RESUMO
The mammalian neocortex is formed by sequential radial migration of newborn excitatory neurons. Migrating neurons undergo a multipolar-to-bipolar transition at the subplate (SP) layer, where extracellular matrix (ECM) components are abundantly expressed. Here, we investigate the role of the ECM at the SP layer. We show that TGF-ß signaling-related ECM proteins, and their downstream effector, p-smad2/3, are selectively expressed in the SP layer. We also find that migrating neurons express a disintegrin and metalloproteinase with thrombospondin motif 2 (ADAMTS2), an ECM metalloproteinase, just below the SP layer. Knockdown and knockout of Adamts2 suppresses the multipolar-to-bipolar transition of migrating neurons and disturbs radial migration. Time-lapse luminescence imaging of TGF-ß signaling indicates that ADAMTS2 activates this signaling pathway in migrating neurons during the multipolar-to-bipolar transition at the SP layer. Overexpression of TGF-ß2 in migrating neurons partially rescues migration defects in ADAMTS2 knockout mice. Our data suggest that ADAMTS2 secreted by the migrating multipolar neurons activates TGF-ß signaling by ECM remodeling of the SP layer, which might drive the multipolar to bipolar transition.
Assuntos
Proteínas ADAMTS , Movimento Celular , Camundongos Knockout , Neocórtex , Neurônios , Transdução de Sinais , Fator de Crescimento Transformador beta , Animais , Neocórtex/metabolismo , Neocórtex/citologia , Proteínas ADAMTS/metabolismo , Proteínas ADAMTS/genética , Camundongos , Fator de Crescimento Transformador beta/metabolismo , Neurônios/metabolismo , Matriz Extracelular/metabolismoRESUMO
DNA-binding response regulators (DBRRs) are a broad class of proteins that operate in tandem with their partner kinase proteins to form two-component signal transduction systems in bacteria. Typical DBRRs are composed of two domains where the conserved N-terminal domain accepts transduced signals and the evolutionarily diverse C-terminal domain binds to DNA. These domains are assumed to be functionally independent, and hence recombination of the two domains should yield novel DBRRs of arbitrary input/output response, which can be used as biosensors. This idea has been proved to be successful in some cases; yet, the error rate is not trivial. Improvement of the success rate of this technique requires a deeper understanding of the linker-domain and inter-domain residue interactions, which have not yet been thoroughly examined. Here, we studied residue coevolution of DBRRs of the two main subfamilies (OmpR and NarL) using large collections of bacterial amino acid sequences to extensively investigate the evolutionary signatures of linker-domain and inter-domain residue interactions. Coevolutionary analysis uncovered evolutionarily selected linker-domain and inter-domain residue interactions of known experimental structures, as well as previously unknown inter-domain residue interactions. We examined the possibility of these inter-domain residue interactions as contacts that stabilize an inactive conformation of the DBRR where DNA binding is inhibited for both subfamilies. The newly gained insights on linker-domain/inter-domain residue interactions and shared inactivation mechanisms improve the understanding of the functional mechanism of DBRRs, providing clues to efficiently create functional DBRR-based biosensors. Additionally, we show the feasibility of applying coevolutionary landscape models to predict the functionality of domain-swapped DBRR proteins. The presented result demonstrates that sequence information can be used to filter out bioengineered DBRR proteins that are predicted to be nonfunctional due to a high negative predictive value.
