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1.
Gene ; 725: 144161, 2020 Jan 30.
Artigo em Inglês | MEDLINE | ID: mdl-31639432

RESUMO

Bivalve species with doubly uniparental inheritance of mitochondria have been shown to contain additional mtDNA-encoded proteins suspected to be involved in sex-specific transmission of the female (F) and male (M) mitochondrial genomes. This is true for freshwater mussels and marine clams but was still unclear for marine mussel Mytilus spp. Here we present evidence that a F mtDNA-specific open reading frame (ORF) identified in the control region of M. edulis codes for a protein. The protein was detected, using western blots, in both female and male mantle tissues, which contain the gonads. The protein was also localized, using immunochemistry, in sperm mitochondria.


Assuntos
DNA Mitocondrial/genética , Mytilus/genética , Animais , Bivalves/genética , Feminino , Genoma Mitocondrial/genética , Masculino , Mitocôndrias/genética , Fases de Leitura Aberta/genética , Fatores Sexuais , Espermatozoides/metabolismo
2.
Arch Virol ; 165(2): 387-396, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31865470

RESUMO

A pathogen of significance in the aquaculture sector, the Gram-negative marine bacterium Vibrio parahaemolyticus causes gastroenteritis associated with consumption of improperly prepared seafood. This bacterium can be controlled using lytic bacteriophages as an alternative to antibiotics. Ï•VP-1 is a lytic phage of V. parahaemolyticus that was isolated from an aquafarm water sample with the aim of assessing its potential as a bio-control agent and determining its physicochemical properties and genomic sequence. Morphological analysis by transmission electron microscopy and phylogenetic analysis based on the large terminase subunit gene showed that this phage belongs to the family Myoviridae. It could infect multiple-drug-resistant (MDR) V. parahaemolyticus and V. alginolyticus strains of mangrove and seafood origin. With a maximum adsorption time of 30 min, ϕVP-1 has a short latent period of 10 min with burst size of 44 particles/cell. Whole-genome sequencing was done using the Illumina platform, and annotation was done using GeneMarkS and Prodigal. The 150,764bp genome with an overall G+C content of 41.84% had 203 putative protein-encoding open reading frames, one tRNA gene, and 66 predicted promoters. A number of putative DNA replication and regulation, DNA packaging and structure, and host lysis genes were identified. Comparison of the ϕVP-1 genome sequence to those of known Vibrio phages indicated little discernible DNA sequence similarity, suggesting that ϕVP-1 is a novel Vibrio phage. Sequence analysis revealed the presence of 64 potential ORFs with a T4-like genomic organization. In silico analysis suggested an obligate lytic life cycle and showed the absence of lysogeny or virulence genes. The complete sequence of Ï•VP-1 was annotated and deposited in the GenBank database (accession no. MH363700). The genetic features of this novel phage suggest that it might be applicable for phage therapy against pathogenic strains of V. parahaemolyticus.


Assuntos
Bacteriófagos/genética , Resistência a Múltiplos Medicamentos/genética , Genoma Viral/genética , Myoviridae/genética , Vibrio parahaemolyticus/virologia , Aquicultura/métodos , Composição de Bases/genética , Biofilmes , Genômica/métodos , Fases de Leitura Aberta/genética , Terapia por Fagos/métodos , Filogenia
3.
BMC Bioinformatics ; 20(1): 559, 2019 Nov 08.
Artigo em Inglês | MEDLINE | ID: mdl-31703551

RESUMO

BACKGROUND: Micropeptides are small proteins with length < = 100 amino acids. Short open reading frames that could produces micropeptides were traditionally ignored due to technical difficulties, as few small peptides had been experimentally confirmed. In the past decade, a growing number of micropeptides have been shown to play significant roles in vital biological activities. Despite the increased amount of data, we still lack bioinformatics tools for specifically identifying micropeptides from DNA sequences. Indeed, most existing tools for classifying coding and noncoding ORFs were built on datasets in which "normal-sized" proteins were considered to be positives and short ORFs were generally considered to be noncoding. Since the functional and biophysical constraints on small peptides are likely to be different from those on "normal" proteins, methods for predicting short translated ORFs must be trained independently from those for longer proteins. RESULTS: In this study, we have developed MiPepid, a machine-learning tool specifically for the identification of micropeptides. We trained MiPepid using carefully cleaned data from existing databases and used logistic regression with 4-mer features. With only the sequence information of an ORF, MiPepid is able to predict whether it encodes a micropeptide with 96% accuracy on a blind dataset of high-confidence micropeptides, and to correctly classify newly discovered micropeptides not included in either the training or the blind test data. Compared with state-of-the-art coding potential prediction methods, MiPepid performs exceptionally well, as other methods incorrectly classify most bona fide micropeptides as noncoding. MiPepid is alignment-free and runs sufficiently fast for genome-scale analyses. It is easy to use and is available at https://github.com/MindAI/MiPepid. CONCLUSIONS: MiPepid was developed to specifically predict micropeptides, a category of proteins with increasing significance, from DNA sequences. It shows evident advantages over existing coding potential prediction methods on micropeptide identification. It is ready to use and runs fast.


