SEARCH · PubMed Health

Results for “insertion sequence”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 1,387 records · Page 77Linked to original sources

PubMed2026

Ecr positively regulates activity of the PhoQ/PhoP signalling system in Klebsiella pneumoniae.

BACKGROUND: The rising prevalence of polymyxin resistance in multidrug-resistant Klebsiella pneumoniae presents a critical situation with limited therapeutic options. METHODS: Methods Genomic sequencing of 15 clinical polymyxin-resistant K. pneumoniae strains with multidrug resistance revealed that MgrB inactivation, predominantly disrupted by insertion sequences (ISs) in the IS1, IS4, and IS5 families, was the leading cause of polymyxin resistance. Comparative transcriptomics of wild-type, ΔmgrB, and ΔmgrBΔphoP were performed to elucidate the MgrB-PhoPQ regulatory network. RESULTS: This study conducted a system-wide analysis of the regulatory network and identified a species-specific PhoPQ regulon in K. pneumoniae.Beyond the classical MgrB-PhoPQ-ArnBCADTEF pathway, we identified a previously unannotated PhoPQ-regulated gene, 144 bp LN739_RS09850, encoding an Ecr homologue from Enterobacter cloacae. This protein has been reported to confer colistin heteroresistance, with the underlying mechanism not yet functionally validated. This study revealed that overexpression of Ecr homologues decreased colistin susceptibility in both K. pneumoniae and E. cloacae, but this phenotype was abolished upon phoP deletion, confirming PhoP's essential role. Consistent with this dependency, comparative transcriptomics of Ecr-overexpressing K. pneumoniae vs. control revealed significant upregulation of mgrB, phoPQ, arnBCADTE, and pmrD. Two-hybrid bacterial assays further demonstrated direct Ecr-PhoQ interaction. Electrophoretic mobility shift assay confirmed that PhoP directly binds to the ecr promoter in vitro, and a β-galactosidase reporter assay demonstrated that PhoP enhanced ecr promoter activity, indicating that PhoP regulates ecr expression by directly controlling its transcription. CONCLUSION: Collectively, these findings suggest that PhoP may directly activate the transcription of Ecr, with Ecr feedback activating the PhoPQ system via interaction with PhoQ, leading to induction of the arn operon and consequent polymyxin resistance.

Klebsiella pneumoniae↗

PubMed2002

Does the proposed DSE motif form the active center in the Hermes transposase?

Donor cleavage and strand transfer are two functions performed by transposases during transposition of class II transposable elements. Within transposable elements, the only active center described, to date, facilitating both functions, is the so-called DDE motif. A second motif, R-K-H/K-R-H/W-Y, is found in the site-specific recombinases of the tyrosine recombinase family. While present in many bacterial insertion sequences as well as in the eukaryotic family of mariner/Tc1 elements, the DDE motif was considered absent in other classes of eukaryotic class II elements such as P, and hAT and piggyBac. Based on sequence alignments of a hobo-like element from the nematode Caenorhabditis elegans, to a variety of other hAT transposases and several members of the mariner/Tc1 group, Bigot et al. [Gene 174 (1996) 265] proposed the presence of a DSE motif in hAT transposases. In the present study we tested if each of these three residues is required for transposition of the Hermes element, a member of the hAT family commonly used for insect transformation. While D402N and E572Q mutations lead to knock-out of Hermes function, mutations S535A and S535D did not affect transposition frequency or the choice of integration sites. These data give the first experimental support that D402 and E572 are indeed required for transposition of Hermes. Furthermore, this study indicates that the active center of the Hermes transposase differs from the proposed DSE motif. It remains to be shown if other residues also form the active site of this transposase.

Amino Acid Motifs↗

PubMed2003

DNA sequence and analysis of the pYVa127/90 virulence plasmid of Yersinia enterocolitica strain A127/90.

The complete nucleotide sequence and organization of the Yersinia enterocolitica strain A127/90 serotype O:8 biotype 1B virulence plasmid (pYVa127/90) were determined. In contrast to other biotype 1B strains, usually isolated in the USA, strain A127/90 was isolated in Japan. The organization of the genes carried by the 66591-bp pYVa127/90 plasmid is globally similar to that of 67720-bp pYVe8081, the virulence plasmid from 8081, an American isolate of serotype O:8 biotype 1B. Differences are noticeable in the amino acid sequences of the SycH chaperone, the YopM cytotoxin and the plague-protective antigen LcrV. Interestingly, pYVa127/90 contains the entire coding sequence of the YlpA lipoprotein, which is missing from pYVe8081. In addition, we identified five potential new genes. Several vestigial insertion sequence elements appeared to be different between the two plasmids. These observations suggest that both pYV plasmids are from a common ancestor but may have evolved independently following the conquest of two distant ecological niches.

Antigens, Bacterial↗

PubMed2000

Linkage of determinants for streptogramin A, macrolide-lincosamide-streptogramin B, and chloramphenicol resistance on a conjugative plasmid in Enterococcus faecium and dissemination of this cluster among streptogramin-resistant enterococci.

