Tardigrades, also known as water bears, are small aquatic animals. Some tardigrade species tolerate almost complete dehydration and exhibit extraordinary tolerance to various physical extremes in the dehydrated state. Here we determine a high-quality genome sequence of Ramazzottius varieornatus, one of the most stress-tolerant tardigrade species. Precise gene repertoire analyses reveal the presence of a small proportion (1.2% or less) of putative foreign genes, loss of gene pathways that promote stress damage, expansion of gene families related to ameliorating damage, and evolution and high expression of novel tardigrade-unique proteins. Minor changes in the gene expression profiles during dehydration and rehydration suggest constitutive expression of tolerance-related genes. Using human cultured cells, we demonstrate that a tardigrade-unique DNA-associating protein suppresses X-ray-induced DNA damage by ∼40% and improves radiotolerance. These findings indicate the relevance of tardigrade-unique proteins to tolerability and tardigrades could be a bountiful source of new protection genes and mechanisms.

Sequencing the large genomes of sharks. We focused on the brownbanded bamboo shark Chiloscyllium punctatum, for which we recently tabled embryonic stages 8 , and the cloudy catshark Scyliorhinus torazame. Their whole genomes, measured to be approximately 4.7 and 6.7 Gbp, respectively, were sequenced de novo to obtain assemblies including megabase-long scaffolds (Supplementary Note 1.1). We also assembled the genome of the whale shark Rhincodon typus using short sequence reads previously generated 3 (Supplementary Note 1.2). Using these genome assemblies, we performed genome-wide gene prediction, assisted by transcript evidence and protein-level homology to other vertebrates. The obtained genome assemblies and gene models exhibit high coverage (Supplementary Fig. 1), and of these, the bamboo shark genome assembly achieved the highest continuity (N50 scaffold length, 1.9 Mbp) and completeness (97% of reference orthologues identified at least partially). Using the novel gene models, we constructed orthologue groups encompassing a diverse array of vertebrate species (see below). Our products outperform existing

MotivationAlong with the increasing accessibility to comprehensive sequence information, such as whole genomes and transcriptomes, the demand for assessing their quality has been multiplied. To this end, metrics based on sequence lengths, such as N50, have become a standard, but they only evaluate one aspect of assembly quality. Conversely, analyzing the coverage of pre-selected reference protein-coding genes provides essential content-based quality assessment, but the currently available pipelines for this purpose, CEGMA and BUSCO, do not have a user-friendly interface to serve as a uniform environment for assembly completeness assessment. ResultsHere, we introduce a brand-new web server, gVolante, which provides an online tool for (i) on-demand completeness assessment of sequence sets by means of the previously developed pipelines CEGMA and BUSCO and (ii) browsing pre-computed completeness scores for publicly available data in its database section. Completeness assessments performed on gVolante report scores based on not just the coverage of reference genes but also on sequence lengths (e.g. N50 scaffold length), allowing quality control in multiple aspects. Using gVolante, one can compare the quality of original assemblies between their multiple versions (obtained through program choice and parameter tweaking, for example) and evaluate them in comparison to the scores of public resources found in the database section. Availability and implementationgVoalte is freely available at https://gvolante.riken.jp/.

Highlights d RNAPII RPB1-K1268 ubiquitination is essential for transcription recovery and DNA repair d Sequential ubiquitination of RNAPII and UVSSA coordinates the recruitment of TFIIH d Strand-specific ChIPseq enables mapping of RNAPII and reveals genome-wide repair kinetics d RNAPII ubiquitination protects against neurodegeneration phenotype in Cockayne syndrome

