Archaeal Transcription Research Articles

Structural basis of archaeal RNA polymerase transcription elongation and Spt4/5 recruitment.

Archaeal transcription is carried out by a multi-subunit RNA polymerase (RNAP) that is highly homologous in structure and function to eukaryotic RNAP II. Among the set of basal transcription factors, only Spt5 is found in all domains of life, but Spt5 has been shaped during evolution, which is also reflected in the heterodimerization of Spt5 with Spt4 in Archaea and Eukaryotes. To unravel the mechanistic basis of Spt4/5 function in Archaea, we performed structure-function analyses using the archaeal transcriptional machinery of Pyrococcus furiosus (Pfu). We report single-particle cryo-electron microscopy reconstructions of apo RNAP and the archaeal elongation complex (EC) in the absence and presence of Spt4/5. Surprisingly, Pfu Spt4/5 also binds the RNAP in the absence of nucleic acids in a distinct super-contracted conformation. We show that the RNAP clamp/stalk module exhibits conformational flexibility in the apo state of RNAP and that the enzyme contracts upon EC formation or Spt4/5 engagement. We furthermore identified a contact of the Spt5-NGN domain with the DNA duplex that stabilizes the upstream boundary of the transcription bubble and impacts Spt4/5 activity in vitro. This study, therefore, provides the structural basis for Spt4/5 function in archaeal transcription and reveals a potential role beyond the well-described support of elongation.

An archaeal transcription factor EnfR with a novel 'eighth note' fold controls hydrogen production of a hyperthermophilic archaeon Thermococcus onnurineus NA1.

Thermococcus onnurineus NA1, a hyperthermophilic carboxydotrophic archaeon, produces H2 through CO oxidation catalyzed by proteins encoded in a carbon monoxide dehydrogenase (CODH) gene cluster. TON_1525 with a DNA-binding helix-turn-helix (HTH) motif is a putative repressor regulating the transcriptional expression of the codh gene cluster. The T55I mutation in TON_1525 led to enhanced H2 production accompanied by the increased expression of genes in the codh cluster. Here, TON_1525 was demonstrated to be a dimer. Monomeric TON_1525 adopts a novel 'eighth note' symbol-like fold (referred to as 'eighth note' fold regulator, EnfR), and the dimerization mode of EnfR is unique in that it has no resemblance to structures in the Protein Data Bank. According to footprinting and gel shift assays, dimeric EnfR binds to a 36-bp pseudo-palindromic inverted repeat in the promoter region of the codh gene cluster, which is supported by an in silico EnfR/DNA complex model and mutational studies revealing the implication of N-terminal loops as well as HTH motifs in DNA recognition. The DNA-binding affinity of the T55I mutant was lowered by ∼15-fold, for which the conformational change of N-terminal loops is responsible. In addition, transcriptome analysis suggested that EnfR could regulate diverse metabolic processes besides H2 production.

Inhibitory Effects of Ammonia on Archaeal 16S rRNA Transcripts in Thermophilic Anaerobic Digester Sludge

High temperatures exacerbate the ammonia inhibition of anaerobic digestion coupled with methanogenesis. The inhibition of methane production by ammonia has been observed in other studies. However, the underlying mechanism is not well understood and requires further investigation. This study explored the effect of ammonia stress on archaeal 16S rRNA transcripts in thermophilic anaerobic digester sludge. Different ammonium concentrations were checked for their influence on the methanogenic rate and hydrogen accumulation. Quantitative PCR was used to compare the changes in total archaeal 16S rRNA expression. A Monte Carlo permutation test within redundancy analysis (RDA) was adopted for exploring the relationship between environmental variables and archaeal 16S rRNA and their transcripts. The results showed that with the increase in ammonium concentration, the methanogenic rate decreased and hydrogen accumulation occurred. The total archaeal 16S rRNA genes and transcripts copy numbers decreased significantly in treatments with higher ammonium concentrations (7 and 10 g NH4+-N/L), but did not change much at lower ammonia concentrations (3 g NH4+-N/L) compared with the 0 g NH4+-N/L treatment. The RDA analysis further revealed that most environmental variables, including ammonia and methane, except for formate, were significantly correlated with the community structure activity of archaeal 16S rRNA transcripts rather than the community structure of their genes. The composition of archaeal 16S rRNA transcripts showed that the hydrogenotrophic methanogen Methanothermobacter dominated the methanogenic community activity in all incubations. It exhibited sensitivity to ammonia stress and should be responsible for the methanogenic inhibition under thermophilic conditions. Our findings suggested that archaeal 16S rRNA transcripts, rather than 16S rRNA genes, are key indicators of ammonia stress and methanogenic activity.

