Origins of major archaeal clades correspond to gene acquisitions from bacteria.

The mechanisms that underlie the origin of major prokaryotic groups are poorly understood. In principle, the origin of both species and higher taxa among prokaryotes should entail similar mechanisms--ecological interactions with the environment paired with natural genetic variation involving lineage-specific gene innovations and lineage-specific gene acquisitions. To investigate the origin of higher taxa in archaea, we have determined gene distributions and gene phylogenies for the 267,568 protein-coding genes of 134 sequenced archaeal genomes in the context of their homologues from 1,847 reference bacterial genomes. Archaeal-specific gene families define 13 traditionally recognized archaeal higher taxa in our sample. Here we report that the origins of these 13 groups unexpectedly correspond to 2,264 group-specific gene acquisitions from bacteria. Interdomain gene transfer is highly asymmetric, transfers from bacteria to archaea are more than fivefold more frequent than vice versa. Gene transfers identified at major evolutionary transitions among prokaryotes specifically implicate gene acquisitions for metabolic functions from bacteria as key innovations in the origin of higher archaeal taxa.

Endosymbiotic origin and differential loss of eukaryotic genes.

Chloroplasts arose from cyanobacteria, mitochondria arose from proteobacteria. Both organelles have conserved their prokaryotic biochemistry, but their genomes are reduced, and most organelle proteins are encoded in the nucleus. Endosymbiotic theory posits that bacterial genes in eukaryotic genomes entered the eukaryotic lineage via organelle ancestors. It predicts episodic influx of prokaryotic genes into the eukaryotic lineage, with acquisition corresponding to endosymbiotic events. Eukaryotic genome sequences, however, increasingly implicate lateral gene transfer, both from prokaryotes to eukaryotes and among eukaryotes, as a source of gene content variation in eukaryotic genomes, which predicts continuous, lineage-specific acquisition of prokaryotic genes in divergent eukaryotic groups. Here we discriminate between these two alternatives by clustering and phylogenetic analysis of eukaryotic gene families having prokaryotic homologues. Our results indicate (1) that gene transfer from bacteria to eukaryotes is episodic, as revealed by gene distributions, and coincides with major evolutionary transitions at the origin of chloroplasts and mitochondria; (2) that gene inheritance in eukaryotes is vertical, as revealed by extensive topological comparison, sparse gene distributions stemming from differential loss; and (3) that continuous, lineage-specific lateral gene transfer, although it sometimes occurs, does not contribute to long-term gene content evolution in eukaryotic genomes.

Metabopolis: scalable network layout for biological pathway diagrams in urban map style.

BACKGROUND: Biological pathways represent chains of molecular interactions in biological systems that jointly form complex dynamic networks. The network structure changes from the significance of biological experiments and layout algorithms often sacrifice low-level details to maintain high-level information, which complicates the entire image to large biochemical systems such as human metabolic pathways. RESULTS: Our work is inspired by concepts from urban planning since we create a visual hierarchy of biological pathways, which is analogous to city blocks and grid-like road networks in an urban area. We automatize the manual drawing process of biologists by first partitioning the map domain into multiple sub-blocks, and then building the corresponding pathways by routing edges schematically, to maintain the global and local context simultaneously. Our system incorporates constrained floor-planning and network-flow algorithms to optimize the layout of sub-blocks and to distribute the edge density along the map domain. We have developed the approach in close collaboration with domain experts and present their feedback on the pathway diagrams based on selected use cases. CONCLUSIONS: We present a new approach for computing biological pathway maps that untangles visual clutter by decomposing large networks into semantic sub-networks and bundling long edges to create space for presenting relationships systematically.

Staphylococcus aureus haem biosynthesis and acquisition pathways are linked through haem monooxygenase IsdG.

Haem is an essential cofactor in central metabolic pathways in the vast majority of living systems. Prokaryotes acquire haem via haem biosynthesis pathways, and some also utilize haem uptake systems, yet it remains unclear how they balance haem requirements with the paradox that free haem is toxic. Here, using the model pathogen Staphylococcus aureus, we report that IsdG, one of two haem oxygenase enzymes in the haem uptake system, inhibits the formation of haem via the internal haem biosynthesis route. More specifically, we show that IsdG decreases the activity of ferrochelatase and that the two proteins interact both in vitro and in vivo. Further, a bioinformatics analysis reveals that a significant number of haem biosynthesis pathway containing organisms possess an IsdG-homologue and that those with both biosynthesis and uptake systems have at least two haem oxygenases. We conclude that IsdG-like proteins control intracellular haem levels by coupling the two pathways. IsdG is thus a target for the treatment of S. aureusinfections.

