Convergent Evolution of Hydrogenosomes from Mitochondria by Gene Transfer and Loss.

Hydrogenosomes are H2-producing mitochondrial homologs found in some anaerobic microbial eukaryotes that provide a rare intracellular niche for H2-utilizing endosymbiotic archaea. Among ciliates, anaerobic and aerobic lineages are interspersed, demonstrating that the switch to an anaerobic lifestyle with hydrogenosomes has occurred repeatedly and independently. To investigate the molecular details of this transition, we generated genomic and transcriptomic data sets from anaerobic ciliates representing three distinct lineages. Our data demonstrate that hydrogenosomes have evolved from ancestral mitochondria in each case and reveal different degrees of independent mitochondrial genome and proteome reductive evolution, including the first example of complete mitochondrial genome loss in ciliates. Intriguingly, the FeFe-hydrogenase used for generating H2 has a unique domain structure among eukaryotes and appears to have been present, potentially through a single lateral gene transfer from an unknown donor, in the common aerobic ancestor of all three lineages. The early acquisition and retention of FeFe-hydrogenase helps to explain the facility whereby mitochondrial function can be so radically modified within this diverse and ecologically important group of microbial eukaryotes.

Complex archaea that bridge the gap between prokaryotes and eukaryotes.

The origin of the eukaryotic cell remains one of the most contentious puzzles in modern biology. Recent studies have provided support for the emergence of the eukaryotic host cell from within the archaeal domain of life, but the identity and nature of the putative archaeal ancestor remain a subject of debate. Here we describe the discovery of 'Lokiarchaeota', a novel candidate archaeal phylum, which forms a monophyletic group with eukaryotes in phylogenomic analyses, and whose genomes encode an expanded repertoire of eukaryotic signature proteins that are suggestive of sophisticated membrane remodelling capabilities. Our results provide strong support for hypotheses in which the eukaryotic host evolved from a bona fide archaeon, and demonstrate that many components that underpin eukaryote-specific features were already present in that ancestor. This provided the host with a rich genomic 'starter-kit' to support the increase in the cellular and genomic complexity that is characteristic of eukaryotes.

Confident phylogenetic identification of uncultured prokaryotes through long read amplicon sequencing of the 16S-ITS-23S rRNA operon.

Amplicon sequencing of the 16S rRNA gene is the predominant method to quantify microbial compositions and to discover novel lineages. However, traditional short amplicons often do not contain enough information to confidently resolve their phylogeny. Here we present a cost-effective protocol that amplifies a large part of the rRNA operon and sequences the amplicons with PacBio technology. We tested our method on a mock community and developed a read-curation pipeline that reduces the overall read error rate to 0.18%. Applying our method on four environmental samples, we captured near full-length rRNA operon amplicons from a large diversity of prokaryotes. The method operated at moderately high-throughput (22286-37,850 raw ccs reads) and generated a large amount of putative novel archaeal 23S rRNA gene sequences compared to the archaeal SILVA database. These long amplicons allowed for higher resolution during taxonomic classification by means of long (∼1000 bp) 16S rRNA gene fragments and for substantially more confident phylogenies by means of combined near full-length 16S and 23S rRNA gene sequences, compared to shorter traditional amplicons (250 bp of the 16S rRNA gene). We recommend our method to those who wish to cost-effectively and confidently estimate the phylogenetic diversity of prokaryotes in environmental samples at high throughput.

More than fishing in the dark: PCR of a dispersed sequence produces simple but ultrasensitive Wolbachia detection.

