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1.
PLoS Pathog ; 16(9): e1008328, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32936835

RESUMO

Candida albicans cells depend on the energy derived from amino acid catabolism to induce and sustain hyphal growth inside phagosomes of engulfing macrophages. The concomitant deamination of amino acids is thought to neutralize the acidic microenvironment of phagosomes, a presumed requisite for survival and initiation of hyphal growth. Here, in contrast to an existing model, we show that mitochondrial-localized NAD+-dependent glutamate dehydrogenase (GDH2) catalyzing the deamination of glutamate to α-ketoglutarate, and not the cytosolic urea amidolyase (DUR1,2), accounts for the observed alkalization of media when amino acids are the sole sources of carbon and nitrogen. C. albicans strains lacking GDH2 (gdh2-/-) are viable and do not extrude ammonia on amino acid-based media. Environmental alkalization does not occur under conditions of high glucose (2%), a finding attributable to glucose-repression of GDH2 expression and mitochondrial function. Consistently, inhibition of oxidative phosphorylation or mitochondrial translation by antimycin A or chloramphenicol, respectively, prevents alkalization. GDH2 expression and mitochondrial function are derepressed as glucose levels are lowered from 2% (~110 mM) to 0.2% (~11 mM), or when glycerol is used as primary carbon source. Using time-lapse microscopy, we document that gdh2-/- cells survive, filament and escape from primary murine macrophages at rates indistinguishable from wildtype. In intact hosts, such as in fly and murine models of systemic candidiasis, gdh2-/- mutants are as virulent as wildtype. Thus, although Gdh2 has a critical role in central nitrogen metabolism, Gdh2-catalyzed deamination of glutamate is surprisingly dispensable for escape from macrophages and virulence. Consistently, using the pH-sensitive dye (pHrodo), we observed no significant difference between wildtype and gdh2-/- mutants in phagosomal pH modulation. Following engulfment of fungal cells, the phagosomal compartment is rapidly acidified and hyphal growth initiates and sustained under consistently acidic conditions within phagosomes. Together, our results demonstrate that amino acid-dependent alkalization is not essential for hyphal growth, survival in macrophages and hosts. An accurate understanding of the microenvironment within macrophage phagosomes and the metabolic events underlying the survival of phagocytized C. albicans cells and their escape are critical to understanding the host-pathogen interactions that ultimately determine the pathogenic outcome.


Assuntos
Candida albicans/imunologia , Candidíase/imunologia , Drosophila melanogaster/imunologia , Glutamato Desidrogenase/metabolismo , Macrófagos/imunologia , Aminoácidos/genética , Aminoácidos/metabolismo , Animais , Candida albicans/patogenicidade , Candidíase/metabolismo , Candidíase/microbiologia , Drosophila melanogaster/metabolismo , Drosophila melanogaster/microbiologia , Feminino , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Glutamato Desidrogenase/genética , Interações Hospedeiro-Patógeno , Concentração de Íons de Hidrogênio , Macrófagos/microbiologia , Camundongos , Camundongos Endogâmicos C57BL , Nitrogênio , Fagossomos/imunologia , Fagossomos/metabolismo , Fagossomos/microbiologia , Virulência
2.
Nat Commun ; 11(1): 4603, 2020 09 14.
Artigo em Inglês | MEDLINE | ID: mdl-32929079

RESUMO

Amino acid propensities at a site change in the course of protein evolution. This may happen for two reasons. Changes may be triggered by substitutions at epistatically interacting sites elsewhere in the genome. Alternatively, they may arise due to environmental changes that are external to the genome. Here, we design a framework for distinguishing between these alternatives. Using analytical modelling and simulations, we show that they cause opposite dynamics of the fitness of the allele currently occupying the site: it tends to increase with the time since its origin due to epistasis ("entrenchment"), but to decrease due to random environmental fluctuations ("senescence"). By analysing the genomes of vertebrates and insects, we show that the amino acids originating at negatively selected sites experience strong entrenchment. By contrast, the amino acids originating at positively selected sites experience senescence. We propose that senescence of the current allele is a cause of adaptive evolution.


Assuntos
Aminoácidos/genética , Evolução Molecular , Alelos , Sequência de Aminoácidos , Animais , Simulação por Computador , Meio Ambiente , Genes Mitocondriais , Aptidão Genética , Heterogeneidade Genética , Insetos/genética , Seleção Genética , Vertebrados/genética
3.
PLoS One ; 15(8): e0237883, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32866169

