Ribosomal protein-sequence block structure suggests complex prokaryotic evolution with implications for the origin of eukaryotes

Amino acid sequence alignments of orthologous ribosomal proteins found in Bacteria, Archaea, and Eukaryota display, relative to one another, an unusual segment or block structure, with major evolutionary implications. Within each of the prokaryotic phylodomains the sequences exhibit substantial similarity, but cross-domain alignments break up into (a) universal blocks (conserved in both phylodomains), (b) bacterial blocks (unalignable with any archaeal counterparts), and (c) archaeal blocks (unalignable with any bacterial counterparts). Sequences of those eukaryotic cytoplasmic riboproteins that have orthologs in both Bacteria and Archaea, exclusively match the archaeal block structure. The distinct blocks do not correlate consistently with any identifiable functional or structural feature including RNA and protein contacts. This phylodomain-specific block pattern also exists in a number of other proteins associated with protein synthesis, but not among enzymes of intermediary metabolism. While the universal blocks imply that modern Bacteria and Archaea (as defined by their translational machinery) clearly have had a common ancestor, the phylodomain-specific blocks imply that these two groups derive from single, phylodomain-specific types that came into existence at some point long after that common ancestor. The simplest explanation for this pattern would be a major evolutionary bottleneck, or other scenario that drastically limited the progenitors of modern prokaryotic diversity at a time considerably after the evolution of a fully functional translation apparatus. The vast range of habitats and metabolisms that prokaryotes occupy today would thus reflect divergent evolution after such a restricting event. Interestingly, phylogenetic analysis places the origin of eukaryotes at about the same time and shows a closer relationship of the eukaryotic ribosome-associated proteins to crenarchaeal rather than euryarchaeal counterparts.     

Bacterial origin for the isoprenoid biosynthesis enzyme HMG-CoA reductase of the archaeal orders Thermoplasmatales and Archaeoglobales.

The enzyme 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMG-CoA reductase or HMGR) fulfills an essential role in archaea, as it is required for the synthesis of isoprenoid ethers, the main component of archaeal cell membranes. There are two clearly homologous but structurally different classes of the enzyme, one found mainly in eukaryotes and archaea (class 1), and the other found in bacteria (class 2). This feature facilitated the identification of several cases of interdomain lateral gene transfer (LGT), in particular, the bacterial origin for the HMGR gene from the archaeon Archaeoglobus fulgidus. In order to investigate if this LGT event was recent and limited in its scope or had a broad and long-term impact on the recipient and its related lineages, the HMGR gene was amplified and sequenced from a variety of archaea. The survey covered close relatives of A. fulgidus, the only archaeon known prior to this study to possess a bacterial-like HMGR; representatives of each main euryarchaeal group were also inspected. All culturable members of the archaeal group Archaeoglobales were found to display an HMGR very similar to the enzyme of the bacterium Pseudomonas mevalonii. Surprisingly, two species of the genus Thermoplasma also harbor an HMGR of bacterial origin highly similar to the enzymes found in the Archaeoglobales. Phylogenetic analyses of the HMGR gene and comparisons to reference phylogenies from other genes confirm a common bacterial origin for the HMGRs of Thermoplasmatales and Archaeoglobales. The most likely explanation of these results includes an initial bacteria-to-archaea transfer, followed by a another event between archaea. Their presence in two divergent archaeal lineages suggests an important adaptive role for these laterally transferred genes.     

Insights into ssDNA recognition by the OB fold from a structural and thermodynamic study of Sulfolobus SSB protein

Information processing pathways such as DNA replication are conserved in eukaryotes and archaea and are significantly different from those found in bacteria. Single-stranded DNA-binding (SSB) proteins (or replication protein A, RPA, in eukaryotes) play a central role in many of these pathways. However, whilst euryarchaea have a eukaryotic-type RPA homologue, crenarchaeal SSB proteins appear much more similar to the bacterial proteins, with a single OB fold for DNA binding and a flexible C-terminal tail that is implicated in protein-protein interactions. We have determined the crystal structure of the SSB protein from the crenarchaeote Sulfolobus solfataricus to 1.26 A. The structure shows a striking and unexpected similarity to the DNA-binding domains of human RPA, providing confirmation of the close relationship between archaea and eukaryotes. The high resolution of the structure, together with thermodynamic and mutational studies of DNA binding, allow us to propose a molecular basis for DNA binding and define the features required for eukaryotic and archaeal OB folds.     

