Histone H1 is an essential chromatin component. Metazoans usually contain multiple stage-specific H1s. In particular, specific variants replace somatic H1s during early embryogenesis. In this regard, Drosophila was an exception because a single dH1 was identified that, starting at cellularization, is detected throughout development in somatic cells. Here, we identify the embryonic H1 of Drosophila, dBigH1. dBigH1 is abundant before cellularization occurs, when somatic dH1 is absent and the zygotic genome is inactive. Upon cellularization, when the zygotic genome is progressively activated, dH1 replaces dBigH1 in the soma, but not in the primordial germ cells (PGCs) that have delayed zygotic genome activation (ZGA). In addition, a loss-of-function mutant shows premature ZGA in both the soma and PGCs. Mutant embryos die at cellularization, showing increased levels of active RNApol II and zygotic transcripts, along with DNA damage and mitotic defects. These results show an essential function of dBigH1 in ZGA regulation.

H3K4me3 is a histone modification that accumulates at the transcription-start site (TSS) of active genes and is known to be important for transcription activation. The way in which H3K4me3 is regulated at TSS and the actual molecular basis of its contribution to transcription remain largely unanswered. To address these questions, we have analyzed the contribution of dKDM5/LID, the main H3K4me3 demethylase in Drosophila, to the regulation of the pattern of H3K4me3. ChIP-seq results show that, at developmental genes, dKDM5/LID localizes at TSS and regulates H3K4me3. dKDM5/LID target genes are highly transcribed and enriched in active RNApol II and H3K36me3, suggesting a positive contribution to transcription. Expression-profiling show that, though weakly, dKDM5/LID target genes are significantly downregulated upon dKDM5/LID depletion. Furthermore, dKDM5/LID depletion results in decreased RNApol II occupancy, particularly by the promoter-proximal Pol lloser5 form. Our results also show that ASH2, an evolutionarily conserved factor that locates at TSS and is required for H3K4me3, binds and positively regulates dKDM5/LID target genes. However, dKDM5/LID and ASH2 do not bind simultaneously and recognize different chromatin states, enriched in H3K4me3 and not, respectively. These results indicate that, at developmental genes, dKDM5/LID and ASH2 coordinately regulate H3K4me3 at TSS and that this dynamic regulation contributes to transcription.

Epigenetics may have an important role in mood stabilizer action. Valproic acid (VPA) is a histone deacetylase inhibitor, and lithium (Li) may have downstream epigenetic actions. To identify genes commonly affected by both mood stabilizers and to assess potential epigenetic mechanisms that may be involved in their mechanism of action, we administered Li (N=12), VPA (N=12), and normal chow (N=12) to Brown Norway rats for 30 days. Genomic DNA and mRNA were extracted from the hippocampus. We used the mRNA to perform gene expression analysis on Affymetrix microarray chips, and for genes commonly regulated by both Li and VPA, we validated expression levels using quantitative real-time PCR. To identify potential mechanisms underlying expression changes, genomic DNA was bisulfite treated for pyrosequencing of key CpG island ‘shores' and promoter regions, and chromatin was prepared from both hippocampal tissue and a hippocampal-derived cell line to assess modifications of histones. For most genes, we found little evidence of DNA methylation changes in response to the medications. However, we detected histone H3 methylation and acetylation in the leptin receptor gene, Lepr, following treatment with both drugs. VPA-mediated effects on histones are well established, whereas the Li effects constitute a novel mechanism of transcriptional derepression for this drug. These data support several shared transcriptional targets of Li and VPA, and provide evidence suggesting leptin signaling as an epigenetic target of two mood stabilizers. Additional work could help clarify whether leptin signaling in the brain has a role in the therapeutic action of Li and VPA in bipolar disorder.

To date, different studies about the genetic basis of hybrid male sterility (HMS), a postzygotic reproductive barrier thoroughly investigated using Drosophila species, have demonstrated that no single major gene can produce hybrid sterility without the cooperation of several genetic factors. Early work using hybrids between Drosophila koepferae (Dk) and Drosophila buzzatii (Db) was consistent with the idea that HMS requires the cooperation of several genetic factors, supporting a polygenic threshold (PT) model. Here we present a genome-wide mapping strategy to test the PT model, analyzing serially backcrossed fertile and sterile males in which the Dk genome was introgressed into the Db background. We identified 32 Dk-specific markers significantly associated with hybrid sterility. Our results demonstrate 1) a strong correlation between the number of segregated sterility markers and males' degree of sterility, 2) the exchangeability among markers, 3) their tendency to cluster into low-recombining chromosomal regions, and 4) the requirement for a minimum number (threshold) of markers to elicit sterility. Although our findings do not contradict a role for occasional major hybrid-sterility genes, they conform more to the view that HMS primarily evolves by the cumulative action of many interacting genes of minor effect in a complex PT architecture.

Protein components of the spliceosome are highly conserved in eukaryotes and can influence several steps of the gene expression process. RSR-2, the Caenorhabditis elegans ortholog of the human spliceosomal protein SRm300/SRRM2, is essential for viability, in contrast to the yeast ortholog Cwc21p. We took advantage of mutants and RNA interference (RNAi) to study rsr-2 functions in C. elegans, and through genetic epistasis analysis found that rsr-2 is within the germline sex determination pathway. Intriguingly, transcriptome analyses of rsr-2(RNAi) animals did not reveal appreciable splicing defects but instead a slight global decrease in transcript levels. We further investigated this effect in transcription and observed that RSR-2 colocalizes with DNA in germline nuclei and coprecipitates with chromatin, displaying a ChIP-Seq profile similar to that obtained for the RNA Polymerase II (RNAPII). Consistent with a novel transcription function we demonstrate that the recruitment of RSR-2 to chromatin is splicing-independent and that RSR-2 interacts with RNAPII and affects RNAPII phosphorylation states. Proteomic analyses identified proteins associated with RSR-2 that are involved in different gene expression steps, including RNA metabolism and transcription with PRP-8 and PRP-19 being the strongest interacting partners. PRP-8 is a core component of the spliceosome and PRP-19 is the core component of the PRP19 complex, which interacts with RNAPII and is necessary for full transcriptional activity. Taken together, our study proposes that RSR-2 is a multifunctional protein whose role in transcription influences C. elegans development.