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Inteins are self-splicing protein elements found in viruses and all three domains of life. How the DNA encoding these selfish elements spreads within and between genomes is poorly understood, particularly in eukaryotes where inteins are scarce. Here, we show that the nuclear genomes of three strains of Anaeramoeba encode between 45 and 103 inteins, in stark contrast to four found in the most intein-rich eukaryotic genome described previously. The Anaeramoeba inteins reside in a wide range of proteins, only some of which correspond to intein-containing proteins in other eukaryotes, prokaryotes, and viruses. Our data also suggest that viruses have contributed to the spread of inteins in Anaeramoeba and the colonization of new alleles. The persistence of Anaeramoeba inteins might be partly explained by intragenomic movement of intein-encoding regions from gene to gene. Our intein dataset greatly expands the spectrum of intein-containing proteins and provides insights into the evolution of inteins in eukaryotes.
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Inteínas , Processamento de Proteína , Inteínas/genética , Eucariotos/genética , Proteínas/genética , GenomaRESUMO
Mitochondrial genomes of apicomplexans, dinoflagellates, and chrompodellids that collectively make up the Myzozoa, encode only three proteins (Cytochrome b [COB], Cytochrome c oxidase subunit 1 [COX1], Cytochrome c oxidase subunit 3 [COX3]), contain fragmented ribosomal RNAs, and display extensive recombination, RNA trans-splicing, and RNA-editing. The early-diverging Perkinsozoa is the final major myzozoan lineage whose mitochondrial genomes remained poorly characterized. Previous reports of Perkinsus genes indicated independent acquisition of non-canonical features, namely the occurrence of multiple frameshifts. To determine both ancestral myzozoan and novel perkinsozoan mitochondrial genome features, we sequenced and assembled mitochondrial genomes of four Perkinsus species. These data show a simple ancestral genome with the common reduced coding capacity but disposition for rearrangement. We identified 75 frameshifts across the four species that occur as distinct types and that are highly conserved in gene location. A decoding mechanism apparently employs unused codons at the frameshift sites that advance translation either +1 or +2 frames to the next used codon. The locations of frameshifts are seemingly positioned to regulate protein folding of the nascent protein as it emerges from the ribosome. The cox3 gene is distinct in containing only one frameshift and showing strong selection against residues that are otherwise frequently encoded at the frameshift positions in cox1 and cob. All genes lack cysteine codons implying a reduction to 19 amino acids in these genomes. Furthermore, mitochondrion-encoded rRNA fragment complements are incomplete in Perkinsus spp. but some are found in the nuclear DNA suggesting import into the organelle. Perkinsus demonstrates further remarkable trajectories of organelle genome evolution including pervasive integration of frameshift translation into genome expression.
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Genoma Mitocondrial , Códon , Cisteína/genética , Citocromos b/genética , DNA Mitocondrial/genética , Complexo IV da Cadeia de Transporte de Elétrons/genéticaRESUMO
Blastocystis is the most prevalent eukaryotic microbe colonizing the human gut, infecting approximately 1 billion individuals worldwide. Although Blastocystis has been linked to intestinal disorders, its pathogenicity remains controversial because most carriers are asymptomatic. Here, the genome sequence of Blastocystis subtype (ST) 1 is presented and compared to previously published sequences for ST4 and ST7. Despite a conserved core of genes, there is unexpected diversity between these STs in terms of their genome sizes, guanine-cytosine (GC) content, intron numbers, and gene content. ST1 has 6,544 protein-coding genes, which is several hundred more than reported for ST4 and ST7. The percentage of proteins unique to each ST ranges from 6.2% to 20.5%, greatly exceeding the differences observed within parasite genera. Orthologous proteins also display extreme divergence in amino acid sequence identity between STs (i.e., 59%-61% median identity), on par with observations of the most distantly related species pairs of parasite genera. The STs also display substantial variation in gene family distributions and sizes, especially for protein kinase and protease gene families, which could reflect differences in virulence. It remains to be seen to what extent these inter-ST differences persist at the intra-ST level. A full 26% of genes in ST1 have stop codons that are created on the mRNA level by a novel polyadenylation mechanism found only in Blastocystis. Reconstructions of pathways and organellar systems revealed that ST1 has a relatively complete membrane-trafficking system and a near-complete meiotic toolkit, possibly indicating a sexual cycle. Unlike some intestinal protistan parasites, Blastocystis ST1 has near-complete de novo pyrimidine, purine, and thiamine biosynthesis pathways and is unique amongst studied stramenopiles in being able to metabolize α-glucans rather than ß-glucans. It lacks all genes encoding heme-containing cytochrome P450 proteins. Predictions of the mitochondrion-related organelle (MRO) proteome reveal an expanded repertoire of functions, including lipid, cofactor, and vitamin biosynthesis, as well as proteins that may be involved in regulating mitochondrial morphology and MRO/endoplasmic reticulum (ER) interactions. In sharp contrast, genes for peroxisome-associated functions are absent, suggesting Blastocystis STs lack this organelle. Overall, this study provides an important window into the biology of Blastocystis, showcasing significant differences between STs that can guide future experimental investigations into differences in their virulence and clarifying the roles of these organisms in gut health and disease.