Assuntos
Biologia Computacional/métodos , Aprendizado de Máquina , Peptídeos/análise , Software , Bases de Dados de Proteínas , Fases de Leitura Aberta/genética
4.
Exp Parasitol ; 207: 107776, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31628895

RESUMO

The availability of high quality genomic DNA in sufficient amounts to perform Next Generation Sequencing (NGS) experiments is challenging for pathogens that cannot be cultivated in vitro, as is the case for many parasites. Therefore, Whole Genome Amplification (WGA) of genomic DNA is used to overcome this limitation. In this study, we evaluated the effect of WGA using the intestinal flagellated protozoan Giardia duodenalis as a model, due to its genome compactness (12 Mb), the presence of two diploid nuclei with variable levels of allelic sequence heterogeneity (ASH), and the availability of reference genomes. We selected one isolate (ZX15) belonging to the same genetic group of the reference isolate WB, namely Assemblage A, sub-Assemblage AI. Genomic DNA from the ZX15 isolate (GEN dataset) and that obtained by WGA of 1 ng of the same genomic DNA (WGA dataset) were sequenced on a HiSeq Illumina platform. Trimmed reads from the GEN and WGA experiments were mapped against the WB reference genome, showing the presence of a very small number of mutations (846 and 752, respectively). The difference in the number of mutations is largely accounted by local variation in coverage and not by bias introduced by WGA. No significant difference were observed in the distribution of mutations in coding and non-coding regions, in the proportion of heterozygous mutations (ASH), or in the transition/transversion ratio of Single Nucleotide Variants within coding sequences. We conclude that the quantitative and qualitative impact of WGA on the identification of mutations is limited, and that this technique can be used to conduct comparative genomics studies.


Assuntos
DNA de Protozoário/genética , Giardia lamblia/genética , Giardíase/parasitologia , Pré-Escolar , Biologia Computacional , República Tcheca , DNA de Protozoário/química , DNA de Protozoário/isolamento & purificação , Feminino , Estudo de Associação Genômica Ampla , Variação Estrutural do Genoma , Humanos , Mutação , Técnicas de Amplificação de Ácido Nucleico , Fases de Leitura Aberta/genética
5.
Biochim Biophys Acta Gene Regul Mech ; 1862(10): 194434, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31655156

RESUMO

The 43-kDa transactive response DNA-binding protein (TDP-43) is an example of an RNA-binding protein that regulates RNA metabolism at multiple levels from transcription and splicing to translation. Its role in post-transcriptional RNA processing has been a primary focus of recent research, but its role in regulating transcription has been studied for only a few human genes. We characterized the effects of TDP-43 on transcription genome-wide and found that TDP-43 broadly affects transcription of protein-coding and noncoding RNA genes. Among protein-coding genes, the effects of TDP-43 were greatest for genes <30 thousand base pairs in length. Surprisingly, we found that the loss of TDP-43 resulted in increased evidence for transcription activity near repetitive Alu elements found within expressed genes. The highest densities of affected Alu elements were found in the shorter genes, whose transcription was most affected by TDP-43. Thus, in addition to its role in post-transcriptional RNA processing, TDP-43 plays a critical role in maintaining the transcriptional stability of protein-coding genes and transposable DNA elements.


