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At least 127 records · Page 7Linked to original sources

PubMed1990

Mapping sequences required for productive replication of beet necrotic yellow vein virus RNA 3.

Of the four genome components of beet necrotic yellow vein virus only RNAs 1 and 2 are essential for viral replication in leaves. We have mapped cis-regulatory elements on RNA 3 by introducing deletions into expressible cDNA clones and inoculating leaves with the altered transcripts along with RNAs 1 and 2. Transcripts carrying internal deletions extending to within 69 residues of the 3' poly(A) tail or to within about 300 residues of the 5' terminus were efficiently amplified and encapsidated in vivo. The 3' terminal cis-essential domain can be folded into a secondary structure which is conserved among all four genomic RNAs and which probably contains the minus-strand promoter. RNA 3 transcripts with 75% of the central core of the sequence deleted or replaced by the beta-glucuronidase (GUS) gene were also viable. GUS activity was detected in infected tissue in the latter case.

Base Sequence↗

PubMed1985

The human T cell antigen Leu-2 (T8) is encoded on chromosome 2.

The locus encoding the human T lymphocyte cell surface antigen Leu-2 has been assigned to chromosome 2 with a DNA mapping panel derived from somatic cell hybrids. The two genomic components identified by a cDNA clone for Leu-2 segregated with human chromosome 2 in all 24 independent hybrid clones examined. The cosegregation of the Leu-2 and immunoglobulin kappa (IgK) loci in hybrids with spontaneous rearrangements of chromosome 2 is consistent with the possibility that the Leu-2 locus is on proximal human 2p near IgK. In the mouse, a locus for a T lymphocyte cell surface antigen with properties similar to Leu-2 is closely linked to the IgK locus on mouse chromosome 6. Hence the syntenic relationship of a gene implicated in T cell killing with the immunoglobulin kappa locus would then be conserved in the mouse and human genomes.

Animals↗

PubMed2003

Limitations on geminivirus genome size imposed by plasmodesmata and virus-encoded movement protein: insights into DNA trafficking.

Animals and plants evolved systems to permit non-cell-autonomous trafficking of RNA, whereas DNA plays a cell-autonomous role. In plants, plasmodesmata serve as the conduit for this phenomenon, and viruses have evolved to use this pathway for the spread of infectious nucleic acids. In this study, a plant DNA virus was used to explore the constraints imposed on the movement of DNA through this endogenous RNA trafficking pathway. The combined properties of the geminivirus-encoded movement protein and plasmodesmata were shown to impose a strict limitation on the size of the viral genome at the level of cell-to-cell movement. Size-increased viral genome components underwent homologous and nonhomologous recombination to overcome this strict limitation. Our results provide insights into the genetic mechanisms that underlie viral evolution and provide a likely explanation for why relatively few types of plant DNA viruses have evolved: they would have had to overcome the constraints imposed by an endogenous system operating to ensure that DNA acts in a cell-autonomous manner.

Cloning, Molecular↗

PubMed2004

The EMBL Nucleotide Sequence Database.

The EMBL Nucleotide Sequence Database (http://www.ebi.ac.uk/embl/), maintained at the European Bioinformatics Institute (EBI), incorporates, organizes and distributes nucleotide sequences from public sources. The database is a part of an international collaboration with DDBJ (Japan) and GenBank (USA). Data are exchanged between the collaborating databases on a daily basis to achieve optimal synchrony. The web-based tool, Webin, is the preferred system for individual submission of nucleotide sequences, including Third Party Annotation (TPA) and alignment data. Automatic submission procedures are used for submission of data from large-scale genome sequencing centres and from the European Patent Office. Database releases are produced quarterly. The latest data collection can be accessed via FTP, email and WWW interfaces. The EBI's Sequence Retrieval System (SRS) integrates and links the main nucleotide and protein databases as well as many other specialist molecular biology databases. For sequence similarity searching, a variety of tools (e.g. FASTA and BLAST) are available that allow external users to compare their own sequences against the data in the EMBL Nucleotide Sequence Database, the complete genomic component subsection of the database, the WGS data sets and other databases. All available resources can be accessed via the EBI home page at http://www.ebi.ac.uk.