Using metatranscriptomics to better understand the role of microbial nitrogen cycling in coastal sediment benthic flux denitrification efficiency.

Spatial and temporal variability in benthic flux denitrification efficiency occurs across Port Phillip Bay, Australia. Here, we assess the capacity for untargeted metatranscriptomics to resolve spatiotemporal differences in the microbial contribution to benthic nitrogen cycling. The most abundant sediment transcripts assembled were associated with the archaeal nitrifier Nitrosopumilus. In sediments close to external inputs of organic nitrogen, the dominant transcripts were associated with Nitrosopumilus nitric oxide nitrite reduction (nirK). The environmental conditions close to organic nitrogen inputs that select for increased transcription in Nitrosopumilus (amoCAB, nirK, nirS, nmo, hcp) additionally selected for increased transcription of bacterial nitrite reduction (nxrB) and transcripts associated with anammox (hzo) but not denitrification (bacterial nirS/nirk). In sediments that are more isolated from external inputs of organic nitrogen dominant transcripts were associated with nitrous oxide reduction (nosZ) and changes in nosZ transcript abundance were uncoupled from transcriptional profiles associated with archaeal nitrification. Coordinated transcription of coupled community-level nitrification-denitrification was not well supported by metatranscriptomics. In comparison, the abundance of archaeal nirK transcripts were site- and season-specific. This study indicates that the transcription of archaeal nirK in response to changing environmental conditions may be an important and overlooked feature of coastal sediment nitrogen cycling.

Potential contributions of nitrifiers and denitrifiers to nitrous oxide sources and sinks in China's estuarine and coastal areas

Abstract. Nitrous oxide (N2O) is an important ozone-depleting greenhouse gas produced and consumed by microbially mediated nitrification and denitrification pathways. Estuaries are intensive N2O emission regions in marine ecosystems. However, the potential contributions of nitrifiers and denitrifiers to N2O sources and sinks in China's estuarine and coastal areas are poorly understood. The abundance and transcription of six key microbial functional genes involved in nitrification and denitrification, as well as the clade II-type nosZ gene-bearing community composition of N2O reducers, were investigated in four estuaries spanning the Chinese coastline. The results showed that the ammonia-oxidizing archaeal amoA genes and transcripts were more dominant in the northern Bohai Sea (BS) and Yangtze River estuaries, which had low nitrogen concentrations, while the denitrifier nirS genes and transcripts were more dominant in the southern Jiulong River (JRE) and Pearl River estuaries, which had high levels of terrestrial nitrogen input. Notably, the nosZ clade II gene was more abundant than the clade I-type throughout the estuaries except for in the JRE and a few sites of the BS, while the opposite transcript distribution pattern was observed in these two estuaries. The gene and transcript distributions were significantly constrained by nitrogen and oxygen concentrations as well as by salinity, temperature, and pH. The nosZ clade II gene-bearing community composition along China's coastline had a high level of diversity and was distinctly different from that in the soil and in marine oxygen-minimum-zone waters. By comparing the gene distribution patterns across the estuaries with the distribution patterns of the N2O concentration and flux, we found that denitrification may principally control the N2O emissions pattern.

The structure and activities of the archaeal transcription termination factor Eta detail vulnerabilities of the transcription elongation complex.

