Project Number: 8042-31000-002-11-R
Project Type: Reimbursable Cooperative Agreement
Start Date: Jan 1, 2019
End Date: Dec 31, 2023
The central hypothesis that the sire has a strong and direct influence on establishment of pregnancy in cattle. The goal is this proposal is to test that hypothesis and better understand the biological and genetic mechanisms underpinning pregnancy establishment and loss using an innovative large animal model combined with a systems biology approach. To achieve this goal, Holstein sires will be selected that possess intrinsic and consistent differences in pregnancy outcomes with respect to early and late embryonic mortality. A series of experiments will utilize these phenotype sires to uncover the biological and genetic mechanisms governing embryonic mortality and develop genetic selection tools to improve pregnancy rates in cattle. Specific aims are to: (1) Identify sires with increased embryo mortality. This aim will identify sires with “inferior” ability to create embryos that establish pregnancy and thus have higher prevalence of early and late embryonic mortality. (2) Investigate biological mechanisms governing early embryonic mortality. This aim will test the hypotheses that inferior sires have defects in embryo development, conceptus elongation, implantation and/or placentation that are the basis for increased early embryonic mortality. (3) Determine genetic mechanisms underlying sire effects on embryonic survival and mortality. This aim will test the hypothesis that genetic loci and genomic rearrangements associated with embryonic survival and mortality can be identified using cutting-edge genomic analyses of DNA from fertile and subfertile sires. Completion of the research is expected to fill a substantial gap in existing knowledge by providing novel insights how the sire influences pregnancy success and loss in dairy cattle. The systems biology approach utilizing functional genomics and discovery-based methods to elucidate biological and genetic mechanisms involved in sire fertility and establishment of pregnancy is also innovative. Expected translational outcomes of this research include genetic markers that can be used as diagnostic tools to identify and select sires that create embryos superior for establishment of pregnancy, which will increase pregnancy rates, production efficiency, and profitability of ruminant agriculture enterprises. In biomedicine, translation of the identified biomarkers to humans is expected to provide novel indicators of male fertility that can be used to diagnose, treat and prevent infertility and subfertility and enhance pregnancy outcomes in natural and assisted reproduction.
Ten experiments will be used to address the three specific objectives outlined above. Two experiments will be used to identify sires with increased embryo mortality, or and “inferior” ability to create embryos that establish pregnancy and thus have higher rates of early and late embryonic mortality. In Experiment 1, Holstein sires with increased early embryonic mortality will be identified through a controlled fertility field trial to identify sires who intrinsically possess higher rates of early embryonic mortality. Candidate Holstein sires will be phenotyped using a controlled fertility field trial in Holstein heifers. Experiment 2 will be used to identify Holstein sires with increased late embryonic mortality. Objective 2 includes experiments designed to test the hypotheses that inferior sires have defects in embryo development, conceptus elongation, implantation, and/or placentation that are the basis for increased embryonic mortality. Experiment 3 will determine the effect of sire on in vitro embryo development using semen from subfertile bulls identified in Experiment 1. Experiment 4 will determine the effect of sire on conceptus elongation by harvesting 16-d embryos and studying conceptus morphology. In Experiment 5, 150 Holstein heifers will be synchronized and inseminated with semen from bulls of varying fertility and used to determine the effect of sire on pregnancy establishment. Finally, Experiment 6 will provide insights into early embryonic mortality by determining the effect of sire on conceptus growth and differentiation. The experiment will concentrate on subfertile sires from Experiment 1 that exhibit defects in embryo and/or placental development based on the outcome of Experiment 4. Objective 3 will test the hypothesis that genetic loci and genomic rearrangements associated with sire competency to create embryos that establish pregnancy can be identified using genomic analyses of DNA from the subfertile sires from Aim 1 and high-fertility sires. Experiment 7 will use a genome-wide association study (GWAS) to identify functional variants that will be characterized by predicted effect on protein function. Experiment 8 will build on Experiment 7 and identify copy number variants (CNV) using a whole-genome shotgun detection pipeline. Validation of CNVs will be performed using digital droplet PCR. The hypothesis that different genes are involved in phenotypic expression in bulls that vary in fertility will be tested in Experiment 9 using a population of 1,000 Holstein bulls. Validation bulls will be genotyped with a custom Illumina assay including variants identified in Experiments 7 and 8 and used for GWAS of SCR and cow fertility. Gene network and gene ontology and MeSH term enrichment will be used to identify differentially enriched pathways. Variants will tested to determine if they increase prediction accuracy. Genotyped bulls will be used as a reference population to impute genotypes to the Holstein bulls available in the National Dairy Database. Experiment 10 will identify harmful recessive haplotypes in each of the three groups of bulls used in Experiment 9 using a custom Illumina Infinium BeadChip assay.