Assuntos
Elementos Alu/genética , Proteínas de Ligação a DNA/genética , Fases de Leitura Aberta/genética , Transcrição Genética , Genoma Humano/genética , Humanos , Processamento de RNA/genética , Proteínas de Ligação a RNA/genética , Retroelementos/genética , Fatores de Transcrição/genética , Ativação Transcricional/genética
6.
BMC Infect Dis ; 19(1): 802, 2019 Sep 12.
Artigo em Inglês | MEDLINE | ID: mdl-31510934

RESUMO

BACKGROUND: Chronic infection with hepatitis B virus (HBV) is a serious global health problem. Persistence of the virus occurs as a result of stability of the replication intermediate comprising covalently closed circular DNA (cccDNA). Development of drugs that are capable of disabling this cccDNA is vital. METHODS: To investigate an epigenetic approach to inactivating viral DNA, we engineered transcriptional repressors that comprise an HBV DNA-binding domain of transcription activator like effectors (TALEs) and a fused Krüppel Associated Box (KRAB). These repressor TALEs (rTALEs) targeted the viral surface open reading frame and were placed under transcription control of constitutively active or liver-specific promoters. RESULTS: Evaluation in cultured cells and following hydrodynamic injection of mice revealed that the rTALEs significantly inhibited production of markers of HBV replication without evidence of hepatotoxicity. Increased methylation of HBV DNA at CpG island II showed that the rTALEs caused intended epigenetic modification. CONCLUSIONS: Epigenetic modification of HBV DNA is a new and effective means of inactivating the virus in vivo. The approach has therapeutic potential and avoids potentially problematic unintended mutagenesis of gene editing.


Assuntos
DNA Viral/genética , Vírus da Hepatite B/crescimento & desenvolvimento , Vírus da Hepatite B/genética , Hepatite B/terapia , Hepatite B/virologia , Proteínas Repressoras/metabolismo , Replicação Viral/genética , Animais , Linhagem Celular , Ilhas de CpG , Metilação de DNA , DNA Circular/genética , DNA Viral/biossíntese , Epigênese Genética , Feminino , Fígado/metabolismo , Fígado/virologia , Camundongos , Fases de Leitura Aberta/genética , Regiões Promotoras Genéticas/genética , Proteínas Repressoras/genética
7.
Nucleic Acids Res ; 47(19): 10439-10451, 2019 11 04.
Artigo em Inglês | MEDLINE | ID: mdl-31511890

RESUMO

One challenge in engineering organisms is taking responsibility for their behavior over many generations. Spontaneous mutations arising before or during use can impact heterologous genetic functions, disrupt system integration, or change organism phenotype. Here, we propose restructuring the genetic code itself such that point mutations in protein-coding sequences are selected against. Synthetic genetic systems so-encoded should fail more safely in response to most spontaneous mutations. We designed fail-safe codes and simulated their expected effects on the evolution of so-encoded proteins. We predict fail-safe codes supporting expression of 20 or 15 amino acids could slow protein evolution to ∼30% or 0% the rate of standard-encoded proteins, respectively. We also designed quadruplet-codon codes that should ensure all single point mutations in protein-coding sequences are selected against while maintaining expression of 20 or more amino acids. We demonstrate experimentally that a reduced set of 21 tRNAs is capable of expressing a protein encoded by only 20 sense codons, whereas a standard 64-codon encoding is not expressed. Our work suggests that biological systems using rationally depleted but otherwise natural translation systems should evolve more slowly and that such hypoevolvable organisms may be less likely to invade new niches or outcompete native populations.


Assuntos
Biologia Computacional , Evolução Molecular , Código Genético/genética , Modelos Teóricos , Aminoácidos/genética , Regulação da Expressão Gênica/genética , Fases de Leitura Aberta/genética , Mutação Puntual/genética , Biossíntese de Proteínas/genética , RNA de Transferência/genética
8.
Genomics Proteomics Bioinformatics ; 17(3): 260-272, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31494267