21 Ribosomal evidence suggests that archaea evolved after fungi

We are exploring the potential to trace species evolution with the ribosomal proteins (RibPs) present in bacterial, eukaryotic, and archaeal ribosomes and to compare the independent trees for consistency. The complete genomes of over 8400 bacteria, eukaryota, and archaea are presently in the SwissPro/TrEMBL (SPT) database. A search of SPT using a vector designed with ScanProsite formats (V1) finds and aligns 8405 sequences (5312 bacterial, 2905 eukaryotic, and 169 archaeal) that are homologous with bone fide bacterial S19 ribosomal proteins(S19s). When the 8405 sequences are perfectly aligned, 15 residues are conserved at 90% identity and 40 are conserved at 70% identity. We are not aware of any previous publication reporting sequence alignment of 8400 members of any single family including all bacteria, eukaryota and archaea, for which complete genomes have been published.A Pro and a Gly separated by 11 residues are 100% conserved in the 8405 S19s. In the position immediately before the fully conserved Gly, two residues (Asp and Asn) are present in 98.3% of the 8405 sequences. The Asp residue is found almost exclusively in 2190 gram-positive bacteria. The Asn residue is found in 3065 gram-negative bacteria, 123 Archaea, 1939 eukaryotes, and 64 specific species of gram-positive bacteria. There is biochemical evidence for the existence of distinct mitochondrial, chloroplast, and cytosolic ribosomes and reports that plants have all three forms and mammals only two. Reliable data concerning how individual ribosomal proteins differ in different types of ribosomes are meager. Examination of the eukaryotic S19s reveals the existence of three distinct types. Two of the distinctly different types are found in most fungi, three of the types are found in some viridiplante, and only one type is found in metazoa and archaea. We demonstrate the sequence homology between the mitochondrial form found in fungi and plants and the S19 proteins of alpha proteobacteria; between the chloroplast S19s and the S19s of cyanobacteria; and among the cytosolic S19s found only in fungi, metazoa, archaea, and in some viridiplantae. Our findings suggest that most archaeal species appeared after a gene duplication event in fungi that correlates with the origin of the cytosolic ribosome.     

Archaeal protein translocation crossing membranes in the third domain of life.

Proper cell function relies on correct protein localization. As a first step in the delivery of extracytoplasmic proteins to their ultimate destinations, the hydrophobic barrier presented by lipid-based membranes must be overcome. In contrast to the well-defined bacterial and eukaryotic protein translocation systems, little is known about how proteins cross the membranes of archaea, the third and most recently described domain of life. In bacteria and eukaryotes, protein translocation occurs at proteinaceous sites comprised of evolutionarily conserved core components acting in concert with other, domain-specific elements. Examination of available archaeal genomes as well as cloning of individual genes from other archaeal strains reveals the presence of homologues to selected elements of the bacterial or eukaryotic translocation machines. Archaeal genomic searches, however, also reveal an apparent absence of other, important components of these two systems. Archaeal translocation may therefore represent a hybrid of the bacterial and eukaryotic models yet may also rely on components or themes particular to this domain of life. Indeed, considering the unique chemical composition of the archaeal membrane as well as the extreme conditions in which archaea thrive, the involvement of archaeal-specific translocation elements could be expected. Thus, understanding archaeal protein translocation could reveal the universal nature of certain features of protein translocation which, in some cases, may not be readily obvious from current comparisons of bacterial and eukaryotic systems. Alternatively, elucidation of archaeal translocation could uncover facets of the translocation process either not yet identified in bacteria or eukaryotes, or which are unique to archaea. In the following, the current status of our understanding of protein translocation in archaea is reviewed.     