YCF1: A Green TIC?

A pivotal step in the transformation of an endosymbiotic cyanobacterium to a plastid some 1.5 billion years ago was the evolution of a protein import apparatus, the TOC/TIC machinery, in the common ancestor of Archaeplastida. Recently, a putative new TIC member was identified in Arabidopsis thaliana: TIC214. This finding is remarkable for a number of reasons: (1) TIC214 is encoded by ycf1, so it would be the first plastid-encoded protein of this apparatus; (2) ycf1 is unique to the green lineage (Chloroplastida) but entirely lacking in glaucophytes (Glaucophyta) and the red lineage (Rhodophyta) of the Archaeplastida; (3) ycf1 has been shown to be one of the few indispensable plastid genes (aside from the ribosomal machinery), yet it is missing in the grasses; and (4) 30 years of previous TOC/TIC research missed it. These observations prompted us to survey the evolution of ycf1. We found that ycf1 is not only lacking in grasses and some parasitic plants, but also for instance in cranberry (Ericaceae). The encoded YCF proteins are highly variable, both in sequence length and in the predicted number of N-terminal transmembrane domains. The evolution of the TOC/TIC machinery in the green lineage experienced specific modifications, but our analysis does not support YCF1 to be a general green TIC. It remains to be explained how the apparent complete loss of YCF1 can be tolerated by some embryophytes and whether what is observed for YCF1 function in a member of the Brassicaceae is also true for, e.g., algal and noncanonical YCF1 homologs.

Energy for two: New archaeal lineages and the origin of mitochondria.

Metagenomics bears upon all aspects of microbiology, including our understanding of mitochondrial and eukaryote origin. Recently, ribosomal protein phylogenies show the eukaryote host lineage - the archaeal lineage that acquired the mitochondrion - to branch within the archaea. Metagenomic studies are now uncovering new archaeal lineages that branch more closely to the host than any cultivated archaea do. But how do they grow? Carbon and energy metabolism as pieced together from metagenome assemblies of these new archaeal lineages, such as the Deep Sea Archaeal Group (including Lokiarchaeota) and Bathyarchaeota, do not match the physiology of any cultivated microbes. Understanding how these new lineages live in their environment is important, and might hold clues about how mitochondria arose and how the eukaryotic lineage got started. Here we look at these exciting new metagenomic studies, what they say about archaeal physiology in modern environments, how they impact views on host-mitochondrion physiological interactions at eukaryote origin.

One step beyond a ribosome: The ancient anaerobic core.

Life arose in a world without oxygen and the first organisms were anaerobes. Here we investigate the gene repertoire of the prokaryote common ancestor, estimating which genes it contained and to which lineages of modern prokaryotes it was most similar in terms of gene content. Using a phylogenetic approach we found that among trees for all 8779 protein families shared between 134 archaea and 1847 bacterial genomes, only 1045 have sequences from at least two bacterial and two archaeal groups and retain the ancestral archaeal-bacterial split. Among those, the genes shared by anaerobes were identified as candidate genes for the prokaryote common ancestor, which lived in anaerobic environments. We find that these anaerobic prokaryote common ancestor genes are today most frequently distributed among methanogens and clostridia, strict anaerobes that live from low free energy changes near the thermodynamic limit of life. The anaerobic families encompass genes for bifunctional acetyl-CoA-synthase/CO-dehydrogenase, heterodisulfide reductase subunits C and A, ferredoxins, and several subunits of the Mrp-antiporter/hydrogenase family, in addition to numerous S-adenosyl methionine (SAM) dependent methyltransferases. The data indicate a major role for methyl groups in the metabolism of the prokaryote common ancestor. The data furthermore indicate that the prokaryote ancestor possessed a rotor stator ATP synthase, but lacked cytochromes and quinones as well as identifiable redox-dependent ion pumping complexes. The prokaryote ancestor did possess, however, an Mrp-type H(+)/Na(+) antiporter complex, capable of transducing geochemical pH gradients into biologically more stable Na(+)-gradients. The findings implicate a hydrothermal, autotrophic, and methyl-dependent origin of life. This article is part of a Special Issue entitled 'EBEC 2016: 19th European Bioenergetics Conference, Riva del Garda, Italy, July 2-6, 2016', edited by Prof. Paolo Bernardi.