Transcription must be properly regulated to ensure dynamic gene expression underlying growth, development, and response to environmental cues. Regulation is imposed throughout the transcription cycle, and while many efforts have detailed the regulation of transcription initiation and early elongation, the termination phase of transcription also plays critical roles in regulating gene expression. Transcription termination can be driven by only a few proteins in each domain of life. Detailing the mechanism(s) employed provides insight into the vulnerabilities of transcription elongation complexes (TECs) that permit regulated termination to control expression of many genes and operons. Here, we describe the biochemical activities and crystal structure of the superfamily 2 helicase Eta, one of two known factors capable of disrupting archaeal transcription elongation complexes. Eta retains a twin-translocase core domain common to all superfamily 2 helicases and a well-conserved C terminus wherein individual amino acid substitutions can critically abrogate termination activities. Eta variants that perturb ATPase, helicase, single-stranded DNA and double-stranded DNA translocase and termination activities identify key regions of the C terminus of Eta that, when combined with modeling Eta-TEC interactions, provide a structural model of Eta-mediated termination guided in part by structures of Mfd and the bacterial TEC. The susceptibility of TECs to disruption by termination factors that target the upstream surface of RNA polymerase and potentially drive termination through forward translocation and allosteric mechanisms that favor opening of the clamp to release the encapsulated nucleic acids emerges as a common feature of transcription termination mechanisms.

Diet and Host Genetics Drive the Bacterial and Fungal Intestinal Metatranscriptome of Gilthead Sea Bream.

The gut microbiota is now recognised as a key target for improving aquaculture profit and sustainability, but we still lack insights into the activity of microbes in fish mucosal surfaces. In the present study, a metatranscriptomic approach was used to reveal the expression of gut microbial genes in the farmed gilthead sea bream. Archaeal and viral transcripts were a minority but, interestingly and contrary to rRNA amplicon-based studies, fungal transcripts were as abundant as bacterial ones, and increased in fish fed a plant-enriched diet. This dietary intervention also drove a differential metatranscriptome in fish selected for fast and slow growth. Such differential response reinforced the results of previously inferred metabolic pathways, enlarging, at the same time, the catalogue of microbial functions in the intestine. Accordingly, vitamin and amino acid metabolism, and rhythmic and symbiotic processes were mostly shaped by bacteria, whereas fungi were more specifically configuring the host immune, digestive, or endocrine processes.

Engineering transcriptional regulation in Escherichia coli using an archaeal TetR-family transcription factor.

Synthetic biology requires well-characterized biological parts that can be combined into functional modules. One type of biological parts are transcriptional regulators and their cognate operator elements, which enable to either generate an input-specific response or are used as actuator modules. A range of regulators has already been characterized and used for orthogonal gene expression engineering, however, previous efforts have mostly focused on bacterial regulators. This work aims to design and explore the use of an archaeal TetR family regulator, FadRSa from Sulfolobus acidocaldarius, in a bacterial system, namely Escherichia coli. This is a challenging objective given the fundamental difference between the bacterial and archaeal transcription machinery and the lack of a native TetR-like FadR regulatory system in E. coli. The synthetic σ70-dependent bacterial promoter proD was used as a starting point to design hybrid bacterial/archaeal promoter/operator regions, in combination with the mKate2 fluorescent reporter enabling a readout. Four variations of proD containing FadRSa binding sites were constructed and characterized. While expressional activity of the modified promoter proD was found to be severely diminished for two of the constructs, constructs in which the binding site was introduced adjacent to the -35 promoter element still displayed sufficient basal transcriptional activity and showed up to 7-fold repression upon expression of FadRSa. Addition of acyl-CoA has been shown to disrupt FadRSa binding to the DNA in vitro. However, extracellular concentrations of up to 2mM dodecanoate, subsequently converted to acyl-CoA by the cell, did not have a significant effect on repression in the bacterial system. This work demonstrates that archaeal transcription regulators can be used to generate actuator elements for use in E. coli, although the lack of ligand response underscores the challenge of maintaining biological function when transferring parts to a phylogenetically divergent host.

ChIP-Seq Occupancy Mapping of the Archaeal Transcription Machinery.

Genome-wide occupancy studies for RNA polymerases and their basal transcription factors deliver information about transcription dynamics and the recruitment of transcription elongation and termination factors in eukaryotes and prokaryotes. The primary method to determine genome-wide occupancies is chromatin immunoprecipitation combined with deep sequencing (ChIP-seq). Archaea possess a transcription machinery that is evolutionarily closer related to its eukaryotic counterpart but it operates in a prokaryotic cellular context. Studies on archaeal transcription brought insight into the evolution of transcription machineries and the universality of transcription mechanisms. Because of the limited resolution of ChIP-seq, the close spacing of promoters and transcription units found in archaeal genomes pose a challenge for ChIP-seq and the ensuing data analysis. The extreme growth temperature of many established archaeal model organisms necessitates further adaptations. This chapter describes a version of ChIP-seq adapted for the basal transcription machinery of thermophilic archaea and some modifications to the data analysis.