Microglia-mediated neuroinflammation has been implicated in the pathogenesis of Alzheimer’s disease (AD). Although microglia in aging and neurodegenerative disease model mice show a loss of homeostatic phenotype and activation of disease-associated microglia (DAM), a correlation between those phenotypes and the degree of neuronal cell loss has not been clarified. In this study, we performed RNA sequencing of microglia isolated from three representative neurodegenerative mouse models, AppNL-G-F/NL-G-F with amyloid pathology, rTg4510 with tauopathy, and SOD1G93A with motor neuron disease by magnetic activated cell sorting. In parallel, gene expression patterns of the human precuneus with early Alzheimer’s change (n = 11) and control brain (n = 14) were also analyzed by RNA sequencing. We found that a substantial reduction of homeostatic microglial genes in rTg4510 and SOD1G93A microglia, whereas DAM genes were uniformly upregulated in all mouse models. The reduction of homeostatic microglial genes was correlated with the degree of neuronal cell loss. In human precuneus with early AD pathology, reduced expression of genes related to microglia- and oligodendrocyte-specific markers was observed, although the expression of DAM genes was not upregulated. Our results implicate a loss of homeostatic microglial function in the progression of AD and other neurodegenerative diseases. Moreover, analyses of human precuneus also suggest loss of microglia and oligodendrocyte functions induced by early amyloid pathology in human.

The phylogenetic position of turtles is a currently controversial issue. Recent molecular studies rejected a traditional view that turtles are basal living reptiles (Hedges, S. B., and L. L. Poling. 1999. A molecular phylogeny. Science 83:998-1001; Kumazawa, Y., and M. Nishida. 1999. Complete mitochondrial DNA sequences of the green turtle and blue-tailed mole skink, statistical evidence for archosaurian affinity of turtles. Mol. Biol. Evol. 16:784-792). Instead, these studies grouped turtles with birds and crocodiles. The relationship among turtles, birds, and crocodiles remained unclear to date. To resolve this issue, we have cloned and sequenced two nuclear genes encoding the catalytic subunit of DNA polymerase alpha and glycinamide ribonucleotide synthetase-aminoimidazole ribonucleotide synthetase-glycinamide ribonucleotide formyltransferase from amniotes and an amphibian. The amino acid sequences of these proteins were subjected to a phylogenetic analysis based on the maximum likelihood method. The resulting tree showed that turtles are the sister group to a monophyletic cluster of archosaurs (birds and crocodiles). All other possible tree topologies were significantly rejected.

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BackgroundRNA-seq enables gene expression profiling in selected spatiotemporal windows and yields massive sequence information with relatively low cost and time investment, even for non-model species. However, there remains a large room for optimizing its workflow, in order to take full advantage of continuously developing sequencing capacity.MethodTranscriptome sequencing for three embryonic stages of Madagascar ground gecko (Paroedura picta) was performed with the Illumina platform. The output reads were assembled de novo for reconstructing transcript sequences. In order to evaluate the completeness of transcriptome assemblies, we prepared a reference gene set consisting of vertebrate one-to-one orthologs.ResultTo take advantage of increased read length of >150 nt, we demonstrated shortened RNA fragmentation time, which resulted in a dramatic shift of insert size distribution. To evaluate products of multiple de novo assembly runs incorporating reads with different RNA sources, read lengths, and insert sizes, we introduce a new reference gene set, core vertebrate genes (CVG), consisting of 233 genes that are shared as one-to-one orthologs by all vertebrate genomes examined (29 species)., The completeness assessment performed by the computational pipelines CEGMA and BUSCO referring to CVG, demonstrated higher accuracy and resolution than with the gene set previously established for this purpose. As a result of the assessment with CVG, we have derived the most comprehensive transcript sequence set of the Madagascar ground gecko by means of assembling individual libraries followed by clustering the assembled sequences based on their overall similarities.ConclusionOur results provide several insights into optimizing de novo RNA-seq workflow, including the coordination between library insert size and read length, which manifested in improved connectivity of assemblies. The approach and assembly assessment with CVG demonstrated here would be applicable to transcriptome analysis of other species as well as whole genome analyses.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-015-2007-1) contains supplementary material, which is available to authorized users.