Biochemical fossils of the ancient transition from geoenergetics to bioenergetics in prokaryotic one carbon compound metabolism.

The deep dichotomy of archaea and bacteria is evident in many basic traits including ribosomal protein composition, membrane lipid synthesis, cell wall constituents, and flagellar composition. Here we explore that deep dichotomy further by examining the distribution of genes for the synthesis of the central carriers of one carbon units, tetrahydrofolate (H4F) and tetrahydromethanopterin (H4MPT), in bacteria and archaea. The enzymes underlying those distinct biosynthetic routes are broadly unrelated across the bacterial-archaeal divide, indicating that the corresponding pathways arose independently. That deep divergence in one carbon metabolism is mirrored in the structurally unrelated enzymes and different organic cofactors that methanogens (archaea) and acetogens (bacteria) use to perform methyl synthesis in their H4F- and H4MPT-dependent versions, respectively, of the acetyl-CoA pathway. By contrast, acetyl synthesis in the acetyl-CoA pathway - from a methyl group, CO2 and reduced ferredoxin - is simpler, uniform and conserved across acetogens and methanogens, and involves only transition metals as catalysts. The data suggest that the acetyl-CoA pathway, while being the most ancient of known CO2 assimilation pathways, reflects two phases in early evolution: an ancient phase in a geochemically confined and non-free-living universal common ancestor, in which acetyl thioester synthesis proceeded spontaneously with the help of geochemically supplied methyl groups, and a later phase that reflects the primordial divergence of the bacterial and archaeal stem groups, which independently invented genetically-encoded means to synthesize methyl groups via enzymatic reactions. This article is part of a Special Issue entitled: 18th European Bioenergetic Conference.

Contact angles and wettability of ionic liquids on polar and non-polar surfaces.

Many applications involving ionic liquids (ILs) require the knowledge of their interfacial behaviour, such as wettability and adhesion. In this context, herein, two approaches were combined aiming at understanding the impact of the IL chemical structures on their wettability on both polar and non-polar surfaces, namely: (i) the experimental determination of the contact angles of a broad range of ILs (covering a wide number of anions of variable polarity, cations, and cation alkyl side chain lengths) on polar and non-polar solid substrates (glass, Al-plate, and poly-(tetrafluoroethylene) (PTFE)); and (ii) the correlation of the experimental contact angles with the cation-anion pair interaction energies generated by the Conductor-like Screening Model for Real Solvents (COSMO-RS). The combined results reveal that the hydrogen-bond basicity of ILs, and thus the IL anion, plays a major role through their wettability on both polar and non-polar surfaces. The increase of the IL hydrogen-bond accepting ability leads to an improved wettability of more polar surfaces (lower contact angles) while the opposite trend is observed on non-polar surfaces. The cation nature and alkyl side chain lengths have however a smaller impact on the wetting ability of ILs. Linear correlations were found between the experimental contact angles and the cation-anion hydrogen-bonding and cation ring energies, estimated using COSMO-RS, suggesting that these features primarily control the wetting ability of ILs. Furthermore, two-descriptor correlations are proposed here to predict the contact angles of a wide variety of ILs on glass, Al-plate, and PTFE surfaces. A new extended list is provided for the contact angles of ILs on three surfaces, which can be used as a priori information to choose appropriate ILs before a given application.

Fusion/fission protein family identification in Archaea.