Assuntos
Bactérias , Transdução de Sinais , Mutação , Bactérias/genética , Transdução de Sinais/genética , Sequência de Aminoácidos , DNA/química , Proteínas de Bactérias/químicaRESUMO
Cleavage and polyadenylation at the 3' end of the pre-mRNA is essential for mRNA function, by regulating its translatability, stability and translocation to the cytoplasm. Cleavage factor I (CFI) is a multi-subunit component of the pre-mRNA 3' end processing machinery in eukaryotes. Here, we report that plant CFI 25 subunit of CFI plays an important role in maintaining the diversity of the 3' ends of mRNA. The genome of Arabidopsis thaliana (L.) Heynh. contained four genes encoding three putative CFI subunits (AtCFI 25, AtCFI 59 and AtCFI 68), orthologous to the mammalian CFI subunits. There were two CFI 25 paralogs (AtCFI 25a and AtCFI 25b) that shared homology with human CFI 25. Two null alleles of AtCFI 25a displayed smaller rosette leaves, longer stigmatic papilla, smaller anther, earlier flowering and lower fertility compared to wild-type plants. Null alleles of AtCFI 25b, as well as, plants ectopically expressing full-length cDNA of AtCFI 25a, displayed no obvious morphological defects. AtCFI 25a was shown to interact with AtCFI 25b, AtCFI 68 and itself, suggesting various forms of CFI in plants. Furthermore, we show that AtCFI 25a function was essential for maintaining proper diversity of the 3' end lengths of transcripts coding for CFI subunits, suggesting a self-regulation of the CFI machinery in plants. AtCFI 25a was also important to maintain 3' ends for other genes to different extent. Collectively, AtCFI 25a, but not AtCFI 25b, seemed to play important roles during Arabidopsis development by maintaining proper diversity of the 3' UTR lengths.
Assuntos
Arabidopsis , Animais , Regiões 3' não Traduzidas/genética , Arabidopsis/genética , Fibrinogênio , Poliadenilação/genéticaRESUMO
Streptomyces incarnatus NRRL8089 produces the antiviral, antifungal, antiprotozoal nucleoside antibiotic sinefungin. To enhance sinefungin production, multiple mutations were introduced to the rpoB gene encoding RNA polymerase (RNAP) ß-subunit at the target residues, D447, S453, H457, and R460. Sparse regression analysis using elastic-net lasso-ridge penalties on previously reported H457X mutations identified a numeric parameter set, which suggested that H457R/Y/F may cause production enhancement. H457R/R460C mutation successfully enhanced the sinefungin production by 3-fold, while other groups of mutations, such as D447G/R460C or D447G/H457Y, made moderate or even negative effects. To identify why the rif cluster residues have diverse effects on sinefungin production, an RNAP/DNA/mRNA complex model was constructed by homology modeling and molecular dynamics simulation. The 4 residues were located near the mRNA strand. Density functional theory-based calculation suggested that D447, H457, and R460 are in direct contact with ribonucleotide, and partially positive charges are induced by negatively charged chain of mRNA.
Assuntos
Adenosina/análogos & derivados , Antibacterianos/biossíntese , Proteínas de Bactérias/genética , RNA Polimerases Dirigidas por DNA/genética , Mutação , Streptomyces/genética , Adenosina/biossíntese , Adenosina/química , Substituição de Aminoácidos , Antibacterianos/química , Antifúngicos/química , Antifúngicos/metabolismo , Antimaláricos/química , Antimaláricos/metabolismo , Antiprotozoários/química , Antiprotozoários/metabolismo , Antivirais/química , Antivirais/metabolismo , Proteínas de Bactérias/química , Proteínas de Bactérias/metabolismo , Sítios de Ligação , DNA/química , DNA/genética , DNA/metabolismo , RNA Polimerases Dirigidas por DNA/química , RNA Polimerases Dirigidas por DNA/metabolismo , Teoria da Densidade Funcional , Regulação Bacteriana da Expressão Gênica , Simulação de Dinâmica Molecular , Ligação Proteica , Conformação Proteica em alfa-Hélice , Conformação Proteica em Folha beta , Domínios e Motivos de Interação entre Proteínas , RNA Mensageiro/química , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Streptomyces/enzimologiaRESUMO
