RESUMO
Water lilies belong to the angiosperm order Nymphaeales. Amborellales, Nymphaeales and Austrobaileyales together form the so-called ANA-grade of angiosperms, which are extant representatives of lineages that diverged the earliest from the lineage leading to the extant mesangiosperms1-3. Here we report the 409-megabase genome sequence of the blue-petal water lily (Nymphaea colorata). Our phylogenomic analyses support Amborellales and Nymphaeales as successive sister lineages to all other extant angiosperms. The N. colorata genome and 19 other water lily transcriptomes reveal a Nymphaealean whole-genome duplication event, which is shared by Nymphaeaceae and possibly Cabombaceae. Among the genes retained from this whole-genome duplication are homologues of genes that regulate flowering transition and flower development. The broad expression of homologues of floral ABCE genes in N. colorata might support a similarly broadly active ancestral ABCE model of floral organ determination in early angiosperms. Water lilies have evolved attractive floral scents and colours, which are features shared with mesangiosperms, and we identified their putative biosynthetic genes in N. colorata. The chemical compounds and biosynthetic genes behind floral scents suggest that they have evolved in parallel to those in mesangiosperms. Because of its unique phylogenetic position, the N. colorata genome sheds light on the early evolution of angiosperms.
Assuntos
Genoma de Planta , Nymphaea/genética , Filogenia , Flores/genética , Flores/metabolismo , Nymphaea/metabolismo , Odorantes/análiseRESUMO
As a midsized gene family conserved more by lineage than function, the typical plant terpene synthases (TPSs) could be a valuable tool to examine plant evolution. TPSs are pivotal in biosynthesis of gibberellins and related phytohormones as well as in formation of the extensive arsenal of specialized plant metabolites mediating ecological interactions whose production is often lineage specific. Yet the origin and early evolution of the TPS family is not well understood. Systematic analysis of an array of transcriptomes and sequenced genomes indicated that the TPS family originated after the divergence of land plants from charophytic algae. Phylogenetic and biochemical analyses support the hypothesis that the ancestral TPS gene encoded a bifunctional class I and II diterpene synthase producing the ent-kaurene required for phytohormone production in all extant lineages of land plants. Moreover, the ancestral TPS gene likely underwent duplication at least twice early in land plant evolution. Together these two gave rise to three TPS lineages leading to the extant TPS-c, TPS-e/f, and the remaining TPS (h/d/a/b/g) subfamilies, with the latter dedicated to secondary rather than primary metabolism while the former two contain those genes involved in ent-kaurene production. Nevertheless, parallel evolution from the ent-kaureneproducing class I and class II diterpene synthases has led to roles for TPS-e/f and -c subfamily members in secondary metabolism as well. These results clarify TPS evolutionary history and provide context for the role of these genes in producing the vast diversity of terpenoid natural products observed today in various land plant lineages.
Assuntos
Alquil e Aril Transferases , Embriófitas , Evolução Molecular , Proteínas de Plantas , Alquil e Aril Transferases/classificação , Alquil e Aril Transferases/genética , Embriófitas/enzimologia , Embriófitas/genética , Duplicação Gênica , Filogenia , Reguladores de Crescimento de Plantas , Proteínas de Plantas/classificação , Proteínas de Plantas/genética , Terpenos/metabolismoRESUMO
Chimeric terpene synthases, which consist of C-terminal prenyltransferase (PT) and N-terminal class I terpene synthase (TS) domains (termed PTTSs here), is unique to fungi and produces structurally diverse di- and sesterterpenes. Prior to this study, 20 PTTSs had been functionally characterized. Our understanding of the origin and functional evolution of PTTS genes is limited. Our systematic search of sequenced fungal genomes among diverse taxa revealed that PTTS genes were restricted to Dikarya. Phylogenetic findings indicated different potential models of the origin and evolution of PTTS genes. One was that PTTS genes originated in the common Dikarya ancestor and then underwent frequent gene loss among various subsequent lineages. To understand their functional evolution, we selected 74 PTTS genes for biochemical characterization in an efficient precursor-providing yeast system employing chassis-based, robot-assisted, high-throughput automatic assembly. We found 34 PTTS genes that encoded active enzymes and collectively produced 24 di- and sesterterpenes. About half of these di- and sesterterpenes were also the products of the 20 known PTTSs, indicating functional conservation, whereas the PTTS products included the previously unknown sesterterpenes, sesterevisene (1), and sesterorbiculene (2), suggesting that a diversity of PTTS products awaits discovery. Separating functional PTTSs into two monophyletic groups implied that an early gene duplication event occurred during the evolution of the PTTS family followed by functional divergence with the characteristics of distinct cyclization mechanisms.