The majority of newly discovered archaeal lineages remain without a cultivated representative, but scarce experimental data from the cultivated organisms show that they harbor distinct functional repertoires. To unveil the ecological as well as evolutionary impact of Archaea from metagenomics, new computational methods need to be developed, followed by in-depth analysis. Among them is the genome-wide protein fusion screening performed here. Natural fusions and fissions of genes not only contribute to microbial evolution but also complicate the correct identification and functional annotation of sequences. The products of these processes can be defined as fusion (or composite) proteins, the ones consisting of two or more domains originally encoded by different genes and split proteins, and the ones originating from the separation of a gene in two (fission). Fusion identifications are required for proper phylogenetic reconstructions and metabolic pathway completeness assessments, while mappings between fused and unfused proteins can fill some of the existing gaps in metabolic models. In the archaeal genome-wide screening, more than 1,900 fusion/fission protein clusters were identified, belonging to both newly sequenced and well-studied lineages. These protein families are mainly associated with different types of metabolism, genetic, and cellular processes. Moreover, 162 of the identified fusion/fission protein families are archaeal specific, having no identified fused homolog within the bacterial domain. Our approach was validated by the identification of experimentally characterized fusion/fission cases. However, around 25% of the identified fusion/fission families lack functional annotations for both composite and split states, showing the need for experimental characterization in Archaea.IMPORTANCEGenome-wide fusion screening has never been performed in Archaea on a broad taxonomic scale. The overlay of multiple computational techniques allows the detection of a fine-grained set of predicted fusion/fission families, instead of rough estimations based on conserved domain annotations only. The exhaustive mapping of fused proteins to bacterial organisms allows us to capture fusion/fission families that are specific to archaeal biology, as well as to identify links between bacterial and archaeal lineages based on cooccurrence of taxonomically restricted proteins and their sequence features. Furthermore, the identification of poorly characterized lineage-specific fusion proteins opens up possibilities for future experimental and computational investigations. This approach enhances our understanding of Archaea in general and provides potential candidates for in-depth studies in the future.

Advances in screening hyperthermic nanomedicines in 3D tumor models.

Hyperthermic nanomedicines are particularly relevant for tackling human cancer, providing a valuable alternative to conventional therapeutics. The early-stage preclinical performance evaluation of such anti-cancer treatments is conventionally performed in flat 2D cell cultures that do not mimic the volumetric heat transfer occurring in human tumors. Recently, improvements in bioengineered 3D in vitro models have unlocked the opportunity to recapitulate major tumor microenvironment hallmarks and generate highly informative readouts that can contribute to accelerating the discovery and validation of efficient hyperthermic treatments. Leveraging on this, herein we aim to showcase the potential of engineered physiomimetic 3D tumor models for evaluating the preclinical efficacy of hyperthermic nanomedicines, featuring the main advantages and design considerations under diverse testing scenarios. The most recent applications of 3D tumor models for screening photo- and/or magnetic nanomedicines will be discussed, either as standalone systems or in combinatorial approaches with other anti-cancer therapeutics. We envision that breakthroughs toward developing multi-functional 3D platforms for hyperthermia onset and follow-up will contribute to a more expedited discovery of top-performing hyperthermic therapies in a preclinical setting before their in vivo screening.

The superfamily of heme-copper oxygen reductases: types and evolutionary considerations.

Heme-copper oxygen reductases (HCO) reduce O(2) to water being the last enzymatic complexes of most aerobic respiratory chains. These enzymes promote energy conservation coupling the catalytic reaction to charge separation and charge translocation across the prokaryotic cytoplasmatic or mitochondrial membrane. In this way they contribute to the establishment and maintenance of the transmembrane difference of electrochemical potential, which is vital for solute/nutrient cell import, synthesis of ATP and motility. The HCO enzymes most probably share with the nitric oxide reductases, NORs, a common ancestor. We have proposed the classification of HCOs into three different types, A, B and C; based on the constituents of their proton channels (Pereira, Santana and Teixeira (2001) Biochim Biophys Acta, 1505, 185-208). This classification was recently challenged by the suggestion of other different types of HCOs. Using an enlarged sampling we performed an exhaustive bioinformatic reanalysis of HCOs family. Our results strengthened our previously proposed classification and showed no need for the existence of more divisions. Now, we analyze the taxonomic distribution of HCOs and NORs and the congruence of their sequence trees with the 16S rRNA tree. We observed that HCOs are widely distributed in the two prokaryotic domains and that the different types of enzymes are not confined to a specific taxonomic group or environmental niche.

Modular structure of complex II: An evolutionary perspective.