COP9 signalosome (CSN) is a nuclear complex composed of eight distinct subunits that governs vast developmental processes in Arabidopsis thaliana (L.) Heynh. The null alleles of csn mutants display pleiotropic phenotypes that result in seedling lethality. To date, several partially complemented transgenic plants, expressing the particular CSN subunit in its corresponding null mutant allele, were utilized to bypass seedling lethality and investigate CSN regulation at later stages of development. One such transgenic plant corresponding to CSN1 subunit, fus6/CSN1-3-4, accumulates wild-type level of CSN1 and displays normal plant architecture at vegetative stage. Here we show through histological analyses that fus6/CSN1-3-4 plants display impairment of pollen development at the bicellular stage. This defect is identical to that observed in RNAi plants of SAP130, encoding a subunit of the multiprotein splicing factor SF3b. We further dissected the previously reported interaction between CSN1 and SAP130, to reveal that approximately 100 amino-acid residues located at the N-terminal end of CSN1 (CSN1NN) were essential for this interaction. In silico structure modeling demonstrated that CSN1NN could swing out towards SAP130 to dock onto its Helical Insertion protruding from the structure. These results support our model that CSN1 embeds itself within CSN protein complex through its C-terminal half and reaches out to targets through its N-terminal portion of the protein. Taken together, this is the first report to document the identical loss-of-function phenotypes of CSN1 and SAP130 during male gametogenesis. Thus, we propose that SAP130 and CSN1 coordinately regulate development of male reproductive organs.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Complexo do Signalossomo COP9 , Gametogênese , Masculino , Plantas Geneticamente ModificadasRESUMO
While CpG dinucleotides are significantly reduced compared to other dinucleotides in mammalian genomes, they can congregate and form CpG islands, which localize around the 5' regions of genes, where they function as promoters. CpG-island promoters are generally unmethylated and are often found in housekeeping genes. However, their nucleotide sequences and existence per se are not conserved between humans and mice, which may be due to evolutionary gain and loss of the regulatory regions. In this study, human and rhesus monkey genomes, with moderately conserved sequences, were compared at base resolution. Using transcription start site data, we first validated our methods' ability to identify orthologous promoters and indicated a limitation using the 5' end of curated gene models, such as NCBI RefSeq, as their transcription start sites. We found that, in addition to deamination mutations, insertions and deletions of bases, repeats, and long fragments contributed to the mutations of CpG dinucleotides. We also observed that the G + C contents tended to change in CpG-poor environments, while CpG content was altered in G + C-rich environments. While loss of CpG islands can be caused by gradual decreases in CpG sites, gain of these islands appear to require two distinct nucleotide altering steps. Taken together, our findings provide novel insights into the process of acquisition and diversification of CpG-island promoters in vertebrates.
Assuntos
Ilhas de CpG , Metilação de DNA , Epigênese Genética , Genoma , Regiões Promotoras Genéticas , Animais , Sequência de Bases , Variação Genética , Genoma Humano , Genômica/métodos , Humanos , Mutação INDEL , Macaca mulatta , Mamíferos/genética , Camundongos , Mutação , Sequências Reguladoras de Ácido Nucleico , Sítio de Iniciação de TranscriçãoRESUMO
BRCA1/2 genetic testing to use PARP inhibitor for breast cancer has a possibility of the "secondary finding" among the younger nonaffected family members of the patient, which turns them into at-risk for hereditary breast cancer. Proper understanding of the background of the hereditary cancer is now required for appropriate acceptance of the risk. Therefore, we investigated the level of knowledge and attitudes of younger women on hereditary breast cancer in Japan. Study subject was Japanese university women between 20 and 30 years of age, without medical history of breast cancer. We conducted the anonymous self-answering questionnaire to them. We received responses from 353 women. The levels of knowledge, awareness, and interest were relatively high. Women with a family history of breast cancer were less likely to undergo testing than women without (92.8% vs. 74.5%, p < 0.001). The rates of positive response toward risk-reducing mastectomy (RRM) and risk-reducing salpingo-oophorectomy (RRSO) was significantly high for medical majors compared with that for other majors (RRM: medical 71.6% vs. science 54.5% vs. humanities 53.8%, p = 0.008, RRSO: 35.4% vs. 36.3% vs. 48.4%, p = 0.027). Approximately half of respondents answered that they would hesitate to get married (45.3%) or to have children (55.4%), if they were a BRCA1/2 mutation carrier. The results may help to establish the methods for supporting the decision-making for reproduction of younger women who are unexpectedly labeled as being at-risk for HBOC.