Assuntos
Alquil e Aril Transferases/genética , Proteínas Fúngicas/genética , Proteínas Mutantes Quiméricas/genética , Alquil e Aril Transferases/metabolismo , Diterpenos/química , Diterpenos/metabolismo , Evolução Molecular , Proteínas Fúngicas/metabolismo , Fungos/classificação , Fungos/enzimologia , Fungos/genética , Genoma Fúngico/genética , Estrutura Molecular , Proteínas Mutantes Quiméricas/metabolismo , Mutação , Filogenia , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Sesterterpenos/química , Sesterterpenos/metabolismoRESUMO
BACKGROUND: Despite preventive measures, infections continue to pose significant risks to pediatric allogeneic hematopoietic cell transplantation (allo-HCT) recipients. The gut microbiota has been linked to clinical outcomes following adult allo-HCT. This study evaluated whether similar disruptions or differing microbiota patterns were associated with infection risk in pediatric allo-HCT. METHODS: In a prospective observational study, fecal samples were obtained from 74 children before conditioning and upon neutrophil recovery. Microbiome signatures identified through sequencing were examined for their associations with infections or acute graft-versus-host disease (aGVHD) in the first-year post-HCT using Cox proportional hazards analysis. RESULTS: Microbiome disruption in adults, did not predict infection risk in pediatric allo-HCT. Unique microbiota signatures were associated with different infections or aGVHD. A ratio of strict and facultative anaerobes (eg, Lachnoclostridium, Parabacteroides) prior to conditioning predicted bacteremia risk (Cox hazard ratio [HR], 3.89). A distinct ratio of oral (eg, Rothia, Veillonella) to intestinal anaerobes (eg, Anaerobutyricum, Romboutsia) at neutrophil recovery predicted likelihood of bacterial infections (Cox HR, 1.81) and viral enterocolitis (Cox HR, 1.96). CONCLUSIONS: Interactions between medical interventions, pediatric hosts, and microbial communities contribute to microbiota signatures that predict infections. Further multicenter study is necessary to validate the generalizability of these ratios as biomarkers.
Assuntos
Microbioma Gastrointestinal , Doença Enxerto-Hospedeiro , Transplante de Células-Tronco Hematopoéticas , Adulto , Humanos , Criança , Transplante de Células-Tronco Hematopoéticas/efeitos adversos , Bactérias/genética , Fezes/microbiologiaRESUMO
Red algae (Rhodophyta) and land plants belong to the monophyletic clade Archaeplastida, and taxa of both groups are rich producers of terpene secondary metabolites. The terpene carbon skeletons of land plants are made by two types of terpene synthases: typical plant terpene synthases and microbial-type terpene synthases (MTPSLs); however, terpene biosynthesis in red algae is poorly understood. By systematic sequence analysis of seven genomes and 34 transcriptomes of red algae, MTPSL homologs were identified within one genome and two transcriptomes, whereas no homolog of typical plant terpene synthase genes was found. Phylogenetic analysis showed that red algae MTPSLs group with bacterial terpene synthases. Analysis of the genome assembly and characterization of neighboring genes demonstrated red algal MTPSLs to be bona fide red algal genes and not microbial contaminants. MTPSL genes from Porphyridium purpureum and Erythrolobus australicus were characterized via heterologous expression in Escherichia coli and demonstrated to have sesquiterpene synthase activities. We detected a number of volatile sesquiterpenes in the headspace of P. purpureum and E. australicus cultures, most identical to the in vitro products of the respective MTPSLs. Expression of the MTPSL gene in P. purpureum was found to be induced by methyl jasmonate, suggesting a role for this gene in host defense. In summary, this study indicates that the formation of terpene carbon skeletons in red algae is carried out by MTPSLs that are phylogenetically unrelated to typical plant terpene synthases and most likely originated in Rhodophyta via horizontal gene transfer from bacteria.