Origin of eukaryotes from within archaea,archaeal eukaryome and bursts of gene gain: eukaryogenesis just made easier?

The origin of eukaryotes is a fundamental, forbidding evolutionary puzzle. Comparative genomic analysis clearly shows that the last eukaryotic common ancestor (LECA) possessed most of the signature complex features of modern eukaryotic cells, in particular the mitochondria, the endomembrane system including the nucleus, an advanced cytoskeleton and the ubiquitin network. Numerous duplications of ancestral genes, e.g. DNA polymerases, RNA polymerases and proteasome subunits, also can be traced back to the LECA. Thus, the LECA was not a primitive organism and its emergence must have resulted from extensive evolution towards cellular complexity. However, the scenario of eukaryogenesis, and in particular the relationship between endosymbiosis and the origin of eukaryotes, is far from being clear. Four recent developments provide new clues to the likely routes of eukaryogenesis. First, evolutionary reconstructions suggest complex ancestors for most of the major groups of archaea, with the subsequent evolution dominated by gene loss. Second, homologues of signature eukaryotic proteins, such as actin and tubulin that form the core of the cytoskeleton or the ubiquitin system, have been detected in diverse archaea. The discovery of this ‘dispersed eukaryome’ implies that the archaeal ancestor of eukaryotes was a complex cell that might have been capable of a primitive form of phagocytosis and thus conducive to endosymbiont capture. Third, phylogenomic analyses converge on the origin of most eukaryotic genes of archaeal descent from within the archaeal evolutionary tree, specifically, the TACK superphylum. Fourth, evidence has been presented that the origin of the major archaeal phyla involved massive acquisition of bacterial genes. Taken together, these findings make the symbiogenetic scenario for the origin of eukaryotes considerably more plausible and the origin of the organizational complexity of eukaryotic cells more readily explainable than they appeared until recently.     

An actin-based cytoskeleton in archaea

In eukaryotic and bacterial cells, spatial organization is dependent upon cytoskeletal filaments. Actin is a main eukaryotic cytoskeletal element, involved in key processes such as cell shape determination, mechanical force generation and cytokinesis. We describe an archaeal cytoskeleton which forms helical structures within Pyrobaculum calidifontis cells, as shown by in situ immunostaining. The core components include an archaeal actin homologue, Crenactin, closely related to the eukaryotic counterpart. The crenactin gene belongs to a conserved gene cluster denoted Arcade (actin-related cytoskeleton in Archaea involved in shape determination). The phylogenetic distribution of arcade genes is restricted to the crenarchaeal Thermoproteales lineage, and to Korarchaeota, and correlates with rod-shaped and filamentous cell morphologies. Whereas Arcadin-1, -3 and -4 form helical structures, suggesting cytoskeleton-associated functions, Arcadin-2 was found to be localized between segregated nucleoids in a cell subpopulation, in agreement with possible involvement in cytokinesis. The results support a crenarchaeal origin of the eukaryotic actin cytoskeleton and, as such, have implications for theories concerning the origin of the eukaryotic cell.     

Crenarchaeota and their role in the nitrogen cycle in a subsurface radioactive thermal spring in the Austrian Central Alps

