Dissecting Tissue-Specific Super-Enhancers by Integrating Genome-Wide Analyses and CRISPR/Cas9 Genome Editing

AbstractGenome-wide association studies (GWAS) have uncovered thousands of genetic variants that influence risk for human diseases and traits. Yet understanding the mechanisms by which these genetic variants, mainly noncoding, have an impact on associated diseases and traits remains a significant hurdle. In this review, we discuss emerging experimental approaches that are being applied for functional studies of causal variants and translational advances from GWAS findings to disease prevention and treatment. We highlight the use of genome editing technologies in GWAS functional studies to modify genomic sequences, with proof-of-principle examples. We discuss the challenges in interrogating causal variants, points for consideration in experimental design and interpretation of GWAS locus mechanisms, and the potential for novel therapeutic opportunities. With the accumulation of knowledge of functional genetics, therapeutic genome editing based on GWAS discoveries will become increasingly feasible.

Download Full-text

E-MAGMA: an eQTL-informed method to identify risk genes using genome-wide association study summary statistics

Bioinformatics ◽

10.1093/bioinformatics/btab115 ◽

2021 ◽

Author(s):

Zachary F Gerring ◽

Angela Mina-Vargas ◽

Eric R Gamazon ◽

Eske M Derks

Keyword(s):

Genome Wide Association ◽

Chromosome 1 ◽

Supplementary Information ◽

Genome Wide Association Studies ◽

Summary Statistics ◽

Tissue Specific ◽

Complex Disorders ◽

Risk Variants ◽

Genome Wide ◽

Causal Genes

Abstract Motivation Genome-wide association studies have successfully identified multiple independent genetic loci that harbour variants associated with human traits and diseases, but the exact causal genes are largely unknown. Common genetic risk variants are enriched in non-protein-coding regions of the genome and often affect gene expression (expression quantitative trait loci, eQTL) in a tissue-specific manner. To address this challenge, we developed a methodological framework, E-MAGMA, which converts genome-wide association summary statistics into gene-level statistics by assigning risk variants to their putative genes based on tissue-specific eQTL information. Results We compared E-MAGMA to three eQTL informed gene-based approaches using simulated phenotype data. Phenotypes were simulated based on eQTL reference data using GCTA for all genes with at least one eQTL at chromosome 1. We performed 10 simulations per gene. The eQTL-h2 (i.e., the proportion of variation explained by the eQTLs) was set at 1%, 2%, and 5%. We found E-MAGMA outperforms other gene-based approaches across a range of simulated parameters (e.g. the number of identified causal genes). When applied to genome-wide association summary statistics for five neuropsychiatric disorders, E-MAGMA identified more putative candidate causal genes compared to other eQTL-based approaches. By integrating tissue-specific eQTL information, these results show E-MAGMA will help to identify novel candidate causal genes from genome-wide association summary statistics and thereby improve the understanding of the biological basis of complex disorders. Availability A tutorial and input files are made available in a github repository: https://github.com/eskederks/eMAGMA-tutorial. Supplementary information Supplementary data are available at Bioinformatics online.

Download Full-text

Nuclear receptor HNF4A transrepresses CLOCK:BMAL1 and modulates tissue-specific circadian networks

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1816411115 ◽

2018 ◽

Vol 115 (52) ◽

pp. E12305-E12312 ◽

Cited By ~ 16

Author(s):

Meng Qu ◽

Tomas Duffy ◽

Tsuyoshi Hirota ◽

Steve A. Kay

Keyword(s):