Assuntos
Proteína BRCA1/genética , Proteína BRCA2/genética , Neoplasias da Mama/epidemiologia , Conhecimentos, Atitudes e Prática em Saúde , Adulto , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/genética , Feminino , Testes Genéticos , Humanos , Mutação/genética , Inibidores de Poli(ADP-Ribose) Polimerases/uso terapêutico , Estudantes , Universidades , Adulto JovemRESUMO
In this study, we present an analysis of metagenome sequences obtained from a filtrate of a siphon tissue homogenate of otter clams (Lutraria rhynchaena) with swollen-siphon disease. The viral signal was mined from the metagenomic data, and a novel circular ssDNA virus was identified. Genomic features and phylogenetic analysis showed that the virus belongs to the phylum Cressdnaviricota, which consists of viruses with circular, single-stranded DNA (ssDNA) genomes. Members of this phylum have been identified in various species and in environmental samples. The newly found virus is distantly related to the currently known members of the phylum Cressdnaviricota.
Assuntos
Bivalves/genética , Vírus de DNA/classificação , DNA Viral/genética , Genoma Viral , Animais , Vírus de DNA/isolamento & purificação , DNA Circular/genética , DNA de Cadeia Simples/genética , Microbiologia Ambiental , Metagenômica , Filogenia , Análise de Sequência de DNARESUMO
BACKGROUND: More than 7000 papers related to "protein refolding" have been published to date, with approximately 300 reports each year during the last decade. Whilst some of these papers provide experimental protocols for protein refolding, a survey in the structural life science communities showed a necessity for a comprehensive database for refolding techniques. We therefore have developed a new resource - "REFOLDdb" that collects refolding techniques into a single, searchable repository to help researchers develop refolding protocols for proteins of interest. RESULTS: We based our resource on the existing REFOLD database, which has not been updated since 2009. We redesigned the data format to be more concise, allowing consistent representations among data entries compared with the original REFOLD database. The remodeled data architecture enhances the search efficiency and improves the sustainability of the database. After an exhaustive literature search we added experimental refolding protocols from reports published 2009 to early 2017. In addition to this new data, we fully converted and integrated existing REFOLD data into our new resource. REFOLDdb contains 1877 entries as of March 17th, 2017, and is freely available at http://p4d-info.nig.ac.jp/refolddb/ . CONCLUSION: REFOLDdb is a unique database for the life sciences research community, providing annotated information for designing new refolding protocols and customizing existing methodologies. We envisage that this resource will find wide utility across broad disciplines that rely on the production of pure, active, recombinant proteins. Furthermore, the database also provides a useful overview of the recent trends and statistics in refolding technology development.
Assuntos
Algoritmos , Bases de Dados de Proteínas , Internet , Redobramento de Proteína , Proteínas/química , Humanos , Interface Usuário-ComputadorRESUMO
Life science research now heavily relies on all sorts of databases for genome sequences, transcription, protein three-dimensional (3D) structures, protein-protein interactions, phenotypes and so forth. The knowledge accumulated by all the omics research is so vast that a computer-aided search of data is now a prerequisite for starting a new study. In addition, a combinatory search throughout these databases has a chance to extract new ideas and new hypotheses that can be examined by wet-lab experiments. By virtually integrating the related databases on the Internet, we have built a new web application that facilitates life science researchers for retrieving experts' knowledge stored in the databases and for building a new hypothesis of the research target. This web application, named VaProS, puts stress on the interconnection between the functional information of genome sequences and protein 3D structures, such as structural effect of the gene mutation. In this manuscript, we present the notion of VaProS, the databases and tools that can be accessed without any knowledge of database locations and data formats, and the power of search exemplified in quest of the molecular mechanisms of lysosomal storage disease. VaProS can be freely accessed at http://p4d-info.nig.ac.jp/vapros/ .