RESUMO

Chromochloris zofingiensis represents an industrially relevant and unique green alga, given its capability of synthesizing triacylglycerol (TAG) and astaxanthin simultaneously for storage in lipid droplets (LDs). To further decipher lipid metabolism, the nitrogen deprivation (ND)-induced LDs from C. zofingiensis were isolated, purified, and subjected to proteomic analysis. Intriguingly, many C. zofingiensis LD proteins had no orthologs present in LD proteome of the model alga Chlamydomonas reinhardtii. Seven novel LD proteins (i.e., two functionally unknown proteins, two caleosins, two lipases, and one l-gulonolactone oxidase) and the major LD protein (MLDP), which were all transcriptionally up-regulated by ND, were selected for further investigation. Heterologous expression in yeast demonstrated that all tested LD proteins were localized to LDs and all except the two functionally unknown proteins enabled yeast to produce more TAG. MLDP could restore the phenotype of mldp mutant strain and enhance TAG synthesis in wild-type strain of C. reinhardtii. Although MLDP and caleosins had a comparable abundance in LDs, they responded distinctly to ND at the transcriptional level. The two lipases, instead of functioning as TAG lipases, likely recycled polar lipids to support TAG synthesis. For the first time, we reported that l-gulonolactone oxidase was abundant in LDs and facilitated TAG accumulation. Moreover, we also proposed a novel working model for C. zofingiensis LDs. Taken together, our work unravels the unique characteristics of C. zofingiensis LDs and provides insights into algal LD biogenesis and TAG synthesis, which would facilitate genetic engineering of this alga for TAG improvement.


Assuntos
Proteínas de Algas/metabolismo , Clorófitas/metabolismo , Gotículas Lipídicas/metabolismo , Metabolismo dos Lipídeos , Proteômica/métodos , Ácidos Graxos/metabolismo , Mutação/genética , Nitrogênio/deficiência , Fases de Leitura Aberta/genética , Fenótipo , Proteoma/metabolismo , Saccharomyces cerevisiae/metabolismo , Triglicerídeos/biossíntese
9.
Genomics Proteomics Bioinformatics ; 17(3): 319-331, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31494268

RESUMO

Proteins usually associate with other molecules physically to execute their functions. Identifying these interactions is important for the functional analysis of proteins. Previously, we reported the parallel analysis of translated ORFs (PLATO) to couple ribosome display of full-length ORFs with affinity enrichment of mRNA/protein/ribosome complexes for the "bait" molecules, followed by the deep sequencing analysis of mRNA. However, the sample processing, from extraction of precipitated mRNA to generation of DNA libraries, includes numerous steps, which is tedious and may cause the loss of materials. Barcoded PLATO (PLATO-BC), an improved platform was further developed to test its application for protein interaction discovery. In this report, we tested the antisera-antigen interaction using serum samples from patients with inclusion body myositis (IBM). Tripartite motif containing 21 (TRIM21) was identified as a potentially new IBM autoantigen. We also expanded the application of PLATO-BC to identify protein interactions for JQ1, single ubiquitin peptide, and NS5 protein of Zika virus. From PLATO-BC analyses, we identified new protein interactions for these "bait" molecules. We demonstrate that Ewing sarcoma breakpoint region 1 (EWSR1) binds to JQ1 and their interactions may interrupt the EWSR1 association with acetylated histone H4. RIO kinase 3 (RIOK3), a newly identified ubiquitin-binding protein, is preferentially associated with K63-ubiquitin chain. We also find that Zika NS5 protein interacts with two previously unreported host proteins, par-3 family cell polarity regulator (PARD3) and chromosome 19 open reading frame 53 (C19orf53), whose attenuated expression benefits the replication of Zika virus. These results further demonstrate that PLATO-BC is capable of identifying novel protein interactions for various types of "bait" molecules.


Assuntos
Fases de Leitura Aberta/genética , Mapeamento de Interação de Proteínas/métodos , Anticorpos/metabolismo , Células HEK293 , Humanos , Peptídeos/metabolismo , Ligação Proteica , Ubiquitina/metabolismo , Zika virus/fisiologia , Infecção por Zika virus/genética , Infecção por Zika virus/virologia
10.
Nat Commun ; 10(1): 4006, 2019 09 05.
Artigo em Inglês | MEDLINE | ID: mdl-31488843

RESUMO

The genomes of many prokaryotes contain substantial fractions of gene pairs with overlapping stop and start codons (ATGA or TGATG). A potential benefit of overlapping gene pairs is translational coupling. In 720 genomes of archaea and bacteria representing all major phyla, we identify substantial, albeit highly variable, fractions of co-directed overlapping gene pairs. Various patterns are observed for the utilization of the SD motif for de novo initiation at upstream genes versus reinitiation at overlapping gene pairs. We experimentally test the predicted coupling in 9 gene pairs from the archaeon Haloferax volcanii and 5 gene pairs from the bacterium Escherichia coli. In 13 of 14 cases, translation of both genes is strictly coupled. Mutational analysis of SD motifs located upstream of the downstream genes indicate that the contribution of the SD to translational coupling widely varies from gene to gene. The nearly universal, abundant occurrence of overlapping gene pairs suggests that tight translational coupling is widespread in archaea and bacteria.


