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Results for “Genome Components”

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PubMed2005

[Antigenome technology: rapid and comprehensive identification and validation of antigens as putative components in vaccines against bacterial pathogens].

The resolution of complete genome sequences of many pathogens has dramatically expanded our perspectives in developing novel vaccines by increasing both the speed and scale of target identification. Genomics-based technologies with their significant advantages can be applied to most pathogens and have exploited genome sequence information in alliance with adjunct technologies, including bioinformatics, expression analyses (microarrays, in vivo expression technologies), random mutagenesis or protein/peptide based selection methods (proteomics and immunoselection using expression libraries). A novel approach that combines the advantages of full genome coverage and serological antigen identification has been developed in the laboratory of the authors for the identification of protein antigens from bacterial pathogens. This technology has been applied to several bacterial pathogens and, as a result, defined the most relevant antigenic proteins that are targeted by the human immune system, including their antibody binding sites (epitopes). This catalogue of antigens is termed the antigenome and the novel technology offers an integrated approach for antigen validation in order to select the most promising candidates for the development of subunit vaccines against the targeted bacterial diseases. Using the antigenome technology, novel protective antigens had been discovered from several important human pathogens.

Antigens, Bacterial↗

PubMed1990

Transcription of the human cytomegalovirus glycoprotein gene family in the short unique component of the viral genome.

The HXLF (HindIII X left reading frame) genes are a group of six open reading frames (ORFs) within the short unique component of the human cytomegalovirus genome. Two or more ORFs of the HXLF gene family code for components of the virion envelope glycoprotein complex designated gcII. Transcription of the various HXLF genes in infected cells was analyzed using specific DNA, RNA, and synthetic oligonucleotide probes. Transcription from each of the six HXLF genes was analyzed at immediate early, early, and late times after infection. Even though an enhancer element in six copies is downstream of HXLF6, immediate early cytoplasmic RNAs from the HXLF genes was not detectable. In contrast, early and late cytoplasmic RNAs were detectable for all six HXLF genes. The HXLF1 and 2 genes and the HXLF3 and 4 genes are transcribed as bicistronic mRNAs of 1.6 and 1.7 kb, respectively. The HXLF5 and 6 genes are transcribed as monocistronic mRNAs. In the HXLF4, 5, and 6 transcription units cytoplasmic viral RNAs were detected from both strands. Bidirectional transcription upstream of the enhancer element occurs at only early and late times after infection. The significance of bicistronic mRNA to the formation of the glycoprotein complex gcII requires further investigation.

Blotting, Northern↗

PubMed2005

Genetic imprinting analysis for alcoholism genes using variance components approach.

Genomic imprinting, which is also known as the parent-of-origin effect, is a mechanism that only expresses one copy of a gene pair depending upon the parental origin. Although many chromosomal regions in the human genome are likely to be imprinted, imprinting is not accounted for in the usual linkage analysis. In this study, using a variance-components approach with a quantitative phenotype ttth-FP1, we found significant evidence of imprinting at two loci, D7S1790 and D1S1631, on chromosome 1 and chromosome 7, respectively. Our results suggest that allowing for the possibility of imprinting can increase the power to detect linkage for localizing genes for alcoholism.

Alcoholism↗

PubMed2003

Evidence for assembly-dependent folding of protein and RNA in an icosahedral virus.

Ordered nucleic acid in an icosahedral virus was first visualized in the X-ray structure of the Picorna-like plant virus, Bean pod mottle virus (BPMV). Virus particles containing the 3500 nucleotide segment of the BPMV bipartite RNA genome (middle component) had nearly 20% of the genome ordered. Here we report the refined structures of the middle component, bottom component (particles containing the 5800 nucleotide segment of the genome), and top component (empty particles of BPMV capsid protein). The bottom component particles contain ordered RNA in the same location as middle component. Although the ordered RNA density in both nucleoprotein particles is the average of the contents of 60 icosahedral asymmetric units, both nucleoprotein components show that the base density for the first two nucleotides is predominantly purine, while the next five appear to be predominantly pyrimidine. The empty capsid demonstrates that RNA dictates the order of the N-terminal 19 residues of the large subunit because these residues are invisible in the top component.