Previous results from a 16S rRNA gene library analysis showed high diversity within the prokaryotic community of a subterranean radioactive thermal spring, the "Franz-Josef-Quelle" (FJQ) in Bad Gastein, Austria, as well as evidence for ammonia oxidation by crenarchaeota. This study reports further characterization of the community by denaturing gradient gel electrophoresis (DGGE) analysis, fluorescence in situ hybridization (FISH), and semiquantitative nitrification measurements. DGGE bands from three types of samples (filtered water, biofilms on glass slides, and naturally grown biofilms), including samples collected at two distinct times (January 2005 and July 2006), were analyzed. The archaeal community consisted mainly of Crenarchaeota of the soil-subsurface-freshwater group (group 1.1b) and showed a higher diversity than in the previous 16S rRNA gene library analysis, as was also found for crenarchaeal amoA genes. No bacterial amoA genes were detected. FISH analysis of biofilms indicated the presence of archaeal cells with an abundance of 5.3% (+/-4.5%) in the total 4',6-diamidino-2-phenylindole (DAPI)-stained community. Microcosm experiments of several weeks in duration showed a decline of ammonium that correlated with an increase of nitrite, the presence of crenarchaeal amoA genes, and the absence of bacterial amoA genes. The data suggested that only ammonia-oxidizing archaea (AOA) perform the first step of nitrification in this 45 degrees C environment. The crenarchaeal amoA gene sequences grouped within a novel cluster of amoA sequences from the database, originating from geothermally influenced environments, for which we propose the designation "thermal spring" cluster and which may be older than most AOA from soils on earth.     

Crystal structure of translation initiation factor 5B from the crenarchaeon Aeropyrum pernix

Initiation factor 5B (IF5B) is a universally conserved translational GTPase that catalyzes ribosomal subunit joining. In eukaryotes, IF5B directly interacts via a groove in its domain IV with initiation factor 1A (IF1A), another universally conserved initiation factor, to accomplish efficient subunit joining. Here, we have determined the first structure of a crenarchaeal IF5B, which revealed that the archaea‐specific region of IF5B (helix α15) binds and occludes the groove of domain IV. Therefore, archaeal IF5B cannot access IF1A in the same manner as eukaryotic IF5B. This fact suggests that different relationships between IF5B and IF1A exist in archaea and eukaryotes. Proteins 2016; 84:712–717. © 2016 Wiley Periodicals, Inc.     

Vertical distribution and phylogenetic characterization of marine planktonic Archaea in the Santa Barbara Channel.

Newly described phylogenetic lineages within the domain Archaea have recently been found to be significant components of marine picoplankton assemblages. To better understand the ecology of these microorganisms, we investigated the relative abundance, distribution, and phylogenetic composition of Archaea in the Santa Barbara Channel. Significant amounts of archaeal rRNA and rDNA (genes coding for rRNA) were detected in all samples analyzed. The relative abundance of archaeal rRNA as measured by quantitative oligonucleotide hybridization experiments was low in surface waters but reached higher values (20 to 30% of prokaryotic rRNA) at depths below 100 m. Probes were developed for the two major groups of marine Archaea detected. rRNA originating from the euryarchaeal group (group II) was most abundant in surface waters, whereas rRNA from the crenarchaeal group (group I) dominated at depth. Clone libraries of PCR-amplified archaeal rRNA genes were constructed with samples from 0 and 200 m deep. Screening of libraries by hybridization with specific oligonucleotide probes, as well as subsequent sequencing of the cloned genes, indicated that virtually all archaeal rDNA clones recovered belonged to one of the two groups. The recovery of cloned rDNA sequence types in depth profiles exhibited the same trends as were observed in quantitative rRNA hybridization experiments. One representative of each of 18 distinct restriction fragment length polymorphism types was partially sequenced. Recovered sequences spanned most of the previously reported phylogenetic diversity detected in planktonic crenarchaeal and euryarchaeal groups. Several rDNA sequences appeared to be harbored in archaeal types which are widely distributed in marine coastal waters. In total, data suggest that marine planktonic crenarchaea and euryarchaea of temperate coastal habitats thrive in different zones of the water column. The relative rRNA abundance of the crenarchaeal group suggests that its members constitute a significant fraction of the prokaryotic biomass in subsurface coastal waters.     