ACPSF1 cooperates with terminator U-tract to dictate archaeal transcription termination efficacy.

Recently, aCPSF1 was reported to function as the long-sought global transcription termination factor of archaea; however, the working mechanism remains elusive. This work, through analyzing transcript-3'end-sequencing data of Methanococcus maripaludis, found genome-wide positive correlations of both the terminator uridine(U)-tract and aCPSF1 with hierarchical transcription termination efficacies (TTEs). In vitro assays determined that aCPSF1 specifically binds to the terminator U-tract with U-tract number-related binding affinity, and in vivo assays demonstrated the two elements are indispensable in dictating high TTEs, revealing that aCPSF1 and the terminator U-tract cooperatively determine high TTEs. The N-terminal KH domains equip aCPSF1 with specific-binding capacity to terminator U-tract and the aCPSF1-terminator U-tract cooperation; while the nuclease activity of aCPSF1 was also required for TTEs. aCPSF1 also guarantees the terminations of transcripts with weak intrinsic terminator signals. aCPSF1 orthologs from Lokiarchaeota and Thaumarchaeota exhibited similar U-tract cooperation in dictating TTEs. Therefore, aCPSF1 and the intrinsic U-rich terminator could work in a noteworthy two-in-one termination mode in archaea, which may be widely employed by archaeal phyla; using one trans-action factor to recognize U-rich terminator signal and cleave transcript 3'-end, the archaeal aCPSF1-dependent transcription termination may represent a simplified archetypal mode of the eukaryotic RNA polymerase II termination machinery.

Taxon-Specific Shifts in Bacterial and Archaeal Transcription of Dissolved Organic Matter Cycling Genes in a Stratified Fjord

ABSTRACTA considerable fraction of organic matter derived from photosynthesis in the euphotic zone settles into the ocean’s interior and, as it progresses, is degraded by diverse microbial consortia that utilize a suite of extracellular enzymes and membrane transporters. Still, the molecular details that regulate carbon cycling across depths remain little explored. As stratification in fjords has made them attractive models to explore patterns in biological oceanography, we here analyzed bacterial and archaeal transcription in samples from five depth layers in the Gullmar Fjord, Sweden. Transcriptional variation over depth correlated with gradients in chlorophyll a and nutrient concentrations. Differences in transcription between sampling dates (summer and early autumn) were strongly correlated with ammonium concentrations, which potentially was linked with a stronger influence of (micro-)zooplankton grazing in summer. Transcriptional investment in carbohydrate-active enzymes (CAZymes) decreased with depth and shifted toward peptidases, partly a result of elevated CAZyme transcription by Flavobacteriales, Cellvibrionales, and Synechococcales at 2 to 25 m and a dominance of peptidase transcription by Alteromonadales and Rhodobacterales from 50 m down. In particular, CAZymes for chitin, laminarin, and glycogen were important. High levels of transcription of ammonium transporter genes by Thaumarchaeota at depth (up to 18% of total transcription), along with the genes for ammonia oxidation and CO2 fixation, indicated that chemolithoautotrophy contributed to the carbon flux in the fjord. The taxon-specific expression of functional genes for processing of the marine pool of dissolved organic matter and inorganic nutrients across depths emphasizes the importance of different microbial foraging mechanisms over spatiotemporal scales for shaping biogeochemical cycles.IMPORTANCE It is generally recognized that stratification in the ocean strongly influences both the community composition and the distribution of ecological functions of microbial communities, which in turn are expected to shape the biogeochemical cycling of essential elements over depth. Here, we used metatranscriptomics analysis to infer molecular detail on the distribution of gene systems central to the utilization of organic matter in a stratified marine system. We thereby uncovered that pronounced shifts in the transcription of genes encoding CAZymes, peptidases, and membrane transporters occurred over depth among key prokaryotic orders. This implies that sequential utilization and transformation of organic matter through the water column is a key feature that ultimately influences the efficiency of the biological carbon pump.