Assuntos
Proteínas de Algas/metabolismo , Alquil e Aril Transferases/metabolismo , Proteínas de Bactérias/metabolismo , Rodófitas/genética , Terpenos/metabolismo , Acetatos/farmacologia , Proteínas de Algas/genética , Alquil e Aril Transferases/genética , Proteínas de Bactérias/genética , Ciclopentanos/farmacologia , Evolução Molecular , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Oxilipinas/farmacologia , Filogenia , Porphyridium/efeitos dos fármacos , Porphyridium/genética , Porphyridium/metabolismo , Rodófitas/citologia , Rodófitas/metabolismo , Sesquiterpenos/análise , Sesquiterpenos/metabolismo , Técnicas de Cultura de Tecidos , Compostos Orgânicos Voláteis/análise , Compostos Orgânicos Voláteis/metabolismoRESUMO
Marchantia polymorpha is a basal terrestrial land plant, which like most liverworts accumulates structurally diverse terpenes believed to serve in deterring disease and herbivory. Previous studies have suggested that the mevalonate and methylerythritol phosphate pathways, present in evolutionarily diverged plants, are also operative in liverworts. However, the genes and enzymes responsible for the chemical diversity of terpenes have yet to be described. In this study, we resorted to a HMMER search tool to identify 17 putative terpene synthase genes from M. polymorpha transcriptomes. Functional characterization identified four diterpene synthase genes phylogenetically related to those found in diverged plants and nine rather unusual monoterpene and sesquiterpene synthase-like genes. The presence of separate monofunctional diterpene synthases for ent-copalyl diphosphate and ent-kaurene biosynthesis is similar to orthologs found in vascular plants, pushing the date of the underlying gene duplication and neofunctionalization of the ancestral diterpene synthase gene family to >400 million years ago. By contrast, the mono- and sesquiterpene synthases represent a distinct class of enzymes, not related to previously described plant terpene synthases and only distantly so to microbial-type terpene synthases. The absence of a Mg2+ binding, aspartate-rich, DDXXD motif places these enzymes in a noncanonical family of terpene synthases.
Assuntos
Alquil e Aril Transferases/metabolismo , Marchantia/enzimologia , Marchantia/metabolismo , Alquil e Aril Transferases/genética , Evolução Molecular , Marchantia/genética , Transcriptoma/genéticaRESUMO
Terpenes are structurally diverse natural products involved in many ecological interactions. The pivotal enzymes for terpene biosynthesis, terpene synthases (TPSs), had been described only in plants and fungi in the eukaryotic domain. In this report, we systematically analyzed the genome sequences of a broad range of nonplant/nonfungus eukaryotes and identified putative TPS genes in six species of amoebae, five of which are multicellular social amoebae from the order of Dictyosteliida. A phylogenetic analysis revealed that amoebal TPSs are evolutionarily more closely related to fungal TPSs than to bacterial TPSs. The social amoeba Dictyostelium discoideum was selected for functional study of the identified TPSs. D. discoideum grows as a unicellular organism when food is abundant and switches from vegetative growth to multicellular development upon starvation. We found that expression of most D. discoideum TPS genes was induced during development. Upon heterologous expression, all nine TPSs from D. discoideum showed sesquiterpene synthase activities. Some also exhibited monoterpene and/or diterpene synthase activities. Direct measurement of volatile terpenes in cultures of D. discoideum revealed essentially no emission at an early stage of development. In contrast, a bouquet of terpenes, dominated by sesquiterpenes including ß-barbatene and (E,E)-α-farnesene, was detected at the middle and late stages of development, suggesting a development-specific function of volatile terpenes in D. discoideum. The patchy distribution of TPS genes in the eukaryotic domain and the evidence for TPS function in D. discoideum indicate that the TPS genes mediate lineage-specific adaptations.
