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Testis specific histone 2B is associated with sperm chromatin dynamics and bull fertility-a pilot study.

ABSTRACT: Bull fertility is the degree of sperm's ability to fertilize and activate the egg and support embryo development, and this is critical for herd reproductive performance. We used the bull as a unique model organism for the study of male fertility because cattle genetics and physiology is similar to those of other mammals including humans. Moreover, reliable fertility data along with well-established in vitro systems are available for bovine. The objective of this original study was to ascertain evolutionary diversification and expression dynamics of Testis Specific Histone 2B (TH2B) in sperm from Holstein bulls with different fertility scores.The intensity of TH2B was determined by using flow cytometry in sperm from 13 high and 13 low fertility bulls. Expression levels of TH2B were measured using immunofluorescence and Western blotting in sperm from five high and five low fertility bulls. Sequence identity, evolutionary distance and interactome of TH2B were evaluated by dotmatcher, STRING and Cytoscape. Data were analyzed using linear mixed effects model and regression plots were drawn.The intensity of TH2B as measured by flow cytometry was significantly affected by an interaction between fertility group and fertility score (P = 0.0182). The intensity of TH2B in sperm from the high fertility group decreased (P = 0.0055) as fertility increased. TH2B was constantly detectable in sperm and expression levels of TH2B decreased in relation to fertility in sperm from the high fertility group (P = 0.018). TH2B biological functions include male gamete generation, chromosome organization, DNA packaging, DNA conformation change, chromatin organization, nucleosome organization, chromatin disassembly, spermatid nucleus elongation, spermatid nucleus differentiation, sperm motility, chromatin organization, chromatin condensation, chromatin silencing, nucleus organization, and chromatin remodeling (P < 0.05).We elucidated the cellular localization and molecular physiology of TH2B using both computational and cell biology approaches. In addition to advancing the fundamental science of mammalian male gamete, the present findings can be potentially used to evaluate semen quality and predict male fertility in the future.This study did not involve any live animals. We did not perform any anesthesia, euthanasia, or any kind of animal sacrifice. The cryopreserved semen samples were obtained from Alta Genetics, Inc., Watertown, WI, USA. All samples were preserved in liquid nitrogen.

SUBMITTER: Kutchy NA

PROVIDER: S-EPMC5539985 | biostudies-literature | 2017 Aug

REPOSITORIES: biostudies-literature

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Testis specific histone 2B is associated with sperm chromatin dynamics and bull fertility-a pilot study.

Kutchy Naseer A NA Velho Ana A Menezes Erika S B ESB Jacobsen Marie M Thibaudeau Giselle G Wills Robert W RW Moura Arlindo A Kaya Abdullah A Perkins Andy A Memili Erdogan E

Reproductive biology and endocrinology : RB&E 20170801 1

<h4>Background</h4>Bull fertility is the degree of sperm's ability to fertilize and activate the egg and support embryo development, and this is critical for herd reproductive performance. We used the bull as a unique model organism for the study of male fertility because cattle genetics and physiology is similar to those of other mammals including humans. Moreover, reliable fertility data along with well-established in vitro systems are available for bovine. The objective of this original study ...[more]

PMID: 28764714

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Project description:Bull fertility, ability of the sperm to fertilize and activate the egg and support embryo development, is vital for cattle reproduction and production. Even though majority of histones are replaced by protamines, some histones are retained in sperm. It is known that chromatin remodeling during spermatogenesis results in dynamic changes in sperm chromatin structure through post-translational modifications (PTM) of sperm histones, which are important for regulation of gene expression. However, amounts of sperm Histone 4 (H4), its acetylated form (H4 acetyl), and to what extent these molecular attributes influence sperm chromatin structure and bull fertility are unknown. These gaps in the knowledge base are important because they are preventing advances in the fundamental science of bovine male gamete and improvement of bull fertility. The objective of this study was to test the hypothesis that expression dynamics as well as PTM of sperm H4 are associated with bull fertility. Flow cytometry was utilized to quantify H4 and H4 acetylated form in sperm from seven high and seven low fertility Holstein bulls. The results indicated that the average number of cells with H4 or H4 acetyl expression in high and low fertility bull sperm were 34.6 ± 20.4, 1.88 ± 1.8, 15.2 ± 20.8, and 1.4 ± 1.2, respectively. However, the sperm enriched in both H4 and H4 acetyl were different between high and low fertility groups (3.5 ± 0.6; 1.8 ± 0.8; P = 0.043). The localization and detection of H4 and H4 acetylation were measured by immunocytochemistry which revealed that H4 and H4 acetylation were equally distributed in the sperm head of high and low fertility sires. Western blotting results confirmed the presence of the H4 and its acetylated form in the sperm. Bioinformatics studies demonstrated that H4 is highly conserved among mammalians, and have significant gene ontology on spermatogenesis, early embryo implantation, and sperm capacitation. The results are significant because it demonstrates the replacement of canonical histone H4 into modified H4 acetylation in sperm and regulate its dynamics which is crucial for bull fertility and reproductive biotechnology. These findings advance fundamental science of mammalian early development and reproductive biotechnology.