Assuntos
Biologia Computacional , Bases de Dados Genéticas , Genoma , Internet , Software , Animais , Humanos , Camundongos , Conformação Proteica , Ratos , Análise de Sequência de DNARESUMO
Phosphorylation and acetylation are the most prevalent post-translational modifications (PTMs) detected in not only eukaryotes but also bacteria. We performed phosphoproteome and acetylome analyses of proteins from an extremely thermophilic eubacterium Thermus thermophilus HB8, and identified numerous phosphorylation and acetylation sites. To facilitate the elucidation of the structural aspects of these PTM events, we mapped the PTM sites on the known tertiary structures for the respective proteins and their homologs. Wu et al. (Mol Cell Proteomics 12:2701-2713, 2013) recently reported phosphoproteome analysis of proteins from T. thermophilus HB27. Therefore, we assessed the structural characteristics of these phosphorylation and acetylation sites on the tertiary structures of the identified proteins or their homologs. Our study revealed that many of the identified phosphosites are in close proximity to bound ligands, i.e., the numbers of 'nearby' and 'peripheral' phosphorylation sites represent 56 % (48/86 sites) of total identified phosphorylation sites. In addition, approximately 60 % of all phosphosites exhibited <10 % accessible surface area of their side chains, suggesting some structural rearrangement is required for phosphoryl transfer by kinases. Our findings also indicate that phosphorylation of a residue occurs more frequently at a flexible region of the protein, whereas lysine acetylation occurs more frequently in an ordered structure.
Assuntos
Proteínas de Bactérias/metabolismo , Processamento de Proteína Pós-Traducional , Thermus thermophilus/metabolismo , Acetilação , Aldeído Liases/metabolismo , Sequência de Aminoácidos , Aspartato Aminotransferases/metabolismo , Fosfopeptídeos/análise , Fosfopeptídeos/metabolismo , Fosforilação , Estrutura Terciária de Proteína , Proteoma/análise , Proteômica , Relação Estrutura-AtividadeRESUMO
Origin recognition complex (ORC), consisting of six subunits ORC1-6, is known to bind to replication origins and function in the initiation of DNA replication in eukaryotic cells. In contrast to the fact that Saccharomyces cerevisiae ORC recognizes the replication origin in a sequence-specific manner, metazoan ORC has not exhibited strict sequence-specificity for DNA binding. Here we report that human ORC binds preferentially to G-quadruplex (G4)-preferable G-rich RNA or single-stranded DNA (ssDNA). We mapped the G-rich RNA-binding domain in the ORC1 subunit, in a region adjacent to its ATPase domain. This domain itself has an ability to preferentially recognize G4-preferable sequences of ssDNA. Furthermore, we found, by structure modeling, that the G-rich RNA-binding domain is similar to the N-terminal portion of AdoMet_MTase domain of mammalian DNA methyltransferase 1. Therefore, in contrast with the binding to double-stranded DNA, human ORC has an apparent sequence preference with respect to its RNA/ssDNA binding. Interestingly, this specificity coincides with the common signature present in most of the human replication origins. We expect that our findings provide new insights into the regulations of function and chromatin binding of metazoan ORCs.