Nuclear Receptor ◽

Metabolic Pathways ◽

Feedback Loop ◽

Transcriptional Activity ◽

Transcript Level ◽

Colon Cells ◽

Tissue Specific ◽

Metabolic Genes ◽

Genome Wide ◽

Circadian Control

Either expression level or transcriptional activity of various nuclear receptors (NRs) have been demonstrated to be under circadian control. With a few exceptions, little is known about the roles of NRs as direct regulators of the circadian circuitry. Here we show that the nuclear receptor HNF4A strongly transrepresses the transcriptional activity of the CLOCK:BMAL1 heterodimer. We define a central role for HNF4A in maintaining cell-autonomous circadian oscillations in a tissue-specific manner in liver and colon cells. Not only transcript level but also genome-wide chromosome binding of HNF4A is rhythmically regulated in the mouse liver. ChIP-seq analyses revealed cooccupancy of HNF4A and CLOCK:BMAL1 at a wide array of metabolic genes involved in lipid, glucose, and amino acid homeostasis. Taken together, we establish that HNF4A defines a feedback loop in tissue-specific mammalian oscillators and demonstrate its recruitment in the circadian regulation of metabolic pathways.

Download Full-text

Short term but highly efficient Cas9 expression mediated by excisional system using adenovirus vector and Cre

Scientific Reports ◽

10.1038/s41598-021-03803-w ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Sayaka Nagamoto ◽

Miyuki Agawa ◽

Emi Tsuchitani ◽

Kazunori Akimoto ◽

Saki Kondo Matsushima ◽

...

Keyword(s):

Hepatitis B ◽

Genome Editing ◽

Adenovirus Vector ◽

Virus Genome ◽

Dna Viruses ◽

Tissue Specific ◽

B Virus ◽

Tissue Specific Promoter ◽

A Cell

AbstractGenome editing techniques such as CRISPR/Cas9 have both become common gene engineering technologies and have been applied to gene therapy. However, the problems of increasing the efficiency of genome editing and reducing off-target effects that induce double-stranded breaks at unexpected sites in the genome remain. In this study, we developed a novel Cas9 transduction system, Exci-Cas9, using an adenovirus vector (AdV). Cas9 was expressed on a circular molecule excised by the site-specific recombinase Cre and succeeded in shortening the expression period compared to AdV, which expresses the gene of interest for at least 6 months. As an example, we chose hepatitis B, which currently has more than 200 million carriers in the world and frequently progresses to liver cirrhosis or hepatocellular carcinoma. The efficiencies of hepatitis B virus genome disruption by Exci-Cas9 and Cas9 expression by AdV directly (Avec) were the same, about 80–90%. Furthermore, Exci-Cas9 enabled cell- or tissue-specific genome editing by expressing Cre from a cell- or tissue-specific promoter. We believe that Exci-Cas9 developed in this study is useful not only for resolving the persistent expression of Cas9, which has been a problem in genome editing, but also for eliminating long-term DNA viruses such as human papilloma virus.

Download Full-text

Efficient CRISPR/Cas9 genome editing in a salmonid fish cell line using a lentivirus delivery system

10.1101/734442 ◽

2019 ◽

Cited By ~ 2

Author(s):

Remi L. Gratacap ◽

Tim Regan ◽

Carola E. Dehler ◽

Samuel A.M. Martin ◽

Pierre Boudinot ◽

...

Keyword(s):

Disease Resistance ◽

Cell Line ◽

Genome Editing ◽

Target Genes ◽

High Efficiency ◽

Genetic Resistance ◽

Model Organisms ◽

Fish Cell ◽

Genome Wide ◽

Lentivirus Transduction

1AbstractGenome editing is transforming bioscience research, but its application to non-model organisms, such as farmed animal species, requires optimisation. Salmonids are the most important aquaculture species by value, and improving genetic resistance to infectious disease is a major goal. However, use of genome editing to evaluate putative disease resistance genes in cell lines, and the use of genome-wide CRISPR screens is currently limited by a lack of available tools and techniques. In the current study, an optimised protocol using lentivirus transduction for efficient integration of constructs into the genome of a Chinook salmon (Oncorhynchus tshwaytcha) cell line (CHSE-214) was developed. As proof-of-principle, two target genes were edited with high efficiency in an EGFP-Cas9 stable CHSE cell line; specifically, the exogenous, integrated EGFP and the endogenous RIG-I locus. Finally, the effective use of antibiotic selection to enrich the successfully edited targeted population was demonstrated. The optimised lentiviral-mediated CRISPR method reported here increases possibilities for efficient genome editing in salmonid cells, in particular for future applications of genome-wide CRISPR screens for disease resistance.