Succinate dehydrogenases (SDHs) and fumarate reductases (FRDs) catalyse the interconversion of succinate and fumarate, a reaction highly conserved in all domains of life. The current classification of SDH/FRDs is based on the structure of the membrane anchor subunits and their cofactors. It is, however, unknown whether this classification would hold in the context of evolution. In this work, a large-scale comparative genomic analysis of complex II addresses the questions of its taxonomic distribution and phylogeny. Our findings report that for types C, D, and F, structural classification and phylogeny go hand in hand, while for types A, B and E the situation is more complex, highlighting the possibility for their classification into subgroups. Based on these findings, we proposed a revised version of the evolutionary scenario for these enzymes in which a primordial soluble module, corresponding to the cytoplasmatic subunits, would give rise to the current diversity via several independent membrane anchor attachment events.

Stepwise pathway for early evolutionary assembly of dissimilatory sulfite and sulfate reduction.

Microbial dissimilatory sulfur metabolism utilizing dissimilatory sulfite reductases (Dsr) influenced the biochemical sulfur cycle during Earth's history and the Dsr pathway is thought to be an ancient metabolic process. Here we performed comparative genomics, phylogenetic, and synteny analyses of several Dsr proteins involved in or associated with the Dsr pathway across over 195,000 prokaryotic metagenomes. The results point to an archaeal origin of the minimal DsrABCMK(N) protein set, having as primordial function sulfite reduction. The acquisition of additional Dsr proteins (DsrJOPT) increased the Dsr pathway complexity. Archaeoglobus would originally possess the archaeal-type Dsr pathway and the archaeal DsrAB proteins were replaced with the bacterial reductive-type version, possibly at the same time as the acquisition of the QmoABC and DsrD proteins. Further inventions of two Qmo complex types, which are more spread than previously thought, allowed microorganisms to use sulfate as electron acceptor. The ability to use the Dsr pathway for sulfur oxidation evolved at least twice, with Chlorobi and Proteobacteria being extant descendants of these two independent adaptations.

The emergence of protein complexes: quaternary structure, dynamics and allostery. Colworth Medal Lecture.

All proteins require physical interactions with other proteins in order to perform their functions. Most of them oligomerize into homomers, and a vast majority of these homomers interact with other proteins, at least part of the time, forming transient or obligate heteromers. In the present paper, we review the structural, biophysical and evolutionary aspects of these protein interactions. We discuss how protein function and stability benefit from oligomerization, as well as evolutionary pathways by which oligomers emerge, mostly from the perspective of homomers. Finally, we emphasize the specificities of heteromeric complexes and their structure and evolution. We also discuss two analytical approaches increasingly being used to study protein structures as well as their interactions. First, we review the use of the biological networks and graph theory for analysis of protein interactions and structure. Secondly, we discuss recent advances in techniques for detecting correlated mutations, with the emphasis on their role in identifying pathways of allosteric communication.

The alternative complex III: a different architecture using known building modules.

Until recently cytochrome bc(1) complexes were the only enzymes known to be able to transfer electrons from reduced quinones to cytochrome c. However, a complex with the same activity and with a unique subunit composition was purified from the membranes of Rhodothermus marinus. This complex, named alternative complex III (ACIII) was then biochemical, spectroscopic and genetically characterized. Later it was observed that the presence of ACIII was not exclusive of R. marinus being the genes coding for ACIII widespread, at least in the Bacteria domain. In this work, a comprehensive description of the current knowledge on ACIII is presented. The relation of ACIII with members of the complex iron-sulfur molybdoenzyme family is investigated by analyzing all the available completely sequenced genomes. It is concluded that ACIII is a new complex composed by a novel combination of modules already identified in other respiratory complexes.

DiSCo: a sequence-based type-specific predictor of Dsr-dependent dissimilatory sulphur metabolism in microbial data.

