Systematic human/zebrafish comparative identification of cis-regulatory activity around vertebrate developmental transcription factor genes

AbstractPan-vertebrate developmental cis-regulatory elements are discernible as highly conserved noncoding elements (HCNEs) and are often dispersed over large areas around the pleiotropic genes whose expression they control. On the loci of two developmental transcription factor genes, SOX3 and PAX6, we demonstrate that HCNEs conserved between human and zebrafish can be systematically and reliably tested for their regulatory function in multiple stable transgenes in zebrafish, and their genomic reach estimated with confidence using synteny conservation and HCNE density along these loci. HCNEs of both human and zebrafish function as specific developmental enhancers in zebrafish. We show that human HCNEs result in expression patterns in zebrafish equivalent to those in mouse, establishing zebrafish as a suitable model for large-scale testing of human developmental enhancers. Orthologous human and zebrafish enhancers underwent functional evolution within their sequence and often directed related but non-identical expression patterns. Despite an evolutionary distance of 450 million years, one pax6 HCNE drove expression in identical areas when comparing zebrafish vs. human HCNEs. HCNEs from the same area often drive overlapping patterns, suggesting that multiple regulatory inputs are required to achieve robust and precise complex expression patterns exhibited by developmental genes

Exonic remnants of whole-genome duplication reveal cis-regulatory function of coding exons

Using a comparative genomics approach to reconstruct the fate of genomic regulatory blocks (GRBs) and identify exonic remnants that have survived the disappearance of their host genes after whole-genome duplication (WGD) in teleosts, we discover a set of 38 candidate cis-regulatory coding exons (RCEs) with predicted target genes. These elements demonstrate evolutionary separation of overlapping protein-coding and regulatory information after WGD in teleosts. We present evidence that the corresponding mammalian exons are still under both coding and non-coding selection pressure, are more conserved than other protein coding exons in the host gene and several control sets, and share key characteristics with highly conserved non-coding elements in the same regions. Their dual function is corroborated by existing experimental data. Additionally, we show examples of human exon remnants stemming from the vertebrate 2R WGD. Our findings suggest that long-range cis-regulatory inputs for developmental genes are not limited to non-coding regions, but can also overlap the coding sequence of unrelated genes. Thus, exonic regulatory elements in GRBs might be functionally equivalent to those in non-coding regions, calling for a re-evaluation of the sequence space in which to look for long-range regulatory elements and experimentally test their activity

Chromatin preparation for ChIP-seq in Oikopleura dioica

<p>Chromatin immunoprecipitation (ChIP) is a widely used molecular technique for binding site identification of DNA-associated proteins in many organisms. Although well established, its use in non-model organisms usually requires extensive protocol optimisation.<br>Here we describe an optimized protocol for collection, fixation, lysis and chromatin sonication of three developmental stages of the chordate model <em>Oikopleura dioica</em>, an emergent model organism for chordate developmental and evolutionary biology studies.</p> <p>Includes troubleshooting section of commonly encountered problems.</p

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\ua0MB 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

Morphogenesis underlying the development of the everted teleost telencephalon

BACKGROUND: Although the mechanisms underlying brain patterning and regionalization are very much conserved, the morphology of different brain regions is extraordinarily variable across vertebrate phylogeny. This is especially manifest in the telencephalon, where the most dramatic variation is seen between ray-finned fish, which have an everted telencephalon, and all other vertebrates, which have an evaginated telencephalon. The mechanisms that generate these distinct morphologies are not well understood. RESULTS: Here we study the morphogenesis of the zebrafish telencephalon from 12 hours post fertilization (hpf) to 5 days post fertilization (dpf) by analyzing forebrain ventricle formation, evolving patterns of gene and transgene expression, neuronal organization, and fate mapping. Our results highlight two key events in telencephalon morphogenesis. First, the formation of a deep ventricular recess between telencephalon and diencephalon, the anterior intraencephalic sulcus (AIS), effectively creates a posterior ventricular wall to the telencephalic lobes. This process displaces the most posterior neuroepithelial territory of the telencephalon laterally. Second, as telencephalic growth and neurogenesis proceed between days 2 and 5 of development, the pallial region of the posterior ventricular wall of the telencephalon bulges into the dorsal aspect of the AIS. This brings the ventricular zone (VZ) into close apposition with the roof of the AIS to generate a narrow ventricular space and the thin tela choroidea (tc). As the pallial VZ expands, the tc also expands over the upper surface of the telencephalon. During this period, the major axis of growth and extension of the pallial VZ is along the anteroposterior axis. This second step effectively generates an everted telencephalon by 5 dpf. CONCLUSION: Our description of telencephalic morphogenesis challenges the conventional model that eversion is simply due to a laterally directed outfolding of the telencephalic neuroepithelium. This may have significant bearing on understanding the eventual organization of the adult fish telencephalon

