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At least 667 records · Page 37Linked to original sources
PubMed1993

A new type of G-->A hypermutation affecting human immunodeficiency virus.

A form of G-->A hypermutation preferentially affecting GA dinucleotides of genomic RNA has been found to occur in retroviral systems ("type 1"). In a detailed longitudinal study of an AIDS patient we have observed a new type of G-->A hypermutation, which preferentially affects one or more 5' G residues in runs of G's. HIV-1 proviral DNA samples obtained at widely separate times during this patient's course contained representatives of this type of G-->A hypermutation, designated "type 2." We propose that G-->A hypermutation is caused by a mutated form of the HIV-1 reverse transcriptase; and that hypermutated DNA may persist for long periods in infected patients, perhaps as proviral DNA in long-lived cell lineages. Like type 1 G-->A hypermutation, type 2 G-->A hypermutation may contribute to the heterogeneity of replicating pools of HIV by recombination.

Acquired Immunodeficiency Syndrome↗
PubMed1995

Molecular characterization of human immunodeficiency virus type 1 isolates from Venezuela.

Eight HIV-1 isolates from Venezuela have been characterized by nucleotide sequencing of the entire reverse transcriptase (RT)- and surface glycoprotein (gp 120)-coding regions. Average mutant frequencies were 2.5 x 10(-2) substitutions per nucleotide (s/nt) for the RT-coding region, and 10 x 10(-2) or 6.8 x 10(-2) s/nt for the gp120-coding region, depending on whether gaps introduced for optimal alignment were or were not, respectively, considered in the calculations. Phylogenetic trees were derived by maximum-likelihood, neighbor-joining, and maximum parsimony methods. In the trees derived from both RT- and gp120-coding regions, Venezuelan isolates cluster with subtype B viruses. However, the relative position of some of the isolates is considerably different in the two trees. Unique V3 loop amino acid sequences, not represented in the current database, have been identified among the Venezuelan isolates. In addition to representing the first molecular characterization of HIV-1 from Venezuela, the extensive genetic heterogeneity observed reinforces the interest in characterizing additional HIV-1 isolates worldwide for adequate vaccine design.

Adult↗
PubMed1993

A tRNA gene transcription initiation site is similar to mRNA and rRNA promoters in plant mitochondria.

The gene for tRNA(Phe) is located 292 nucleotides upstream of the tRNA(Pro) gene in the Oenothera mitochondrial genome. Hybridization with in vitro capped primary transcripts indicates a transcription initiation site in the 5' region of the gene for tRNA(Phe). Primer extension experiments show the presence of precursor transcripts covering tRNA(Phe) and adjacent sequences up to a transcription initiation site 181 or 180 nucleotides upstream of the tRNA gene. The genomic sequence at this transcription initiation site contains the consensus motif derived for putative promoters of mitochondrial protein and rRNA coding genes in dicotyledonous plants. This sequence similarity suggests that tRNAs, rRNAs and mRNAs can be transcribed from homologous promoters in plant mitochondria.

Base Sequence↗
PubMed1996

Strand transfer is enhanced by mismatched nucleotides at the 3' primer terminus: a possible link between HIV reverse transcriptase fidelity and recombination.

Strand transfer catalyzed by HIV reverse transcriptase (RT) was examined. The system consisted of a 142 nt RNA (donor) to which a 50 nt DNA primer was hybridized. The primer bound such that its 3' terminal nucleotide hybridized to the 12th nt from the 5' end of the donor. The 3' terminal nucleotide on the primer was either a G, A or T residue. Since the corresponding nucleotide of the donor was a C, the G formed a matched terminus and the A or T a mismatched terminus. The efficiency with which DNA bound to the donor transferred to a second RNA, termed acceptor, was monitored. The acceptor was homologous to the donor for all but the last 9 nt at the 5' end of the donor. Therefore, homologous strand transfer could occur at any point prior to the DNA being extended into the nonhomologous region on the donor. Strand transfer occurred approximately twice as efficiently with the mismatched versus matched substrates. The mismatched nucleotide was fixed into transfer products indicating that excision of the mismatch was not required for RT extension or transfer. Results suggest that base misincorporations by RT may promote recombination by enhancing strand transfer.

Base Sequence↗
PubMed2006

The Rhizobium etli sigma70 (SigA) factor recognizes a lax consensus promoter.

