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RNA sequencing reveals candidate genes and polymorphisms related to sperm DNA integrity in testis tissue from boars.

ABSTRACT: Sperm DNA is protected against fragmentation by a high degree of chromatin packaging. It has been demonstrated that proper chromatin packaging is important for boar fertility outcome. However, little is known about the molecular mechanisms underlying differences in sperm DNA fragmentation. Knowledge of sequence variation influencing this sperm parameter could be beneficial in selecting the best artificial insemination (AI) boars for commercial production. The aim of this study was to identify genes differentially expressed in testis tissue of Norwegian Landrace and Duroc boars, with high and low sperm DNA fragmentation index (DFI), using transcriptome sequencing.Altogether, 308 and 374 genes were found to display significant differences in expression level between high and low DFI in Landrace and Duroc boars, respectively. Of these genes, 71 were differentially expressed in both breeds. Gene ontology analysis revealed that significant terms in common for the two breeds included extracellular matrix, extracellular region and calcium ion binding. Moreover, different metabolic processes were enriched in Landrace and Duroc, whereas immune response terms were common in Landrace only. Variant detection identified putative polymorphisms in some of the differentially expressed genes. Validation showed that predicted high impact variants in RAMP2, GIMAP6 and three uncharacterized genes are particularly interesting for sperm DNA fragmentation in boars.We identified differentially expressed genes between groups of boars with high and low sperm DFI, and functional annotation of these genes point towards important biochemical pathways. Moreover, variant detection identified putative polymorphisms in the differentially expressed genes. Our results provide valuable insights into the molecular network underlying DFI in pigs.

SUBMITTER: van Son M

PROVIDER: S-EPMC5706377 | biostudies-literature | 2017 Nov

REPOSITORIES: biostudies-literature

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RNA sequencing reveals candidate genes and polymorphisms related to sperm DNA integrity in testis tissue from boars.

van Son Maren M Tremoen Nina Hårdnes NH Gaustad Ann Helen AH Myromslien Frøydis Deinboll FD Våge Dag Inge DI Stenseth Else-Berit EB Zeremichael Teklu Tewoldebrhan TT Grindflek Eli E

BMC veterinary research 20171128 1

<h4>Background</h4>Sperm DNA is protected against fragmentation by a high degree of chromatin packaging. It has been demonstrated that proper chromatin packaging is important for boar fertility outcome. However, little is known about the molecular mechanisms underlying differences in sperm DNA fragmentation. Knowledge of sequence variation influencing this sperm parameter could be beneficial in selecting the best artificial insemination (AI) boars for commercial production. The aim of this study ...[more]

PMID: 29183316

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Project description:BackgroundSperm hyperactive motility has previously been shown to influence litter size in pigs, but little is known about the underlying biological mechanisms. The aim of this study was to use RNA sequencing to investigate gene expression differences in testis tissue from Landrace and Duroc boars with high and low levels of sperm hyperactive motility. Boars with divergent phenotypes were selected based on their sperm hyperactivity values at the day of ejaculation (day 0) (contrasts (i) and (ii) for Landrace and Duroc, respectively) and on their change in hyperactivity between day 0 and after 96 h liquid storage at 18 °C (contrast (iii)).ResultsRNA sequencing was used to measure gene expression in testis. In Landrace boars, 3219 genes were differentially expressed for contrast (i), whereas 102 genes were differentially expressed for contrast (iii). Forty-one differentially expressed genes were identified in both contrasts, suggesting a functional role of these genes in hyperactivity regardless of storage. Zinc finger DNLZ was the most up-regulated gene in contrasts (i) and (iii), whereas the most significant differentially expressed gene for the two contrasts were ADP ribosylation factor ARFGAP1 and solute carrier SLC40A1, respectively. For Duroc (contrast (ii)), the clustering of boars based on their gene expression data did not reflect their difference in sperm hyperactivity phenotypes. No results were therefore obtained for this breed. A case-control analysis of variants identified in the Landrace RNA sequencing data showed that SNPs in NEU3, CHRDL2 and HMCN1 might be important for sperm hyperactivity.ConclusionsDifferentially expressed genes were identified in Landrace boars with high and low levels of sperm hyperactivity at the day of ejaculate collection and high and low change in hyperactivity after 96 h of sperm storage. The results point towards important candidate genes, biochemical pathways and sequence variants underlying sperm hyperactivity in pigs.