Animals↗

PubMed2006

Drosophila melanogaster as a model for studying protein-encoding genes that are resident in constitutive heterochromatin.

The organization of chromosomes into euchromatin and heterochromatin is one of the most enigmatic aspects of genome evolution. For a long time, heterochromatin was considered to be a genomic wasteland, incompatible with gene expression. However, recent studies--primarily conducted in Drosophila melanogaster--have shown that this peculiar genomic component performs important cellular functions and carries essential genes. New research on the molecular organization, function and evolution of heterochromatin has been facilitated by the sequencing and annotation of heterochromatic DNA. About 450 predicted genes have been identified in the heterochromatin of D. melanogaster, indicating that the number of active genes is higher than had been suggested by genetic analysis. Most of the essential genes are still unknown at the molecular level, and a detailed functional analysis of the predicted genes is difficult owing to the lack of mutant alleles. Far from being a peculiarity of Drosophila, heterochromatic genes have also been found in Saccharomyces cerevisiae, Schizosaccharomyces pombe, Oryza sativa and Arabidopsis thaliana, as well as in humans. The presence of expressed genes in heterochromatin seems paradoxical because they appear to function in an environment that has been considered incompatible with gene expression. In the future, genetic, functional genomic and proteomic analyses will offer powerful approaches with which to explore the functions of heterochromatic genes and to elucidate the mechanisms driving their expression.

Animals↗

PubMed1987

Inhibition of estrous behavior in rats by intrahypothalamic application of agents that disrupt nuclear binding of estrogen-receptor complexes.

This study tested the hypothesis that estrogen facilitation of reproductive behavior in female rats requires the binding of estrogen-receptor complexes to the genomic components of hypothalamic cell nuclei. Female rats were implanted stereotaxically with bilateral guide cannulae aimed at the ventromedial nucleus of the hypothalamus (VMH). Animals were ovariectomized following recovery from the implant surgery and randomly assigned to receive one of four drug treatments: actinomycin-D, ethidium bromide, netropsin, or 4', 6-diamidino-2-phenylindole. Each female received at least two tests for estrous behavior 48 hr after estrogen priming. On one test, drug-filled cannulae were lowered into the VMH 1 hr prior to a subcutaneous injection of 2-3 micrograms of estradiol benzoate (EB); on the other test blank cannulae were inserted 1 hr prior to EB treatment. Intracranial administration of all four compounds, which disrupt estrogen-receptor binding to hypothalamic nuclei, inhibited both the quantity and the quality of lordosis responses to systemic injections of EB. The results support the hypothesis that specific receptor interactions with the genome of hypothalamic cells mediate estrogen facilitation of estrous behavior in female rats.

Animals↗

PubMed2026

A functional atlas of transposon-encoded products and their integration into host networks.

Transposable elements (TEs) are pervasive genomic components that propagate via self-encoded factors, yet the nature, regulation, and function of these factors remain largely unresolved. Here, we integrated extensive long- and short-read transcriptome data, regulatory network analyses, deep proteomics, and structural predictions to construct a comprehensive atlas of TE products in Arabidopsis. We show that TE expression is embedded within host regulatory circuits, with DNA methylation and transcription factors jointly shaping TE transcriptional activity. Proteomic analyses confirm the production of over a hundred of high-confidence TE-encoded proteins, and structure-guided analyses of the transcript-informed TE proteome predict previously uncharacterized structural folds, multimerization capacity, and host protein interaction potential. Structural alignments further uncover cryptic homologies between TE-encoded proteins and host factors, including cases of domestications and co-options. Together, our study reveals the functional integration of TEs into cellular pathways and underscores the role of TEs as active drivers of genome function and innovation.

Arabidopsis↗

PubMed1977

Characterization of RNA from equine infectious anemia virus.