BACKGROUND: Detecting intracellular bacterial symbionts can be challenging when they persist at very low densities. Wolbachia, a widespread bacterial endosymbiont of invertebrates, is particularly challenging. Although it persists at high titers in many species, in others its densities are far below the detection limit of classic end-point Polymerase Chain Reaction (PCR). These low-titer infections can be reliably detected by combining PCR with DNA hybridization, but less elaborate strategies based on end-point PCR alone have proven less sensitive or less general. RESULTS: We introduce a multicopy PCR target that allows fast and reliable detection of A-supergroup Wolbachia--even at low infection titers--with standard end-point PCR. The target is a multicopy motif (designated ARM: A-supergroup repeat motif) discovered in the genome of wMel (the Wolbachia in Drosophila melanogaster). ARM is found in at least seven other Wolbachia A-supergroup strains infecting various Drosophila, the wasp Muscidifurax and the tsetse fly Glossina. We demonstrate that end-point PCR targeting ARM can reliably detect both high- and low-titer Wolbachia infections in Drosophila, Glossina and interspecific hybrids. CONCLUSIONS: Simple end-point PCR of ARM facilitates detection of low-titer Wolbachia A-supergroup infections. Detecting these infections previously required more elaborate procedures. Our ARM target seems to be a general feature of Wolbachia A-supergroup genomes, unlike other multicopy markers such as insertion sequences (IS).

Close encounters of the third domain: the emerging genomic view of archaeal diversity and evolution.

The Archaea represent the so-called Third Domain of life, which has evolved in parallel with the Bacteria and which is implicated to have played a pivotal role in the emergence of the eukaryotic domain of life. Recent progress in genomic sequencing technologies and cultivation-independent methods has started to unearth a plethora of data of novel, uncultivated archaeal lineages. Here, we review how the availability of such genomic data has revealed several important insights into the diversity, ecological relevance, metabolic capacity, and the origin and evolution of the archaeal domain of life.

Novel TPR::ROS1 Fusion Gene Activates MAPK, PI3K and JAK/STAT Signaling in an Infant-type Pediatric Glioma.

BACKGROUND/AIM: Although fusion genes involving the proto-oncogene receptor tyrosine kinase ROS1 are rare in pediatric glioma, targeted therapies with small inhibitors are increasingly being approved for histology-agnostic fusion-positive solid tumors. PATIENT AND METHODS: Here, we present a 16-month-old boy, with a brain tumor in the third ventricle. The patient underwent complete resection but relapsed two years after diagnosis and underwent a second operation. The tumor was initially classified as a low-grade glioma (WHO grade 2); however, methylation profiling suggested the newly WHO-recognized type: infant-type hemispheric glioma. To further refine the molecular background, and search for druggable targets, whole genome (WGS) and whole transcriptome (RNA-Seq) sequencing was performed. RESULTS: Concomitant WGS and RNA-Seq analysis revealed several segmental gains and losses resulting in complex structural rearrangements and fusion genes. Among the top-candidates was a novel TPR::ROS1 fusion, for which only the 3' end of ROS1 was expressed in tumor tissue, indicating that wild type ROS1 is not normally expressed in the tissue of origin. Functional analysis by Western blot on protein lysates from transiently transfected HEK293 cells showed the TPR::ROS1 fusion gene to activate the MAPK-, PI3K- and JAK/STAT- pathways through increased phosphorylation of ERK, AKT, STAT and S6. The downstream pathway activation was also confirmed by immunohistochemistry on tumor tissue slides from the patient. CONCLUSION: We have mapped the activated oncogenic pathways of a novel ROS1-fusion gene and broadened the knowledge of the newly recognized infant-type glioma subtype. The finding facilitates suitable targeted therapies for the patient in case of relapse.

Hikarchaeia demonstrate an intermediate stage in the methanogen-to-halophile transition.

Halobacteria (henceforth: Haloarchaea) are predominantly aerobic halophiles that are thought to have evolved from anaerobic methanogens. This remarkable transformation most likely involved an extensive influx of bacterial genes. Whether it entailed a single massive transfer event or a gradual stream of transfers remains a matter of debate. To address this, genomes that descend from methanogen-to-halophile intermediates are necessary. Here, we present five such near-complete genomes of Marine Group IV archaea (Hikarchaeia), the closest known relatives of Haloarchaea. Their inclusion in gene tree-aware ancestral reconstructions reveals an intermediate stage that had already lost a large number of genes, including nearly all of those involved in methanogenesis and the Wood-Ljungdahl pathway. In contrast, the last Haloarchaea common ancestor gained a large number of genes and expanded its aerobic respiration and salt/UV resistance gene repertoire. Our results suggest that complex and gradual patterns of gain and loss shaped the methanogen-to-halophile transition.