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Blastocystis/genética , Genoma de Protozoário , Blastocystis/metabolismo , Metabolismo dos Carboidratos , Códon de Terminação , Microbioma Gastrointestinal , Humanos , Íntrons , Especificidade da EspécieRESUMO
PREMISE OF THE STUDY: The Bahamas archipelago is formed by young, tectonically stable carbonate banks that harbor direct geological evidence of global ice-volume changes. We sought to detect signatures of major changes on gene flow patterns and reconstruct the phylogeographic history of the monophyletic Zamia pumila complex across the Bahamas. METHODS: Nuclear molecular markers with both high and low mutation rates were used to capture two different time scale signatures and test several gene flow and demographic hypotheses. KEY RESULTS: Single-copy nuclear genes unveiled apparent ancestral admixture on Andros, suggesting a significant role of this island as main hub of diversity of the archipelago. We detected demographic and spatial expansion of the Zamia pumila complex on both paleo-provinces around the Piacenzian (Pliocene)/Gelasian (Pleistocene). Populations evidenced signatures of different migration models that have occurred at two different times. Populations on Long Island (Z. lucayana) may either represent a secondary colonization of the Bahamas by Zamia or a rapid and early-divergence event of at least one population on the Bahamas. CONCLUSIONS: Despite changes in migration patterns with global climate, expected heterozygosity with both marker systems remains within the range reported for cycads, but with significant levels of increased inbreeding detected by the microsatellites. This finding is likely associated with reduced gene flow between and within paleo-provinces, accompanied by genetic drift, as rising seas enforced isolation. Our study highlights the importance of the maintenance of the predominant direction of genetic exchange and the role of overseas dispersion among the islands during climate oscillations.
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Biodiversidade , Filogenia , Zamiaceae/genética , Bahamas , Variação Genética , Ilhas , Repetições de Microssatélites , FilogeografiaRESUMO
Arecaceae tribe Cocoseae is the most economically important tribe of palms, including both coconut and African oil palm. It is mostly represented in the Neotropics, with one and two genera endemic to South Africa and Madagascar, respectively. Using primers for six single copy WRKY gene family loci, we amplified DNA from 96 samples representing all genera of the palm tribe Cocoseae as well as outgroup tribes Reinhardtieae and Roystoneae. We compared parsimony (MP), maximum likelihood (ML), and Bayesian (B) analysis of the supermatrix with three species-tree estimation approaches. Subtribe Elaeidinae is sister to the Bactridinae in all analyses. Within subtribe Attaleinae, Lytocaryum, previously nested in Syagrus, is now positioned by MP and ML as sister to the former, with high support; B maintains Lytocaryum embedded within Syagrus. Both MP and ML resolve Cocos as sister to Syagrus; B positions Cocos as sister to Attalea. Bactridineae is composed of two sister clades, Bactris and Desmoncus in one, for which there is morphological support, and a second comprising Acrocomia, Astrocaryum, and Aiphanes. Two B and one ML gene tree-species estimation approaches are incongruent with the supermatrix in a few critical intergeneric clades, but resolve the same infrageneric relationships. The biogeographic history of the Cocoseae is dominated by dispersal events. The tribe originated in the late Cretaceous in South America. Evaluated together, the supermatrix and species tree analyses presented in this paper provide the most accurate picture of the evolutionary history of the tribe to date, with more congruence than incongruence among the various methodologies.