The genome of equine infectious anemia virus, a nononcogenic retrovirus, has been characterized by velocity sedimentation, electrophoresis in polyacrylamide gels, buoyant density in CS2SO4, and susceptibility to nuclease digestion. The nucleic acid of purified virus was resolved by sedimentation analysis into a fast-sedimenting genome component, which comprises about two-thirds of the virion RNA, and a slow-sedimenting RNA, which is probably comprised of host-derived tRNA and a trace amount of 5S RNA. The fast-sedimenting RNA had a sedimentation coefficient of 62S and a molecular weight of 5.4 X 10(6) to 5.6 X 10(6), as determined by sedimentation velocity and electrophoretic mobility. Upon heat denaturation, [3H]uridine-labeled 62S RNA dissociated into material comprised of 90 to 95% single-stranded species, sedimenting predominantly at 34S, with a molecular weight of 2.7 X 10(6) to 2.9 X 10(6) and 5 to 10% 4S RNA. The 62S RNA was predominantly single-stranded but contained double-stranded regions, as indicated by partial resistance to RNase IA and SI nuclease and by a lower buoyant density in CS2SO4 than that of the single-stranded 34S RNA derived by heat denaturation. These data indicated that the viral genome consisted of two 34S subunits of single-stranded RNA held in a high-molecular-weight complex with 4S RNA by a mechanism involving a small degree of base pairing. Thus, the structure of equine infectious anemia virus RNA is similar to that of other retroviruses.

Centrifugation, Density Gradient↗

PubMed1992

The equine herpesvirus type 1 (EHV-1) homolog of herpes simplex virus type 1 US9 and the nature of a major deletion within the unique short segment of the EHV-1 KyA strain genome.

The DNA sequence of the short (S) genomic component of the equine herpesvirus type 1 (EHV-1)KyA strain has been determined recently in our laboratory. Analysis of a 1353-bp BamHI/PvuII clone mapping at the unique short/terminal inverted repeat (Us/TR) junction revealed 507 bp of Us and 846 bp of TR sequences as well as an open reading frame (ORF) that is contained entirely within the Us. This ORF encodes a potential polypeptide of 219 amino acids that shows significant homology to the US9 proteins of herpes simplex virus type 1 (HSV-1), EHV-4, pseudorabies virus (PRV), and varicella zoster virus (VZV). The US9 polypeptides of the two equine herpesviruses exhibit 50% identity but are twice as large as their counterparts in HSV-1, PRV, and VZV. All five US9 proteins are enriched for serine and threonine residues and share a conserved domain of highly basic residues followed by a region of nonpolar amino acids. DNA sequence and Southern blot hybridization analyses revealed that the Us of EHV-1 KyA differs from the Us of EHV-1 KyD and AB1 in that the ORFs encoding glycoproteins I and E and a unique 10-kDa polypeptide are deleted from the KyA genome. These data demonstrate that the predicted 10-kDa protein unique to EHV-1 is nonessential for replication in vitro and that EHV-1 glycoproteins I and E, like their equivalents in HSV-1 and PRV, are also nonessential. These findings and those reported previously by this laboratory and others reveal that the Us segment of EHV-1 comprises nine ORFs, two of which, US4 and 10-kDa ORF, are unique to EHV-1. The gene order of the Us is US2, protein kinase, gG, US4, gD, gI, gE, 10 kDa, and US9.

Amino Acid Sequence↗

PubMed2004

An intragenic guide RNA location suggests a complex mechanism for mitochondrial gene expression in Trypanosoma brucei.

In Trypanosoma brucei, two classes of transcripts are produced from two distinct mitochondrial genome components. Guide RNAs (gRNAs) are usually minicircle encoded and exist as primary transcripts, while the maxicircle-encoded rRNAs and mRNAs are processed from a polycistronic precursor. The genes for the gRNAs gMURF2-II and gCYb(560) each have uncommon kinetoplast DNA (kDNA) locations that are not typically associated with transcription initiation events. We demonstrate that the conserved maxicircle gRNA gMURF2-II has an unusual location within the ND4 gene. This is the first report of a completely intragenic gene in kDNA. In addition, the gMURF2-II and ND4 transcripts are generated by distinctly different events; the ND4 mRNA is processed from a polycistronic precursor, while transcription of the gRNA initiates downstream of the 5' end of the ND4 gene. The gCYb(560) gene has an atypical minicircle location in that it is not flanked by the inverted repeat sequences that surround the majority of minicircle gRNA genes. Our data indicate that the mature gCYb(560) gRNA is also a primary transcript and that the 5'-end heterogeneity previously observed for this gRNA is a result of multiple transcription initiation sites and not of imprecise 5'-end processing. Together, these data indicate that gRNA genes represent individual transcription units, regardless of their genomic context, and suggest a complex mechanism for mitochondrial gene expression in T. brucei.