A collection of Rhizobium etli promoters was isolated from a genomic DNA library constructed in the promoter-trap vector pBBMCS53, by their ability to drive the expression of a gusA reporter gene. Thirty-seven clones were selected, and their transcriptional start-sites were determined. The upstream sequence of these 37 start-sites, and the sequences of seven previously identified promoters were compared. On the basis of sequence conservation and mutational analysis, a consensus sequence CTTGACN16-23TATNNT was obtained. In this consensus sequence, nine on of twelve bases are identical to the canonical Escherichia coli sigma70 promoter, however the R.etli promoters only contain 6.4 conserved bases on average. We show that the R.etli sigma factor SigA recognizes all R.etli promoters studied in this work, and that E.coli RpoD is incapable of recognizing them. The comparison of the predicted structure of SigA with the known structure of RpoD indicated that regions 2.4 and 4.2, responsible for promoter recognition, are different only by a single amino acid, whereas the region 1 of SigA contains 72 extra residues, suggesting that the differences contained in this region could be related to the lax promoter recognition of SigA.

Bacteria↗
PubMed1999

Conservation of putative promoter sequences located upstream of chlamydial major sigma factor gene, sigA among Chlamydia spp.

A highly conserved 40-nucleotide sequence was identified. Two completely conserved sequences, TAGATT and TAAACT, separated by 17 nucleotides resemble the consensus sequence recognized by the Escherichia coli major sigma factor and sequence found in other chlamydial promoters. In addition, the adenine-rich sequence present in many chlamydial promoters was also conserved upstream of the putative -35 element. These findings suggest that the conserved sequence may play a role in the regulatory function at the transcriptional level. Multiple ATG codons were found at the 5'-terminal region of the chlamydial sigA ORFs except for Chlamydia pneumoniae, although the putative Shine-Dargarno sequence was absent.

Bacterial Outer Membrane Proteins↗
PubMed1997

Molecular systematic studies of eubacteria, using sigma70-type sigma factors of group 1 and group 2.

Sigma factors of the sigma70 family were used as a phylogenetic tool to compare evolutionary relationships among eubacteria. Several new sigma factor genes were cloned and sequenced to increase the variety of available sequences. Forty-two group 1 sigma factor sequences of various species were analyzed with the help of a distance matrix method to establish a phylogenetic tree. The tree derived by using sigma factors yielded subdivisions, including low-G+C and high-G+C gram-positive bacteria, cyanobacteria, and the alpha, beta, gamma, and delta subdivisions of proteobacteria, consistent with major bacterial groups found in trees derived from analyses with other molecules. However, some groupings (e.g., the chlamydiae, mycoplasmas, and green sulfur bacteria) are found in different positions than for trees obtained by using other molecular markers. A direct comparison to the most extensively used molecule in systematic studies, small-subunit rRNA, was made by deriving trees from essentially the same species set and using similar phylogenetic methods. Differences and similarities based on the two markers are discussed. Additionally, 31 group 2 sigma factors were analyzed in combination with the group 1 proteins in order to detect functional groupings of these alternative sigma factors. The data suggest that promoters recognized by the major vegetative sigma factors of eubacteria will contain sequence motifs and spacing very similar to those for the sigma70 sigma factors of Escherichia coli.

Amino Acid Sequence↗
PubMed2003

Identification of Bacillus anthracis by rpoB sequence analysis and multiplex PCR.

Comparative sequence analysis was performed upon Bacillus anthracis and its closest relatives, B. cereus and B. thuringiensis. Portions of rpoB DNA from 10 strains of B. anthracis, 16 of B. cereus, 10 of B. thuringiensis, 1 of B. mycoides, and 1 of B. megaterium were amplified and sequenced. The determined rpoB sequences (318 bp) of the 10 B. anthracis strains, including five Korean isolates, were identical to those of Ames, Florida, Kruger B, and Western NA strains. Strains of the "B. cereus group" were separated into two subgroups, in which the B. anthracis strains formed a separate clade in the phylogenetic tree. However, B. cereus and B. thuringiensis could not be differentiated. Sequence analysis confirmed the five Korean isolates as B. anthracis. Based on the rpoB sequences determined in the present study, multiplex PCR generating either B. anthracis-specific amplicons (359 and 208 bp) or cap DNA (291 bp) in a virulence plasmid could be used for the rapid differential detection and identification of virulent B. anthracis.

Anthrax↗
PubMed1999

Two nucleotides immediately upstream of the essential A6G3 slippery sequence modulate the pattern of G insertions during Sendai virus mRNA editing.

