Source: UNIVERSITY OF VERMONT submitted to NRP
ESTABLISHING A PAN-EPIGENOME FOR CATTLE AND SHEEP
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
ACTIVE
Funding Source
AFRI COMPETITIVE GRANT
Reporting Frequency
Annual
Accession No.
1027849
Grant No.
2022-67016-36216
Cumulative Award Amt.
$1,300,000.00
Proposal No.
2021-07077
Multistate No.
(N/A)
Project Start Date
Jan 1, 2022
Project End Date
Dec 31, 2025
Grant Year
2022
Program Code
[A1201]- Animal Health and Production and Animal Products: Animal Breeding, Genetics, and Genomics
Recipient Organization
UNIVERSITY OF VERMONT
(N/A)
BURLINGTON,VT 05405
Performing Department
Animal and Veterinary Sciences
Non Technical Summary
Epigenetic modifications include chemical changes to DNA and RNA base pairs that can influence gene function and subsequently affect economically important traits in livestock species. The collection of sequences representative of the DNA in a population or pangenome, is important to capture the breadth of genomic variation between breeds and populations of livestock species. However, to fully comprehend diversity in livestock species, one must also exhaustively characterize a variety of epigenetic modifications so that we can truly understand variation at a molecular level. The proposed project is designed to establish a cattle and sheep pan-epigenome. To accomplish this goal, we will comprehensively characterize epigenomic conservation and diversity through examination of the amount and location of methylation on DNA and RNA, histone modifications, open chromatin, chromatin accessibility in the liver, muscle, and prefrontal cortex of multiple breeds of cattle and sheep. Finally, comparative epigenomics will be used to identify orthologous regions of the sheep and bovine epigenomes that harbor conserved epigenetic modifications and variants. Cataloging epigenetic diversity across cattle and sheep will increase our understanding of how different genomes, epigenomes, and gene products from diverse breeds of cattle and sheep affect a variety of important biological phenotypes, supporting more accurate prediction of traits and improving breeding strategies.
Animal Health Component
0%
Research Effort Categories
Basic
50%
Applied
0%
Developmental
50%
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
3043399108050%
3043620108025%
3043699108025%
Knowledge Area
304 - Animal Genome;

Subject Of Investigation
3620 - Meat, sheep; 3699 - Sheep and wool, general/other; 3399 - Beef cattle, general/other;

Field Of Science
1080 - Genetics;
Goals / Objectives
The overarching goal of this project is to establish a cattle and sheep pan-epigenome. To accomplish this goal, we plan to accomplish the following specific aims: 1) Comprehensive characterization of epigenetic diversity in multiple breeds of cattle and sheep which will serve as the basis for the bovine and ovine pan-epigenome. 2.) Identify epigenetic variation with potential to impact phenotype. 3.) Employ comparative epigenomics to identify consensus epigenomes shared between sheep and cattle.
Project Methods
The following methods will be used to accomplish the previously stated specific aims:Tissue samples will be collected from four male calves from each of Angus, Holstein, and Brahman cattle, as well as Suffolk, Rambouillet and Katahdin sheep.Liver, muscle, and prefrontal cortex will be subjected to six types of next generation sequencing. Specifically, whole genome bisulfite sequencing (WGBS), oxidative reduced representation bisulfite sequencing (oxRRBS), chromatin immunoprecipitation sequencing (ChIP-Seq, methylated RNA immunoprecipitation sequencing (MeRIP-Seq), assay for transposase-accessible chromatin using sequencing (ATAC-Seq) and whole genome sequencing (WGS).Ensuing data analysis will permit characterization of 5-methylcytosine (5-mC), 5-hydroxymethylcytosine (5hmC), histone marks H3K4me3, H3K27ac, H3K4me1 and H3K27me3, chromatin accessibility and N6-methyladenosine (N6-A) in each of the previously mentioned tissues.Bismark and Bowtie2 will be used to align WGBS and oxRRBS reads to the most recent species appropriate reference genome.The level of 5mC for each cytosine in the genome will be called using BSSeeker2. Methpipe will used to identify differentially methylated regions and differentially methylated regions will be annotated using the program GenomicRanges.The level of 5hmC and differentially hydroxymethylated regions will be called using Methpipe. Annotation of differentially hydroxymethylated regions will be performed with GenomicRanges.ChIP-Seq analysis will be performed as described in Davenport et al 2021.ATAC-Seq data will be analyzed using a standardized software pipeline developed by ENCODE DCC fo the ENCODE Consortium to perform quality-control analysis and read alignmentA pipeline for analysis of MeRIP-Seq data will be established. We expect to utilized Bowtie for alignment of the most recent species reference genome and expression levels of mRNA will be measured with StringTie. Further, differential expression analysis will be performed using edgeR and the software exomePeak will be used to call and visualize peaks. Peaks will be annotated using CHIPseeker and motif search will be conducted with MEME and Homer.Genomic variants will be called using the GATK Germline Variant Prediction pipeline and CGmapTools will be employed to identify variants in methylation data.