Assuntos
DNA de Cadeia Simples/química , Complexos Multiproteicos/química , Ácidos Nucleicos Heteroduplexes/química , Complexo de Reconhecimento de Origem/química , RNA/química , Animais , Metilases de Modificação do DNA/química , Metilases de Modificação do DNA/genética , Metilases de Modificação do DNA/metabolismo , DNA de Cadeia Simples/genética , DNA de Cadeia Simples/metabolismo , Humanos , Complexos Multiproteicos/genética , Complexos Multiproteicos/metabolismo , Ácidos Nucleicos Heteroduplexes/genética , Ácidos Nucleicos Heteroduplexes/metabolismo , Complexo de Reconhecimento de Origem/genética , Complexo de Reconhecimento de Origem/metabolismo , Estrutura Terciária de Proteína , RNA/genética , RNA/metabolismo , Saccharomyces cerevisiae , Xenopus laevisRESUMO
Lysosomal ß-galactosidase (ß-Gal) deficiency causes a group of disorders that include neuronopathic GM1 gangliosidosis and non-neuronopathic Morquio B disease. We have previously proposed the use of small molecule ligands of ß-Gal as pharmacological chaperones (PCs) for the treatment of GM1 gangliosidosis brain pathology. Although it is still under development, PC therapy has yielded promising preclinical results in several lysosomal diseases. In this study, we evaluated the effect of bicyclic 1-deoxygalactonojirimycin (DGJ) derivative of the sp(2)-iminosugar type, namely 5N,6S-(N'-butyliminomethylidene)-6-thio-1- deoxygalactonojirimycin (6S-NBI-DGJ), as a novel PC for human mutant ß-Gal. In vitro, 6S-NBI-DGJ had the ability to inhibit the activity of human ß-Gal in a competitive manner and was able to protect this enzyme from heat-induced degradation. Computational analysis supported that the rigid glycone bicyclic core of 6S-NBI-DGJ binds to the active site of the enzyme, with the aglycone N'-butyl substituent, in a precise E-orientation, located at a hydrophobic region nearby. Chaperone potential profiling indicated significant increases of enzyme activity in 24 of 88 ß-Gal mutants, including four common mutations. Finally, oral administration of 6S-NBI-DGJ ameliorated the brain pathology of GM1 gangliosidosis model mice. These results suggest that 6S-NBI-DGJ is a novel PC that may be effective on a broad range of ß-Gal mutants.
Assuntos
1-Desoxinojirimicina/análogos & derivados , Gangliosidose GM1/tratamento farmacológico , Chaperonas Moleculares/farmacologia , 1-Desoxinojirimicina/farmacologia , Administração Oral , Animais , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Células Cultivadas , Biologia Computacional , Modelos Animais de Doenças , Inibidores Enzimáticos/farmacologia , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Gangliosidose GM1/genética , Imino Açúcares/química , Imino Açúcares/farmacologia , Lisossomos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mucopolissacaridose IV/tratamento farmacológico , Mucopolissacaridose IV/genética , Mutação , Recombinação Genética , beta-Galactosidase/química , beta-Galactosidase/genéticaRESUMO
BACKGROUND: Lynch syndrome is a hereditary cancer predisposition syndrome caused by a mutation in one of the DNA mismatch repair (MMR) genes. About 24% of the mutations identified in Lynch syndrome are missense substitutions and the frequency of missense variants in MSH6 is the highest amongst these MMR genes. Because of this high frequency, the genetic testing was not effectively used in MSH6 so far. We, therefore, developed CoDP (Combination of the Different Properties), a bioinformatics tool to predict the impact of missense variants in MSH6. METHODS: We integrated the prediction results of three methods, namely MAPP, PolyPhen-2 and SIFT. Two other structural properties, namely solvent accessibility and the change in the number of heavy atoms of amino acids in the MSH6 protein, were further combined explicitly. MSH6 germline missense variants classified by their associated clinical and molecular data were used to fit the parameters for the logistic regression model and to assess the prediction. The performance of CoDP was compared with those of other conventional tools, namely MAPP, SIFT, PolyPhen-2 and PON-MMR. RESULTS: A total of 294 germline missense variants were collected from the variant databases and literature. Of them, 34 variants were available for the parameter training and the prediction performance test. We integrated the prediction results of MAPP, PolyPhen-2 and SIFT, and two other structural properties, namely solvent accessibility and the change in the number of heavy atoms of amino acids in the MSH6 protein, were further combined explicitly. Variants data classified by their associated clinical and molecular data were used to fit the parameters for the logistic regression model and to assess the prediction. The values of the positive predictive value (PPV), the negative predictive value (NPV), sensitivity, specificity and accuracy of the tools were compared on the whole data set. PPV of CoDP was 93.3% (14/15), NPV was 94.7% (18/19), specificity was 94.7% (18/19), sensitivity was 93.3% (14/15) and accuracy was 94.1% (32/34). Area under the curve of CoDP was 0.954, that of MAPP for MSH6 was 0.919, of SIFT was 0.864 and of PolyPhen-2 HumVar was 0.819. The power to distinguish between pathogenic and non-pathogenic variants of these methods was tested by Wilcoxon rank sum test (p < 8.9 × 10(-6) for CoDP, p < 3.3 × 10(-5) for MAPP, p < 3.1 × 10(-4) for SIFT and p < 1.2 × 10(-3) for PolyPhen-2 HumVar), and CoDP was shown to outperform other conventional methods. CONCLUSION: In this paper, we provide a human curated data set for MSH6 missense variants, and CoDP, the prediction tool, which achieved better accuracy for predicting the impact of missense variants in MSH6 than any other known tools. CoDP is available at http://cib.cf.ocha.ac.jp/CoDP/.