Single-Molecule/Cell Analyses Reveal Principles of Genome-Folding Mechanisms in the Three Domains of Life.

Comparative structural/molecular biology by single-molecule analyses combined with single-cell dissection, mass spectroscopy, and biochemical reconstitution have been powerful tools for elucidating the mechanisms underlying genome DNA folding. All genomes in the three domains of life undergo stepwise folding from DNA to 30–40 nm fibers. Major protein players are histone (Eukarya and Archaea), Alba (Archaea), and HU (Bacteria) for fundamental structural units of the genome. In Euryarchaeota, a major archaeal phylum, either histone or HTa (the bacterial HU homolog) were found to wrap DNA. This finding divides archaea into two groups: those that use DNA-wrapping as the fundamental step in genome folding and those that do not. Archaeal transcription factor-like protein TrmBL2 has been suggested to be involved in genome folding and repression of horizontally acquired genes, similar to bacterial H-NS protein. Evolutionarily divergent SMC proteins contribute to the establishment of higher-order structures. Recent results are presented, including the use of Hi-C technology to reveal that archaeal SMC proteins are involved in higher-order genome folding, and the use of single-molecule tracking to reveal the detailed functions of bacterial and eukaryotic SMC proteins. Here, we highlight the similarities and differences in the DNA-folding mechanisms in the three domains of life.

Enhanced H2 production by deletion of the Tfx family DNA-binding protein in the hyperthermophilic archaeon Thermococcus onnurineus NA1

A gene encoding an archaeal transcription regulator (TON_1525) homologous to the Tfx family DNA-binding protein has been identified to affect gene expression of carbon monoxide metabolism in the hyperthermophilic archaeon Thermococcus onnurineus NA1. To broadly understand gene regulation by the TON_1525 gene, a gene deletion mutant (Δ1525) was constructed and transcriptome changes were investigated. A total of 351 genes were found to be differentially expressed in the Δ1525 mutant. The expression levels of the hydrogenase genes were significantly changed; mbh (membrane-bound hydrogenase) and mch (membrane-bound carbon monoxide-dependent hydrogenase) genes were increased, and mfh2 (membrane-bound formate-dependent hydrogenase) and soluble hydrogenase genes were decreased. The Δ1525 mutant was cultured in a medium supplemented with maltodextrin or carbon monoxide to investigate whether changes in gene expression lead to physiological changes. The maximum H2 production rates of the Δ1525 mutant were significantly enhanced on both substrates compared to the wild-type strain. These results indicate that the TON_1525 gene can affect the expression of various genes including mbh and mch, consequently regulating the metabolism of T. onnurineus NA1. This study expands our understanding of the functional role of the Tfx family DNA-binding proteins.

Early Evolution of Transcription Systems and Divergence of Archaea and Bacteria.

DNA template-dependent multi-subunit RNA polymerases (RNAPs) found in all three domains of life and some viruses are of the two-double-Ψ-β-barrel (DPBB) type. The 2-DPBB protein format is also found in some RNA template-dependent RNAPs and a major replicative DNA template-dependent DNA polymerase (DNAP) from Archaea (PolD). The 2−DPBB family of RNAPs and DNAPs probably evolved prior to the last universal common cellular ancestor (LUCA). Archaeal Transcription Factor B (TFB) and bacterial σ factors include homologous strings of helix-turn-helix units. The consequences of TFB-σ homology are discussed in terms of the evolution of archaeal and bacterial core promoters. Domain-specific DPBB loop inserts functionally connect general transcription factors to the RNAP active site. Archaea appear to be more similar to LUCA than Bacteria. Evolution of bacterial σ factors from TFB appears to have driven divergence of Bacteria from Archaea, splitting the prokaryotic domains.

