Paternal Poly (ADP-ribose) Metabolism Modulates Retention of Inheritable Sperm Histones and Early Embryonic Gene Expression

To achieve the extreme nuclear condensation necessary for sperm function, most histones are replaced with protamines during spermiogenesis in mammals. Mature sperm retain only a small fraction of nucleosomes, which are, in part, enriched on gene regulatory sequences, and recent findings suggest that these retained histones provide epigenetic information that regulates expression of a subset of genes involved in embryo development after fertilization. We addressed this tantalizing hypothesis by analyzing two mouse models exhibiting abnormal histone positioning in mature sperm due to impaired poly(ADP-ribose) (PAR) metabolism during spermiogenesis and identified altered sperm histone retention in specific gene loci genome-wide using MNase digestion-based enrichment of mononucleosomal DNA. We then set out to determine the extent to which expression of these genes was altered in embryos generated with these sperm. For control sperm, most genes showed some degree of histone association, unexpectedly suggesting that histone retention in sperm genes is not an all-or-none phenomenon and that a small number of histones may remain associated with genes throughout the genome. The amount of retained histones, however, was altered in many loci when PAR metabolism was impaired. To ascertain whether sperm histone association and embryonic gene expression are linked, the transcriptome of individual 2-cell embryos derived from such sperm was determined using microarrays and RNA sequencing. Strikingly, a moderate but statistically significant portion of the genes that were differentially expressed in these embryos also showed different histone retention in the corresponding gene loci in sperm of their fathers. These findings provide new evidence for the existence of a linkage between sperm histone retention and gene expression in the embryo

Personalization in Skipforward, an Ontology-Based Distributed Annotation System

Abstract. Skipforward is a distributed annotation system allowing users to enter and browse statements about items and their features. Items can be things such as movies or books; item features are the genre of a movie or the storytelling pace of a book. Whenever multiple users annotate the same item with a statement about the same feature, these individual statements get aggregated by the system. For aggregation, individual user statements are weighted according to a competence metric based on the constrained Pearson correlation, adapted for Skipforward data: A user gets assigned high competence with regard to the feature in question if, for other items and the same feature type, he had a similar opinion to the current user. Since the competence metric is dependent on the user currently viewing the data, the user’s view of the data is completely personalized. In this paper, the personalization aspect as well as the item and expert recommender are presented.

Aberrant chromatin composition in mouse models of altered PAR metabolism.

<p>Chromomycin A3 (CMA3) intercalation into the DNA indicates incomplete chromatin condensation in <i>s</i>perm from <i>Parg</i>(110)^−/− (A) and PJ34-treated (C) males with histone retention. (B, D) Histogram of sperm CMA3-staining intensities reflects that severity of CM3A staining varied at the level of individual sperm and individual fathers (n>200 nuclei/sample, 3 males/group). (E) Immunoblot analyses of sperm protein lysates showing increase in histone retention in PJ34 treated males. TUBA1A: alpha tubulin loading control. (F) Overlaps of genes identified as differentially histone associated in sperm from 3 individual <i>Parg</i>(110)^−/− males (“PargA”, “PargB”, “PargC”, the fathers of the embryos analyzed below) by micrococcal nuclease digests (MND) compared to the wild-type controls. The “PargAll” data set contains all genes commonly identified as differentially MNase-sensitive across 10 <i>Parg</i>(110)^−/− males compared with 9 wild-type control animals. The red circle indicates common genes that were differentially histone associated in all groups (1604+216 = 1820, red circle) compared with wild-type. (G) PJ34: differentially MNase-sensitive genes in three different males (like in E) and overlap with a surrogate dataset (“PJ34All”) consisting of data from all 4 PJ34-treated males compared with 9 wild-type control males. The overlap of 2,489 genes that were commonly differentially histone associated in sperm samples is indicated (blue circle). (H) Overlap of genes commonly affected by differential histone association between the <i>Parg</i>(110)^−/− and the PJ34 models compared to wild-type controls (red and blue circles in F and G). A Pearson correlation examining significance of this overlap using a genetic background of 19,472 genes was calculated with a resulting P<0.0001, dismissing the null hypothesis that the observed overlap is coincidental (predicted number). The list and GO-term analysis of the 583 genes is contained in <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1004317#pgen.1004317.s004" target="_blank">Dataset S4</a> (MS Excel).</p

Perturbing PAR metabolism results in differential sperm histone association of gene loci with either excessive or reduced retention of nucleosomes.

<p>A) Functional GO-term enrichment of genes affected by elevated histone association (MAT(+)) or local failure to retain histones in regulatory gene sequences (MAT(−)) in sperm from <i>Parg</i>(110)^−/− (left panels) and PJ34-treated males (right hand panels). The y-axis shows GO terms and logarithmic scale indicates their p-values of GO-terms returned by DAVID. False discovery rates (FDR) are indicated above the graphs. The numbers of genes in a given GO-term are in parentheses. (B) Overlaps of relative histone enrichment or deficiency in <i>Parg</i>(110)^−/− or PJ34-treated mouse models compared to wild-type controls. (C, D) Comparison of genes that are differentially histone associated in <i>Parg</i>(110)^−/− or PJ34 sperm with known maternal transcripts or newly expressed embryonic transcripts or spermatogenesis-specific genes indicates the potential relevance of aberrant histone association on genes expressed in the 2-cell embryo (Embryo). Maternal: transcripts present in 1-cell embryos prior to the major wave of genome activation <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1004317#pgen.1004317-Zeng1" target="_blank">[36]</a>. The genes in the maternal, embryonic and spermatogenic groups are listed in <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1004317#pgen.1004317.s004" target="_blank">Dataset S4</a> (MS Excel).</p

Genes associated with MNase-sensitivity.

<p><i>Parg</i>(110)^−/− (PargKO), PJ34, Wildtype controls: groups of sperm samples analyzed for gene histone association by tiling arrays with genomic “input” correction. Numbers of genes with significant histone binding in sperm are indicated. ♂ [n]: numbers of males. PargA/Wt, PJ34A/Wt, etc.: sperm samples of single males (♂, n = 1) analyzed for sperm histone association after pair-wise comparison to wild-type samples. Total genes [n]: number of sperm genes (differentially) associated with histones, indiscriminate whether sensitivity was increased (MAT+ genes [n]) or decreased (MAT− genes [n]). Please note that a number of genes were associated with both (MAT+ and MAT−) fractions. All Parg/Wt, All PJ34/Wt: comparison of all <i>Parg</i>(110)^−/− males (n = 10) with wild-type (n = 9) males or males treated with PJ34 (n = 4) with wild-type control males (n = 9).</p

Genes with differential expression in 2-cell embryos.

<p>Separating differentially expressed genes in 2-cell embryos (2CEDE) into up-regulated (ratio>1) and down-regulated genes (ratio<1) reveals a strong bias of differentially-expressed genes towards up-regulation or illegitimate activation of genes across all experimental groups/platforms used. The numbers in brackets indicate percentage of genes in a category, e.g., 75.4% of all genes detected in the microarrays of <i>Parg</i>(110)^−/− 2CE DE were up-regulated and 24.6% were down-regulated.</p

Top network of Vk2 PIC/NIC discriminatory genes generated by Ingenuity Pathway Analysis.

<p>Red/pink color indicates upregulation of the genes (microarray data). Connections of NFkB complex with other genes is shown in blue color.</p

Genes differenially expressed in VK2 cells treated with proinflammatory/immunomodulatory compounds.

<p>Genes differenially expressed in VK2 cells treated with proinflammatory/immunomodulatory compounds.</p