Progress 01/01/24 to 12/31/24

Outputs
Target Audience:This project is designed to establish a cattle and sheep pan-epigenome and when combined with pangenome data will articulate a more comprehensive view of diversity in livestock species. Target audiences reached during this reporting period includes individuals and groups interested in comparative epigenomics and epitranscriptomics. This includes individuals from the scientific community and industry. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?Students and postdoctoral researchers working on components of the grant were able to present their data at national meetings. How have the results been disseminated to communities of interest?Dissemination has occurred via multiple invited presentations, posters and publications are in preparation. What do you plan to do during the next reporting period to accomplish the goals?A postdoctoral fellow responsible for data analysis, will be hired by Dr. McKay. Data from all samples will be completed.

Impacts
What was accomplished under these goals? A new methodology for simultaneously generating 5mC, 5hmC and CNV data was tested and found to be less than optimal. Cut and tag libraries for Angus cattle have been completed.

Publications

  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2024 Citation: McKay, SD. 2024. Demystifying Epitranscriptomics in Livestock. Breeding for Resiliency  2024 Resilient Dairy Genome Project. Guelph, ON, Canada.
  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2024 Citation: 2. McKay, SD. 2024. Examining the Potential Influence of Epigenetic Effects on Genetic Evaluations. Beef Improvement Federation, Knoxville, TN
  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2024 Citation: 3. McKay, SD. 2024. A Comparative Epigenomics Approach Towards Elucidating the Functional Impact of DNA and RNA Methylation on Gene Expression in Sheep. International Plant and Animal Genome 31, San Diego, California.
  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2024 Citation: 4. Xie, S., M Stegemiller, G. Becker, K. Davenport, K Shira, D. Konetchy, P Villamediana, S. McKay and B Murdoch. 2024. Effects of DNA md RNA methylation on gene expression in cattle and sheep. International Plant and Animal Genome 31, San Diego, California.
  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2024 Citation: 5. Khilji, S., S. Xie, G. Becker, K. Shira, M. Stegemiller, J. Smitchger, J Williams, L Christensen, R. Pace, D. Konetchy, D Hagen, S. McKay and B. Murdoch. 2024. Transcript Expression Profiles in Bovine Tissues: Insights from RNA-Seq in Angus Cattle. International Plant and Animal Genome 31, San Diego, California.
  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2024 Citation: 8. Xie, S., M Stegemiller, G. Becker, K. Davenport, K Shira, D. Konetchy, P Villamediana, B Murdoch and S. McKay. 2024. Constructing a cattle and sheep pan-epigenome to elucidate epigenetic impacts upon gene expression and phenotypic variation. Society for Molecular Biology and Evolution. Puerto Vallarta, Mexico.
  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2024 Citation: 6. Khilji, S., S. Xie, G. Becker, K. Shira, M. Stegemiller, J. Smitchger, J Williams, L Christensen, R. Pace, D. Konetchy, D Hagen, S. McKay and B. Murdoch. 2024. Deciphering Chromatin Accessibility and Gene Expression Interplay in Bovine Tissues. Advances in Genome Biology and Technology  Agricultural Meeting, Phoenix, AZ
  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2024 Citation: 7. Xie, S., M Stegemiller, G. Becker, K. Davenport, K Shira, D. Konetchy, P Villamediana, B Murdoch and S. McKay. 2024. Towards a cattle and sheep pan-epigenome: Elucidating the functional impact of DNA and RNA methylation on gene expression. Advances in Genome Biology and Technology  Agricultural Meeting, Phoenix, AZ


Progress 01/01/23 to 12/31/23

Outputs
Target Audience:This project is designed to establish a cattle and sheep pan-epigenome and when combined with pangenome data will articulate a more comprehensive view of diversity in livestock species. Target audiences reached during this reporting period includes individuals and groups interested in comparative epigenomics and epitranscriptomics. This includes individuals from the scientific community and industry. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?None. Additional students and postdoctoral fellows are being sought to work on the grant and we foresee professional development opportunities in the coming years. How have the results been disseminated to communities of interest?Dissemination has occurred via two invited presentations and one poster presentation. What do you plan to do during the next reporting period to accomplish the goals?Tissues for Brahman and Holstein cattle as well as Katahdin sheep will be collected in the first half of 2023. Tissues from Brahman and Holstein cattle as well as Katahdin sheep will be sent out for ATAC-Seq and RNA-Seq. Tissues from all breeds and species will be sent out for 5mC sequencing and ChIP-Seq. Additionally, a new method for 5hmC library construction will be tested and if adopted will permit whole genome analysis as opposed to the initially planned reduced representation approach. Analysis will being and/or continue for each sequencing type and methodologies for comparative epigenomics analysis will continue to be explored.