RESUMO

Although whole-genome sequencing has provided novel insights into Neisseria meningitidis, many open reading frames have only been annotated as hypothetical proteins with unknown biological functions. Our previous genetic analyses revealed that the hypothetical protein, NMB1345, plays a crucial role in meningococcal infection in human brain microvascular endothelial cells; however, NMB1345 has no homology to any identified protein in databases and its physiological function could not be elucidated using pre-existing methods. Among the many biological technologies to examine transient protein-protein interaction in vivo, one of the developed methods is genetic code expansion with non-canonical amino acids (ncAAs) utilizing a pyrrolysyl-tRNA synthetase/tRNAPyl pair from Methanosarcina species: However, this method has never been applied to assign function-unknown proteins in pathogenic bacteria. In the present study, we developed a new method to genetically incorporate ncAAs-encoded photocrosslinking probes into N. meningitidis by utilizing a pyrrolysyl-tRNA synthetase/tRNAPyl pair and elucidated the biological function(s) of the NMB1345 protein. The results revealed that the NMB1345 protein directly interacts with PilE, a major component of meningococcal pili, and further physicochemical and genetic analyses showed that the interaction between the NMB1345 protein and PilE was important for both functional pilus formation and meningococcal infectious ability in N. meningitidis. The present study using this new methodology for N. meningitidis provides novel insights into meningococcal pathogenesis by assigning the function of a hypothetical protein.


Assuntos
Aminoácidos/genética , Reagentes para Ligações Cruzadas/metabolismo , Luz , Neisseria meningitidis/genética , Proteínas de Bactérias/genética , Proteínas de Bactérias/isolamento & purificação , Encéfalo/irrigação sanguínea , Endocitose , Células Endoteliais/microbiologia , Fímbrias Bacterianas/metabolismo , Humanos , Microvasos/patologia , Mutação/genética , Plasmídeos/genética
4.
BMC Bioinformatics ; 21(1): 340, 2020 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-32738892

RESUMO

BACKGROUND: Ribosome profiling has been widely used for studies of translation under a large variety of cellular and physiological contexts. Many of these studies have greatly benefitted from a series of data-mining tools designed for dissection of the translatome from different aspects. However, as the studies of translation advance quickly, the current toolbox still falls in short, and more specialized tools are in urgent need for deeper and more efficient mining of the important and new features of the translation landscapes. RESULTS: Here, we present RiboMiner, a bioinformatics toolset for mining of multi-dimensional features of the translatome with ribosome profiling data. RiboMiner performs extensive quality assessment of the data and integrates a spectrum of tools for various metagene analyses of the ribosome footprints and for detailed analyses of multiple features related to translation regulation. Visualizations of all the results are available. Many of these analyses have not been provided by previous methods. RiboMiner is highly flexible, as the pipeline could be easily adapted and customized for different scopes and targets of the studies. CONCLUSIONS: Applications of RiboMiner on two published datasets did not only reproduced the main results reported before, but also generated novel insights into the translation regulation processes. Therefore, being complementary to the current tools, RiboMiner could be a valuable resource for dissections of the translation landscapes and the translation regulations by mining the ribosome profiling data more comprehensively and with higher resolution. RiboMiner is freely available at https://github.com/xryanglab/RiboMiner and https://pypi.org/project/RiboMiner .


Assuntos
Biologia Computacional/métodos , Biossíntese de Proteínas , Ribossomos/metabolismo , Software , Motivos de Aminoácidos , Sequência de Aminoácidos , Aminoácidos/genética , Códon/genética , Análise de Dados , Mineração de Dados
5.
PLoS Genet ; 16(8): e1008966, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32776922

RESUMO

The vacuole of the yeast Saccharomyces cerevisiae plays an important role in nutrient storage. Arginine, in particular, accumulates in the vacuole of nitrogen-replete cells and is mobilized to the cytosol under nitrogen starvation. The arginine import and export systems involved remain poorly characterized, however. Furthermore, how their activity is coordinated by nitrogen remains unknown. Here we characterize Vsb1 as a novel vacuolar membrane protein of the APC (amino acid-polyamine-organocation) transporter superfamily which, in nitrogen-replete cells, is essential to active uptake and storage of arginine into the vacuole. A shift to nitrogen starvation causes apparent inhibition of Vsb1-dependent activity and mobilization of stored vacuolar arginine to the cytosol. We further show that this arginine export involves Ypq2, a vacuolar protein homologous to the human lysosomal cationic amino acid exporter PQLC2 and whose activity is detected only in nitrogen-starved cells. Our study unravels the main arginine import and export systems of the yeast vacuole and suggests that they are inversely regulated by nitrogen.