Project description:BackgroundLinker histone H1 has been studied in vivo and using reconstituted chromatin, but there have been few systematic studies of the effects of the cellular environment on its function. Due to the presence of many other chromatin factors and specific chaperones such as RanBP7/importin beta that regulate histone H1, linker histones likely function differently in vivo than in purified systems.Methodology/principal findingsWe have directly compared H1 binding to sperm nuclei in buffer versus Xenopus egg extract cytoplasm, and monitored the effects of adding nuclear import chaperones. In buffer, RanBP7 decondenses sperm nuclei, while H1 binds tightly to the chromatin and rescues RanBP7-mediated decondensation. H1 binding is reduced in cytoplasm, and H1 exhibits rapid FRAP dynamics in cytoplasm but not in buffer. RanBP7 decreases H1 binding to chromatin in both buffer and extract but does not significantly affect H1 dynamics in either condition. Importin beta has a lesser effect than RanBP7 on sperm chromatin decondensation and H1 binding, while a combination of RanBP7/importin beta is no more effective than RanBP7 alone. In extracts supplemented with RanBP7, H1 localizes to chromosomal foci, which increase after DNA damage. Unlike somatic H1, the embryonic linker histone H1M binds equally well to chromatin in cytoplasm compared to buffer. Amino-globular and carboxyl terminal domains of H1M bind chromatin comparably to the full-length protein in buffer, but are inhibited ?10-fold in cytoplasm. High levels of H1 or its truncations distort mitotic chromosomes and block their segregation during anaphase.Conclusion/significanceRanBP7 can decondense sperm nuclei and decrease H1 binding, but the rapid dynamics of H1 on chromatin depend on other cytoplasmic factors. Cytoplasm greatly impairs the activity of individual H1 domains, and only the full-length protein can condense chromatin properly. Our findings begin to bridge the gap between purified and in vivo chromatin systems.

Project description:BackgroundBiological relevance of the major testis specific histone H2B variant (TH2B) in sperm is not fully understood. Studies in TH2A/TH2B double knockout male mice indicate its role in chromatin compaction and male fertility. Additionally, the presence of TH2B and TH2A reportedly generates more dynamic nucleosomes, leading to an open chromatin structure characteristic of transcriptionally active genome. Given that mature human sperm are transcriptionally and translationally inactive, the presence of TH2B in mature sperm is intriguing. To address its role in sperm, we investigated the genome-wide localization of TH2B in sperm of fertile men.ResultsWe have identified the genomic loci associated with TH2B in fertile human sperm by ChIP-seq analysis. Bioinformatic analysis revealed ~ 5% sperm genome and 5527 genes to be associated with TH2B. Out of these 105 (1.9%) and 144 (2.6%) genes showed direct involvement in sperm function and early embryogenesis, respectively. Chromosome wide analysis for TH2B distribution indicated its least distribution on X and Y chromosomes and varied distribution on autosomes. TH2B showed relatively higher percentage of gene association on chromosome 4, 18, 3 and 2. TH2B enrichment was more in promoter and gene body region. Gene Ontology (GO) analysis revealed signal transduction and associated kinase activity as the most enriched biological and molecular function, respectively. We also observed the enrichment of TH2B at developmentally important loci, such as HOXA and HOXD and on genes required for normal sperm function, few of which were validated by ChIP-qPCR. The relative expression of these genes was altered in particular subgroup of infertile men showing abnormal chromatin packaging. Chromatin compaction positively correlated with sperm- motility, concentration, viability and with transcript levels of PRKAG2 and CATSPER B.ConclusionChIP-seq analysis of TH2B revealed a putative role of TH2B in sperm function and embryo development. Altered expression of TH2B associated genes in infertile individuals with sperm chromatin compaction defects indicates involvement of TH2B in transcriptional regulation of these genes in post meiotic male germ cells. This altered transcriptome may be a consequence or cause of abnormal nuclear remodeling during spermiogenesis.