Assuntos
Archaea/genética , Bactérias/genética , Genes Arqueais/genética , Genes Bacterianos/genética , Iniciação Traducional da Cadeia Peptídica/fisiologia , Terminação Traducional da Cadeia Peptídica/fisiologia , Biossíntese de Proteínas/fisiologia , Archaea/metabolismo , Bactérias/metabolismo , Sequência de Bases , Códon de Iniciação , Escherichia coli/genética , Homologia de Genes , Genes Reporter , Fases de Leitura Aberta/genética , RNA Mensageiro , Regiões Terminadoras Genéticas
11.
Aquat Toxicol ; 215: 105289, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31491707

RESUMO

Mifepristone (RU486), a clinical abortion agent and potential endocrine disruptor, binds to progestin and glucocorticoid receptors and has multiple functional importance in reproductive physiology. A long-term exposure of RU486 resulted in masculinization of female fish, however, the epigenetic landscape remains elusive. Recent studies demonstrated that long non-coding RNAs (lncRNAs) might play potential roles in epigenetic modulation of sex differentiation, ovarian cancer and germline stem cell survival. To further understand the influence of RU486 exposure on epigenetic regulation, we performed a comparative investigation on sex-biased gonadal lncRNAs profiles using control XX/XY and RU486-induced sex reversed XX Nile tilapia (Oreochromis niloticus) by RNA-seq. In total, 962 sexually differentially expressed lncRNAs and their target genes were screened from the gonads of control and sex reversed fish. In comparison with the control XX group, sex reversal induced by RU486 treatment led to significant up-regulation of 757 lncRNAs and down-regulation of 221 lncRNAs. Hierarchical clustering analysis revealed that global lncRNA expression profiles in RU486-treated XX group clustered into the same branch with the control XY, whereas XX control group formed a separate branch. The KEGG pathway enrichment analysis showed that the cis-target genes between RU486-XX and control-XX were concentrated in NOD - like receptor signaling pathway, Cell adhesion molecules (CAMs) and Biosynthesis of amino acids. Real-time PCR and in situ hybridization experiments demonstrate that lncRNAs showing intense fluctuation during RU486 treatment are also sexually dimorphic during early sex differentiation, which further proves the intimate relationship between lncRNAs and sex differentiation and sexual transdifferentiation. Taken together, our data strongly indicates that a long-term exposure of RU486 resulted in sex reversal of XX female fish and the altered expression of sexually dimorphic lncRNAs might partially account for the sex reversal via epigenetic modification.


Assuntos
Ciclídeos/genética , Ciclídeos/fisiologia , Regulação da Expressão Gênica/efeitos dos fármacos , Gônadas/metabolismo , Mifepristona/toxicidade , Progestinas/antagonistas & inibidores , RNA Longo não Codificante/genética , Caracteres Sexuais , Animais , Feminino , Genoma , Gônadas/efeitos dos fármacos , Masculino , Fases de Leitura Aberta/genética , Ovário/efeitos dos fármacos , Ovário/metabolismo , RNA Longo não Codificante/metabolismo , Reprodutibilidade dos Testes , Testículo/efeitos dos fármacos , Testículo/metabolismo , Fatores de Tempo , Distribuição Tecidual/efeitos dos fármacos , Transcrição Genética/efeitos dos fármacos , Poluentes Químicos da Água/toxicidade
12.
Nat Commun ; 10(1): 4066, 2019 09 06.
Artigo em Inglês | MEDLINE | ID: mdl-31492846

RESUMO

Human enteroviruses (HEVs) of the family Picornaviridae, which comprises non-enveloped RNA viruses, are ubiquitous worldwide. The majority of EV proteins are derived from viral polyproteins encoded by a single open reading frame (ORF). Here, we characterize a second ORF in HEVs that is crucial for viral intestinal infection. Disruption of ORF2p expression decreases the replication capacity of EV-A71 in human intestinal epithelial cells (IECs). Ectopic expression of ORF2p proteins derived from diverse enteric enteroviruses sensitizes intestinal cells to the replication of ORF2p-defective EV-A71 and respiratory enterovirus EV-D68. We show that the highly conserved WIGHPV domain of ORF2p is important for ORF2p-dependent viral intestinal infection. ORF2p expression is required for EV-A71 particle release from IECs and can support productive EV-D68 infection in IECs by facilitating virus release. Our results indicate that ORF2p is a determining factor for enteric enterovirus replication in IECs.