Animals↗

PubMed1991

Putative 65 kDa protein of beet yellows closterovirus is a homologue of HSP70 heat shock proteins.

A portion of the RNA genome of beet yellows closterovirus (BYV) has been sequenced encompassing a complete long open reading frame (ORF) potentially encoding a 65 kDa protein. The sequence of this putative protein was strikingly similar to those of HSP70-related heat shock proteins. The counterparts of all the eight segments strongly conserved in HSP70s could be confidently identified in the BYV 65 kDa protein. It is suggested that some of these segments might be the ATP-binding site(s) and that, similarly to the heat shock proteins, the 65 kDa is probably ATP-binding. Generally, however, the divergence between the 65 kDa sequence and the sequences of the HSP70s was much more pronounced than that between any two members of the latter family, allowing a clearer delineation of clusters of conserved residues that might be crucial for protein function. It is suggested that these observations will be helpful in functional dissection of the proteins of the HSP70 family. Analysis of the sequence of a portion of the ORF found upstream from the 65 kDa ORF showed that the C-terminal domain of the encoded protein could be an RNA-dependent RNA polymerase closely related to those of tricornaviruses, a family of RNA plant viruses with three component genomes.

Adenosine Triphosphate↗

PubMed1993

Contributions of the brome mosaic virus RNA-3 3'-nontranslated region to replication and translation.

Sequences upstream of the 3'-terminal tRNA-like structure of brome mosaic virus RNAs have been predicted to fold into several stem-loop and pseudoknot structures. To elucidate the functional role of this upstream region, a series of deletions was made in cDNA clones of RNA-3, a genomic component not required for replication. These deletion mutants were transcribed in vitro and cotransfected with RNA-1 and RNA-2 into barley protoplasts. Deletion of single stem-loop structures gave progeny retaining near-wild-type accumulation levels. Constructions representing deletion of two or three stem-loops substantially lowered the accumulation of progeny RNA-3 relative to wild-type levels. RNA-3 mutants bearing deletions of longer sequences or of the entire region (delta PsKs RNA-3) replicated poorly, yielding no detectable RNA-3 or RNA-4 progeny. Levels of RNA-1 and RNA-2, in the presence of a mutant RNA-3, were found to increase relative to the accumulation observed in a complete wild-type transfection. The stability of delta PsKs RNA-3 in protoplasts was somewhat lower than that of wild-type RNA during the first 3 h postinoculation. Little difference in translatability in vitro of wild-type and RNA-3 constructs bearing deletions within the stem-loop region was observed, and Western immunoblot analysis of viral coat protein produced in transfected protoplasts showed that protein accumulation paralleled the amount of RNA-4 message produced from the various sequences evaluated. These results indicate that the RNA-3 pseudoknot region plays a minor role in translational control but contributes substantially to the overall replication of the brome mosaic virus genome.

Base Sequence↗

PubMed1999

Molecular changes at Rrn loci in barley (Hordeum vulgare L.) hybrids with H. bulbosum (L.)

Southern blots of restriction fragments of genomic DNAs from Hordeum vulgare (L.), H. bulbosum (L.), and interspecific hybrids and their derivatives were hybridized with rDNA probe to identify locus-specific modifications at Rrn loci. H. bulbosum rDNA revealed a single EcoRV site per repeat compared with two sites in H. vulgare rDNA repeats. H. bulbosum accessions possessed at least two rDNA repeat lengths, indicating heterozygosity at the Rrn locus. Hybrids possessed both H. vulgare and H. bulbosum rDNA repeats. Two of the hybrid derivatives possessed bulbosum-specific Sau3AI and HaeIII rDNA fragments, while amphiploid and doubled haploid derivatives lacked H. bulbosum rDNA repeat units and (or) fragments. Two hybrid derivatives, one amphiploid and a doubled haploid derived from the same parental combination, lacked the vulgare Rrn2-specific 9.0-kb rDNA repeat. This is the first conclusive evidence for the elimination of vulgare genetic material in vulgare-bulbosum hybrids. The ratios of 9.0- to 9.9-kb vulgare repeats and H. vulgare to H. bulbosum rDNA repeats indicate partial loss of the vulgare-specific 9.0-kb rDNA repeat among the hybrids. Differences in MboI and Sau3AI fragments and the ratios of 9.0 to 9.9 kb vulgare rDNA repeats revealed differential methylation at Rrn1 and Rrn2 loci. Hybrids and derivatives showed differential distribution of methylation of EcoRI, BglII, and SacI sites at the Rrn1 locus. Two of the hybrid derivatives exhibited extensive CpG-biased methylation. Data presented here are indicative of the differences in the onset of events triggered by the interaction of the component genomes and enabled detection of differential methylation among Rrn loci, loss of H. vulgare genetic material, and development of doubled haploids with the Rrn1 locus.