Assuntos
Arginina/metabolismo , Nitrogênio/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/genética , Aminoácidos/genética , Transporte Biológico/genética , Humanos , Membranas Intracelulares/metabolismo , Lisossomos/genética , Proteínas de Membrana Transportadoras/genética , Proteínas de Membrana Transportadoras/metabolismo , Saccharomyces cerevisiae/metabolismo , Vacúolos/genética , Vacúolos/metabolismo
6.
PLoS Pathog ; 16(8): e1008818, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32776973

RESUMO

Leprosy is a chronic disease caused by Mycobacterium leprae. Worldwide, more than 200,000 new patients are affected by leprosy annually, making it the second most common mycobacterial disease after tuberculosis. The MHC/HLA region has been consistently identified as carrying major leprosy susceptibility variants in different populations at times with inconsistent results. To establish the unambiguous molecular identity of classical HLA class I and class II leprosy susceptibility factors, we applied next-generation sequencing to genotype with high-resolution 11 HLA class I and class II genes in 1,155 individuals from a Vietnamese leprosy case-control sample. HLA alleles belonging to an extended haplotype from HLA-A to HLA-DPB1 were associated with risk to leprosy. This susceptibility signal could be reduced to the HLA-DRB1*10:01~ HLA-DQA1*01:05 alleles which were in complete linkage disequilibrium (LD). In addition, haplotypes containing HLA-DRB3~ HLA-DRB1*12:02 and HLA-C*07:06~ HLA-B*44:03~ HLA-DRB1*07:01 alleles were found as two independent protective factors for leprosy. Moreover, we replicated the previously associated HLA-DRB1*15:01 as leprosy risk factor and HLA-DRB1*04:05~HLA-DQA1*03:03 as protective alleles. When we narrowed the analysis to the single amino acid level, we found that the associations of the HLA alleles were largely captured by four independent amino acids at HLA-DRß1 positions 57 (D) and 13 (F), HLA-B position 63 (E) and HLA-A position 19 (K). Hence, analyses at the amino acid level circumvented the ambiguity caused by strong LD of leprosy susceptibility HLA alleles and identified four distinct leprosy susceptibility factors.


Assuntos
Aminoácidos/genética , Predisposição Genética para Doença , Antígenos de Histocompatibilidade Classe II/genética , Antígenos de Histocompatibilidade Classe I/genética , Hanseníase/patologia , Mutação , Adolescente , Adulto , Feminino , Haplótipos , Humanos , Hanseníase/genética , Masculino , Adulto Jovem
7.
RNA ; 26(10): 1431-1447, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32611709

RESUMO

RNA structure influences numerous processes in all organisms. In bacteria, these processes include transcription termination and attenuation, small RNA and protein binding, translation initiation, and mRNA stability, and can be regulated via metabolite availability and other stresses. Here we use Structure-seq2 to probe the in vivo RNA structurome of Bacillus subtilis grown in the presence and absence of amino acids. Our results reveal that amino acid starvation results in lower overall dimethyl sulfate (DMS) reactivity of the transcriptome, indicating enhanced protection owing to protein binding or RNA structure. Starvation-induced changes in DMS reactivity correlated inversely with transcript abundance changes. This correlation was particularly pronounced in genes associated with the stringent response and CodY regulons, which are involved in adaptation to nutritional stress, suggesting that RNA structure contributes to transcript abundance change in regulons involved in amino acid metabolism. Structure-seq2 accurately reported on four known amino acid-responsive riboswitches: T-box, SAM, glycine, and lysine riboswitches. Additionally, we discovered a transcription attenuation mechanism that reduces yfmG expression when amino acids are added to the growth medium. We also found that translation of a leader peptide (YfmH) encoded just upstream of yfmG regulates yfmG expression. Our results are consistent with a model in which a slow rate of yfmH translation caused by limitation of the amino acids encoded in YfmH prevents transcription termination in the yfmG leader region by favoring formation of an overlapping antiterminator structure. This novel RNA switch offers a way to simultaneously monitor the levels of multiple amino acids.


Assuntos
Aminoácidos/genética , Bacillus subtilis/genética , Proteínas de Bactérias/genética , RNA Bacteriano/genética , Regulação Bacteriana da Expressão Gênica/genética , Conformação de Ácido Nucleico , Estabilidade de RNA/genética , Transcrição Genética/genética , Transcriptoma/genética
8.
J Infect Public Health ; 13(10): 1544-1550, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32654959

RESUMO

BACKGROUND: Middle East respiratory syndrome coronavirus (MERS-CoV) has continued to cause sporadic outbreaks of severe respiratory tract infection over the last 8 years. METHODS: Complete genome sequencing using next-generation sequencing was performed for MERS-CoV isolates from cases that occurred in Riyadh between 2015 and 2019. Phylogenetic analysis and molecular mutational analysis were carried out to investigate disease severity. RESULTS: A total of eight MERS-CoV isolates were subjected to complete genome sequencing. Phylogenetic analysis resulted in the assembly of 7/8 sequences within lineage 3 and one sequence within lineage 4 showing complex genomic recombination. The isolates contained a variety of unique amino acid substitutions in ORF1ab (41), the N protein (10), the S protein (9) and ORF4b (5). CONCLUSION: Our study shows that MERS-CoV is evolving. The emergence of new variants carries the potential for increased virulence and could impose a challenge to the global health system. We recommend the sequencing every new MERS-CoV isolate to observe the changes in the virus and relate them to clinical outcomes.


