Project description:Epidemiological studies in humans suggest that acquired paternal traits, such as obesity, are associated with a higher risk of fathering small for gestational age offspring. Studies in non-human mammals suggest that such associations could be mediated by DNA methylation changes in spermatozoa that influence offspring development in utero. Human obesity is associated with differential DNA methylation in peripheral blood. It is unclear, however, whether this differential DNA methylation is reflected in less readily available tissues such as spermatozoa. In this study, we profiled genome-wide DNA methylation with the Infinium MethylationEPIC array in matched samples of human blood and sperm from lean (discovery n = 47; replication n = 21) and obese (n = 22) healthy males of proven fertility. To characterize sperm-specific DNA methylation signatures, we compared spermatozoal DNA methylation data to that of nearly 6,000 somatic tissue samples available on the Gene Expression Omnibus database. We studied covariation patterns between whole blood and sperm and investigated consistent obesity-associated DNA methylation differences.
Project description:BACKGROUND: In previous studies using candidate gene approaches, low sperm count (oligospermia) has been associated with altered sperm mRNA content and DNA methylation in both imprinted and non-imprinted genes. We performed a genome-wide analysis of sperm DNA methylation and mRNA content to test for associations with sperm function. METHODS AND RESULTS: Sperm DNA and mRNA were isolated from 21 men with a range of semen parameters presenting to a tertiary male reproductive health clinic. DNA methylation was measured with the Illumina Infinium array at 27,000 CpG loci. Unsupervised clustering of methylation data differentiated the 21 sperm samples by their motility values. Recursively partitioned mixture modeling (RPMM) of methylation data resulted in four distinct methylation profiles that were significantly associated with sperm motility (P=0.01). Linear models of microarray analysis (LIMMA) was performed based on motility and identified 9,189 CpG loci with significantly altered methylation (Q<0.05) in the low motility samples, with many loci located in genes associated with subfertility and epigenetic regulation. In the low motility samples, the majority of disrupted CpG loci (80%) were hypomethylated. Of the aberrantly methylated CpGs, 194 were associated with imprinted genes almost equally distributed into hypermethylated (predominantly paternally expressed) and hypomethylated (predominantly maternally expressed) groups. Sperm mRNA was measured with the Human Gene 1.0 ST Affymetrix GeneChip Array. LIMMA analysis based on motility identified 20 candidate transcripts as differentially expressed in low motility sperm, including HDAC1 (NCBI 3065), SIRT3 (NCBI 23410), and DNMT3A (NCBI 1788). Altered expression of these epigenetic regulatory genes was associated with RPMM DNA methylation class. CONCLUSIONS: Using integrative genome-wide approaches to study epigenetic and gene expression patterns in human sperm we identified CpG methylation profiles and mRNA alterations associated with low sperm motility, and that low motility sperm may have aberrant genome-wide hypomethylation due to excess HDAC1 activity. See summary above
Project description:BACKGROUND: In previous studies using candidate gene approaches, low sperm count (oligospermia) has been associated with altered sperm mRNA content and DNA methylation in both imprinted and non-imprinted genes. We performed a genome-wide analysis of sperm DNA methylation and mRNA content to test for associations with sperm function. METHODS AND RESULTS: Sperm DNA and mRNA were isolated from 21 men with a range of semen parameters presenting to a tertiary male reproductive health clinic. DNA methylation was measured with the Illumina Infinium array at 27,000 CpG loci. Unsupervised clustering of methylation data differentiated the 21 sperm samples by their motility values. Recursively partitioned mixture modeling (RPMM) of methylation data resulted in four distinct methylation profiles that were significantly associated with sperm motility (P=0.01). Linear models of microarray analysis (LIMMA) was performed based on motility and identified 9,189 CpG loci with significantly altered methylation (Q<0.05) in the low motility samples, with many loci located in genes associated with subfertility and epigenetic regulation. In the low motility samples, the majority of disrupted CpG loci (80%) were hypomethylated. Of the aberrantly methylated CpGs, 194 were associated with imprinted genes almost equally distributed into hypermethylated (predominantly paternally expressed) and hypomethylated (predominantly maternally expressed) groups. Sperm mRNA was measured with the Human Gene 1.0 ST Affymetrix GeneChip Array. LIMMA analysis based on motility identified 20 candidate transcripts as differentially expressed in low motility sperm, including HDAC1 (NCBI 3065), SIRT3 (NCBI 23410), and DNMT3A (NCBI 1788). Altered expression of these epigenetic regulatory genes was associated with RPMM DNA methylation class. CONCLUSIONS: Using integrative genome-wide approaches to study epigenetic and gene expression patterns in human sperm we identified CpG methylation profiles and mRNA alterations associated with low sperm motility, and that low motility sperm may have aberrant genome-wide hypomethylation due to excess HDAC1 activity.