Assuntos
Enterovirus/genética , Fases de Leitura Aberta/genética , Vírus de RNA/genética , Replicação Viral/genética , Sequência de Aminoácidos , Sequência de Bases , Enterovirus/fisiologia , Infecções por Enterovirus/transmissão , Infecções por Enterovirus/virologia , Células Epiteliais/virologia , Fezes/virologia , Células HT29 , Interações Hospedeiro-Patógeno/genética , Humanos , Intestinos/citologia , Intestinos/virologia , Vírus de RNA/fisiologia , Proteínas Virais/genética , Proteínas Virais/metabolismo
13.
BMC Genomics ; 20(1): 686, 2019 Aug 30.
Artigo em Inglês | MEDLINE | ID: mdl-31470809

RESUMO

BACKGROUND: Mutations in minor spliceosome components such as U12 snRNA (cerebellar ataxia) and U4atac snRNA (microcephalic osteodysplastic primordial dwarfism type 1 (MOPD1)) result in tissue-specific symptoms. Given that the minor spliceosome is ubiquitously expressed, we hypothesized that these restricted phenotypes might be caused by the tissue-specific regulation of the minor spliceosome targets, i.e. minor intron-containing genes (MIGs). The current model of inefficient splicing is thought to apply to the regulation of the ~ 500 MIGs identified in the U12DB. However this database was created more than 10 years ago. Therefore, we first wanted to revisit the classification of minor introns in light of the most recent reference genome. We then sought to address specificity of MIG expression, minor intron retention, and alternative splicing (AS) across mouse and human tissues. RESULTS: We employed position-weight matrices to obtain a comprehensive updated list of minor introns, consisting of 722 mouse and 770 human minor introns. These can be found in the Minor Intron DataBase (MIDB). Besides identification of 99% of the minor introns found in the U12DB, we also discovered ~ 150 new MIGs. We then analyzed the RNAseq data from eleven different mouse tissues, which revealed tissue-specific MIG expression and minor intron retention. Additionally, many minor introns were efficiently spliced compared to their flanking major introns. Finally, we identified several novel AS events across minor introns in both mouse and human, which were also tissue-dependent. Bioinformatics analysis revealed that several of the AS events could result in the production of novel tissue-specific proteins. Moreover, like the major introns, we found that these AS events were more prevalent in long minor introns, while retention was favoured in shorter introns. CONCLUSION: Here we show that minor intron splicing and AS across minor introns is a highly organised process that might be regulated in coordination with the major spliceosome in a tissue-specific manner. We have provided a framework to further study the impact of the minor spliceosome and the regulation of MIG expression. These findings may shed light on the mechanism underlying tissue-specific phenotypes in diseases associated with minor spliceosome inactivation. MIDB can be accessed at https://midb.pnb.uconn.edu .


Assuntos
Processamento Alternativo , Íntrons , Animais , Biologia Computacional/métodos , Bases de Dados Genéticas , Genes , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Fases de Leitura Aberta/genética , Especificidade de Órgãos/genética , Especificidade de Órgãos/fisiologia , Processamento de RNA , RNA Nuclear Pequeno/genética , RNA Nuclear Pequeno/metabolismo , Spliceossomos/genética
14.
Arch Virol ; 164(11): 2865-2871, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31401694

RESUMO

Phage Vp_R1 belongs to the family Podoviridae and has a C3 morphotype, with an elongated head with a diameter of 190 ± 1.1 nm and an ultrashort tail with a length of 9 ± 1.2 nm. The double-stranded DNA genome is 112.1 kb long, has a mol% G + C content of 40.3, contains 129 ORFs, and encodes four tRNAs. Phylogenetic analysis suggests that phage Vp_R1 is a novel member of the genus Kuravirus.