Single-Molecule Fret Analysis of Key Protein Conformational Changes During Promoter Escape by RNA Polymerase

Bacterial RNA Polymerases (RNAPs) bind to transcription initiation factors called σ factors to start promoter-specific transcription. Within the σ factor lies a highly conserved structural module, the “σ-finger”, which forms a loop that protrudes towards the RNAP active-center and that interacts with the template strand DNA. The close proximity of the σ-finger to the “heart” of transcription is implicated in the pre-organisation of the template strand DNA, in the synthesis of the first short RNAs, and in the pausing of transcription upon synthesis of a 6-mer RNA. However, the σ-finger also blocks entry of the nascent RNA to the RNAP exit channel and must be displaced during initial transcription as the first step in promoter escape. Despite recent structural studies, σ-finger conformational changes during late transcription initiation and promoter escape are still unknown. Here we report a novel single-molecule FRET ruler that monitors the conformational dynamics of the E.coli σ70-finger from the early stages of transcription initiation to promoter escape. Our results using the σ-finger FRET ruler on transcription complexes formed on a derivative of the lac promoter show that the σ-finger adopts three conformations, which interconvert with rates 0.01-1.5s−1. Intriguingly, we find the σ-finger first displaced in complexes trapped in synthesis of abortive transcripts up to a 10-mer RNA. This observation is likely driven by collision of the σ-finger with the 5’-end of the growing nascent RNA. Based on our results, we propose a new model describing the relation of the conformational changes in the σ-finger to the dynamics of transcription initiation and promoter escape. Archaeal and eukaryotic transcription initiation complexes also contain structural modules that block the RNAP exit channel, so it is therefore likely that this model applies to all kingdoms of life.

The conserved ribonuclease aCPSF1 triggers genome-wide transcription termination of Archaea via a 3'-end cleavage mode.

Transcription termination defines accurate transcript 3′-ends and ensures programmed transcriptomes, making it critical to life. However, transcription termination mechanisms remain largely unknown in Archaea. Here, we reported the physiological significance of the newly identified general transcription termination factor of Archaea, the ribonuclease aCPSF1, and elucidated its 3′-end cleavage triggered termination mechanism. The depletion of Mmp-aCPSF1 in Methanococcus maripaludis caused a genome-wide transcription termination defect and disordered transcriptome. Transcript-3′end-sequencing revealed that transcriptions primarily terminate downstream of a uridine-rich motif where Mmp-aCPSF1 performed an endoribonucleolytic cleavage, and the endoribonuclease activity was determined to be essential to the in vivo transcription termination. Co-immunoprecipitation and chromatin-immunoprecipitation detected interactions of Mmp-aCPSF1 with RNA polymerase and chromosome. Phylogenetic analysis revealed that the aCPSF1 orthologs are ubiquitously distributed among the archaeal phyla, and two aCPSF1 orthologs from Lokiarchaeota and Thaumarchaeota could replace Mmp-aCPSF1 to terminate transcription of M. maripaludis. Therefore, the aCPSF1 dependent termination mechanism could be widely employed in Archaea, including Lokiarchaeota belonging to Asgard Archaea, the postulated archaeal ancestor of Eukaryotes. Strikingly, aCPSF1-dependent archaeal transcription termination reported here exposes a similar 3′-cleavage mode as the eukaryotic RNA polymerase II termination, thus would shed lights on understanding the evolutionary linking between archaeal and eukaryotic termination machineries.

Prokaryotic sigma factors and their transcriptional counterparts in Archaea and Eukarya.

RNA polymerases (RNAPs) carry out transcription in the three domains of life, Bacteria, Archaea, and Eukarya. Transcription initiation is highly regulated by a variety of transcription factors, whose number and subunit complexity increase during evolution. This process is regulated in Bacteria by the σ factor, while the three eukaryotic RNAPs require a complex set of transcription factors (TFs) and a TATA-binding protein (TBP). The archaeal transcription system appears to be an ancestral version of the eukaryotic RNAPII, requiring transcription factor B (TFB), TBP, and transcription factor E (TFE). The function of the bacterial sigma (σ) factor has been correlated to the roles played by the eukaryotic RNAP II and the archaeal RNAP. In addition, σ factors, TFB, and TFIIB all contain multiple DNA binding helix-turn-helix (HTH) structural motifs; although TFIIB and TFB display two HTH domains, while the bacterial σ factor spans 4 HTH motifs. The sequence similarities and structure alignments of the bacterial σ factor, eukaryotic TFIIB, and archaeal TFB evidence that these three proteins are homologs.Key Points• Transcription initiation is highly regulated by TFs.• Transcription is finely regulated in all domains of life by different sets of TFs.• Specific TFs in Bacteria, Eukarya and Archaea are homologs.