Assuntos
Infecções por Coronavirus/virologia , Coronavírus da Síndrome Respiratória do Oriente Médio/genética , Mutação , RNA Viral/análise , Adulto , Idoso , Aminoácidos/genética , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Proteínas do Nucleocapsídeo/genética , Filogenia , Arábia Saudita , Virulência/genética , Sequenciamento Completo do Genoma
9.
Sheng Wu Gong Cheng Xue Bao ; 36(5): 891-898, 2020 May 25.
Artigo em Chinês | MEDLINE | ID: mdl-32567272

RESUMO

Unnatural amino acid orthogonal translation machinery can insert unnatural amino acids at desired sites of protein through stop codon by means of foreign orthogonal translation system composed of aminoacyl-tRNA synthetase and orthogonal tRNA genes. This new genetic engineering technology is not only a new tool for biochemical researches of proteins, but also an epoch-making technology for the development of new-type live viral vaccines. The mutated virus containing premature termination codon in genes necessary for replication can be propagated in transgenic cells harboring unnatural amino acid orthogonal translation machinery in media with corresponding unnatural amino acid, but it cannot replicate in conventional host cells. This replication-deficient virus is a new-type of live viral vaccine that possesses advantages of high efficacy of traditional attenuated vaccine and high safety of killed vaccine. This article reviews the application and prospect of unnatural amino acid orthogonal translation machinery in the development of novel replication-deficient virus vaccines.


Assuntos
Aminoácidos , Aminoacil-tRNA Sintetases , Engenharia Genética , Vacinas Virais , Aminoácidos/genética , Engenharia de Proteínas , RNA de Transferência
10.
Proc Natl Acad Sci U S A ; 117(27): 15731-15739, 2020 07 07.
Artigo em Inglês | MEDLINE | ID: mdl-32561643

RESUMO

De novo emergence demands a transition from disordered polypeptides into structured proteins with well-defined functions. However, can polypeptides confer functions of evolutionary relevance, and how might such polypeptides evolve into modern proteins? The earliest proteins present an even greater challenge, as they were likely based on abiotic, spontaneously synthesized amino acids. Here we asked whether a primordial function, such as nucleic acid binding, could emerge with ornithine, a basic amino acid that forms abiotically yet is absent in modern-day proteins. We combined ancestral sequence reconstruction and empiric deconstruction to unravel a gradual evolutionary trajectory leading from a polypeptide to a ubiquitous nucleic acid-binding protein. Intermediates along this trajectory comprise sequence-duplicated functional proteins built from 10 amino acid types, with ornithine as the only basic amino acid. Ornithine side chains were further modified into arginine by an abiotic chemical reaction, improving both structure and function. Along this trajectory, function evolved from phase separation with RNA (coacervates) to avid and specific double-stranded DNA binding. Our results suggest that phase-separating polypeptides may have been an evolutionary resource for the emergence of early proteins, and that ornithine, together with its postsynthesis modification to arginine, could have been the earliest basic amino acids.


Assuntos
Arginina/química , Nucleoproteínas/genética , Ornitina/química , Peptídeos/genética , Sequência de Aminoácidos/genética , Aminoácidos/química , Aminoácidos/genética , Arginina/genética , DNA/química , DNA/genética , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/genética , Nucleoproteínas/química , Ornitina/genética , Peptídeos/química , Proteínas/química , Proteínas/genética , RNA/química , RNA/genética
11.
Genes (Basel) ; 11(6)2020 05 29.
Artigo em Inglês | MEDLINE | ID: mdl-32486006

RESUMO

In the last 5 years, frequent outbreaks of infectious bronchitis virus (IBV) are observed in both broiler and layer chicken flocks in the Kingdom of Saudi Arabia (KSA) in spite of extensive usage of vaccines. The IBV is a widespread avian coronavirus affecting both vaccinated and unvaccinated chicken flocks and is attributed to significant economic losses, around the globe. In the present study, 58 (n = 58) samples were collected from four different commercial poultry flocks from 8 KSA districts during 2019. A total of nine positive isolates (9/58; 15.5%), based on real-time reverse transcriptase PCR targeting nucleocapsid (N) gene, were used for further genetic characterization and evolutionary analysis. Genetic characterization of the partial spike (S1) gene revealed the clustering of the reported isolates into three different genotypes, whereas four additional isolates were grouped within 4/91 genotype, two isolates within IS/885 genotype, one isolate was closely related to IS/1494/06, and two isolates were grouped within classic serotype (vaccine-like strains). Phylodynamic revealed clustering of four isolated viruses within GI-13 lineage, three isolates within GI-23 lineage, and two isolates within GI-1 lineage. Results indicate that there are high evolutionary distances between the newly identified IBV strains in this study and the commercially used vaccines (GI-1), suggesting that IBV strains circulating in the KSA are under constant evolutionary pressures. Selective pressure biostatistics analyses consistently demonstrate the presence of a higher positive score which highlights the role of natural selection, a mechanism of virus evolution on sites located on the protein surface, within or nearby domains involved in viral attachment or related functions. Recombination analysis revealed emergence of two isolates through recombination events resulting in new recombinant viruses. Taken together, these finding demonstrate the genetic and evolutionary insights into the currently circulating IBV genotypes in KSA, which could help to better understand the origin, spread, and evolution of infectious bronchitis viruses, and to ascertain the importance of disease monitoring as well as re-evaluation for the currently used vaccines and vaccination programs.