Assuntos
Alquil e Aril Transferases/genética , Dictyostelium/genética , Genoma de Protozoário , Filogenia , Proteínas de Protozoários/genética , Terpenos/metabolismo , Adaptação Fisiológica , Alquil e Aril Transferases/classificação , Alquil e Aril Transferases/metabolismo , Evolução Biológica , Clonagem Molecular , Dictyostelium/classificação , Dictyostelium/enzimologia , Escherichia coli/genética , Escherichia coli/metabolismo , Expressão Gênica , Regulação da Expressão Gênica no Desenvolvimento , Isoenzimas/classificação , Isoenzimas/genética , Isoenzimas/metabolismo , Família Multigênica , Proteínas de Protozoários/classificação , Proteínas de Protozoários/metabolismo , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , VolatilizaçãoRESUMO
The vast abundance of terpene natural products in nature is due to enzymes known as terpene synthases (TPSs) that convert acyclic prenyl diphosphate precursors into a multitude of cyclic and acyclic carbon skeletons. Yet the evolution of TPSs is not well understood at higher levels of classification. Microbial TPSs from bacteria and fungi are only distantly related to typical plant TPSs, whereas genes similar to microbial TPS genes have been recently identified in the lycophyte Selaginella moellendorffii The goal of this study was to investigate the distribution, evolution, and biochemical functions of microbial terpene synthase-like (MTPSL) genes in other plants. By analyzing the transcriptomes of 1,103 plant species ranging from green algae to flowering plants, putative MTPSL genes were identified predominantly from nonseed plants, including liverworts, mosses, hornworts, lycophytes, and monilophytes. Directed searching for MTPSL genes in the sequenced genomes of a wide range of seed plants confirmed their general absence in this group. Among themselves, MTPSL proteins from nonseed plants form four major groups, with two of these more closely related to bacterial TPSs and the other two to fungal TPSs. Two of the four groups contain a canonical aspartate-rich "DDxxD" motif. The third group has a "DDxxxD" motif, and the fourth group has only the first two "DD" conserved in this motif. Upon heterologous expression, representative members from each of the four groups displayed diverse catalytic functions as monoterpene and sesquiterpene synthases, suggesting these are important for terpene formation in nonseed plants.
Assuntos
Alquil e Aril Transferases/genética , Evolução Molecular , Filogenia , Transcriptoma/genética , Clorófitas/genética , Mapeamento Cromossômico , Embriófitas/genética , Perfilação da Expressão Gênica , Proteínas de Plantas/genética , Sementes/genéticaRESUMO
Circadian control of gene expression is well characterized at the transcriptional level, but little is known about diel or circadian control of translation. Genome-wide translation state profiling of mRNAs in Arabidopsis thaliana seedlings grown in long day was performed to estimate ribosome loading per mRNA. The experiments revealed extensive translational regulation of key biological processes. Notably, translation of mRNAs for ribosomal proteins and mitochondrial respiration peaked at night. Central clock mRNAs are among those subject to fluctuations in ribosome loading. There was no consistent phase relationship between peak translation states and peak transcript levels. The overlay of distinct transcriptional and translational cycles can be expected to alter the waveform of the protein synthesis rate. Plants that constitutively overexpress the clock gene CCA1 showed phase shifts in peak translation, with a 6-h delay from midnight to dawn or from noon to evening being particularly common. Moreover, cycles of ribosome loading that were detected under continuous light in the wild type collapsed in the CCA1 overexpressor. Finally, at the transcript level, the CCA1-ox strain adopted a global pattern of transcript abundance that was broadly correlated with the light-dark environment. Altogether, these data demonstrate that gene-specific diel cycles of ribosome loading are controlled in part by the circadian clock.