Assuntos
Biologia Computacional/métodos , Proteínas de Ligação a DNA/genética , Variação Genética , Software , Neoplasias Colorretais Hereditárias sem Polipose/genética , Reparo de Erro de Pareamento de DNA , Humanos , Mutação de Sentido IncorretoRESUMO
In thermophilic microorganisms, c-type cytochrome (cyt) proteins mainly function in the respiratory chain as electron carriers. Genome analyses at the beginning of this century revealed a variety of genes harboring the heme c motif. Here, we describe the results of surveying genes with the heme c motif, CxxCH, in a genome database comprising four strains of Thermus thermophilus, including strain HB8, and the confirmation of 19 c-type cytochromes among 27 selected genes. We analyzed the 19 genes, including the expression of four, by a bioinformatics approach to elucidate their individual attributes. One of the approaches included an analysis based on the secondary structure alignment pattern between the heme c motif and the 6th ligand. The predicted structures revealed many cyt c domains with fewer ß-strands, such as mitochondrial cyt c, in addition to the ß-strand unique to Thermus inserted in cyt c domains, as in T. thermophilus cyt c552 and caa3 cyt c oxidase subunit IIc. The surveyed thermophiles harbor potential proteins with a variety of cyt c folds. The gene analyses led to the development of an index for the classification of cyt c domains. Based on these results, we propose names for T. thermophilus genes harboring the cyt c fold.
Assuntos
Citocromos , Thermus thermophilus , Thermus thermophilus/genética , Thermus thermophilus/metabolismo , Transporte de Elétrons , Citocromos/metabolismo , Thermus/metabolismo , Complexo IV da Cadeia de Transporte de Elétrons/genética , Complexo IV da Cadeia de Transporte de Elétrons/química , Complexo IV da Cadeia de Transporte de Elétrons/metabolismoRESUMO
ThermusQ is a website (https://www.thermusq.net/) that aims to gather all the molecular information on Thermus thermophilus and to provide a platform to easily access the whole view of the bacterium. ThermusQ comprises the genome sequences of 22 strains from T. thermophilus and T. oshimai strains, plus the sequences of known Thermus phages. ThermusQ also contains information and map diagrams of pathways unique to Thermus strains. The website provides tools to retrieve sequence data in different ways. By gathering the whole data of T. thermophilus strains, the strainspecific characteristics was found. This bird's-eye view of the whole data will lead the research community to identify missing important data and the integration will provide a platform to conduct future biochemical simulations of the bacterium.