Assuntos
Variação Genética , Vírus da Bronquite Infecciosa/genética , Doenças das Aves Domésticas/virologia , Recombinação Genética , Aminoácidos/genética , Animais , Galinhas/virologia , Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/veterinária , Evolução Molecular , Vírus da Bronquite Infecciosa/isolamento & purificação , Epidemiologia Molecular , Filogenia , Doenças das Aves Domésticas/epidemiologia , Arábia Saudita/epidemiologia , Seleção Genética , Proteínas Virais/genética , Proteínas Virais/metabolismo , Vacinas Virais/uso terapêutico
12.
RNA ; 26(9): 1094-1103, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32385138

RESUMO

N 6-threonylcarbamoyl adenosine (t6A) is a nucleoside modification found in all kingdoms of life at position 37 of tRNAs decoding ANN codons, which functions in part to restrict translation initiation to AUG and suppress frameshifting at tandem ANN codons. In Bacteria the proteins TsaB, TsaC (or C2), TsaD, and TsaE, comprise the biosynthetic apparatus responsible for t6A formation. TsaC(C2) and TsaD harbor the relevant active sites, with TsaC(C2) catalyzing the formation of the intermediate threonylcarbamoyladenosine monophosphate (TC-AMP) from ATP, threonine, and CO2, and TsaD catalyzing the transfer of the threonylcarbamoyl moiety from TC-AMP to A37 of substrate tRNAs. Several related modified nucleosides, including hydroxynorvalylcarbamoyl adenosine (hn6A), have been identified in select organisms, but nothing is known about their biosynthesis. To better understand the mechanism and structural constraints on t6A formation, and to determine if related modified nucleosides are formed via parallel biosynthetic pathways or the t6A pathway, we carried out biochemical and biophysical investigations of the t6A systems from E. coli and T. maritima to address these questions. Using kinetic assays of TsaC(C2), tRNA modification assays, and NMR, our data demonstrate that TsaC(C2) exhibit relaxed substrate specificity, producing a variety of TC-AMP analogs that can differ in both the identity of the amino acid and nucleotide component, whereas TsaD displays more stringent specificity, but efficiently produces hn6A in E. coli and T. maritima tRNA. Thus, in organisms that contain modifications such as hn6A in their tRNA, we conclude that their origin is due to formation via the t6A pathway.


Assuntos
Adenosina/análogos & derivados , Vias Biossintéticas/genética , Nucleosídeos/genética , RNA de Transferência/genética , Adenosina/genética , Monofosfato de Adenosina/genética , Trifosfato de Adenosina/genética , Aminoácidos/genética , Domínio Catalítico/genética , Escherichia coli/genética , Conformação Proteica , Especificidade por Substrato/genética , Thermotoga maritima/genética , Treonina/genética
13.
Virus Res ; 283: 197976, 2020 07 02.
Artigo em Inglês | MEDLINE | ID: covidwho-46070

RESUMO

An outbreak of atypical pneumonia caused by a novel Betacoronavirus (ßCoV), named SARS-CoV-2 has been declared a public health emergency of international concern by the World Health Organization. In order to gain insight into the emergence, evolution and adaptation of SARS-CoV-2 viruses, a comprehensive analysis of genome composition and codon usage of ßCoV circulating in China was performed. A biased nucleotide composition was found for SARS-CoV-2 genome. This bias in genomic composition is reflected in its codon and amino acid usage patterns. The overall codon usage in SARS-CoV-2 is similar among themselves and slightly biased. Most of the highly frequent codons are A- and U-ending, which strongly suggests that mutational bias is the main force shaping codon usage in this virus. Significant differences in relative synonymous codon usage frequencies among SARS-CoV-2 and human cells were found. These differences are due to codon usage preferences.