Editing of paramyxovirus P gene mRNAs occurs cotranscriptionally and functions to fuse an alternate downstream open reading frame to the N-terminal half of the P protein. G residues are inserted into a short G run contained within a larger purine run (AnGn) in this process, by a mechanism whereby the transcribing polymerase stutters (i.e., reads the same template cytosine more than once). Although Sendai virus (SeV) and bovine parainfluenza virus type 3 (bPIV3) are closely related, the G insertions in their P mRNAs are distributed differently. SeV predominantly inserts a single G residue within the G run of the sequence 5' AACAAAAAAGGG, whereas bPIV3 inserts one to six G's at roughly equal frequency within the sequence 5' AUUAAAAAAGGGG (differences are underlined). We have examined how the cis-acting editing sequence determines the number of G's inserted, both in a transfected cell system using minigenome analogues and by generating recombinant viruses. We found that the presence of four rather than three G's in the purine run did not affect the distribution of G insertions. However, when the underlined AC of the SeV sequence was replaced by the UU found in bPIV3, the editing phenotype from both the minigenome and the recombinant virus resembled that found in natural bPIV3 infections (i.e., a significant fraction of the mRNAs contained two to six G insertions). The two nucleotides located just upstream of the polypurine tract are thus key determinants of the editing phenotype of these viruses. Moreover, the minimum number of A residues that will promote SeV editing phenotype is six but can be reduced to five when the upstream AC is replaced by UU. A model for how the upstream dinucleotide controls the insertion phenotype is presented.

Base Sequence↗
PubMed1995

Four structurally distinct, non-DNA-binding subunits of human nuclear respiratory factor 2 share a conserved transcriptional activation domain.

Nuclear respiratory factor 2 (NRF-2) was previously purified to near homogeneity from HeLa cells on the basis of its ability to bind tandem recognition sites in the rat cytochrome oxidase subunit IV (RCO4) promoter. It consisted of five subunits, alpha, beta 1, beta 2, gamma 1, and gamma 2. Sequencing of tryptic peptides from alpha and from mixtures of the two beta or two gamma subunits revealed sequence identities with subunits of the mouse GA-binding protein (GABP), a ubiquitously expressed ETS domain activator composed of three subunits, alpha, beta 1, and beta 2. To understand the precise relationship between NRF-2 and GABP, cDNAs for all five NRF-2 subunits have now been cloned and their products have been overexpressed. The results establish that the two additional NRF-2 subunits are molecular variants that differ from GABP beta 1 and beta 2 by having a 12-amino-acid insertion containing two serine doublets. PCR and RNase protection assays show that mRNAs for these variants are expressed in the human but not the rodent cells and tissues examined. The insertion did not alter the ability of the beta and gamma subunits to associate with alpha, the DNA-binding subunit, nor did it affect the ability of NRF-2 beta 1 or beta 2 to direct high-affinity binding of alpha to tandem sites in the RCO4 promoter. In addition, the four NRF-2 beta and gamma subunits were equally proficient in activating transcription in transfected cells when fused to a GAL4 DNA-binding domain. The domain responsible for this transcriptional activation was localized by deletion mapping to a region of approximately 70 amino acids that is conserved in all four NRF-2 beta and gamma subunits. The repeated glutamine-containing hydrophobic clusters within this region bear a strong resemblance to those recently implicated in protein-protein interactions within the transcriptional apparatus.

Amino Acid Sequence↗
PubMedDate not supplied

[Distribution and functional significance of A/T tracts in promotor sequences of Escherichia coli].

Distribution of the A/T tracts described in earlier publications in the region extending from nucleotide -250 to +150 relative to the transcription initiation site of gene transcribed regions adjacent to promoter was studied. Upstream of the -35 region a succession of A/T tracts was discovered distributed at a shorter distance one from another than in analogous elements of the transcribed region (1 and 1.5 helix turns, respectively). Such a positional dependence suggests different functional manifestation of A/T tracts at different transcription steps. Single initiation using the T7D promoter mutant derivatives devoid of A/T tracts in two critical positions, +41 and , yielded shortened products of the corresponding length. One might speculate that such elements adjacent to promoter region play a significant role in transcription complex functioning.

AT Rich Sequence↗
PubMed1992

Characterization of HIV isolates arising after prolonged zidovudine therapy.