Project description:Fertility is one of the most economically important traits in both beef and dairy cattle production; however, only female fertility is typically subjected to selection. Male and female fertility have only a small positive genetic correlation which is likely due to the existence of a relatively small number of genetic variants within each breed that cause embryonic and developmental losses. Genomic tools have been developed that allow the identification of lethal recessive loci based upon marker haplotypes. Selection against haplotypes harbouring lethal alleles in conjunction with selection to improve female fertility will result in an improvement in male fertility. Genomic selection has resulted in a two to fourfold increase in the rate of genetic improvement of most dairy traits in US Holstein cattle, including female fertility. Considering the rapidly increasing rate of adoption of high-throughput single nucleotide polymorphism genotyping in both the US dairy and beef industries, genomic selection should be the most effective of all currently available approaches to improve male fertility. However, male fertility phenotypes are not routinely recorded in natural service mating systems and when artificial insemination is used, semen doses may be titrated to lower post-thaw progressively motile sperm numbers for high-merit and high-demand bulls. Standardization of sperm dosages across bull studs for semen distributed from young bulls would allow the capture of sire conception rate phenotypes for young bulls that could be used to generate predictions of genetic merit for male fertility in both males and females. These data would allow genomic selection to be implemented for male fertility in addition to female fertility within the US dairy industry. While the rate of use of artificial insemination is much lower within the US beef industry, the adoption of sexed semen in the dairy industry has allowed dairy herds to select cows from which heifer replacements are produced and cows that are used to produce terminal crossbred bull calves sired by beef breed bulls. Capture of sire conception rate phenotypes in dairy herds utilizing sexed semen will contribute data enabling genomic selection for male fertility in beef cattle breeds. As the commercial sector of the beef industry increasingly adopts fixed-time artificial insemination, sire conception rate phenotypes can be captured to facilitate the development of estimates of genetic merit for male fertility within US beef breeds.

Project description:BackgroundDuring epididymal transit, spermatozoa go through several functional maturation steps, resulting from interactions with epididymal secretomes specific to each region. In particular, the sperm membrane is under constant remodeling, with sequential attachment and shedding of various molecules provided by the epididymal lumen fluid and epididymosomes, which also deliver sncRNA cargo to sperm. As a result, the payload of sperm sncRNAs changes during the transit from the epididymis caput to the cauda. This work was designed to study the dynamics of cattle sperm sncRNAs from spermatogenesis to final maturation.ResultsComprehensive catalogues of sperm sncRNAs were obtained from testicular parenchyma, epididymal caput, corpus and cauda, as well as ejaculated semen from three Holstein bulls. The primary cattle sncRNA sperm content is markedly remodeled as sperm mature along the epididymis. Expression of piRNAs, which are abundant in testis parenchyma, decreases dramatically at epididymis. Conversely, sperm progressively acquires miRNAs, rsRNAs, and tsRNAs along epididymis, with regional specificities. For instance, miRNAs and tsRNAs are enriched in epididymis cauda and ejaculated sperm, while rsRNA expression peaks at epididymis corpus. In addition, epididymis corpus contains mainly 20 nt long piRNAs, instead of 30 nt in all other locations. Beyond the bulk differences in abundance of sncRNAs classes, K-means clustering was performed to study their spatiotemporal expression profile, highlighting differences in specific sncRNAs and providing insights into their putative biological role at each maturation stage. For instance, Gene Ontology analyses using miRNA targets highlighted enriched processes such as cell cycle regulation, response to stress and ubiquitination processes in testicular parenchyma, protein metabolism in epididymal sperm, and embryonic morphogenesis in ejaculated sperm.ConclusionsOur findings confirm that the sperm sncRNAome does not simply reflect a legacy of spermatogenesis. Instead, sperm sncRNA expression shows a remarkable level of plasticity resulting probably from the combination of multiple factors such as loss of the cytoplasmic droplet, interaction with epididymosomes, and more surprisingly, the putative in situ production and/or modification of sncRNAs by sperm. Given the suggested role of sncRNA in epigenetic trans-generational inheritance, our detailed spatiotemporal analysis may pave the way for a study of sperm sncRNAs role in embryo development.

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Testis specific histone 2B is associated with sperm chromatin dynamics and bull fertility-a pilot study.

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Testis specific histone 2B is associated with sperm chromatin dynamics and bull fertility-a pilot study.

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