Origin and evolution of eukaryotic chaperonins: phylogenetic evidence for ancient duplications in CCT genes

Chaperonins are oligomeric protein-folding complexes which are divided into two distantly related structural classes. Group I chaperonins (called GroEL/cpn60/hsp60) are found in bacteria and eukaryotic organelles, while group II chaperonins are present in archaea and the cytoplasm of eukaryotes (called CCT/TriC). While archaea possess one to three chaperonin subunit-encoding genes, eight distinct CCT gene families (paralogs) have been characterized in eukaryotes. We are interested in determining when during eukaryotic evolution the multiple gene duplications producing the CCT subunits occurred. We describe the sequence and phylogenetic analysis of five CCT genes from TRICHOMONAS: vaginalis and seven from GIARDIA: lamblia, representatives of amitochondriate protist lineages thought to have diverged early from other eukaryotes. Our data show that the gene duplications producing the eight CCT paralogs took place prior to the organismal divergence of TRICHOMONAS: and GIARDIA: from other eukaryotes. Thus, these divergent protists likely possess completely hetero-oligomeric CCT complexes like those in yeast and mammalian cells. No close phylogenetic relationship between the archaeal chaperonins and specific CCT subunits was observed, suggesting that none of the CCT gene duplications predate the divergence of archaea and eukaryotes. The duplications producing the CCTdelta and CCTepsilon subunits, as well as CCTalpha, CCTbeta, and CCTeta, are the most recent in the CCT gene family. Our analyses show significant differences in the rates of evolution of archaeal chaperonins compared with the eukaryotic CCTs, as well as among the different CCT subunits themselves. We discuss these results in light of current views on the origin, evolution, and function of CCT complexes.     

Evidence of independent gene duplications during the evolution of archaeal and eukaryotic family B DNA polymerases

Eukaryotes and archaea both possess multiple genes coding for family B DNA polymerases. In animals and fungi, three family B DNA polymerases, alpha, delta, and epsilon, are responsible for replication of nuclear DNA. We used a PCR-based approach to amplify and sequence phylogenetically conserved regions of these three DNA polymerases from Giardia intestinalis and Trichomonas vaginalis, representatives of early-diverging eukaryotic lineages. Phylogenetic analysis of eukaryotic and archaeal paralogs suggests that the gene duplications that gave rise to the three replicative paralogs occurred before the divergence of the earliest eukaryotic lineages, and that all eukaryotes are likely to possess these paralogs. One eukaryotic paralog, epsilon, consistently branches within archaeal sequences to the exclusion of other eukaryotic paralogs, suggesting that an epsilon-like family B DNA polymerase was ancestral to both archaea and eukaryotes. Because crenarchaeote and euryarchaeote paralogs do not form monophyletic groups in phylogenetic analysis, it is possible that archaeal family B paralogs themselves evolved by a series of gene duplications independent of the gene duplications that gave rise to eukaryotic paralogs.      

Growth of ammonia-oxidizing archaea in soil microcosms is inhibited by acetylene

Autotrophic ammonia-oxidizing bacteria were considered to be responsible for the majority of ammonia oxidation in soil until the recent discovery of the autotrophic ammonia-oxidizing archaea. To assess the relative contributions of bacterial and archaeal ammonia oxidizers to soil ammonia oxidation, their growth was analysed during active nitrification in soil microcosms incubated for 30 days at 30 °C, and the effect of an inhibitor of ammonia oxidation (acetylene) on their growth and soil nitrification kinetics was determined. Denaturing gradient gel electrophoresis (DGGE) analysis of bacterial ammonia oxidizer 16S rRNA genes did not detect any change in their community composition during incubation, and quantitative PCR (qPCR) analysis of bacterial amoA genes indicated a small decrease in abundance in control and acetylene-containing microcosms. DGGE fingerprints of archaeal amoA and 16S rRNA genes demonstrated changes in the relative abundance of specific crenarchaeal phylotypes during active nitrification. Growth was also indicated by increases in crenarchaeal amoA gene copy number, determined by qPCR. In microcosms containing acetylene, nitrification and growth of the crenarchaeal phylotypes were suppressed, suggesting that these crenarchaea are ammonia oxidizers. Growth of only archaeal but not bacterial ammonia oxidizers occurred in microcosms with active nitrification, indicating that ammonia oxidation was mostly due to archaea in the conditions of the present study.     