Assuntos
Genoma Viral/genética , Podoviridae/genética , Vibrio parahaemolyticus/virologia , Sequência de Aminoácidos , Composição de Bases/genética , DNA Viral/genética , Fases de Leitura Aberta/genética , Podoviridae/isolamento & purificação , Análise de Sequência de DNA , Proteínas Virais/genética
15.
Arch Virol ; 164(11): 2859-2863, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31385115

RESUMO

In this study, we report the molecular characterization of a novel mycovirus in a phytopathogenic fungus of the species Colletotrichum gloeosporioides, which we named "Colletotrichum gloeosporioides RNA virus 1" (CgRV1). The virus has a dsRNA genome of 2,975 bp and possesses two non-overlapping open reading frames (ORFs 1 and 2). The smaller ORF1 encodes a protein of unknown function, and the larger ORF2 encodes the RNA-dependent RNA polymerase (RdRp). Phylogenetic analysis based on the RdRp sequence showed that CgRV1 clustered with and is closely related to unclassified mycoviruses that are distinct from members of the family Partitiviridae.


Assuntos
Colletotrichum/virologia , Micovírus/genética , Genoma Viral/genética , Vírus não Classificados/genética , Sequência de Aminoácidos , Micovírus/isolamento & purificação , Fases de Leitura Aberta/genética , Doenças das Plantas/microbiologia , RNA Viral/genética , Vírus não Classificados/isolamento & purificação
16.
Arch Virol ; 164(11): 2877-2880, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31451964

RESUMO

A temperate bacteriophage, IME1320_01, was induced by mitomycin C treatment from Corynebacterium striatum. This phage possesses a double-stranded DNA genome of 40,086 bp with a G+C content of 58%. A total of 53 putative open reading frames (ORFs) were identified in its genome. BLASTn analysis revealed that IME1320_01 had the highest sequence similarity to Corynebacterium striatum strain 216, with a genome homology coverage of 44% and highest sequence identity of 95%. The termini of the phage genome was non-redundant, with a 13-nt 3'-protruding cohesive end. To the best of our knowledge, phage IME1320_01 is the first inducible phage to be identified in Corynebacterium striatum.


Assuntos
Corynebacterium/virologia , Genoma Viral/genética , Siphoviridae/genética , Ativação Viral/efeitos dos fármacos , Composição de Bases/genética , DNA Viral/genética , Mitomicina/farmacologia , Fases de Leitura Aberta/genética , Análise de Sequência de DNA , Siphoviridae/classificação , Siphoviridae/isolamento & purificação
17.
Arch Virol ; 164(11): 2671-2682, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31399875

RESUMO

Rodents host different orthohepeviruses, namely orthohepevirus C genotype HEV-C1 (rat hepatitis E virus, HEV) and the additional putative genotypes HEV-C3 and HEV-C4. Here, we screened 2,961 rodents from Central Europe by reverse transcription polymerase chain reaction (RT-PCR) and identified HEV RNA in 13 common voles (Microtus arvalis) and one bank vole (Myodes glareolus) with detection rates of 2% (95% confidence interval [CI]: 1-3.4) and 0.08% (95% CI: 0.002-0.46), respectively. Sequencing of a 279-nucleotide RT-PCR amplicon corresponding to a region within open reading frame (ORF) 1 showed a high degree of similarity to recently described common vole-associated HEV (cvHEV) sequences from Hungary. Five novel complete cvHEV genome sequences from Central Europe showed the typical HEV genome organization with ORF1, ORF2 and ORF3 and RNA secondary structure. Uncommon features included a noncanonical start codon in ORF3, multiple insertions and deletions within ORF1 and ORF2/ORF3, and the absence of a putative ORF4. Phylogenetic analysis showed all of the novel cvHEV sequences to be monophyletic, clustering most closely with an unassigned bird-derived sequence and other sequences of the species Orthohepevirus C. The nucleotide and amino acid sequence divergence of the common vole-derived sequences was significantly correlated with the spatial distance between the trapping sites, indicating mostly local evolutionary processes. Detection of closely related HEV sequences in common voles in multiple localities over a distance of 800 kilometers suggested that common voles are infected by cvHEV across broad geographic distances. The common vole-associated HEV strain is clearly divergent from HEV sequences recently found in narrow-headed voles (Microtus gregalis) and other cricetid rodents.