Assuntos
Betacoronavirus/classificação , Betacoronavirus/genética , Uso do Códon/genética , Doenças Transmissíveis Emergentes/virologia , Regulação Viral da Expressão Gênica/genética , Genoma Viral/genética , Genômica , Aminoácidos/genética , Animais , Betacoronavirus/isolamento & purificação , China/epidemiologia , Quirópteros/virologia , Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/veterinária , Infecções por Coronavirus/virologia , Evolução Molecular , Furões/virologia , Humanos , Mutagênese/genética , Fases de Leitura Aberta/genética , Viverridae/virologia
14.
Virus Res ; 283: 197976, 2020 07 02.
Artigo em Inglês | MEDLINE | ID: mdl-32294518

RESUMO

An outbreak of atypical pneumonia caused by a novel Betacoronavirus (ßCoV), named SARS-CoV-2 has been declared a public health emergency of international concern by the World Health Organization. In order to gain insight into the emergence, evolution and adaptation of SARS-CoV-2 viruses, a comprehensive analysis of genome composition and codon usage of ßCoV circulating in China was performed. A biased nucleotide composition was found for SARS-CoV-2 genome. This bias in genomic composition is reflected in its codon and amino acid usage patterns. The overall codon usage in SARS-CoV-2 is similar among themselves and slightly biased. Most of the highly frequent codons are A- and U-ending, which strongly suggests that mutational bias is the main force shaping codon usage in this virus. Significant differences in relative synonymous codon usage frequencies among SARS-CoV-2 and human cells were found. These differences are due to codon usage preferences.


Assuntos
Betacoronavirus/classificação , Betacoronavirus/genética , Uso do Códon/genética , Doenças Transmissíveis Emergentes/virologia , Regulação Viral da Expressão Gênica/genética , Genoma Viral/genética , Genômica , Aminoácidos/genética , Animais , Betacoronavirus/isolamento & purificação , China/epidemiologia , Quirópteros/virologia , Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/veterinária , Infecções por Coronavirus/virologia , Evolução Molecular , Furões/virologia , Humanos , Mutagênese/genética , Fases de Leitura Aberta/genética , Viverridae/virologia
15.
Gene ; 747: 144673, 2020 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-32304783

RESUMO

Krabbe disease is one of the rarest autosomal recessive disorders in human, caused by mutation in the GALC (ß-galactosylceramidase) gene, resulting in several mental and physical health issues. Due to its rarity and phenotypic heterogeneity, diagnosis rate of this disease is very low. This study generated information on the recessive allele frequency dynamics of GALC gene across 15 global populations, with the highest frequency detected in Druze (Israel) population and the lowest frequency in Turkey and the United States. The recessive allele would take more time period (about 24,975 years) to be completely removed from the population having the lowest frequency and vice versa. The codon usage patterns of four isoforms of GALC gene revealed that a few synonymous codons were used more frequently than others in the isoforms. The codon AGA (arginine) was found to be overrepresented in GALC gene, except for galactocerebrosidase isoform a precursor. Further, GALC gene showed low codon usage bias (CUB) as evident from high ENC values (55.7-58.2), with A/T ending codons more preferred to G/C ending codons. CUB analysis elucidated the dual role of mutational pressure (major role) and natural selection (minor role) in GALC gene evolution.


Assuntos
Uso do Códon/genética , Galactosilceramidase/genética , Frequência do Gene/genética , Leucodistrofia de Células Globoides/enzimologia , Leucodistrofia de Células Globoides/genética , Aminoácidos/genética , Composição de Bases/genética , Códon/genética , Evolução Molecular , Galactosilceramidase/metabolismo , Humanos , Filogenia , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo
16.
Gene ; 747: 144682, 2020 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-32304786

RESUMO

Human Papillomavirus 16 (HPV16) is the most oncogenic HPV and the most associated genotype with cervical cancer development and progression. Currently, all developed vaccines are targeting HPV16 and were designed based on the major L1 capsid protein. Thus, evaluation of the diversity of HPV16 L1 sequence, mainly in the antigenic regions, will be of a great interest to assess the efficacy of the prophylactic vaccines and to predict the impact of genetic variations in these regions on the vaccination-induced immunity. A total of 377 HPV16 L1 sequences, published in public domain GenBank database, from the Americas, Africa, Asia, and Europe were collected and assembled. A total of 626 mutation events affecting 83 distinct nucleotides into the five antigenic regions of L1 gene of HPV16 were reported, and most SNPs were located in DE (27.38%, 23/83) and FG (31%, 26/83) loops. Overall, 4 mutations were frequently found in HPV16 sequences: T176N and N181T in EF loop; A266T in the FG loop and T353P/I/N HI loop. Of particular interest, some SNPs are ubiquitous and were found in all populations whereas others were population specific and their presence was limited to one or 2 at the maximum. Association between mutations in the antigenic regions and ethnicity was also investigated and showed that mutations in BC and DE loops were present with no significant difference in sequences from Europe, Asia, America and Africa. However, most mutations in FG loop are reported in sequences from European cases and are less pronounced in cases from America and Asia, whereas mutations EF and HI loops prevail in Asian cases. These data highlight a high number of variant amino acid residues that could affect the vaccination-induced immunity and impact the effectiveness of the prophylactic vaccination to fight against HPV, warranting the need of further investigation for vaccines and natural history studies of HPV16.


