BackgroundHansenula polymorpha DL1 is a methylotrophic yeast, widely used in fundamental studies of methanol metabolism, peroxisome biogenesis and function, and also as a microbial cell factory for production of recombinant proteins and metabolic engineering towards the goal of high temperature ethanol production.ResultsWe have sequenced the 9 Mbp H. polymorpha DL1 genome and performed whole-genome analysis for the H. polymorpha transcriptome obtained from both methanol- and glucose-grown cells. RNA-seq analysis revealed the complex and dynamic character of the H. polymorpha transcriptome under the two studied conditions, identified abundant and highly unregulated expression of 40% of the genome in methanol grown cells, and revealed alternative splicing events. We have identified subtelomerically biased protein families in H. polymorpha, clusters of LTR elements at G + C-poor chromosomal loci in the middle of each of the seven H. polymorpha chromosomes, and established the evolutionary position of H. polymorpha DL1 within a separate yeast clade together with the methylotrophic yeast Pichia pastoris and the non-methylotrophic yeast Dekkera bruxellensis. Intergenome comparisons uncovered extensive gene order reshuffling between the three yeast genomes. Phylogenetic analyses enabled us to reveal patterns of evolution of methylotrophy in yeasts and filamentous fungi.ConclusionsOur results open new opportunities for in-depth understanding of many aspects of H. polymorpha life cycle, physiology and metabolism as well as genome evolution in methylotrophic yeasts and may lead to novel improvements toward the application of H. polymorpha DL-1 as a microbial cell factory.
A metagenomic analysis of the dynamic changes of the composition of the intestinal microbiome of five participants of the MARS-500 experiment was performed. DNA samples were isolated from the feces of the participants taken just before the experiment, upon 14, 30, 210, 363 and 510 days of isolation in the experimental module, and two weeks upon completion of the experiment. The taxonomic composition of the microbiome was analyzed by pyrosequencing of 16S rRNA gene fragments. Both the taxonomic and functional gene content of the microbiome of one participant were analyzed by whole metagenome sequencing using the SOLiD technique. Each participant had a specific microbiome that could be assigned to one of three recognized enterotypes. Two participants had enterotype I microbiomes characterized by the prevalence of Bacteroides, while the microbiomes of two others, assigned to type II, were dominated by Prevotella. One participant had a microbiome of mixed type. It was found that (1) changes in the taxonimic composition of the microbiomes occurred in the course of the experiment, but the enterotypes remained the same; (2) significant changes in the compositions of the microbiomes occurred just 14-30 days after the beginning of the experiment, presumably indicating the influence of stress factors in the first stage of the experiment; (3) a tendency toward a reversion of the microbiomes to their initial composition was observed two weeks after the end of the experiment, but complete recovery was not achieved. The metagenomic analysis of the microbiome of one of the participants showed that in spite of variations in the taxonomic compositions of microbiomes, the “functional” genetic composition was much more stable for most of the functional gene categories. Probably in the course of the experiment the taxonomic composition of the gut microbiome was adaptively changed to reflect the individual response to the experimental conditions. A new, balanced taxonomic composition of the microbiome was formed to ensure a stable gene content of the community as a whole without negative consequences for the health of the participants.
Two new thermophilic branched chain amino acid transaminases have been identified within the genomes of different hyper-thermophilic archaea, Geoglobus acetivorans, and Archaeoglobus fulgidus. These enzymes belong to the class IV of transaminases as defined by their structural fold. The enzymes have been cloned and over-expressed in Escherichia coli and the recombinant enzymes have been characterized both biochemically and structurally. Both enzymes showed high thermostability with optimal temperature for activity at 80 and 85°C, respectively. They retain good activity after exposure to 50% of the organic solvents, ethanol, methanol, DMSO and acetonitrile. The enzymes show a low activity to (R)-methylbenzylamine but no activity to (S)-methylbenzylamine. Both enzymes have been crystallized and their structures solved in the internal aldimine form, to 1.9 Å resolution for the Geoglobus enzyme and 2.0 Å for the Archaeoglobus enzyme. Also the Geoglobus enzyme structure has been determined in complex with the amino acceptor α-ketoglutarate and the Archaeoglobus enzyme in complex with the inhibitor gabaculine. These two complexes have helped to determine the conformation of the enzymes during enzymatic turnover and have increased understanding of their substrate specificity. A comparison has been made with another (R) selective class IV transaminase from the fungus Nectria haematococca which was previously studied in complex with gabaculine. The subtle structural differences between these enzymes has provided insight regarding their different substrate specificities.