Capsid Proteins↗

PubMed2006

Domain organization within repeated DNA sequences: application to the study of a family of transposable elements.

MOTIVATION: The analysis of repeated elements in genomes is a fascinating domain of research that is lacking relevant tools for transposable elements (TEs), the most complex ones. The dynamics of TEs, which provides the main mechanism of mutation in some genomes, is an essential component of genome evolution. In this study we introduce a new concept of domain, a segmentation unit useful for describing the architecture of different copies of TEs. Our method extracts occurrences of a terminus-defined family of TEs, aligns the sequences, finds the domains in the alignment and searches the distribution of each domain in sequences. After a classification step relative to the presence or the absence of domains, the method results in a graphical view of sequences segmented into domains. RESULTS: Analysis of the new non-autonomous TE AtREP21 in the model plant Arabidopsis thaliana reveals copies of very different sizes and various combinations of domains which show the potential of our method. AVAILABILITY: DomainOrganizer web page is available at www.irisa.fr/symbiose/DomainOrganizer/.

Algorithms↗

PubMed1992

Hagfish humoral defense protein exhibits structural and functional homology with mammalian complement components.

A genomic clone and cDNA fragment encoding a portion of a humoral recognition molecule from the hagfish were isolated and sequenced. The serum protein has previously been described as having structural features that are immunoglobulin-like. Amino acid sequence obtained from the 77-kDa H1 heavy chain facilitated the isolation of a genomic clone containing at least two coding regions. Through use of primers derived from the genomic sequences, a 231-base-pair cDNA fragment was obtained by PCR from liver RNA. Comparison of the deduced 120-amino acid sequence from the N terminus of H1 with known protein sequences revealed substantial sequence similarity with the beta chain of the murine fourth complement component C4 and with the related third and fifth complement molecules C5 and C3 and the major histocompatibility complex-encoded sex-limited protein. Observation of structural and functional similarities associated with the sequence similarity indicate that these molecules share an evolutionary relationship: the polypeptide chain structure of hagfish complement-like protein (CLP) resembles that of C4; CLP contains a hidden thioester group on the 70-kDa chain; CLP binds to streptococcal cells and enhances the phagocytosis of yeast by hagfish leukocytes. These data suggest that CLP forms part of a non-clonally-derived complement-related humoral defense system in the hagfish.

Amino Acid Sequence↗

PubMed1986

Synthesis of viral DNA forms in Nicotiana plumbaginifolia protoplasts inoculated with cassava latent virus (CLV); evidence for the independent replication of one component of the CLV genome.

Totipotent leaf mesophyll protoplasts of Nicotiana plumbaginifolia, Viviani were inoculated with cassava latent virus (CLV) or with full length copies of CLV genomic DNAs 1 and 2 excised from replicative forms of M13 clones. Virus specific DNAs began to appear 48-72h after inoculation with virus or cloned DNAs, coincident with the onset of host cell division. Infected cells accumulated supercoiled forms of DNAs 1 and 2 as well as progeny single-stranded (ss) virion (+) sense DNAs representing each component of the genome. Both supercoiled and ss molecules were synthesised by cells inoculated with cloned DNA 1 alone but DNA 2 failed to replicate independently.

Cell Division↗

PubMed1996

Accessory human cytomegalovirus glycoprotein US9 in the unique short component of the viral genome promotes cell-to-cell transmission of virus in polarized epithelial cells.

Human cytomegalovirus (CMV) encodes accessory glycoproteins that are dispensable for virus growth in nonpolarized cells in culture. We report that CMV deletion mutants lacking the gene for accessory glycoprotein US9 in the unique short component of the viral genome are impaired in plaque formation in polarized human retinal pigment epithelial (ARPE-19) cells. Comparison of CMV deletion mutants in US9 with herpes simplex virus type 1 deletion mutants lacking glycoproteins gE and gI showed that both of these mutants are impaired in altering junctional complexes and increasing paracellular permeability in polarized ARPE-19 cells cultured on permeable filter supports. Results of functional studies indicate that CMV US9 and homologs of gE have analogous roles in promoting virus spread across lateral membranes of polarized epithelial cells.

Animals↗

PubMed1999

Domain organization and molecular characterization of 13 two-component systems identified by genome sequencing of Streptococcus pneumoniae.