Assuntos
Proteínas do Capsídeo/genética , Proteínas do Capsídeo/imunologia , Variação Genética , Papillomavirus Humano 16/genética , Imunidade , Proteínas Oncogênicas Virais/genética , Proteínas Oncogênicas Virais/imunologia , Vacinação , Aminoácidos/genética , Antígenos Virais/imunologia , Sequência de Bases , Grupos Étnicos/genética , Humanos , Modelos Moleculares , Mutação/genética , Filogenia , Polimorfismo de Nucleotídeo Único/genética
17.
Proc Natl Acad Sci U S A ; 117(16): 8845-8849, 2020 04 21.
Artigo em Inglês | MEDLINE | ID: mdl-32253306

RESUMO

The genetic incorporation of noncanonical amino acids (ncAAs) into proteins has been realized in bacteria, yeast, and mammalian cells, and recently, in multicellular organisms including plants and animals. However, the addition of new building blocks to the genetic code of tissues from human origin has not yet been achieved. To this end, we report a self-replicating Epstein-Barr virus-based episomal vector for the long-term encoding of ncAAs in human hematopoietic stem cells and reconstitution of this genetically engineered hematopoietic system in mice.


Assuntos
Aminoácidos/genética , Diferenciação Celular/genética , Vetores Genéticos/genética , Células-Tronco Hematopoéticas/fisiologia , Engenharia de Proteínas/métodos , Animais , Sangue Fetal/citologia , Técnicas de Transferência de Genes , Código Genético , Células HEK293 , Transplante de Células-Tronco Hematopoéticas , Herpesvirus Humano 4/genética , Humanos , Camundongos , Camundongos Endogâmicos NOD , Plasmídeos/genética , Cultura Primária de Células/métodos , Transfecção/métodos , Quimeras de Transplante , Transplante Heterólogo/métodos
18.
Infect Dis Poverty ; 9(1): 33, 2020 Mar 25.
Artigo em Inglês | MEDLINE | ID: covidwho-13772

RESUMO

BACKGROUND: Coronavirus can cross the species barrier and infect humans with a severe respiratory syndrome. SARS-CoV-2 with potential origin of bat is still circulating in China. In this study, a prediction model is proposed to evaluate the infection risk of non-human-origin coronavirus for early warning. METHODS: The spike protein sequences of 2666 coronaviruses were collected from 2019 Novel Coronavirus Resource (2019nCoVR) Database of China National Genomics Data Center on Jan 29, 2020. A total of 507 human-origin viruses were regarded as positive samples, whereas 2159 non-human-origin viruses were regarded as negative. To capture the key information of the spike protein, three feature encoding algorithms (amino acid composition, AAC; parallel correlation-based pseudo-amino-acid composition, PC-PseAAC and G-gap dipeptide composition, GGAP) were used to train 41 random forest models. The optimal feature with the best performance was identified by the multidimensional scaling method, which was used to explore the pattern of human coronavirus. RESULTS: The 10-fold cross-validation results showed that well performance was achieved with the use of the GGAP (g = 3) feature. The predictive model achieved the maximum ACC of 98.18% coupled with the Matthews correlation coefficient (MCC) of 0.9638. Seven clusters for human coronaviruses (229E, NL63, OC43, HKU1, MERS-CoV, SARS-CoV, and SARS-CoV-2) were found. The cluster for SARS-CoV-2 was very close to that for SARS-CoV, which suggests that both of viruses have the same human receptor (angiotensin converting enzyme II). The big gap in the distance curve suggests that the origin of SARS-CoV-2 is not clear and further surveillance in the field should be made continuously. The smooth distance curve for SARS-CoV suggests that its close relatives still exist in nature and public health is challenged as usual. CONCLUSIONS: The optimal feature (GGAP, g = 3) performed well in terms of predicting infection risk and could be used to explore the evolutionary dynamic in a simple, fast and large-scale manner. The study may be beneficial for the surveillance of the genome mutation of coronavirus in the field.


Assuntos
Betacoronavirus/imunologia , Infecções por Coronavirus , Coronavirus/imunologia , Reservatórios de Doenças/virologia , Pandemias , Peptidil Dipeptidase A/metabolismo , Pneumonia Viral , Receptores Virais/genética , Glicoproteína da Espícula de Coronavírus/imunologia , Algoritmos , Aminoácidos/genética , Animais , Betacoronavirus/genética , China , Chlorocebus aethiops , Coronavirus/genética , Coronavirus/isolamento & purificação , Infecções por Coronavirus/genética , Infecções por Coronavirus/transmissão , Infecções por Coronavirus/virologia , Endopeptidases/genética , Endopeptidases/metabolismo , Genoma/genética , Genoma Viral/genética , Humanos , Pandemias/prevenção & controle , Peptidil Dipeptidase A/genética , Filogenia , Pneumonia Viral/genética , Pneumonia Viral/transmissão , Pneumonia Viral/virologia , Receptores Virais/metabolismo , Medição de Risco
19.
Infect Dis Poverty ; 9(1): 33, 2020 Mar 25.
Artigo em Inglês | MEDLINE | ID: mdl-32209118