Complex epigenomes of endocycling ovaries vs. mitotic testis in the closest living relative of vertebrates

<p><strong>Abstract</strong></p> <p>The animal epigenome is composed of interconnected distinct functions, where the key is a chromatin structure and dynamics. Epigenetic information is highly specific for certain cell type and developmental stage and dynamically changes with the cell cycle. Here we present thorough analysis of epigenome components of maturing gonads of Oikopleura dioica. Use of ovary and testes samples enabled us to compare between mitotic male germline and endocycling female nurse cells. The urochordate Oikopleura is considered the closest relative to vertebrates with a great proportion of its extremely compact genome being splice-led and/or transcribed in operons. Genome-wide ChIP-chip profiles of numerous posttranslational histone modifications, along with RNA polymerase II, CTCF and p300 binding sites were generated from two homogeneous cell population samples. To capture the combinatorial action of the histone modifications we applied unsupervised learning method and annotated the genome with 15 distinct chromatin states and related those to numerous genomic features. Besides many conserved histone modification functions, we found some exceptions, which could be attributed to sex chromosomes, cell cycle mode or germ line-specificity. Interestingly, the nurse nuclei pattern is in agreement to what is known from other metazoan epigenomes, but testis epigenome revealed quite different histone code. In addition, to gain more insight into the cell cycle and developmental stage specificities of the histone modifications, we analyzed expression profiles of panel of histone modifier enzymes and address the complexity of the chromatin dynamics from this point of view. Overall, we provide a detailed structure of an early chordate epigenome.</p> <p><strong>References:</strong></p> <p>Jenuwein, T., & Allis, C. D. (2001). Translating the histone code. Science (New York, N.Y.), 293(5532), 1074–1080Ho, J. W. K., Jung, Y. L., Liu, T., Alver, B. H., Lee, S., Ikegami, K., … Park, P. J. (2014). Comparative analysis of metazoan chromatin organization. Nature, 512(7515), 449–452. doi:10.1038/nature13415Fuchs, J., Demidov, D., Houben, A., & Schubert, I. (2006). Chromosomal histone modification patterns--from conservation to diversity. Trends in Plant Science, 11(4), 199–208Delsuc, F., Brinkmann, H., Chourrout, D., & Philippe, H. (2006). Tunicates and not cephalochordates are the closest living relatives of vertebrates. Nature, 439(7079), 965–8. doi:10.1038/nature04336Denoeud, F., Henriet, S., Mungpakdee, S., Aury, J.-M., Da Silva, C., … Chourrout, D. (2010). Plasticity of animal genome architecture unmasked by rapid evolution of a pelagic tunicate. Science (New York, N.Y.), 330(6009), 1381–5.Bouquet, J.-M., Spriet, E., Troedsson, C., Otterå, H., Chourrout, D., & Thompson, E. M. (2009). Culture optimization for the emergent zooplanktonic model organism Oikopleura dioica. Journal of Plankton Research, 31(4), 359–370.Moosmann, A., Campsteijn, C., Jansen, P. W., Nasrallah, C., Raasholm, M., Stunnenberg, H. G., & Thompson, E. M. (2011). Histone variant innovation in a rapidly evolving chordate lineage. BMC Evolutionary Biology, 11(1), 208.Danks, G., Campsteijn, C., Parida, M., Butcher, S.,… Manak, J. R. (2013). OikoBase: a genomics and developmental transcriptomics resource for the urochordate Oikopleura dioica. Nucleic Acids Research, 41, D845–53.Ganot, P., Bouquet, J.-M., Kallesøe, T., & Thompson, E. M. (2007). The Oikopleura coenocyst, a unique chordate germ cell permitting rapid, extensive modulation of oocyte production. Developmental Biology, 302(2), 591–600.Ernst, J., & Kellis, M. (2012). ChromHMM: automating chromatin-state discovery and characterization. Nature Methods, 9(3), 215–6.</p> <p><strong><br></strong></p

Additional file 1 of Distinct core promoter codes drive transcription initiation at key developmental transitions in a marine chordate

Supplementary Material. Supplemental Results, Supplemental Methods, Table S1, Table S2, Figures S1 - S12 and Supplemental References. (PDF 13 464 kb