In bacteria, adaptive responses to environmental stimuli are often initiated by two-component signal transduction systems (TCS). The prototypical TCS comprises two proteins: a histidine kinase (HK, hk) and a response regulator (RR rr). Recent research has suggested that compounds that inhibit two-component systems might have good antibacterial activity. In order to identify TCS that are crucial for growth or virulence of Streptococcus pneumoniae, we have examined the genomic sequence of a virulent S. pneumoniae strain for genes that are related to known histidine kinases or response regulators. Altogether 13 histidine kinases and 13 response regulators have been identified. The protein sequences encoded by these genes were compared with sequences deposited in public databases. This analysis revealed that two of the 13 pneumococcal TCSs have been described before (ciaRH and comDE) and two are homologous to the yycFG and the phoRP genes of Bacillus subtilis. All the pneumococcal response regulators contain putative DNA binding motifs within the C-terminal output domain, implying that they are involved in transcriptional control. Two of these response regulators are obviously the first representatives of a new subfamily containing an AraC-type DNA-binding effector domain. To assess the regulatory role of these transcription factors, we disrupted each of the 13 response regulator genes by insertional mutagenesis. All the viable mutant strains with disrupted response regulator genes were further characterized with regard to growth in vitro, competence, and experimental virulence. Two response regulator genes could not be inactivated, indicating that they may regulate essential cellular functions. The possibility of using these systems as targets for the development of novel antibacterials will be discussed.

Amino Acid Motifs↗

PubMed1998

Asymmetric infectivity of pseudorecombinants of cabbage leaf curl virus and squash leaf curl virus: implications for bipartite geminivirus evolution and movement.

The bipartite geminiviruses squash leaf curl virus (SqLCV) and cabbage leaf curl virus (CLCV) have distinct host ranges. SqLCV infects a broad range of plants within the Cucurbitaceae, including pumpkin and squash, and CLCV has a broad host range within Brassicaceae that includes cabbage and Arabidopsis thaliana. Despite this, the genomic A components of these viruses share a high degree of sequence identity, particularly in the gene encoding the replication protein AL1, and their common regions are 77% identical. However, there is unexpected sequence diversity in the common regions of the two CLCV genomic A and B components, these being only 80% identical. Based on these sequence similarities, we investigated the host range properties of pseudorecombinants of SqLCV and CLCV. We found that in a pseudorecombinant virus consisting of the A component of CLCV and the B component of SqLCV, both components replicated in tobacco protoplasts, and this pseudorecombinant was infectious and caused systemic disease in Nicotiana benthamiana, a common host to all bipartite geminiviruses. However, this pseudorecombinant did not move systemically in pumpkin or Arabidopsis, despite the demonstrated replication compatibility of the genome components. As a result of the greater sequence differences between the common regions, the pseudorecombinant of SqLCV A and CLCV B components neither replicated the CLCV B component nor systemically infected any of the hosts tested. These findings demonstrate that for different geminiviruses with distinct host ranges, the replication origins and AL1 proteins can be sufficiently similar to permit infectious pseudorecombinants, but replication alone is not sufficient to cause systemic disease, and host range may ultimately be limited at the level of movement. The results of this study further suggest that CLCV is an evolving virus that can provide insights into how new bipartite geminiviruses arise from mixed infections.

Base Sequence↗

PubMed2000

A genomic analysis of two-component signal transduction in Streptococcus pneumoniae.

A genomics-based approach was used to identify the entire gene complement of putative two-component signal transduction systems (TCSTSs) in Streptococcus pneumoniae. A total of 14 open reading frames (ORFs) were identified as putative response regulators, 13 of which were adjacent to genes encoding probable histidine kinases. Both the histidine kinase and response regulator proteins were categorized into subfamilies on the basis of phylogeny. Through a systematic programme of mutagenesis, the importance of each novel TCSTS was determined with respect to viability and pathogenicity. One TCSTS was identified that was essential for the growth of S. pneumoniaeThis locus was highly homologous to the yycFG gene pair encoding the essential response regulator/histidine kinase proteins identified in Bacillus subtilis and Staphylococcus aureus. Separate deletions of eight other loci led in each case to a dramatic attenuation of growth in a mouse respiratory tract infection model, suggesting that these signal transduction systems are important for the in vivo adaptation and pathogenesis of S. pneumoniae. The identification of conserved TCSTSs important for both pathogenicity and viability in a Gram-positive pathogen highlights the potential of two-component signal transduction as a multicomponent target for antibacterial drug discovery.