Journal Article↗

PubMed1988

Genetic analysis of tomato golden mosaic virus: the coat protein is not required for systemic spread or symptom development.

The geminiviruses are a unique group of higher plant viruses that are composed of twin isometric particles which contain circular, single-stranded DNA. Tomato golden mosaic virus (TGMV), a whitefly-transmitted agent, belongs to the subgroup of geminiviruses whose members possess a bipartite genome. The TGMV A genome component has the capacity to encode at least four proteins. One of these is the viral coat protein, as inferred by homology with coat-protein, genes of other geminiviruses and by the observation of typical geminate particles in transgenic plants that contain inserts of TGMV A DNA. We have investigated the role of the coat protein in TGMV replication and report here that its coding sequence may be interrupted or substantially deleted without loss of infectivity. However, certain coat-protein mutants showed reproducible delays in time of symptom appearance as well as reduced symptom development, when inoculated onto transgenic Nicotiana benthamiana plants containing the TGMV B component. The most attenuated symptoms were seen with a mutant in which the coat-protein coding sequence was almost entirely deleted. The significance of these findings for the development of plant vectors from TGMV DNA is discussed.

Journal Article↗

PubMed2003

Dehydroepiandrosterone upregulates neural androgen receptor level and transcriptional activity.

The mechanism of action of dehydroepiandrosterone (DHEA), a neuroactive neurosteroid synthesized in the brains of humans and other mammals, has not been fully characterized in the adult brain. Although well known for modulatory effects on GABA(A), NMDA, and sigma(1) receptors, studies in both CNS and peripheral target cells suggest that DHEA also may exert genomic effects via the androgen receptor (AR). The current study tested the hypothesis that DHEA was capable of producing androgenic effects in the CNS by assaying its ability to induce three characteristic effects of an androgenic compound. These included the ability to upregulate neural AR protein level in mouse brain and immortalized GT1-7 hypothalamic cells, the capacity to induce transcriptional activity through AR in CV-1 cells transfected with an MMTV-ARE-CAT reporter, and competition for recombinant AR binding in a radioligand binding assay. The results showed that DHEA treatment significantly augmented AR both in vivo and in vitro, and that this effect was not blocked by trilostane (TRIL), a known 3beta-hydroxysteroid dehydrogenase (3beta-HSD) inhibitor. DHEA also promoted AR-mediated CAT reporter expression and competed with dihydrotestosterone (DHT) for binding to recombinant AR in a cell-free system. These data indicate that DHEA possesses intrinsic androgenic activity that is potentially independent of metabolic conversion to other androgens, and that it can affect gene function through the AR. In combination with its modulation of neurotransmitter receptors at the cell membrane level, the findings suggest that the mechanism of action of DHEA in the brain can involve a "crosstalk" cellular signaling system that involves both nongenomic and genomic components.

Animals↗

PubMed1993

Detection and possible functions of African cassava mosaic virus DNA B gene products.