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Metamonads are a diverse group of heterotrophic microbial eukaryotes adapted to living in hypoxic environments. All metamonads but one harbour metabolically altered 'mitochondrion-related organelles' (MROs) with reduced functions, however the degree of reduction varies. Here, we generate high-quality draft genomes, transcriptomes, and predicted proteomes for five recently discovered free-living metamonads. Phylogenomic analyses placed these organisms in a group we name the 'BaSk' (Barthelonids+Skoliomonads) clade, a deeply branching sister group to the Fornicata, a phylum that includes parasitic and free-living flagellates. Bioinformatic analyses of gene models shows that these organisms are predicted to have extremely reduced MRO proteomes in comparison to other free-living metamonads. Loss of the mitochondrial iron-sulfur cluster assembly system in some organisms in this group appears to be linked to the acquisition in their common ancestral lineage of a SUF-like minimal system Fe/S cluster pathway by lateral gene transfer. One of the isolates, Skoliomonas litria, appears to have lost all other known MRO pathways. No proteins were confidently assigned to the predicted MRO proteome of this organism suggesting that the organelle has been lost. The extreme mitochondrial reduction observed within this free-living anaerobic protistan clade demonstrates that mitochondrial functions may be completely lost even in free-living organisms.
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Mitocôndrias , Filogenia , Proteoma , Mitocôndrias/metabolismo , Mitocôndrias/genética , Proteoma/metabolismo , Proteoma/genética , Transcriptoma , Eucariotos/genética , Eucariotos/metabolismo , Eucariotos/classificação , Transferência Genética Horizontal , Proteínas Ferro-Enxofre/metabolismo , Proteínas Ferro-Enxofre/genéticaRESUMO
BACKGROUND AND AIMS: Despite a recent new classification, a stable phylogeny for the cycads has been elusive, particularly regarding resolution of Bowenia, Stangeria and Dioon. In this study, five single-copy nuclear genes (SCNGs) are applied to the phylogeny of the order Cycadales. The specific aim is to evaluate several gene tree-species tree reconciliation approaches for developing an accurate phylogeny of the order, to contrast them with concatenated parsimony analysis and to resolve the erstwhile problematic phylogenetic position of these three genera. METHODS: DNA sequences of five SCNGs were obtained for 20 cycad species representing all ten genera of Cycadales. These were analysed with parsimony, maximum likelihood (ML) and three Bayesian methods of gene tree-species tree reconciliation, using Cycas as the outgroup. A calibrated date estimation was developed with Bayesian methods, and biogeographic analysis was also conducted. KEY RESULTS: Concatenated parsimony, ML and three species tree inference methods resolve exactly the same tree topology with high support at most nodes. Dioon and Bowenia are the first and second branches of Cycadales after Cycas, respectively, followed by an encephalartoid clade (Macrozamia-Lepidozamia-Encephalartos), which is sister to a zamioid clade, of which Ceratozamia is the first branch, and in which Stangeria is sister to Microcycas and Zamia. CONCLUSIONS: A single, well-supported phylogenetic hypothesis of the generic relationships of the Cycadales is presented. However, massive extinction events inferred from the fossil record that eliminated broader ancestral distributions within Zamiaceae compromise accurate optimization of ancestral biogeographical areas for that hypothesis. While major lineages of Cycadales are ancient, crown ages of all modern genera are no older than 12 million years, supporting a recent hypothesis of mostly Miocene radiations. This phylogeny can contribute to an accurate infrafamilial classification of Zamiaceae.
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Núcleo Celular/genética , Cycadopsida/classificação , Cycadopsida/genética , Dosagem de Genes/genética , Genes de Plantas/genética , Filogenia , Árvores/genética , Funções Verossimilhança , Filogeografia , Polimorfismo Genético , Especificidade da Espécie , Fatores de TempoRESUMO
BACKGROUND: Blastocystis is one of the most common eukaryotic microorganisms colonizing the intestines of both humans and animals, but the conditions under which it may be a pathogen are unclear. METHODS: To study the genomic characteristics of circulating subtypes (ST) in Colombia, we established nine xenic cultures from Blastocystis isolated from human fecal samples, we identified 10 different subtypes, since one sample had a mixed infection. Thus, the genomes of the subtypes ST1 (n = 3), ST2 (n = 1), ST3 (n = 2), ST6 (n = 1), ST7 (n = 1), and ST8 (n = 2) were sequenced using Illumina and Oxford Nanopore Technologies (ONT). RESULTS: Analyses of these draft nuclear genomes indicated remarkable diversity in terms of genome size and guanine-cytosine (GC) content among the compared STs. Illumina sequencing-only draft genomes contained 824 to 2077 scaffolds, with total genome size ranging from 12 to 13.2 Mb and N50 values ranging from 10,585 to 29,404 base pairs (bp). The genome of one ST1 isolate was sequenced using ONT. This assembly was more contiguous, with a size of 20 million base pairs (Mb) spread over 116 scaffolds, and an N50 of 248,997 bp. CONCLUSION: This work represents one of the few large-scale comparative genomic analyses of Blastocystis isolates, providing an additional glimpse into its genomic diversity.