Assuntos
Arabidopsis/genética , Relógios Circadianos/genética , Regulação da Expressão Gênica de Plantas , RNA Mensageiro/metabolismo , Ribossomos/genética , Proteínas de Arabidopsis/genética , Ritmo Circadiano/genética , Ontologia Genética , Luz , Biossíntese de Proteínas , RNA Mensageiro/genética , Ribossomos/metabolismo , Fatores de Transcrição/genéticaRESUMO
KEY MESSAGE: Conifers contain P450 enzymes from the CYP79 family that are involved in cyanogenic glycoside biosynthesis. Cyanogenic glycosides are secondary plant compounds that are widespread in the plant kingdom. Their biosynthesis starts with the conversion of aromatic or aliphatic amino acids into their respective aldoximes, catalysed by N-hydroxylating cytochrome P450 monooxygenases (CYP) of the CYP79 family. While CYP79s are well known in angiosperms, their occurrence in gymnosperms and other plant divisions containing cyanogenic glycoside-producing plants has not been reported so far. We screened the transcriptomes of 72 conifer species to identify putative CYP79 genes in this plant division. From the seven resulting full-length genes, CYP79A118 from European yew (Taxus baccata) was chosen for further characterization. Recombinant CYP79A118 produced in yeast was able to convert L-tyrosine, L-tryptophan, and L-phenylalanine into p-hydroxyphenylacetaldoxime, indole-3-acetaldoxime, and phenylacetaldoxime, respectively. However, the kinetic parameters of the enzyme and transient expression of CYP79A118 in Nicotiana benthamiana indicate that L-tyrosine is the preferred substrate in vivo. Consistent with these findings, taxiphyllin, which is derived from L-tyrosine, was the only cyanogenic glycoside found in the different organs of T. baccata. Taxiphyllin showed highest accumulation in leaves and twigs, moderate accumulation in roots, and only trace accumulation in seeds and the aril. Quantitative real-time PCR revealed that CYP79A118 was expressed in plant organs rich in taxiphyllin. Our data show that CYP79s represent an ancient family of plant P450s that evolved prior to the separation of gymnosperms and angiosperms. CYP79A118 from T. baccata has typical CYP79 properties and its substrate specificity and spatial gene expression pattern suggest that the enzyme contributes to the formation of taxiphyllin in this plant species.
Assuntos
Sistema Enzimático do Citocromo P-450/metabolismo , Nitrilas/metabolismo , Taxus/enzimologia , Sequência de Aminoácidos , Sistema Enzimático do Citocromo P-450/química , Sistema Enzimático do Citocromo P-450/genética , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Nitrilas/química , Especificidade de Órgãos/genética , Proteínas de Plantas/química , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Alinhamento de Sequência , Taxus/genética , Transcriptoma/genéticaRESUMO
Due to an unfortunate turn of events, the funding note for Open Access publication was not properly provided in the original publication. Hence, the original article has been corrected. The opening line of the Acknowledgement section should read.
RESUMO
BACKGROUND: Terpenoids constitute the largest class of secondary metabolites made by plants and display vast chemical diversity among and within species. Terpene synthases (TPSs) are the pivotal enzymes for terpenoid biosynthesis that create the basic carbon skeletons of this class. Functional divergence of paralogous and orthologous TPS genes is a major mechanism for the diversification of terpenoid biosynthesis. However, little is known about the evolutionary forces that have shaped the evolution of plant TPS genes leading to terpenoid diversity. RESULTS: The orthologs of Oryza Terpene Synthase 1 (OryzaTPS1), a rice terpene synthase gene involved in indirect defense against insects in Oryza sativa, were cloned from six additional Oryza species. In vitro biochemical analysis showed that the enzymes encoded by these OryzaTPS1 genes functioned either as (E)-ß-caryophyllene synthases (ECS), or (E)-ß-caryophyllene & germacrene A synthases (EGS), or germacrene D & germacrene A synthases (DAS). Because the orthologs of OryzaTPS1 in maize and sorghum function as ECS, the ECS activity was inferred to be ancestral. Molecular evolutionary detected the signature of positive Darwinian selection in five codon substitutions in the evolution from ECS to DAS. Homology-based structure modeling and the biochemical analysis of laboratory-generated protein variants validated the contribution of the five positively selected sites to functional divergence of OryzaTPS1. The changes in the in vitro product spectra of OryzaTPS1 proteins also correlated closely to the changes in in vivo blends of volatile terpenes released from insect-damaged rice plants. CONCLUSIONS: In this study, we found that positive Darwinian selection is a driving force for the functional divergence of OryzaTPS1. This finding suggests that the diverged sesquiterpene blend produced by the Oryza species containing DAS may be adaptive, likely in the attraction of the natural enemies of insect herbivores.