Complete chloroplast genome of diatom alga Synedra acus possesses canonical quadripartite structure with two inverted repeats containing ribosomal RNA gene loci that separate small and large single-copy regions. Chloroplast genome maps as a circular molecule of 116 251 bp. It encodes 27 tRNAs, three rRNAs, two small RNA genes, and 128 protein-coding genes. Comparison of the genic features across diatom chloroplast genomes reveals the absence of an overlap between atpD and atpF gene coding sequences that is present in other plastid genomes of diatom origin. This feature is a clear synapomorphy of S. acus plastid genome that is likely a result of either relaxed constraints or extensive selection forces acting upon atpF gene. We also characterized nuclear-encoded acyl-carrier protein gene with chloroplastic targeting in S. acus. The transfer of acpp gene into the nuclear host genome is hypothesized to have occurred independently in several lineages of diatoms.
Sclerotinia borealis is a necrotrophic phytopathogenic fungus notable for its wide host range and environmental persistence. It grows at low temperatures, causing snow mold disease of crop plants. To understand the molecular mechanisms of its pathogenesis and adaptation to the psychrophilic lifestyle, we determined the 39.3-Mb draft genome sequence of S. borealis F-4128.
Methanotrophic verrucomicrobia of the order Methylacidiphilales are known as extremely acidophilic, thermophilic or mesophilic bacteria that inhabit acidic geothermal ecosystems. The occurrence of verrucomicrobial methanotrophs in other types of acidic environments remains an open question. Notably, Methylacidiphilales-affiliated 16S rRNA gene sequences are commonly retrieved from acidic (pH 3.5–5.5) peatlands. In this study, we compared the patterns of verrucomicrobial diversity in four acidic raised bogs and six neutral fens located in European North Russia. Methylacidiphilales-like 16S rRNA gene reads displaying 83–86% similarity to 16S rRNA gene sequences of currently described verrucomicrobial methanotrophs were recovered exclusively from raised bogs. Laboratory incubation of peat samples with 10% methane for 3 weeks resulted in the pronounced increase of a relative abundance of alphaproteobacterial methanotrophs, while no response was detected for Methylacidiphilales-affiliated bacteria. Three metagenome-assembled genomes (MAGs) of peat-inhabiting Methylacidiphilales bacteria were reconstructed and examined for the presence of genes encoding methane monooxygenase enzymes and autotrophic carbon fixation pathways. None of these genomic determinants were detected in assembled MAGs. Metabolic reconstructions predicted a heterotrophic metabolism, with a potential to hydrolyze several plant-derived polysaccharides. As suggested by our analysis, peat-inhabiting representatives of the Methylacidiphilales are acidophilic aerobic heterotrophs, which comprise a sister family of the methanotrophic Methylacidiphilaceae.
The vast majority of multicellular organisms coexist with bacterial symbionts that may play various roles during their life cycle. Parasitoid wasp Megaphragma amalphitanum (Hymenoptera: Trichogrammatidae) belongs to the smallest known insects whose size is comparable with some bacteria. Using 16S rRNA gene sequencing and Whole Genome Sequencing (WGS), we described microbiota diversity for this arthropod and its potential impact on their lifecycle. Metagenomic sequences were deposited to SRA database which is available at NCBI with accession number SRX2363723 and SRX2363724. We found that small body size and limited lifespan do not lead to a significant reduction of bacterial symbionts diversity. At the same time, we show here a specific feature of microbiota composition in M. amalphitanum – the absence of the Rickettsiaceae family representatives that are known to cause sex-ratio distortion in arthropods and well represented in other populations of parasitoid wasps.
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