ABSTRACT
MOTIVATION: The advent of next-generation sequencing technologies has increased the accuracy and quantity of sequence data, opening the door to greater opportunities in genomic research. RESULTS: In this article, we present GNUMAP (Genomic Next-generation Universal MAPper), a program capable of overcoming two major obstacles in the mapping of reads from next-generation sequencing runs. First, we have created an algorithm that probabilistically maps reads to repeat regions in the genome on a quantitative basis. Second, we have developed a probabilistic Needleman-Wunsch algorithm which utilizes _prb.txt and _int.txt files produced in the Solexa/Illumina pipeline to improve the mapping accuracy for lower quality reads and increase the amount of usable data produced in a given experiment. AVAILABILITY: The source code for the software can be downloaded from http://dna.cs.byu.edu/gnumap.
Subject(s)
Algorithms , Chromosome Mapping/methods , DNA/genetics , Sequence Analysis, DNA/methods , Software , Base Sequence , Data Interpretation, Statistical , Molecular Sequence DataABSTRACT
OBJECTIVE: To evaluate the associations between proper protamine incorporation and DNA methylation at imprinted loci. DESIGN: Experimental research study. SETTING: Research laboratory. PATIENT(S): Three populations were tested-abnormal protamine patients, oligozoospermic patients, and fertile donors. INTERVENTION(S): The CpG methylation patterns were examined at seven imprinted loci sequenced: LIT1, MEST, SNRPN, PLAGL1, PEG3, H19, and IGF2. MAIN OUTCOME MEASURE(S): The DNA methylation patterns were analyzed using bisulfite sequencing. The percentage of methylation was compared between fertile and infertile patients displaying abnormal protamination. RESULT(S): At six of the seven imprinted genes, the overall DNA methylation patterns at their respective differentially methylated regions were significantly altered in both infertile patient populations. When comparing the severity of methylation alterations among infertile patients, the oligozoospermic patients were significantly affected at mesoderm-specific transcript (MEST), whereas abnormal protamine patients were affected at KCNQ1, overlapping transcript 1 (LIT1), and at small nuclear ribonucleoprotein polypeptide N (SNRPN). CONCLUSION(S): Patients with male factor infertility had significantly increased methylation alteration at six of seven imprinted loci tested, with differences in significance observed between oligozoospermic and abnormal protamine patients. This could suggest that risk of transmission of epigenetic alterations may be different with diagnoses. However, this study does not provide a causal link for epigenetic inheritance of imprinting diseases, but does show significant association between male factor infertility and alterations in sperm DNA methylation at imprinted loci.
Subject(s)
DNA Methylation/genetics , DNA/genetics , Genetic Loci/genetics , Genomic Imprinting/genetics , Infertility, Male/classification , Infertility, Male/genetics , Spermatozoa , CpG Islands/genetics , DNA/analysis , Humans , Infertility, Male/metabolism , Insulin-Like Growth Factor II/genetics , Kruppel-Like Transcription Factors/genetics , Male , Oligospermia/genetics , Oligospermia/metabolism , Potassium Channels, Voltage-Gated/genetics , Protamines/metabolism , Proteins/genetics , Spermatozoa/chemistry , snRNP Core Proteins/geneticsABSTRACT
Because nucleosomes are widely replaced by protamine in mature human sperm, the epigenetic contributions of sperm chromatin to embryo development have been considered highly limited. Here we show that the retained nucleosomes are significantly enriched at loci of developmental importance, including imprinted gene clusters, microRNA clusters, HOX gene clusters, and the promoters of stand-alone developmental transcription and signalling factors. Notably, histone modifications localize to particular developmental loci. Dimethylated lysine 4 on histone H3 (H3K4me2) is enriched at certain developmental promoters, whereas large blocks of H3K4me3 localize to a subset of developmental promoters, regions in HOX clusters, certain noncoding RNAs, and generally to paternally expressed imprinted loci, but not paternally repressed loci. Notably, trimethylated H3K27 (H3K27me3) is significantly enriched at developmental promoters that are repressed in early embryos, including many bivalent (H3K4me3/H3K27me3) promoters in embryonic stem cells. Furthermore, developmental promoters are generally DNA hypomethylated in sperm, but acquire methylation during differentiation. Taken together, epigenetic marking in sperm is extensive, and correlated with developmental regulators.
