Sex-specific chromatin landscapes in an ultra-compact chordate genome

Abstract

Background: In multicellular organisms, epigenome dynamics are associated with transitions in the cell cycle, development, germline specification, gametogenesis and inheritance. Evolutionarily, regulatory space has increased in complex metazoans to accommodate these functions. In tunicates, the sister lineage to vertebrates, we examine epigenome adaptations to strong secondary genome compaction, sex chromosome evolution and cell cycle modes.

Results: Across the 70 MB Oikopleura dioica genome, we profiled 19 histone modifications, and RNA polymerase II, CTCF and p300 occupancies, to define chromatin states within two homogeneous tissues with distinct cell cycle modes: ovarian endocycling nurse nuclei and mitotically proliferating germ nuclei in testes. Nurse nuclei had active chromatin states similar to other metazoan epigenomes, with large domains of operon-associated transcription, a general lack of heterochromatin, and a possible role of Polycomb PRC2 in dosage compensation. Testis chromatin states reflected transcriptional activity linked to spermatogenesis and epigenetic marks that have been associated with establishment of transgenerational inheritance in other organisms. We also uncovered an unusual chromatin state specific to the Y-chromosome, which combined active and heterochromatic histone modifications on specific transposable elements classes, perhaps involved in regulating their activity.

Conclusions: Compacted regulatory space in this tunicate genome is accompanied by reduced heterochromatin and chromatin state domain widths. Enhancers, promoters and protein-coding genes have conserved epigenomic features, with adaptations to the organization of a proportion of genes in operon units. We further identified features specific to sex chromosomes, cell cycle modes, germline identity and dosage compensation, and unusual combinations of histone PTMs with opposing consensus functions.

Keywords: Dosage compensation; Endocycle; Enhancer; Heterochromatin; Histone; Polycomb; Spermatogenesis; Transposable elements.

Fig. 1

Percentage of genome, autosomes, X- and Y-chromosomes covered by ChIP-enriched regions for each ChIP sample, as well as percentages of the genome covered by transcriptionally active regions (TARs) (using previously published tiling array data from [32], and their definition of a TAR as “any stretch of consecutive positive probes in a particular sample”), in the O. dioica ovary and testis

Fig. 2

Chromatin states in the O. dioica ovary and testis reveal two distinct epigenetic landscapes. Heatmap of model emission parameters (a) for 15 chromatin states learned across the genome, using both samples (see also Additional file 3: Table S3). Proportions of the genome in the ovary and testis covered by each chromatin state (b) show sex/tissue specificity of chromatin states. Heatmaps (c) visualize the fold enrichments of each chromatin state (columns) for a set of genomic features (rows), as listed, in the ovary and testis. This facilitates assignment of putative biological function(s) of each chromatin state in each tissue. Gray shading indicates states that cover below 0.1% of the genome (70,000 bp) in each tissue. These low-coverage states nevertheless have large fold enrichments over certain genomic features, e.g., state 3 has low coverage in the testis but is found more often than expected by chance at active operons. Features are clustered according to their correlation in the ovary, and this ordering is used for the testis heatmap to allow comparison. The table gives the numbers of each feature in each sample. Gene bodies (orange) and promoter regions (purple) are split into active (green) and silent (red) states in the respective ovary and testis tissues. Txn, transcription; TF, transcription factor; ZF, zinc finger protein; HD, homeodomain protein, high- and low-specificity genes according to breadth of expression across development (see “Methods” section); TAR, transcriptionally active region; TE, transposable element; unannotated, regions lacking annotation or transcription; 5meC, methylcytosine; RNApolII-S5P, serine 5 phosphorylated RNA polymerase II

Fig. 3

Gene ontology chromatin state enrichments (light to dark blue = low to high enrichment) in promoters (purple), gene bodies (orange), TSSs (yellow) and TESs (black) of active (green) and silent (red) genes grouped by GO term and clustered. Selected clusters are labeled with representative functions and numbered, to facilitate reference in “Results” section and in Additional file 3: Table S5

Fig. 4

Compact chromatin state domains in the O. dioica epigenome. a Distributions of all chromatin state domain widths for each cell type in O. dioica compared to those in human embryonic stem cells (H1-hESC) [92]. O. dioica domain widths were adjusted for the difference in resolution (50 vs. 200 bp) by rounding O. dioica widths up to the nearest 200 bp. b Comparison of the ratios of human state domain widths for active promoters, transcriptional elongation and heterochromatin (see Additional file 2: Fig. S6) to corresponding state domains in O. dioica. Each box summarizes the ratios of median domain widths (y-axis) for the human cell lines relative to the similar domains in each O. dioica tissue (x-axis). The red line indicates the ratio of the human genome size to that of O. dioica (44-fold smaller). Relative to the genome compaction, heterochromatin domains in O. dioica are disproportionately more compact

Fig. 5

Expression profiles of O. dioica chromatin modifier genes showing the main histone methyltransferases, demethylases, acetyltransferases, deacetylases, kinases and phosphatases (indicated by colored bars) at relevant stages of development, where blue is low and yellow is high expression. Log₂ fold expression difference between ovary and testis is indicated by a graded bar where darkest blue is testis specific and darkest red is ovary specific (see also Additional file 3: Table S6)

Fig. 6

Mosaic plots of Pearson Chi-squared tests for the association of the type of transposable element (TE) on the Y-chromosome and its “bi-modal” overlap with regions of enrichment of heterochromatic H3K9me3 and one of the active marks H3K4me3 (a) or H3K36me3 (b). DIRS, Dictyostelium intermediate repeat sequence; LINE, long interspersed nuclear element; LTR, long terminal repeats; MAV, maverick; MITE, miniature inverted-repeat transposable elements; PLE, PiggyBac-like element; REP, repetitive extragenic palindromic