Project description:It has been thought that epigenetic changes underlie the evolutionary divergence of phenotype between closely related species. However, the manner in which epigenetic changes are generated remains unknown. Although whole-genome DNA methylation profiles in some somatic tissues and sperm have been reported for humans and chimpanzees, a systematic analysis of these data has been lacking. In this manuscript, therefore, we analyzed these methylomes in detail, identified genomic regions with different DNA methylation levels, and examined the cell-type specificity and its association with changes in genomic sequence. Moreover, we generated a methylation map of Japanese macaque sperm and used it as an out-group to infer the evolutional history of methylation in these regions.
Project description:Genome wide DNA methylation profiling of 2 distinct subpopulations of sperm within a single ejaculate. The Illumina Infinium 450k Human DNA methylation Beadchip was used to obtain DNA methylation profiles across approximately 450,000 CpGs. Samples were provided by 20 normozoospermic individuals. A density gradient centrifugation was performed on each samples to yeild two distict populations for each ejaculate (one from the 90% isolate layer high quality sperm and one from the 35% isolate layer low quality sperm of the column). Bisulphite converted DNA from the 40 samples were hybridised to the Illumina Infinium 450k Human Methylation Beadchip
Project description:DNA methylation in sperm is among the most important factors shaping evolution of the mammalian genome. By directly altering germline mutation rates, the DNA methylation system has shaped the CpG landscape of mammalian genomes, resulting in the CpG island phenomenon. Yet little is known about how this system itself has co-evolved with its substrate during mammalian evolution. We analyzed full-genome single-CpG DNA methylation profiles in sperm from human, chimp, gorilla, rhesus, mouse, rat and dog. Our results point to an evolutionary expansion of the unmethylated portion of mammalian genomes. Within the mutually orthologous genome this trend is driven both by the birth of unmethylated regions and by widening of intervals that are unmethylated in ancestors. We find strikingly divergent global features in rodents. At the same time, we observe the evolutionary emergence of methylome features in mouse sperm bears association with similar pathways to those found in human. Together,these results revealed general principles in mammalian epigenome evolution.
Project description:Genome wide DNA methylation profiling of 2 distinct subpopulations of sperm within a single ejaculate. The Illumina Infinium 450k Human DNA methylation Beadchip was used to obtain DNA methylation profiles across approximately 450,000 CpGs. Samples were provided by 20 normozoospermic individuals. A density gradient centrifugation was performed on each samples to yeild two distict populations for each ejaculate (one from the 90% isolate layer high quality sperm and one from the 35% isolate layer low quality sperm of the column).
Project description:IAP is one of the most transpositinally active retrotransposons in the mouse. Its transcription starts from the 5' long terminal repeat (LTR), and the expression level varied between cell types. This variation is thought to arise from differences in the epigenetic states, such as DNA methylation, of some copies of the 5' LTR between the cells. However, due to the high copy number and high sequence similarity, it was difficult to comprehensively and individually analyze epigenetic state of the IAP LTRs. Here, we developed a method called TEPBAT (Target Enrichment after Post-Bisulfite Adaptor Tagging) to analyze DNA methylation of individual retrotransposon copies. Using the method, we determined DNA methylation levels of >8,500 copies of genomic IAP LTRs (>97% of the copies targeted by the PCR primer) in sperm and tail. This revealed that the vast majority of the LTRs were heavily methylated both in sperm and tail, but hypomethylated copies were more frequently found in sperm than in tail. Interestingly, most of these hypomethylated LTRs were solo-type, belonged to specific subfamilies, and carried binding sites for transcription factors (TFs) active in male germ cells. We discuss possible roles for these TF-binding sites and the IAP internal sequence in regulation of LTR methylation.
Project description:Cord blood DNA methylation is associated with numerous health outcomes and environmental exposures. Whole cord blood DNA reflects all nucleated blood cell types, while centrifuging whole blood separates red blood cells by generating a white blood cell buffy coat. Both sample types are used in DNA methylation studies. Cell types have unique methylation patterns and processing can impact cell distributions, which may influence comparability. To evaluate differences in cell composition and DNA methylation between buffy coat and whole cord blood samples, cord blood DNA methylation was measured with the Infinium EPIC BeadChip (Illumina) in 8 individuals, each contributing buffy coat and whole blood samples.
Project description:DNA methylation profile of mouse sperm from conventionally-raised mice and gut dysbiosis experienced mice were characterized using whole-genome bisulfite sequencing. Genome-wide DNA methylation changes between control and dysbiotic male�s sperm were highly comparable, with no change in DNAme globally or at genomic features, only 21 differentially methylated regions (DMR) were identified, which did not overlap known regulatory elements. Epididymal sperm samples were harvested from 11 weeks old inbred male mice that were experiencing gut microbiota dysbiosis for 6-week (antibiotics treated, n=5), or drink sterilized water (control, n=5).