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Infecções por Blastocystis , Blastocystis , Animais , Humanos , Blastocystis/genética , Colômbia , Variação Genética , Filogenia , DNA de Protozoário/genética , FezesRESUMO
To analyze the effect of levofloxacin-induced intestinal microbiota modifications on intestinal, joint, and systemic inflammation in the DBA/1 mice with spontaneous arthritis. The study included two groups of mice, one of which received levofloxacin. The composition and structure of the microbiota were determined in the mice's stool using 16S rRNA sequencing; the differential taxa and metabolic pathway between mice treated with levofloxacin and control mice were also defied. The effect of levofloxacin was evaluated in the intestines, hind paws, and spines of mice through DNA microarray transcriptome and histopathological analyses; systemic inflammation was measured by flow cytometry. Levofloxacin decreased the pro-inflammatory bacteria, including Prevotellaceae, Odoribacter, and Blautia, and increased the anti-inflammatory Muribaculaceae in mice's stool. Histological analysis confirmed the intestinal inflammation in control mice, while in levofloxacin-treated mice, inflammation was reduced; in the hind paws and spines, levofloxacin also decreased the inflammation. Microarray showed the downregulation of genes and signaling pathways relevant in spondyloarthritis, including several cytokines and chemokines. Levofloxacin-treated mice showed differential transcriptomic profiles between peripheral and axial joints and intestines. Levofloxacin decreased the expression of TNF-α, IL-23a, and JAK3 in the three tissues, but IL-17 behaved differently in the intestine and the joints. Serum TNF-α was also reduced in levofloxacin-treated mice. Our results suggest that the microbiota modification aimed at reducing pro-inflammatory and increasing anti-inflammatory bacteria could potentially be a coadjuvant in treating inflammatory arthropathies.
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Levofloxacino , Espondilartrite , Camundongos , Animais , Levofloxacino/farmacologia , Transcriptoma , Fator de Necrose Tumoral alfa/genética , Disbiose/microbiologia , RNA Ribossômico 16S/genética , Camundongos Endogâmicos DBA , Inflamação/tratamento farmacológico , Inflamação/genética , Inflamação/patologia , Camundongos Endogâmicos C57BLRESUMO
Adaptations of ciliates to hypoxic environments have arisen independently several times. Studies on mitochondrion-related organelle (MRO) metabolisms from distinct anaerobic ciliate groups provide evidence for understanding the transitions from mitochondria to MROs within eukaryotes. To deepen our knowledge about the evolutionary patterns of ciliate anaerobiosis, mass-culture and single-cell transcriptomes of two anaerobic species, Metopus laminarius (class Armophorea) and Plagiopyla cf. narasimhamurtii (class Plagiopylea), were sequenced and their MRO metabolic maps were compared. In addition, we carried out comparisons using publicly available predicted MRO proteomes from other ciliate classes (i.e., Armophorea, Litostomatea, Muranotrichea, Oligohymenophorea, Parablepharismea and Plagiopylea). We found that single-cell transcriptomes were similarly comparable to their mass-culture counterparts in predicting MRO metabolic pathways of ciliates. The patterns of the components of the MRO metabolic pathways might be divergent among anaerobic ciliates, even among closely related species. Notably, our findings indicate the existence of group-specific functional relics of electron transport chains (ETCs). Detailed group-specific ETC functional patterns are as follows: full oxidative phosphorylation in Oligohymenophorea and Muranotrichea; only electron-transfer machinery in Armophorea; either of these functional types in Parablepharismea; and ETC functional absence in Litostomatea and Plagiopylea. These findings suggest that adaptation of ciliates to anaerobic conditions is group-specific and has occurred multiple times. Our results also show the potential and the limitations of detecting ciliate MRO proteins using single-cell transcriptomes and improve the understanding of the multiple transitions from mitochondria to MROs within ciliates. Supplementary Information: The online version contains supplementary material available at 10.1007/s42995-022-00147-w.
