Mississippi State University
Gordon, Donna M.
Lemley, Caleb O.
Perkins, Andy D.
Date of Degree
Original embargo terms
Dissertation - Open Access
Doctor of Philosophy
College of Agriculture and Life Sciences
Department of Animal and Dairy Sciences
Male fertility, ability to fertilize and activate the egg and support early embryo development, is crucial for mammalian reproduction and development. Testis specific histone 2B (TH2B) of sperm, protamines (PRM1/2), and posttranslational modifications of histone 3 (H3K27me3 and H3k27ac) are involved in spermatogenesis and male fertility. However, molecular and cellular mechanisms by which TH2B regulates histone to protamine replacement is poorly defined. Immunocytochemistry, western blotting, flow cytometry, computer-assisted sperm analysis (CASA) and bioinformatic approaches were applied to analyze sperm from Holstein bulls with different in vivo fertility. Results from the immunocytochemistry experiments showed that while TH2B and H3K27me3 were localized predominantly at the equatorial and post acrosomal (localized as a crown around the sperm head) parts, respectively. The H3K27ac was also detectable in the bovine sperm head. Signal intensities of TH2B (mean ± SEM) were higher in sperm from the low fertility bulls (220.56 ± 9.20) as compared to those from the high fertility bulls (198.39 ± 10.0). Signal intensities of H3K27me3 (16.25 ± 1.69) were significantly different than those of H3K27ac (4.74 ± 0.88) in bull spermatozoa. Using the bioinformatic tools, including Clustal Omega, Cytoscape, Emboss Dotmatcher, InterProScan, and STRING, we demonstrated that TH2B has the conserved histone H2B domain which has a strong association with proteins involved in chromosome organization and histone ubiquitination. Intensities of PRM1 and PRM2 were significantly associated with one another (p < 0.0001), but neither were significantly associated with fertility. Results from CASA revealed significant differences between high and low fertility bulls regarding average sperm pathway velocity, amplitude of lateral head displacement and straightness (p < 0.05). The interacting proteins of H3 are involved in subcellular processes such as regulation of H3K27 methylation, nucleosome assembly, regulation of DNA replication, and chromatin assembly. These results are significant because they help advance fundamental knowledge in sperm physiology involving epigenetic and genetic determinants. The new knowledge can be used to enhance reproductive biotechnology to improve fertility. In addition, the data generated using the unique bull model can be applied to study mammalian reproduction and development due to similarities in genetics and physiology between bovine and other mammals.
Kutchy, Naseer Ahmad, "Sperm Genetic and Epigenetic Mechanisms Regulating Male Fertility" (2017). Theses and Dissertations. 4030.