Current methods in comparative genomic analyses for metabolic potential prediction of proteins involved in, or associated with the Dsr (dissimilatory sulphite reductase)-dependent dissimilatory sulphur metabolism are both time-intensive and computationally challenging, especially when considering metagenomic data. We developed DiSCo, a Dsr-dependent dissimilatory sulphur metabolism classification tool, which automatically identifies and classifies the protein type from sequence data. It takes user-supplied protein sequences and lists the identified proteins and their classification in terms of protein family and predicted type. It can also extract the sequence data from user-input to serve as basis for additional downstream analyses. DiSCo provides the metabolic functional prediction of proteins involved in Dsr-dependent dissimilatory sulphur metabolism with high levels of accuracy in a fast manner. We ran DiSCo against a dataset composed of over 190 thousand (meta)genomic records and efficiently mapped Dsr-dependent dissimilatory sulphur proteins in 1798 lineages across both prokaryotic domains. This allowed the identification of new micro-organisms belonging to Thaumarchaeota and Spirochaetes lineages with the metabolic potential to use the Dsr-pathway for energy conservation. DiSCo is implemented in Perl 5 and freely available under the GNU GPLv3 at https://github.com/Genome-Evolution-and-Ecology-Group-GEEG/DiSCo.

Energy at Origins: Favorable Thermodynamics of Biosynthetic Reactions in the Last Universal Common Ancestor (LUCA).

Though all theories for the origin of life require a source of energy to promote primordial chemical reactions, the nature of energy that drove the emergence of metabolism at origins is still debated. We reasoned that evidence for the nature of energy at origins should be preserved in the biochemical reactions of life itself, whereby changes in free energy, ΔG, which determine whether a reaction can go forward or not, should help specify the source. By calculating values of ΔG across the conserved and universal core of 402 individual reactions that synthesize amino acids, nucleotides and cofactors from H2, CO2, NH3, H2S and phosphate in modern cells, we find that 95-97% of these reactions are exergonic (ΔG ≤ 0 kJ⋅mol-1) at pH 7-10 and 80-100°C under nonequilibrium conditions with H2 replacing biochemical reductants. While 23% of the core's reactions involve ATP hydrolysis, 77% are ATP-independent, thermodynamically driven by ΔG of reactions involving carbon bonds. We identified 174 reactions that are exergonic by -20 to -300 kJ⋅mol-1 at pH 9 and 80°C and that fall into ten reaction types: six pterin dependent alkyl or acyl transfers, ten S-adenosylmethionine dependent alkyl transfers, four acyl phosphate hydrolyses, 14 thioester hydrolyses, 30 decarboxylations, 35 ring closure reactions, 31 aromatic ring formations, and 44 carbon reductions by reduced nicotinamide, flavins, ferredoxin, or formate. The 402 reactions of the biosynthetic core trace to the last universal common ancestor (LUCA), and reveal that synthesis of LUCA's chemical constituents required no external energy inputs such as electric discharge, UV-light or phosphide minerals. The biosynthetic reactions of LUCA uncover a natural thermodynamic tendency of metabolism to unfold from energy released by reactions of H2, CO2, NH3, H2S, and phosphate.

Dual-Crosslinked Dynamic Hydrogel Incorporating {Mo154 } with pH and NIR Responsiveness for Chemo-Photothermal Therapy.

Polyoxometalates are an emerging class of molecular clusters, with well-defined structures and chemical compositions that are produced through simple, low-cost, and highly reproducible methods. In particular, the wheel-shaped cluster {Mo154 } is a promising photothermal agent due to its intervalence charge transfer transitions. However, its toxicity hinders its systemic administration, being the development of a localized delivery system still incipient. Herein, an injectable and self-healing hydrogel of easy preparation and administration is developed, incorporating both {Mo154 } and doxorubicin for synergistic photothermal and chemotherapy applications. The hydrogel is composed of benzylaldehyde functionalized polyethylene glycol, poly(N-isopropylacrylamide) functionalized chitosan and {Mo154 }. The gelation occurs within 60 s at room temperature, and the dual crosslinking by Schiff base and electrostatic interactions generates a dynamic network, which enables self-healing after injection. Moreover, the hydrogel delivers chemotherapeutic drugs, with a release triggered by dual near infra-red (NIR) radiation and pH changes. This stimuli-responsive release system along with the photothermal conversion ability of the hydrogel allows the simultaneous combination of photothermal and chemotherapy. This synergic system efficiently ablates the cancer tumor in vivo with no systemic toxicity. Overall, this work paves the way for the development of novel {Mo154 }-based systems, incorporated in self-healing and injectable hydrogels for dual chemo-photothermal therapy.