RESUMO

BACKGROUND: Coronavirus can cross the species barrier and infect humans with a severe respiratory syndrome. SARS-CoV-2 with potential origin of bat is still circulating in China. In this study, a prediction model is proposed to evaluate the infection risk of non-human-origin coronavirus for early warning. METHODS: The spike protein sequences of 2666 coronaviruses were collected from 2019 Novel Coronavirus Resource (2019nCoVR) Database of China National Genomics Data Center on Jan 29, 2020. A total of 507 human-origin viruses were regarded as positive samples, whereas 2159 non-human-origin viruses were regarded as negative. To capture the key information of the spike protein, three feature encoding algorithms (amino acid composition, AAC; parallel correlation-based pseudo-amino-acid composition, PC-PseAAC and G-gap dipeptide composition, GGAP) were used to train 41 random forest models. The optimal feature with the best performance was identified by the multidimensional scaling method, which was used to explore the pattern of human coronavirus. RESULTS: The 10-fold cross-validation results showed that well performance was achieved with the use of the GGAP (g = 3) feature. The predictive model achieved the maximum ACC of 98.18% coupled with the Matthews correlation coefficient (MCC) of 0.9638. Seven clusters for human coronaviruses (229E, NL63, OC43, HKU1, MERS-CoV, SARS-CoV, and SARS-CoV-2) were found. The cluster for SARS-CoV-2 was very close to that for SARS-CoV, which suggests that both of viruses have the same human receptor (angiotensin converting enzyme II). The big gap in the distance curve suggests that the origin of SARS-CoV-2 is not clear and further surveillance in the field should be made continuously. The smooth distance curve for SARS-CoV suggests that its close relatives still exist in nature and public health is challenged as usual. CONCLUSIONS: The optimal feature (GGAP, g = 3) performed well in terms of predicting infection risk and could be used to explore the evolutionary dynamic in a simple, fast and large-scale manner. The study may be beneficial for the surveillance of the genome mutation of coronavirus in the field.


Assuntos
Betacoronavirus/imunologia , Infecções por Coronavirus , Coronavirus/imunologia , Reservatórios de Doenças/virologia , Pandemias , Peptidil Dipeptidase A/metabolismo , Pneumonia Viral , Receptores Virais/genética , Glicoproteína da Espícula de Coronavírus/imunologia , Algoritmos , Aminoácidos/genética , Animais , Betacoronavirus/genética , China , Chlorocebus aethiops , Coronavirus/genética , Coronavirus/isolamento & purificação , Infecções por Coronavirus/genética , Infecções por Coronavirus/transmissão , Infecções por Coronavirus/virologia , Endopeptidases/genética , Endopeptidases/metabolismo , Genoma/genética , Genoma Viral/genética , Humanos , Pandemias/prevenção & controle , Peptidil Dipeptidase A/genética , Filogenia , Pneumonia Viral/genética , Pneumonia Viral/transmissão , Pneumonia Viral/virologia , Receptores Virais/metabolismo , Medição de Risco
20.
PLoS Comput Biol ; 16(3): e1007630, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-32119660

RESUMO

In allosteric proteins, the binding of a ligand modifies function at a distant active site. Such allosteric pathways can be used as target for drug design, generating considerable interest in inferring them from sequence alignment data. Currently, different methods lead to conflicting results, in particular on the existence of long-range evolutionary couplings between distant amino-acids mediating allostery. Here we propose a resolution of this conundrum, by studying epistasis and its inference in models where an allosteric material is evolved in silico to perform a mechanical task. We find in our model the four types of epistasis (Synergistic, Sign, Antagonistic, Saturation), which can be both short or long-range and have a simple mechanical interpretation. We perform a Direct Coupling Analysis (DCA) and find that DCA predicts well the cost of point mutations but is a rather poor generative model. Strikingly, it can predict short-range epistasis but fails to capture long-range epistasis, in consistence with empirical findings. We propose that such failure is generic when function requires subparts to work in concert. We illustrate this idea with a simple model, which suggests that other methods may be better suited to capture long-range effects.


Assuntos
Sítio Alostérico/genética , Biologia Computacional/métodos , Epistasia Genética/genética , Regulação Alostérica/fisiologia , Aminoácidos/genética , Animais , Domínio Catalítico/fisiologia , Simulação por Computador , Desenho de Fármacos , Humanos , Ligantes , Modelos Moleculares , Modelos Teóricos , Conformação Proteica , Proteínas/química
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