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Cells replicate and segregate their DNA with precision. Previous studies showed that these regulated cell-cycle processes were present in the last eukaryotic common ancestor and that their core molecular parts are conserved across eukaryotes. However, some metamonad parasites have secondarily lost components of the DNA processing and segregation apparatuses. To clarify the evolutionary history of these systems in these unusual eukaryotes, we generated a genome assembly for the free-living metamonad Carpediemonas membranifera and carried out a comparative genomics analysis. Here, we show that parasitic and free-living metamonads harbor an incomplete set of proteins for processing and segregating DNA. Unexpectedly, Carpediemonas species are further streamlined, lacking the origin recognition complex, Cdc6 and most structural kinetochore subunits. Carpediemonas species are thus the first known eukaryotes that appear to lack this suite of conserved complexes, suggesting that they likely rely on yet-to-be-discovered or alternative mechanisms to carry out these fundamental processes.
Assuntos
Evolução Biológica , Eucariotos/genética , Genoma , Genômica , Animais , DNA/metabolismo , Células Eucarióticas/metabolismo , Microbiologia , Parasitos/genética , Proteínas/genética , Proteínas/metabolismoRESUMO
Comparative phylogeography offers a unique opportunity to understand the interplay between past environmental events and life-history traits on diversification of unrelated but co-distributed species. Here, we examined the effects of the quaternary climate fluctuations and palaeomarine currents and present-day marine currents on the extant patterns of genetic diversity in the two most conspicuous mangrove species of the Neotropics. The black (Avicennia germinans, Avicenniaceae) and the red (Rhizophora mangle, Rhizophoraceae) mangroves have similar geographic ranges but are very distantly related and show striking differences on their life-history traits. We sampled 18 Atlantic and 26 Pacific locations for A. germinans (N = 292) and R. mangle (N = 422). We performed coalescence simulations using microsatellite diversity to test for evidence of population change associated with quaternary climate fluctuations. In addition, we examined whether patterns of genetic variation were consistent with the directions of major marine (historical and present day) currents in the region. Our demographic analysis was grounded within a phylogeographic framework provided by the sequence analysis of two chloroplasts and one flanking microsatellite region in a subsample of individuals. The two mangrove species shared similar biogeographic histories including: (1) strong genetic breaks between Atlantic and Pacific ocean basins associated with the final closure of the Central American Isthmus (CAI), (2) evidence for simultaneous population declines between the mid-Pleistocene and early Holocene, (3) asymmetric historical migration with higher gene flow from the Atlantic to the Pacific oceans following the direction of the palaeomarine current, and (4) contemporary gene flow between West Africa and South America following the major Atlantic Ocean currents. Despite the remarkable differences in life-history traits of mangrove species, which should have had a strong influence on seed dispersal capability and, thus, population connectivity, we found that vicariant events, climate fluctuations and marine currents have shaped the distribution of genetic diversity in strikingly similar ways.
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Several individuals of the Caribbean Zamia clade and other cycad genera were used to identify single-copy nuclear genes for phylogeographic and phylogenetic studies in Cycadales. Two strategies were employed to select target loci: (i) a tblastX search of Arabidopsis conserved ortholog sequence (COS) set and (ii) a tblastX search of Arabidopsis-Populus-Vitis-Oryza Shared Single-Copy genes (APVO SSC) against the EST Zamia databases in GenBank. From the first strategy, 30 loci were selected, and from the second, 16 loci. In both cases, the matching GenBank accessions of Zamia were used as a query for retrieving highly similar sequences from Cycas, Picea, Pinus species or Ginkgo biloba. After retrieving and aligning all the sequences in each locus, intron predictions were completed to assist in primer design. PCR was carried out in three rounds to detect paralogous loci. A total of 29 loci were successfully amplified as a single band of which 20 were likely single-copy loci. These loci showed different diversity and divergence levels. A preliminary screening allowed us to select 8 promising loci (40S, ATG2, BG, GroES, GTP, LiSH, PEX4 and TR) for the Zamia pumila complex and 4 loci (COS26, GroES, GTP and HTS) for all other cycad genera.