Animals↗

PubMed2003

Two-component systems in Prochlorococcus MED4: genomic analysis and differential expression under stress.

Two-component signal transduction systems, composed of histidine sensory kinases and response regulators, constitute a key element of the mechanism by which bacteria sense and acclimatize to changes in their environment. The availability of whole genome sequences permits a detailed analysis of these genes in cyanobacteria. In the present paper, we focus mainly on Prochlorococcus MED4, a strain adapted to surface oceanic conditions, for which six putative response regulators (rer) and five putative histidine kinases (hik) were identified. These numbers are comparable to those found in the other marine picocyanobacteria but much lower than those found in freshwater cyanobacteria. Moreover, the diversity of these genes is low in Prochlorococcus since most histidine kinases are related to a single group (type I) and most response regulators to a single family (OmpR). Under standard conditions, quantitative reverse transcription polymerase chain reaction revealed that one hik (hik03) and two rer (rer04 and rer05) genes were expressed at relatively high levels compared to the other two-component system genes. In response to high light exposure, a moderate increase (>5-fold) was observed in the expression of some putative rer genes (rer01, rer04, rer05, and rer06), whereas a smaller increase (<3-fold) in hik03 and hik04 mRNA levels was detected. In contrast, both cold and heat shocks decreased rather than increased the expression of most hik and rer genes.

Amino Acid Sequence↗

PubMed2001

High-copy-number expression of Sub2p, a member of the RNA helicase superfamily, suppresses hpr1-mediated genomic instability.

We report on a novel role for a pre-mRNA splicing component in genome stability. The Hpr1 protein, a component of an RNA polymerase II complex and required for transcription elongation, is also required for genome stability. Deletion of HPR1 results in a 1,000-fold increase in genome instability, detected as direct-repeat instability. This instability can be suppressed by the high-copy-number SUB2 gene, which is the Saccharomyces cerevisiae homologue of the human splicing factor hUAP56. Although SUB2 is essential, conditional alleles grown at the permissive temperature complement the essential function of SUB2 yet reveal nonessential phenotypes. These studies have uncovered a role for SUB2 in preventing genome instability. The genomic instability observed in sub2 mutants can be suppressed by high-copy-number HPR1. A deletion mutant of CDC73, a component of a PolII complex, is also unstable for direct repeats. This too is suppressed by high-copy-number SUB2. Thus, defects in both the transcriptional machinery and the pre-mRNA splicing machinery can be sources of genome instability. The ability of a pre-mRNA splicing factor to suppress the hyperrecombination phenotype of a defective PolII complex raises the possibility of integrating transcription, RNA processing, and genome stability or a second role for SUB2.

Adenosine Triphosphatases↗

PubMed2025

Genome size estimation from long read overlaps.

MOTIVATION: Accurate genome size estimation is an important component of genomic analyses such as assembly and coverage calculation, though existing tools are primarily optimized for short-read data. RESULTS: We present LRGE, a novel tool that uses read-to-read overlap information to estimate genome size in a reference-free manner. LRGE calculates per-read genome size estimates by analysing the expected number of overlaps for each read, considering read lengths and a minimum overlap threshold. The final size is taken as the median of these estimates, ensuring robustness to outliers such as reads with no overlaps. Additionally, LRGE provides an expected confidence range for the estimate. We validate LRGE on a large, diverse bacterial dataset and confirm it generalizes to eukaryotic datasets. On bacterial genomes, LRGE outperforms k-mer-based methods in both accuracy and computational efficiency and produces genome size estimates comparable to those from assembly-based approaches, like Raven, while using significantly less computational resources. AVAILABILITY AND IMPLEMENTATION: Our method, LRGE (Long Read-based Genome size Estimation from overlaps), is implemented in Rust and is available as a precompiled binary for most architectures, a Bioconda package, a prebuilt container image, and a crates.io package as a binary (lrge) or library (liblrge). The source code is available at https://github.com/mbhall88/lrge and an archive at https://doi.org/10.5281/zenodo.17183812 under an MIT license.