Project description:Chromatin packaging in sperm protects it against DNA fragmentation, and the importance of proper chromatin packaging for boar fertility outcome has become increasingly evident. Little is known however about the molecular mechanisms underlying differences in sperm DNA fragmentation and an understanding of the genes controlling this sperm parameter could help in selecting the best boars for AI use. The aim of this study was to identify differentially expressed genes in testis of Norsvin Landrace and Duroc boars with good and bad sperm DNA fragmentation using transcriptome sequencing and to use the data for polymorphism search. RNA sequence reads were obtained using Illumina technology and mapped by TopHat using the Ensembl pig database. Differentially expressed genes and pathways were analyzed using the R Bioconductor packages edgeR and goseq respectively. Using a false discovery rate of 0.05, 309 and 375 genes were found displaying significant differences in expression level between the good and bad condition in Landrace and Duroc respectively. Of the differentially expressed genes, 72 were found in common for the two breeds. Gene ontology analysis revealed that terms common for the two breeds included extracellular matrix, extracellular region and calcium ion binding. Additionally, different metabolic processes were enriched in Landrace and Duroc, whereas immune response ontologies were found to be important in Landrace. SNP detection in Landrace/Duroc identified 53182/53931 variants in 10924/10748 transcripts and of these, 1573/1827 SNPs occurred in 189/241 unique genes that were also differentially expressed. Possible high impact variants were detected using SnpEff. Transcriptome sequencing identified differentially expressed genes and nucleotide variants related to differences in sperm DNA fragmentation, and functional annotation of the genes pointed towards important biochemical pathways. This study provides insights into the genetic network underlying this trait and is a first step towards using sperm DNA fragmentation for predicting boar fertility.

Project description:BackgroundTibetan pigs (TP) exhibit heritable adaptations to their hypoxic environments as a result of natural selection. However, candidate proteins that affect the sperm quality of boars on plateaus have not yet been clearly investigated.MethodsIn this study, to reveal the candidate proteins that affect the quality of spermatozoa of boars on plateaus, we analyzed the sperm quality using computer-assisted semen analysis (CASA) system and reactive oxygen species (ROS) levels. We also compared the proteomes of sperm proteomes between TP and Yorkshire pigs (YP) raised at high altitudes using the isobaric tags for relative and absolute quantitation (iTRAQ) in combination with the liquid chromatography-tandem mass spectrometry (LC-MS/MS) proteomic method, and confirmed the relative expression levels of the four proteins by western blotting.ResultsThe sperm quality of the TP was superior to that of the YP on plateaus. Of the 1,555 quantified proteins, 318 differentially expressed proteins (DEPs) were identified. Gene ontology (GO) analysis revealed that the DEPs were predominantly associated with the sorbitol metabolic process, removal of superoxide radicals, cellular response to superoxide, response to superoxide and regulation of the mitotic spindle assembly. The Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways were mainly enriched in pathways involved in the regulation of the actin cytoskeleton, glutathione metabolism, oxidative phosphorylation, and estrogen signaling. Based on the protein-protein interaction (PPI) network analysis, we identified 8 candidate proteins (FN1, EGF, HSP90B1, CFL1, GPX4, NDUFA6, VDAC2, and CP) that might play important roles and affect the sperm quality of boars on plateaus. Moreover, the relative expression levels of four proteins (CFL1, EGF, FN1, and GPX4) were confirmed by western blot analysis.ConclusionsOur study revealed 8 candidate proteins (FN1, EGF, HSP90B1, CFL1, GPX4, NDUFA6, VDAC2, and CP) that affect the sperm quality of boar on plateaus and provide a reference for further studies on improving sperm quality and the molecular breeding of boars on plateaus.

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RNA sequencing reveals candidate genes and polymorphisms related to sperm DNA integrity in testis tissue from boars.

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RNA sequencing reveals candidate genes and polymorphisms related to sperm DNA integrity in testis tissue from boars.

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