Human immunodeficiency virus type 1 (HIV-1) was isolated from five patients with late-stage disease treated with zidovudine (ZDV) for more than 1 year. Peripheral blood mononuclear cells (PBMCs) were used for all virus isolations and to assay for drug resistance. The isolates exhibited a 10- to 100-fold decrease in ZDV susceptibility compared to pretreatment isolates. Multiple clones of a 618 bp segment of the HIV reverse transcriptase gene encompassing codons 60-250 were sequenced for each isolate. The association of alterations at codons Asp67----Asn, Lys70----Arg, Thr215----Phe or Tyr, and Lys219----Gln with ZDV resistance has been previously noted (ref. 5). In this study, the most frequent alterations was Thr215----Tyr although genotypic mixtures of Thr/Tyr and Phe/Tyr were also observed. One isolate with a Tyr215 alteration and unaltered codons at 67, 70, and 219 had high-level ZDV resistance. Alterations at codons 67, 70, and 219 did not appear to increase resistance when seen in combination with Tyr215. Virus isolates obtained from each patient by cultivation with either 0 or 4 microM ZDV were compared and found to have similar alterations at codons 67, 70, 215, and 219, although one instance of apparent in vitro selection for Tyr215 over Phe215 was observed. Assays using PBMCs for virus propagation will permit susceptibility testing of HIV isolates from most patients on antiretroviral drugs to investigate the clinical significance of drug resistance.

Amino Acid Sequence↗
PubMed2000

RNA polymerase (L) gene and genome terminal sequences of ephemeroviruses bovine ephemeral fever virus and Adelaide River virus indicate a close relationship to vesiculoviruses.

The sequence of the RNA genome of bovine ephemeral fever virus (BEFV) was determined from the start of the L (polymerase) gene to the end of the untranslated 5' trailer sequence, completing the sequence of the 14900 nucleotide (nt) genome. The 6470 nt L gene encodes a single long ORF of 2144 amino acids with a deduced molecular weight of 249766 Da. The 70 nt BEFV 5' trailer region displays partial terminal complementarity with the 3' leader sequence and contains a 26 nt direct repeat of the U-rich domain of the 3' leader region. The 47 nt 5' trailer region of Adelaide River virus (ARV) displays terminal sequence similarity to the BEFV trailer and partial terminal complementarity with the ARV 3' leader sequence, but does not contain the direct repeat sequence. The BEFV L protein contains all characteristic sequence motifs of amino acid blocks I-VI, conserved among RNA polymerase proteins of single-stranded (-) RNA viruses, separated by regions of lower homology. Phylogenetic analysis using the complete BEFV L protein sequence indicated a closer relationship to vesicular stomatitis virus than to rabies virus. Sequence comparison of two conserved central domains encompassing blocks II and III and block VI of the BEFV and ARV L proteins indicated they are closely related. An extended phylogenetic analysis using the block III sequence, confirmed the relationship of these ephemeroviruses to vesiculo- and lyssaviruses and to other single-stranded (-) RNA viruses.

Amino Acid Sequence↗
PubMed1979

Site specific enzymatic cleavage of RNA.

The hybridization of a DNA oligonucleotide a specific tetramer or longer) will direct a cleavage by RNase H (EC 3.1.4.34) to a specific site in RNA. The resulting fragments can then be labeled at their 5' or 3' ends, purified, and sequenced directly. This procedure is demonstrated with two RNA molecules of known sequence: 5.8S rRNA from yeast (158 nucleotides) and satellite tobacco necrosis virus (STNV) RNA (1240 nucleotides).

Animals↗
PubMed1992

Characterization of a 7-kilodalton subunit of vaccinia virus DNA-dependent RNA polymerase with structural similarities to the smallest subunit of eukaryotic RNA polymerase II.

A previously unrecognized 7-kDa polypeptide copurified with the DNA-dependent RNA polymerase of vaccinia virus virions. Internal amino acid sequences of the small protein matched a viral genomic open reading frame of 63 codons. Antipeptide antiserum was used to confirm the specific and complete association of the 7-kDa protein with RNA polymerase. The amino acid sequence predicted from the viral gene, named rpo7, was 23% identical to that of the smallest subunit of Saccharomyces cerevisiae RNA polymerase II, and a metal-binding motif, Cys-X-X-Cys-Gly, was located at precisely the same location near the N terminus in the two proteins. RNA analyses demonstrated early transcriptional initiation and termination signals in the rpo7 gene sequence. The viral RNA polymerase subunit was synthesized during the early phase of infection and continued to accumulate during the late phase.

Amino Acid Sequence↗