Assuntos
Thermus thermophilus , Thermus , Thermus thermophilus/genética , Thermus/genética , Thermus/metabolismoRESUMO
A Thermus thermophilus lytic phage was isolated from a Japanese hot spring using a type IV pili-deficient strain as an indicator host, and designated as φMN1. Electron microscopic (EM) examination revealed that φMN1 had an icosahedral head and a contractile tail, suggesting that φMN1 belonged to Myoviridae. An EM analysis focused on φMN1 adsorption to the Thermus host cell showed that the receptor molecules for the phage were uniformly distributed on the outer surface of the cells. The circular double-stranded DNA of φMN1 was 76,659 base pairs in length, and the guanine and cytosine content was 61.8%. It was predicted to contain 99 open reading frames, and its putative distal tail fiber protein, which is essential for non-piliated host cell surface receptor recognition, was dissimilar in terms of sequence and length with its counterpart in the type IV pili-dependent φYS40. A phage proteomic tree revealed that φMN1 and φYS40 are in the same cluster, but many genes had low sequence similarities and some seemed to be derived from both mesophilic and thermophilic organisms. The gene organization suggested that φMN1 evolved from a non-Thermus phage through large-scale recombination events of the genes determining the host specificity, followed by gradual evolution by recombination of both the thermophilic and mesophilic DNAs assimilated by the host Thermus cells. This newly isolated phage will provide evolutionary insights into thermophilic phages.
Assuntos
Bacteriófagos , Fontes Termais , Bacteriófagos/genética , Thermus thermophilus/genética , Proteômica , Japão , Fases de Leitura AbertaRESUMO
The authors sequenced the complete mitochondrial (mt) genomes of the band-legged ground cricket (Dianemobius fascipes nigrofasciatus Matsumura, 1904) and a temperate form of the lawn ground cricket (Polionemobius taprobanensis Walker, 1869), collected in Japan. The length of the mt genome sequences was 15,354 bp in D. fascipes nigrofasciatus and 16,063 bp in P. taprobanensis. Annotation of the mt genome sequences revealed 13 protein-coding genes, two rRNA genes, and 22 tRNA genes. The orientation of the genes was the same as in other Grylloidea species, and the order was the same as in other Trigonidiidae species. In our phylogenetic analysis, D. fascipes nigrofasciatus formed a clade with D. fascipes collected in China, and the temperate form of P. taprobanensis formed a clade with P. taprobanensis collected in China. Comparison of the numbers of positions with different amino acid residues encoded by the protein-coding genes implied the separate species status of each member of each of the two pairs of ground crickets. The mt genome sequences of D. fascipes nigrofasciatus and P. taprobanensis will contribute to phylogenetic and taxonomic studies of the Trigonidiidae.
RESUMO
Thermus thermophilus has several minor lipid molecules with structures that have not been described yet. In this study, we identified a new lipid molecule in T. thermophilus HB8 with an amino group at the polar head, by detecting lipid spots with HPTLC and mass spectrometry. The structure of the lipid resembles an amino sugar phospholipid, except for the glucosamine that lacks an acetyl group. We named this amino phosphoglycolipid PGLN, and proposed its synthetic pathway from a precursor, phosphatidyl-glyceric alkylamine. The primary amine structure of PGLN may contribute to high temperature adaptation through electrostatic interactions between the head groups.
RESUMO
Significant advances in biophysical methods such as next-generation sequencing technologies have now opened the way to conduct evolutionary and applied research based on the genomic information of greatly diverse insects. Crickets belonging to Orthoptera (Insecta: Polyneoptera), one of the most flourishing groups of insects, have contributed to the development of multiple scientific fields including developmental biology and neuroscience and have been attractive targets in evolutionary ecology for their diverse ecological niches. In addition, crickets have recently gained recognition as food and feed. However, the genomic information underlying their biological basis and application research toward breeding is currently underrepresented. In this review, we summarize the progress of genomics of crickets. First, we outline the phylogenetic position of crickets in insects and then introduce recent studies on cricket genomics and transcriptomics in a variety of fields. Furthermore, we present findings from our analysis of polyneopteran genomes, with a particular focus on their large genome sizes, chromosome number, and repetitive sequences. Finally, how the cricket genome can be beneficial to the food industry is discussed. This review is expected to enhance greater recognition of how important the cricket genomes are to the multiple biological fields and how basic research based on cricket genome information can contribute to tackling global food security. Supplementary Information: The online version contains supplementary material available at 10.1007/s12551-021-00924-4.