Assuntos
Alquil e Aril Transferases/genética , Alquil e Aril Transferases/metabolismo , Oryza/enzimologia , Oryza/genética , Seleção Genética/genética , Terpenos/metabolismoRESUMO
Streptococcus pneumoniae is a major human pathogen of global health concern and the rapid emergence of antibiotic resistance poses a serious public health problem worldwide. Fluoroquinolone resistance in S. pneumoniae is an intriguing case because the prevalence of fluoroquinolone resistance does not correlate with increasing usage and has remained rare. Our data indicate that deleterious fitness costs in the mammalian host constrain the emergence of fluoroquinolone resistance both by de novo mutation and recombination. S. pneumoniae was able to circumvent such deleterious fitness costs via the development of antibiotic tolerance through metabolic adaptation that reduced the production of reactive oxygen species, resulting in a fitness benefit during infection of mice treated with fluoroquinolones. These data suggest that the emergence of fluoroquinolone resistance is tightly constrained in S. pneumoniae by fitness tradeoffs and that mutational pathways involving metabolic networks to enable tolerance phenotypes are an important contributor to the evasion of antibiotic-mediated killing.IMPORTANCEThe increasing prevalence of antibiotic resistant bacteria is a major global health concern. While many species have the potential to develop antibiotic resistance, understanding the barriers to resistance emergence in the clinic remains poorly understood. A prime example of this is fluroquinolone resistance in Streptococcus pneumoniae, whereby, despite continued utilization, resistance to this class of antibiotic remains rare. In this study, we found that the predominant pathways for developing resistance to this antibiotic class severely compromised the infectious capacity of the pneumococcus, providing a key impediment for the emergence of resistance. Using in vivo models of experimental evolution, we found that S. pneumoniae responds to repeated fluoroquinolone exposure by modulating key metabolic pathways involved in the generation of redox molecules, which leads to antibiotic treatment failure in the absence of appreciable shifts in resistance levels. These data underscore the complex pathways available to pathogens to evade antibiotic mediating killing via antibiotic tolerance.
Assuntos
Fluoroquinolonas , Infecções Pneumocócicas , Humanos , Animais , Camundongos , Fluoroquinolonas/farmacologia , Streptococcus pneumoniae/metabolismo , Infecções Pneumocócicas/tratamento farmacológico , Infecções Pneumocócicas/microbiologia , Farmacorresistência Bacteriana/genética , Testes de Sensibilidade Microbiana , Antibacterianos/farmacologia , Antibacterianos/metabolismo , MamíferosRESUMO
MISTRG is an immunodeficient mouse strain that expresses multiple human cytokines that support hematopoietic stem cell maintenance and myelopoiesis. While establishing a breeding colony of MISTRG mice in a dedicated barrier room, 6 cases of death or disease occurred in pregnant or postpartum mice. Clinically, this manifested as hunched posture, dyspnea, and 1 case of emaciation with ataxia. Pathologic analysis of 7 mice revealed multisystemic necrosuppurative inflammation variably affecting the uterus and placenta, joints, meninges, inner and middle ears, kidneys, and small intestine. Bacteria cultured from the blood of septic mice were identified with 89% probability by the Vitek 2 identification system as Streptococcus sanguinus with atypical biochemical parameters; the API 20E/NE system fully differentiated the isolates as a novel Streptococcus species. MALDI Biotyper-based mass spectrometry also indicated that the phenotype represented a novel Streptococcus spp. Sequencing revealed that the full-length 16S rRNA gene identity was below 97% with known Streptococcus species, including the 2 closest species Streptococcus acidominimus and Streptococcus azizii. We propose the name Streptococcus murisepticum spp. nov to our novel isolates. All male mice in this colony remained healthy despite their association with diseased female mice. Overall, 19% of the colony carried the novel Streptococcus in their oral cavity, but it could not be detected in feces. The organism was sensitive to amoxicillin, which was administered via drinking water throughout pregnancy and weaning to establish a colony of pathogen-negative future breeders. The colony remained disease-free and culture-negative for Streptococcus murisepticum spp. nov after treatment with amoxicillin. We suspect that oral colonization of MISTRG mice with the novel Streptococcus species and its associated unique pathology in periparturient mice is potentially the principal cause of loss of this strain at several institutions. Therefore, screening the oral cavity for α-hemolytic streptococci followed by targeted antibiotic treatment may be necessary when establishing MISTRG and allied immunodeficient mouse strains.