Spatial variations in bacterial and archaeal abundance and community composition in boreal forest pine mycorrhizospheres

Mycorrhizal fungi have a strong impact on soil biota. In this study, bacterial and archaeal populations in different parts of Suillus bovinus - Pinus sylvestris mycorrhizospheres in boreal forest were quantified and identified by DNA analysis. The numbers of bacterial and archaeal 16S rRNA gene copies were highest in uncolonized humus and lowest in fruiting bodies. The numbers of bacterial 16S rRNA gene copies varied from 1.3 × 107 to 3.1 × 109 copies g−1 fw and archaeal copies from 4.1 × 107 to 9.6 × 108 copies g−1 fw. The relatively high number of archaeal 16S rRNA gene copies was likely due to the cold and highly organic habitat. The presence of hyphae appeared to further promote archaeal numbers and the archaea:bacteria ratio was over one in samples containing only fungal material. Most detected archaea belonged to terrestrial Thaumarchaeota. Proteobacteria, Actinobacteria and Acidobacteria were predictably the dominating bacterial taxa in the samples with clear trend of Betaproteobacteria preferring the pine root habitats.

Microbial metagenomes and metatranscriptomes during a coastal phytoplankton bloom

Metagenomic and metatranscriptomic time-series data covering a 52-day period in the fall of 2016 provide an inventory of bacterial and archaeal community genes, transcripts, and taxonomy during an intense dinoflagellate bloom in Monterey Bay, CA, USA. The dataset comprises 84 metagenomes (0.8 terabases), 82 metatranscriptomes (1.1 terabases), and 88 16S rRNA amplicon libraries from samples collected on 41 dates. The dataset also includes 88 18S rRNA amplicon libraries, characterizing the taxonomy of the eukaryotic community during the bloom. Accompanying the sequence data are chemical and biological measurements associated with each sample. These datasets will facilitate studies of the structure and function of marine bacterial communities during episodic phytoplankton blooms.

Fecal metatranscriptomics of macaques with idiopathic chronic diarrhea reveals altered mucin degradation and fucose utilization

BackgroundIdiopathic chronic diarrhea (ICD) is a common cause of morbidity and mortality among juvenile rhesus macaques. Characterized by chronic inflammation of the colon and repeated bouts of diarrhea, ICD is largely unresponsive to medical interventions, including corticosteroid, antiparasitic, and antibiotic treatments. Although ICD is accompanied by large disruptions in the composition of the commensal gut microbiome, no single pathogen has been concretely identified as responsible for the onset and continuation of the disease.ResultsFecal samples were collected from 12 ICD-diagnosed macaques and 12 age- and sex-matched controls. RNA was extracted for metatranscriptomic analysis of organisms and functional annotations associated with the gut microbiome. Bacterial, fungal, archaeal, protozoan, and macaque (host) transcripts were simultaneously assessed. ICD-afflicted animals were characterized by increased expression of host-derived genes involved in inflammation and increased transcripts from bacterial pathogens such as Campylobacter and Helicobacter and the protozoan Trichomonas. Transcripts associated with known mucin-degrading organisms and mucin-degrading enzymes were elevated in the fecal microbiomes of ICD-afflicted animals. Assessment of colon sections using immunohistochemistry and of the host transcriptome suggests differential fucosylation of mucins between control and ICD-afflicted animals. Interrogation of the metatranscriptome for fucose utilization genes reveals possible mechanisms by which opportunists persist in ICD. Bacteroides sp. potentially cross-fed fucose to Haemophilus whereas Campylobacter expressed a mucosa-associated transcriptome with increased expression of adherence genes.ConclusionsThe simultaneous profiling of bacterial, fungal, archaeal, protozoan, and macaque transcripts from stool samples reveals that ICD of rhesus macaques is associated with increased gene expression by pathogens, increased mucin degradation, and altered fucose utilization. The data suggest that the ICD-afflicted host produces fucosylated mucins that are leveraged by potentially pathogenic microbes as a carbon source or as adhesion sites.