Polyclonal antisera raised against synthetic oligopeptides have been used to detect the DNA B gene products BV1 and BC1 of the geminivirus African cassava mosaic virus following SDS-PAGE fractionation of Nicotiana benthamiana extracts. BV1 antiserum detected a soluble protein of 29 kDa, consistent with the size predicted from sequence data. BC1 antiserum detected proteins of 37, 39, and 42 kDa in addition to variable, less abundant species, all of which are larger than the predicted size of 34 kDa. BC1 antiserum detected a single protein of 35-36 kDa following in vitro translation in reticulocyte lysate, suggesting that BC1 is post-translationally modified in plants. The nature of the modification was not resolved, although neither glycosylation nor association with nucleic acids is involved. In common with putative spread proteins of several other plants viruses, BC1 co-fractionated with the cell wall. The replication of both genomic components in N. tabacum protoplasts was unaffected by the introduction of frameshift mutations into BV1 and BC1 coding regions. In inoculated N. benthamiana leaves, however, the accumulation of a BV1 mutant was significantly reduced compared to the levels attained by co-inoculated, complementing BV1 and BC1 mutants. In contrast, the accumulation of a BC1 mutant was unaffected, although symptom induction in inoculated leaves and systemic infection occurred only in the presence of both BV1 and BC1. The results are consistent with a role for BV1 in localized cell-to-cell spread and for BC1, possibly together with BV1, in long-distance vascular spread of the virus.

Blotting, Western↗

PubMed2001

Pathogenicity of a natural recombinant associated with ageratum yellow vein disease: implications for geminivirus evolution and disease aetiology.

Yellow vein disease of Ageratum conyzoides is caused by a viral DNA complex consisting of the genomic component (DNA A) of the monopartite begomovirus Ageratum yellow vein virus (AYVV, family: Geminiviridae) and a small satellite-like DNA beta component. AYVV DNA A is unable to induce symptoms in this host alone but can systemically infect A. conyzoides in which it accumulates to low levels. Here, we demonstrate that the yellow vein phenotype can also be produced by co-inoculating A. conyzoides with AYVV DNA A and recDNA-Abeta17, a naturally occurring recombinant of approximately the same size as DNA beta that contains sequences from both DNA A and DNA beta. Symptoms induced by DNA A and recDNA-Abeta17 in A. conyzoides and Nicotiana glutinosa are qualitatively similar to those associated with DNA A and DNA beta although milder. Recombination between DNA A and DNA beta to produce a chimera resembling recDNA-Abeta17 was observed after whitefly transmission of the disease in A. conyzoides. Hence, such recombination events are likely to occur frequently, implying that recombinants will normally be associated with this type of disease complex in the field. Possible implications of these findings for the evolution of begomoviruses and the aetiology of their diseases are discussed.

Asteraceae↗

PubMed2001

Exchange of three amino acids in the coat protein results in efficient whitefly transmission of a nontransmissible Abutilon mosaic virus isolate.

Geminiviruses are transmitted in a circulative manner by whiteflies, leafhoppers, or treehoppers. The whitefly species Bemisia tabaci (Genn.) is the vector for members of the genus Begomovirus. The closely related bipartite Central American begomoviruses Abutilon mosaic virus (AbMV), Sida golden mosaic virus originating from Costa Rica (SiGMV-CR), and Sida golden mosaic virus originating from Honduras (SiGMV-Hoyv) were used to study transmission by their insect vector. The AbMV isolate is defective in transmission, whereas the two Sida-infecting viruses are readily transmitted by B. tabaci. These three viruses are able to form pseudorecombinant viruses by exchange of genomic components. The pseudorecombinant virus SiGMV-Hoyv A/AbMV B was transmissible, whereas the reciprocal pseudorecombinant virus AbMV A/SiGMV-Hoyv B was not transmitted, indicating that DNA B is not involved in the transmission defect. However, the uptake of the pseudorecombinant virus AbMV A/SiGMV-Hoyv B was much better than AbMV itself, indicating that DNA B or DNA B gene products enhance uptake of viral DNA. Exchange of AbMV coat protein with that of SiGMV-CR resulted in a transmissible chimeric AbMV. Mutagenesis of the AbMV coat protein showed that the exchange of two amino acids, at positions 124 and 149, was sufficient to obtain a whitefly-transmissible AbMV mutant. However, when amino acid 174 was altered in addition to amino acids 124 and 149 AbMV was readily transmitted by B. tabaci. From this we conclude that it is not a concise motif, such as the amino acid triplet, aspartate-alanine-glycine (DAG), involved in aphid transmission of potyviruses, that determines transmissibility of begomoviruses by B. tabaci. Instead it is the composition of the coat protein domain from amino acid 123 to 149, as a minimal transmission domain, with the contribution of amino acids 149 to 174 for efficient transmission.

Amino Acid Sequence↗