Genome Size↗

PubMed2005

Absence of a coding region for the helper component-proteinase in the genome of cucumber vein yellowing virus, a whitefly-transmitted member of the Potyviridae.

The complete nucleotide sequence of isolates of Cucumber vein yellowing virus (CVYV) has been determined. The viral genome comprises 9734 nucleotides, excluding a 3'-terminal poly(A) sequence. The genome of CVYV has a 5'-non coding and a 3' non coding region of respectively 67 and 240 nucleotides. The RNA of CVYV encodes a single polyprotein of 3148 amino acid residues and has a deduced genome organization and motifs typical for a member of the family Potyviridae. However, CVYV is atypical because it lacks a coding sequence region for the putative helper-component as well as conserved helper-component-proteinase motifs which may account for its vector relations. All the present coding regions were compared to those from several members of the Potyviridae family. CVYV is most closely related to Sweetpotato mild mottle virus confirming its assignation to the genus Ipomovirus, despite similarities with tritimoviruses.

Animals↗

PubMed2003

The methylated component of the Neurospora crassa genome.

Cytosine methylation is common, but not ubiquitous, in eukaryotes. Mammals and the fungus Neurospora crassa have about 2-3% of cytosines methylated. In mammals, methylation is almost exclusively in the under-represented CpG dinucleotides, and most CpGs are methylated whereas in Neurospora, methylation is not preferentially in CpG dinucleotides and the bulk of the genome is unmethylated. DNA methylation is essential in mammals but is dispensable in Neurospora, making this simple eukaryote a favoured organism in which to study methylation. Recent studies indicate that DNA methylation in Neurospora depends on one DNA methyltransferase, DIM-2 (ref. 6), directed by a histone H3 methyltransferase, DIM-5 (ref. 7), but little is known about its cellular and evolutionary functions. As only four methylated sequences have been reported previously in N. crassa, we used methyl-binding-domain agarose chromatography to isolate the methylated component of the genome. DNA sequence analysis shows that the methylated component of the genome consists almost exclusively of relics of transposons that were subject to repeat-induced point mutation--a genome defence system that mutates duplicated sequences.

Chromatography, Affinity↗

PubMed1999

The complete genome sequence of the major component of a mild citrus tristeza virus isolate.

The genome of the Spanish mild isolate T385 of citrus tristeza virus (CTV) was completely sequenced and compared with the genomes of the severe isolates T36 (Florida), VT (Israel) and SY568 (California). The genome of T385 was 19,259 nt in length, 37 nt shorter than the genome of T36, and 33 and 10 nt longer than those of VT and SY568, respectively, but their organization was identical. T385 had mean nucleotide identities of 81.3, 89.3 and 94% with T36, VT and SY568, respectively. The 3' UTR had over 97% identity in all isolates, whereas the 5' UTR of T385 had 67% identity with VT, 66.3% with SY568 and only 42.5% with T36. In the coding regions, the nucleotide differences between T385 and VT were evenly distributed along the genome (around 90% identity); this was not observed between T385 and the other isolates. T385 and T36 had nucleotide identities around 90% in the eight 3'-terminal ORFs of the genome, but only 72.3% in ORF 1a, a divergence pattern similar to that reported previously for T36 and VT. T385 and SY568 had nucleotide identities close to 90% in the 5'- and 3'-terminal regions of the genome, whereas the central region had over 99% identity. Our data suggest that the central region in the SY568 genome results from RNA recombination between two CTV genomes, one of which was almost identical to T385.

Base Sequence↗

PubMed1981

The major components of the mouse and human genomes. 2. Reassociation kinetics.

The reassociation kinetics of DNA fragments obtained from the major components of the mouse and human genomes (recently isolated in our laboratory) have been investigated. It has been found that the relative amounts of interspersed repeated and unique sequences strikingly differ in the different major components of each genome and in the corresponding major components of the two genomes. Furthermore, within each major component, the interspersed repeated and unique sequences do not differ in dG + dC contents. These findings lead to the general conclusion that the sequence organization of mammalian genomes is not uniform in different chromosomal regions and that it exhibits remarkable variations in different mammals.

Animals↗