Impacts
What was accomplished under these goals? We have collected 20 tissues from 4 male juvenile Angus cattle, Suffolk sheep and Rambouillet sheep but only liver, muscle and prefrontal cortex tissues will be utilized for the purposes of this project. Additional tissues collected but not utilized for sequencing were immediately placed in cryotubes and frozen in liquid nitrogen storage for potential future research. Liver, muscle and prefrontal cortex were sent out for ATAC-Seq and RNA-Seq. Existing pipelines for ATAC-Seq and RNA-Seq analysis are being utilized.

Publications

  • Type: Conference Papers and Presentations Status: Published Year Published: 2022 Citation: McKay, SD, DE Hagen, RD Schnabel and BM Murdoch. 2021. The Future of Animal Genetics: Pan-Genomics, PanEpigenomics and High-Throughput Phenotyping. Visions III Star Gazing the Galaxy of Animal Genetics and Genomics. Ames, Iowa
  • Type: Conference Papers and Presentations Status: Published Year Published: 2022 Citation: McKay, S, D Hagen, R Schnabel, and B Murdoch. 2022. Cattle Genome to Herd Phenotyping for Precision Agriculture. AG2PI & NIFA Workshop: Visualizing the Future of AG2PI (Thinking Big). Ames, Iowa.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2023 Citation: 1. McKay, SD. 2023. Ascertaining the Functional Impact of DNA and RNA Methylation in Livestock. Iowa State University.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2023 Citation: 2. McKay, SD, BM Murdoch, and DE Hagen. 2023. Working towards a Pan-Epigenome in Cattle and Sheep. American Society of Animal Science. Albuquerque, New Mexico
  • Type: Conference Papers and Presentations Status: Published Year Published: 2023 Citation: 3. McKay, SD, S Xie and BM Murdoch. 2023. RNA Methylation as a Mechanistic Link between Epigenotype and Phenotype. 39th International Society of Animal Genetics Conference, Cape Town, South Africa
  • Type: Conference Papers and Presentations Status: Published Year Published: 2023 Citation: 4. McKay, SD and BM Murdoch. 2023. RNA Methylation as a Mechanistic Link between Genotype and Phenotype. International Plant and Animal Genome XXX, San Diego, California.


Progress 01/01/22 to 12/31/22

Outputs
Target Audience:This project is designed to establish a cattle and sheep pan-epigenome and when combined with pangenome data will articulate a more comprehensive view of diversity in livestock species. Target audiences reached during this reporting period includes individuals and groups interested in comparative epigenomics and epitranscriptomics. This includes individuals from the scientific community andindustry. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided? Nothing Reported How have the results been disseminated to communities of interest?Dissemination has occurred via two invited presentations and one poster presentation. What do you plan to do during the next reporting period to accomplish the goals?Tissues for Brahman and Holstein cattle as well as Katahdin sheep will be collected in the first half of 2023. Tissues from Brahman and Holstein cattle as well as Katahdin sheep will be sent out for ATAC-Seq and RNA-Seq. Tissues from all breeds and species will be sent out for 5mC sequencing and ChIP-Seq. Additionally, a new method for 5hmC library construction will be tested and if adopted will permit whole genome analysis as opposed to the initially planned reduced representation approach. Analysis will being and/or continue for each sequencing type and methodologies for comparative epigenomics analysis will continue to be explored.

Impacts
What was accomplished under these goals? Tissues were collected for Angus cattle and Suffolk and Merino sheep. ATAC-Seq and RNA-Seq data has been obtained for Angus tissue samples. Analysis pipelines have been established and comparative epigenomics methodology has been investigated.

Publications

  • Type: Conference Papers and Presentations Status: Published Year Published: 2022 Citation: McKay, S, D Hagen, R Schnabel, and B Murdoch. 2022. Cattle Genome to Herd Phenotyping for Precision Agriculture. AG2PI & NIFA Workshop: Visualizing the Future of AG2PI (Thinking Big). Ames, Iowa.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2022 Citation: Poster: McKay, SD, DE Hagen, RD Schnabel and BM Murdoch. 2021. The Future of Animal Genetics: Pan-Genomics, Pan-Epigenomics and High-Throughput Phenotyping. Visions III Star Gazing the Galaxy of Animal Genetics and Genomics. Ames, Iowa
  • Type: Conference Papers and Presentations Status: Published Year Published: 2022 Citation: McKay, SD, DE Hagen, RD Schnabel and BM Murdoch. 2021. The Future of Animal Genetics: Pan-Genomics, Pan-Epigenomics and High-Throughput Phenotyping. Visions III Star Gazing the Galaxy of Animal Genetics and Genomics. Ames, Iowa