Assuntos
Arvicolinae/virologia , Hepatite Viral Animal/virologia , Hepevirus/classificação , Hepevirus/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Europa (Continente) , Genoma Viral/genética , Hepevirus/isolamento & purificação , Fases de Leitura Aberta/genética
18.
Arch Virol ; 164(11): 2853-2857, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31377887

RESUMO

A double-stranded RNA (dsRNA) segment was extracted from the ectomycorrhizal fungus Geopora sumneriana (Cooke) M. Torre, and its full-length cDNA sequence, comprising 3146 nucleotides, was determined. Sequence analysis revealed the presence of a large open reading frame (ORF) on the positive strand of this dsRNA segment when the mold mitochondrial genetic code was applied. The ORF encodes a putative RNA-dependent RNA polymerase (RdRp), which shares the highest degree of similarity with Tuber excavatum mitovirus, with 37.52% identity. This dsRNA segment represents the genome replication intermediate of a novel mitovirus that was tentatively designated as "Geopora sumneriana mitovirus 1" (GsMV1). Phylogenetic analysis further suggested that GsMV1 is a member of the family Narnaviridae. This is the first study reporting on a mitovirus genome sequence in the ectomycorrhizal fungus G. sumneriana.


Assuntos
Colletotrichum/virologia , Micovírus/classificação , Micovírus/genética , Genoma Viral/genética , Sequência de Aminoácidos , Sequência de Bases , Micovírus/isolamento & purificação , Fases de Leitura Aberta/genética , RNA Replicase/genética , RNA Viral/genética , Análise de Sequência de RNA , Sequenciamento Completo do Genoma
19.
Nucleic Acids Res ; 47(17): 9358-9367, 2019 09 26.
Artigo em Inglês | MEDLINE | ID: mdl-31392980

RESUMO

Translation regulation plays an important role in eukaryotic gene expression. Upstream open reading frames (uORFs) are potent regulatory elements located in 5' mRNA transcript leaders. Translation of uORFs usually inhibit the translation of downstream main open reading frames, but some enhance expression. While a minority of uORFs encode conserved functional peptides, the coding regions of most uORFs are not conserved. Thus, the importance of uORF coding sequences on their regulatory functions remains largely unknown. We investigated the impact of an uORF coding region on gene regulation by assaying the functions of thousands of variants in the yeast YAP1 uORF. Varying uORF codons resulted in a wide range of functions, including repressing and enhancing expression of the downstream ORF. The presence of rare codons resulted in the most inhibitory YAP1 uORF variants. Inhibitory functions of such uORFs were abrogated by overexpression of complementary tRNA. Finally, regression analysis of our results indicated that both codon identity and position impact uORF function. Our results support a model in which a uORF coding sequence impacts its regulatory functions by altering the speed of uORF translation.


Assuntos
Biossíntese de Proteínas , Processamento de Proteína Pós-Traducional/genética , RNA Mensageiro/genética , Ribossomos/genética , Regiões 5' não Traduzidas/genética , Códon/genética , Regulação da Expressão Gênica/genética , Fases de Leitura Aberta/genética , Sequências Reguladoras de Ácido Nucleico/genética , Saccharomyces cerevisiae/genética
20.
Nucleic Acids Res ; 47(15): 8111-8125, 2019 09 05.
Artigo em Inglês | MEDLINE | ID: mdl-31340039

RESUMO

It has been a long debate whether the 98% 'non-coding' fraction of human genome can encode functional proteins besides short peptides. With full-length translating mRNA sequencing and ribosome profiling, we found that up to 3330 long non-coding RNAs (lncRNAs) were bound to ribosomes with active translation elongation. With shotgun proteomics, 308 lncRNA-encoded new proteins were detected. A total of 207 unique peptides of these new proteins were verified by multiple reaction monitoring (MRM) and/or parallel reaction monitoring (PRM); and 10 new proteins were verified by immunoblotting. We found that these new proteins deviated from the canonical proteins with various physical and chemical properties, and emerged mostly in primates during evolution. We further deduced the protein functions by the assays of translation efficiency, RNA folding and intracellular localizations. As the new protein UBAP1-AST6 is localized in the nucleoli and is preferentially expressed by lung cancer cell lines, we biologically verified that it has a function associated with cell proliferation. In sum, we experimentally evidenced a hidden human functional proteome encoded by purported lncRNAs, suggesting a resource for annotating new human proteins.


Assuntos
Biossíntese de Proteínas , Proteoma/genética , Proteômica/métodos , RNA Longo não Codificante/genética , Células A549 , Animais , Linhagem Celular Tumoral , Evolução Molecular , Perfilação da Expressão Gênica/métodos , Código Genético , Humanos , Fases de Leitura Aberta/genética , Peptídeos/genética , Primatas/genética , Proteoma/metabolismo , RNA Longo não Codificante/química , RNA Longo não Codificante/metabolismo , Ribossomos/genética
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