DNA Repair Systems in Archaea: Mementos from the Last Universal Common Ancestor?

DNA repair in the Archaea is relevant to the consideration of genome maintenance and replication fidelity in the last universal common ancestor (LUCA) from two perspectives. First, these prokaryotes embody a mix of bacterial and eukaryal molecular features. Second, DNA repair proteins would have been essential in LUCA to maintain genome integrity, regardless of the environmental temperature. Yet we know very little of the basic molecular mechanisms of DNA damage and repair in the Archaea in general. Many studies on DNA repair in archaea have been conducted with hyperthermophiles because of the additional stress imposed on their macromolecules by high temperatures. In addition, of the six complete archaeal genome sequences published so far, five are thermophilic archaea. We have recently shown that the hyperthermophile Pyrococcus furiosus has an extraordinarily high capacity for repair of radiation-induced double-strand breaks and we have identified and sequenced several genes involved in DNA repair in P. furiosus. At the sequence level, only a few genes share homology with known bacterial repair genes. For instance, our phylogenetic analysis indicates that archaeal recombinases occur in two paralogous gene families, one of which is very deeply branched, and both recombinases are more closely related to the eukaryotic RAD51 and Dmc1 gene families than to the Escherichia coli recA gene. We have also identified a gene encoding a repair endo/exonuclease in the genomes of several Archaea. The archaeal sequences are highly homologous to those of the eukaryotic Rad2 family and they cluster with genes of the FEN-1 subfamily, which are known to be involved in DNA replication and repair in eukaryotes. We argue that there is a commonality of mechanisms and protein sequences, shared between prokaryotes and eukaryotes for several modes of DNA repair, reflecting diversification from a minimal set of genes thought to represent the genome of the LUCA.     

古菌ESCRT系统研究进展

内体分拣转运复合体（ESCRT,endosomal sorting complex required for transport）曾被认为是真核生物特有的系统,涉及膜重塑、泛素化蛋白质分拣等重要细胞生命过程。近年的研究显示,TACK（包括Thaumarchaeota、Aigarchaeota、Crenarchaeota和Korarchaeota门）古菌超门中存在着一类与分泌膜囊泡、古菌病毒出胞以及细胞分裂过程等膜重塑过程相关的细胞分裂（Cdv,cell division）系统,该系统中的CdvB和CdvC是真核生物ESCRT-III和Vps4的同源蛋白,提示真核生物ESCRT系统可能起源自古菌。然而,由于TACK古菌中缺少真核生物ESCRT系统的其他关键成分,这一假设仍有争议。最近发现的阿斯加德（Asgard）古菌是一类被认为与真核生物最近缘的古菌,其基因组具有较完整的ESCRT相关蛋白的编码基因,提示真核生物的ESCRT很可能起源于阿斯加德古菌。本文首先简要介绍真核生物ESCRT系统的组成及生物学功能,然后分别总结TACK古菌的Cdv系统和阿斯加德古菌的ESCRT系统的研究进展,重点讨论它们的组成及生物学功能,为进一步了解古菌ESCRT系统与真核生物起源的关系提供参考。     

Origins and evolution of eukaryotic RNA interference

Small interfering RNAs (siRNAs) and genome-encoded microRNAs (miRNAs) silence genes via complementary interactions with mRNAs. With thousands of miRNA genes identified and genome sequences of diverse eukaryotes available for comparison, the opportunity emerges for insights into the origin and evolution of RNA interference (RNAi). The miRNA repertoires of plants and animals appear to have evolved independently. However, conservation of the key proteins involved in RNAi suggests that the last common ancestor of modern eukaryotes possessed siRNA-based mechanisms. Prokaryotes have an RNAi-like defense system that is functionally analogous but not homologous to eukaryotic RNAi. The protein machinery of eukaryotic RNAi seems to have been pieced together from ancestral archaeal, bacterial and phage proteins that are involved in DNA repair and RNA processing.