Genomes of two archaeal endosymbionts show convergent adaptations to an intracellular lifestyle.

Endosymbiosis is a widespread phenomenon in the microbial world and can be based on diverse interactions between endosymbiont and host cell. The vast majority of the known endosymbiotic interactions involve bacteria that have invaded eukaryotic host cells. However, methanogenic archaea have been found to thrive in anaerobic, hydrogenosome-containing protists and it was suggested that this symbiosis is based on the transfer of hydrogen. Here, we used culture-independent genomics approaches to sequence the genomes of two distantly related methanogenic endosymbionts that have been acquired in two independent events by closely related anaerobic ciliate hosts Nyctotherus ovalis and Metopus contortus, respectively. The sequences obtained were then validated as originating from the ciliate endosymbionts by in situ probing experiments. Comparative analyses of these genomes and their closest free-living counterparts reveal that the genomes of both endosymbionts are in an early stage of adaptation towards endosymbiosis as evidenced by the large number of genes undergoing pseudogenization. For instance, the observed loss of genes involved in amino acid biosynthesis in both endosymbiont genomes indicates that the endosymbionts rely on their hosts for obtaining several essential nutrients. Furthermore, the endosymbionts appear to have gained significant amounts of genes of potentially secreted proteins, providing targets for future studies aiming to elucidate possible mechanisms underpinning host-interactions. Altogether, our results provide the first genomic insights into prokaryotic endosymbioses from the archaeal domain of life.

Genomic inference of the metabolism of cosmopolitan subsurface Archaea, Hadesarchaea.

The subsurface biosphere is largely unexplored and contains a broad diversity of uncultured microbes(1). Despite being one of the few prokaryotic lineages that is cosmopolitan in both the terrestrial and marine subsurface(2-4), the physiological and ecological roles of SAGMEG (South-African Gold Mine Miscellaneous Euryarchaeal Group) Archaea are unknown. Here, we report the metabolic capabilities of this enigmatic group as inferred from genomic reconstructions. Four high-quality (63-90% complete) genomes were obtained from White Oak River estuary and Yellowstone National Park hot spring sediment metagenomes. Phylogenomic analyses place SAGMEG Archaea as a deeply rooting sister clade of the Thermococci, leading us to propose the name Hadesarchaea for this new Archaeal class. With an estimated genome size of around 1.5 Mbp, the genomes of Hadesarchaea are distinctly streamlined, yet metabolically versatile. They share several physiological mechanisms with strict anaerobic Euryarchaeota. Several metabolic characteristics make them successful in the subsurface, including genes involved in CO and H2 oxidation (or H2 production), with potential coupling to nitrite reduction to ammonia (DNRA). This first glimpse into the metabolic capabilities of these cosmopolitan Archaea suggests they are mediating key geochemical processes and are specialized for survival in the subsurface biosphere.

An archaeal origin for the actin cytoskeleton: Implications for eukaryogenesis.

A hallmark of the eukaryotic cell is the actin cytoskeleton, involved in a wide array of processes ranging from shape determination and phagocytosis to intracellular transport and cytokinesis. Recently, we reported the discovery of an actin-based cytoskeleton also in Archaea. The archaeal actin ortholog, Crenactin, was shown to belong to a conserved operon, Arcade (actin-related cytoskeleton in Archaea involved in shape determination), encoding an additional set of cytoskeleton-associated proteins. Here, we elaborate on the implications of these findings for the evolutionary relation between archaea and eukaryotes, with particular focus on the possibility that eukaryotic actin and actin-related proteins have evolved from an ancestral archaeal actin gene. Archaeal actin could thus have played an important role in cellular processes essential for the origin and early evolution of the eukaryotic lineage. Further exploration of uncharacterized archaeal lineages is necessary to find additional missing pieces in the evolutionary trajectory that ultimately gave rise to present-day organisms.