Download Full-text

A transcriptome-wide Mendelian randomization study to uncover tissue-dependent regulatory mechanisms across the human phenome

10.1101/563379 ◽

2019 ◽

Cited By ~ 2

Author(s):

Tom G Richardson ◽

Gibran Hemani ◽

Tom R Gaunt ◽

Caroline L Relton ◽

George Davey Smith

Keyword(s):

Gene Expression ◽

Genetic Variants ◽

Complex Traits ◽

Mendelian Randomization ◽

Drug Repositioning ◽

Association Studies ◽

Thyroid Tissue ◽

Genome Wide Association Studies ◽

Tissue Specific ◽

Genome Wide

AbstractBackgroundDeveloping insight into tissue-specific transcriptional mechanisms can help improve our understanding of how genetic variants exert their effects on complex traits and disease. By applying the principles of Mendelian randomization, we have undertaken a systematic analysis to evaluate transcriptome-wide associations between gene expression across 48 different tissue types and 395 complex traits.ResultsOverall, we identified 100,025 gene-trait associations based on conventional genome-wide corrections (P < 5 × 10−08) that also provided evidence of genetic colocalization. These results indicated that genetic variants which influence gene expression levels in multiple tissues are more likely to influence multiple complex traits. We identified many examples of tissue-specific effects, such as genetically-predicted TPO, NR3C2 and SPATA13 expression only associating with thyroid disease in thyroid tissue. Additionally, FBN2 expression was associated with both cardiovascular and lung function traits, but only when analysed in heart and lung tissue respectively.We also demonstrate that conducting phenome-wide evaluations of our results can help flag adverse on-target side effects for therapeutic intervention, as well as propose drug repositioning opportunities. Moreover, we find that exploring the tissue-dependency of associations identified by genome-wide association studies (GWAS) can help elucidate the causal genes and tissues responsible for effects, as well as uncover putative novel associations.ConclusionsThe atlas of tissue-dependent associations we have constructed should prove extremely valuable to future studies investigating the genetic determinants of complex disease. The follow-up analyses we have performed in this study are merely a guide for future research. Conducting similar evaluations can be undertaken systematically at http://mrcieu.mrsoftware.org/Tissue_MR_atlas/.

Download Full-text

Genome-wide identification of tissue-specific long non-coding RNA in three farm animal species

BMC Genomics ◽

10.1186/s12864-018-5037-7 ◽

2018 ◽

Vol 19 (1) ◽

Cited By ~ 36

Author(s):

Colin Kern ◽

Ying Wang ◽

James Chitwood ◽

Ian Korf ◽

Mary Delany ◽

...

Keyword(s):

Animal Species ◽

Farm Animal ◽

Tissue Specific ◽

Non Coding Rna ◽

Genome Wide ◽

Long Non Coding Rna

Download Full-text

Genome editing's potential target diseases in the cardiovascular field

10.31219/osf.io/gc23p ◽

2021 ◽

Author(s):

Moataz Dowaidar

Keyword(s):

Cardiovascular Diseases ◽

Genome Editing ◽

Target Cells ◽

Clinical Use ◽

Editing Efficiency ◽

Potential Target ◽

Genome Wide ◽

Significant Barrier

Two types of cardiovascular diseases can be cured or prevented using genome editing. The liver is the organ that has received the most attention in terms of clinical genome editing for cardiovascular diseases. Off-target mutagenesis is a concern of any form of genome editing. Off-target mutations in target cells or tissues may lead to undesirable functional phenotypes, including cancer. For therapeutic editing of the heart, the authors claim it's critical to achieve high editing efficiency at a chosen genomic site in a desired tissue. Off-target editing can be tested genome-wide unbiasedly using newer cell-based methods. For low off-target impact, well-designed gRNAs are important. The delivery of genome editors to target tissues and cells is a significant barrier to clinical use.

Download Full-text