ; spatiotemporal ; phase 1 study ; Phase I Study ; ChIP-seq ; ChIP Sequencing ; chromatin immunoprecipitation-sequencing ; transcriptome sequencing ; RNA Seq ; RNA sequencing ; RNAseq ; epigenome ; multiple omics ; multiomics ; transcriptome ; global gene expression ; global transcription profile ; experimental study ; experiment ; experimental research ; preservation ;

Disruption of gene regulation is a major causal factor in heritable disease, developmental disorders,and oncogenesis. While chromatin immunoprecipitation followed by sequencing (ChIP-seq) and Tn5-transposase based tagging (Cut&Tag) enable analysis of transcription factor binding and epigenetic stateprofiling in bulk tissue samples and tumor biopsies, they produce only population average signals. Yetregulatory networks and perturbations are heterogeneous between cell classes and types. Single-celltechnologies can overcome the challenge of cellular heterogeneity and provide deeper insight into celltype-specific gene regulatory programs in healthy, diseased, and cancerous tissues. In prior work, wedeveloped a single-cell joint assay of histone modification and RNA expression (Paired-Tag), enablingcell-type-stratified epigenetic profiling from bulk samples. This technology has attracted customers inboth academic and nonprofit research. In the proposed study, we will develop automated protocols andeffect laboratory informatics systems to establish a Paired-Tag services laboratory, we will refine ourprotocol to improve experimental throughput and reduce cost, and we will develop a Paired-Tag "TF"protocol for profiling transcription factor binding profiles. If successful, the research would enable next-generation multi-omic analysis of tumor or disease samples at comparable cost to single-omictechnologies.

Public Health Relevance Statement:
We proposed to expand our research laboratory into a Paired-Tag services laboratory by developing automated protocols and integrating these with a laboratory information management system. We further proposed to reduce experimental costs by expanding multiplexing and exploring microfluidic single-cell platforms, to make our novel technology easier to use by publishing our processing and analysis pipelines, and to expand our technology's impact by adapting it to profile transcription factors.

Project Terms:
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