Assuntos
Infecções Estreptocócicas , Gravidez , Masculino , Feminino , Humanos , Animais , Camundongos , Infecções Estreptocócicas/diagnóstico , RNA Ribossômico 16S/genética , Streptococcus/genética , Amoxicilina , BocaRESUMO
As opposed to de novo mutation, ß-lactam resistance in S. pneumoniae is often conferred via homologous recombination during horizontal gene transfer. We hypothesize that ß-lactam resistance in pathogenic streptococci is restricted to naturally competent species via intra-/interspecies recombination due to in vivo fitness trade-offs of de novo penicillin-binding protein (PBP) mutations. We show that de novo mutant populations have abrogated invasive disease capacity and are difficult to evolve in vivo. Conversely, serially transformed recombinant strains efficiently integrate resistant oral streptococcal DNA, gain penicillin resistance and tolerance, and retain virulence in mice. Large-scale changes in pbp2X, pbp2B, and non-PBP-related genes occur in recombinant isolates. Our results indicate that horizontal transfer of ß-lactam resistance engenders initially favorable or minimal cost changes in vivo compared with de novo mutation(s), underscoring the importance of recombination in the emergence of ß-lactam resistance and suggesting why some pathogenic streptococci lacking innate competence remain universally susceptible.
Assuntos
Infecções Pneumocócicas , Streptococcus pneumoniae , Camundongos , Animais , Streptococcus pneumoniae/genética , Transferência Genética Horizontal , Virulência/genética , Testes de Sensibilidade Microbiana , Resistência beta-Lactâmica/genética , Proteínas de Ligação às Penicilinas/genética , Proteínas de Ligação às Penicilinas/metabolismo , Mutação/genética , Proteínas de Bactérias/metabolismo , Antibacterianos/farmacologia , Antibacterianos/metabolismoRESUMO
Thirty-one potential miRNAs that belong to 16 miRNA families were discovered from more than 324 000 EST sequences of apple (Malus domestica). In addition, precise sequences, especially terminal nucleotides of the 16 apple miRNAs (mdo-miRNAs) in 16 families were validated by miR-RACE, a newly developed method for the determination of the potential miRNAs predicted computationally. The expression of these 16 microRNAs could be detected in apple young leaf, old leaf, young stem, flower bud, flower and developing fruits by quantitative RT-PCR (qRT-PCR) and some of them showed tissue-specific expression. Fifty-six potential targets were identified for the 16 apple miRNAs, most of which were transcription factors that play important roles in apple development. Twelve target genes were experimentally verified by qRT-PCR, with some exhibiting different expression trends from their corresponding microRNAs, indicating the cleavage mode of miRNAs on their target genes.
Assuntos
Biologia Computacional/métodos , Etiquetas de Sequências Expressas , Regulação da Expressão Gênica de Plantas , Malus/genética , MicroRNAs/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Análise de Sequência de RNA/métodos , Sequência de Bases , Clonagem Molecular , Biblioteca Gênica , Genes de Plantas/genética , MicroRNAs/metabolismo , Dados de Sequência Molecular , Conformação de Ácido Nucleico , Reprodutibilidade dos TestesRESUMO
Piwi genes play an important role in regulating spermatogenesis and oogenesis because they participate in the biogenesis of piRNAs, a new class of noncoding RNAs. However, these genes are not well understood in most insects. To understand the function of piwi genes in honeybee reproduction, we amplified two full-length piwi-like genes, Am-aub and Am-ago3. Both the cloned Am-aub and Am-ago3 genes contained typical PAZ and PIWI domains and active catalytic motifs "Asp-Asp-Asp/His/Glu/Lys," suggesting that the two piwi-like genes possessed slicer activity. We examined the expression levels of Am-aub and Am-ago3 in workers, queens, drones, and female larvae by quantitative PCR. Am-aub was more abundant than Am-ago3 in all the tested samples. Both Am-aub and Am-ago3 were highly expressed in drones but not in workers and queens. The significant finding was that the larval food stream influenced the expression of Piwi genes in adult honeybees. This helps to understand the nutritional control of reproductive status in honeybees at the molecular level.
Assuntos
Abelhas/genética , Abelhas/fisiologia , Regulação da Expressão Gênica/fisiologia , Proteínas de Insetos/genética , Proteínas de Insetos/metabolismo , Animais , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Larva/genética , Larva/metabolismo , Masculino , Dados de Sequência Molecular , Filogenia , ReproduçãoRESUMO
Fungi are successful eukaryotes of wide distribution. They are known as rich producers of secondary metabolites, especially terpenoids, which are important for fungi-environment interactions. Horizontal gene transfer (HGT) is an important mechanism contributing to genetic innovation of fungi. However, it remains unclear whether HGT has played a role in creating the enormous chemical diversity of fungal terpenoids. Here we report that fungi have acquired terpene synthase genes (TPSs), which encode pivotal enzymes for terpenoid biosynthesis, from bacteria through HGT. Phylogenetic analysis placed the majority of fungal and bacterial TPS genes from diverse taxa into two clades, indicating ancient divergence. Nested in the bacterial TPS clade is a number of fungal TPS genes that are inferred as the outcome of HGT. These include a monophyletic clade of nine fungal TPS genes, designated as BTPSL for bacterial TPS-like genes, from eight species of related entomopathogenic fungi, including seven TPSs from six species in the genus Metarhizium. In vitro enzyme assays demonstrate that all seven BTPSL genes from the genus Metarhizium encode active enzymes with sesquiterpene synthase activities of two general product profiles. By analyzing the catalytic activity of two resurrected ancestral BTPSLs and one closely related bacterial TPS, the trajectory of functional evolution of BTPSLs after HGT from bacteria to fungi and functional divergence within Metarhizium could be traced. Using M. brunneum as a model species, both BTPSLs and typical fungal TPSs were demonstrated to be involved in the in vivo production of terpenoids, illustrating the general importance of HGT of TPS genes from bacteria as a mechanism contributing to terpenoid diversity in fungi.
Assuntos
Alquil e Aril Transferases/genética , Bactérias/genética , Transferência Genética Horizontal , Hypocreales/genética , Hypocreales/metabolismo , Terpenos/metabolismo , Alquil e Aril Transferases/metabolismo , Bactérias/enzimologia , Biocatálise , Genoma Fúngico/genética , FilogeniaRESUMO
Terpenes constitute a large class of plant secondary metabolites. It was once presumed that these compounds are biosynthesized by typical plant terpene synthases in all land plants. This view has changed with the identification of a new group of terpene synthase genes called MTPSLs for microbial terpene synthase-like genes. MTPSLs are structurally and phylogenetically more related to bacterial and fungal terpene synthases than to typical plant terpene synthases. They are widely distributed in nonseed plants but absent in seed plants and green algae. Much of the terpene diversity in nonseed plants is presumed to be determined by MTPSLs. Phylogenetic analysis suggests that ancestral MTPSL genes were acquired by early land plants from bacteria and fungi through horizontal gene transfer.
Assuntos
Alquil e Aril Transferases/genética , Alquil e Aril Transferases/metabolismo , Evolução Molecular , Proteínas de Plantas/metabolismo , Plantas/metabolismo , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Clorófitas/enzimologia , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Transferência Genética Horizontal , Filogenia , Proteínas de Plantas/genética , Plantas/genética , Metabolismo Secundário , Sementes , Terpenos/metabolismoRESUMO
Obesity is a risk factor for heart disease, stroke, diabetes, high blood pressure, and other chronic diseases. Some drugs, including fenofibrate, are used to treat obesity or excessive weight by lowering the level of specific triglycerides. However, different groups have different drug sensitivities and, consequently, there are differences in drug effects. In this study, we assessed both genetic and nongenetic factors that influence drug responses and stratified patients into groups based on differential drug effect and sensitivity. Our methodology of investigating genetic factors and nongenetic factors is applicable to studying differential effects of other drugs, such as statins, and provides an approach to the development of personalized medicine.