A new streptogramin A resistance gene, satG (= vatE), has been recently identified in Enterococcus faecium UW1965 (Werner and Witte 1999. Antimicrob. Agents Chemother. 43: 1813-1814). Further sequence analysis of this plasmid revealed that vatE is in a cluster together with other resistance genes. The identified ORFs were nearly identical with the already known genes ermB and cat. The ermB fragment exhibited more than 99% identity with a resistance region from the streptococcal plasmid pIP501, whereas the cat fragment also contained a truncated rep gene homologue with more than 99% identity to sequences in small staphylococcal plasmids. The cat-rep and the ermB-vatE segments were linked by an IS1216V insertion sequence widely distributed among enterococci. PCR analysis of additional 76 streptogramin-resistant isolates possessing vatE and ermB revealed a linkage of both genes in 45 isolates (59%); 15 of them with a gene arrangement, cat-repU-IS1216V-ermB-vatE, identical to the reference strain UW1965. An identical linkage of IS1216V-ermB-vatE was found among isolates from poultry manure, poultry meat, stool samples of humans, and hospital patients indicating a possible spread of the resistance gene cluster via the food chain to humans.

Anti-Bacterial Agents↗

PubMed1997

Evaluation of the resolving power of three different DNA fingerprinting methods to discriminate among isolates of a natural Rhizobium meliloti population.

In a comparative study, the PCR-based RAPD and ERIC fingerprint methods were evaluated for their resolving power to discriminate among 21 isolates of a natural Rhizobium meliloti population. PCR fingerprint patterns were analysed by using an automated laser fluorescent (ALF) DNA sequencer, thus allowing the automated on-line storage of data. Results obtained were compared to a classification system using insertion sequence (IS) fingerprinting. Both PCR fingerprint methods were comparable in their ability to resolve differences amongst Rh. meliloti isolates. Grouping of strains on the basis of their RAPD as well as their ERIC fingerprints correlated with grouping of strains according to their IS fingerprints. Moreover, strains displaying identical PCR patterns could be further differentiated according to their IS fingerprints, thus allowing a detailed insight into phylogenetic relationship among strains. The automated evaluation of strain-specific fingerprint patterns has the potential to become a valuable tool for studies of bacterial population genetics. Moreover, the rapid identification of single strains, e.g. pathogens in epidemiological studies seems feasible.

Bacterial Typing Techniques↗

PubMed1983

Nucleotide sequence of Abelson murine leukemia virus genome: structural similarity of its transforming gene product to other onc gene products with tyrosine-specific kinase activity.

The nucleotide sequence of the proviral genome of Abelson murine leukemia virus (A-MuLV), an acute transforming virus of murine origin, has been determined. Like other transforming viruses, A-MuLV contains sequences derived from its helper virus, Moloney murine leukemia virus (M-MuLV), and a cell-derived protooncogene (abl) insertion sequence. By comparison of the A-MuLV sequence with that of M-MuLV, it was possible to precisely localize and define sequences contributed by the host cellular DNA. From the nucleotide sequence, we have predicted the amino acid sequence of p120gag-abl, the product of the A-MuLV gag-abl hybrid gene. The amino acid sequence of the putative abl gene, when compared with the sequences of other tyrosine-specific protein kinases (src, fes, fps, and yes), revealed significant homologies, indicating that all these functionally related transforming genes are derived from divergent members of the same protooncogene family. In addition to the gag-abl sequence, the proviral genome was found to contain an additional open reading frame that could code for an 18,000-dalton protein, whose role is at present undetermined.

Abelson murine leukemia virus↗

PubMed1983

Nucleotide sequence of the simian sarcoma virus genome: demonstration that its acquired cellular sequences encode the transforming gene product p28sis.

The complete nucleotide sequence of the proviral genome of simian sarcoma virus (SSV), an acute transforming retrovirus of primate origin, has been determined. Like other transforming viruses, SSV contains sequences derived from its helper virus, simian sarcoma-associated virus (SSAV), and a cell-derived (v-sis) insertion sequence. By comparison with the sequence of Moloney murine leukemia virus, it was possible to precisely localize and define sequences contributed by SSAV during the generation of SSV. Comparative sequence analysis of SSV and SSAV showed that SSAV provides regulatory sequences for initiation and termination of transcription of the SSV transforming gene. Moreover, coding sequences for the putative protein product of this gene appear to initiate from the amino terminus of the SSAV env gene. Antibodies to synthetic peptides derived from the carboxy and amino termini of the putative protein predicted by the open reading frame identified within v-sis specifically detect a Mr 28,000 protein, p28sis, in SSV-transformed cells. These and other findings confirm the predicted amino acid sequence of this protein and localize it to the coding region of the SSV transforming gene.

Amino Acid Sequence↗

PubMed2002

Tail-anchored and signal-anchored proteins utilize overlapping pathways during membrane insertion.

Tail-anchored proteins are a distinct class of membrane proteins that are characterized by a C-terminal membrane insertion sequence and a capacity for post-translational integration. Although it is now clear that tail-anchored proteins are inserted into the membrane at the endoplasmic reticulum (ER), the molecular basis for their integration is poorly understood. We have used a cross-linking approach to identify ER components that may be involved in the membrane insertion of tail-anchored proteins. We find that several newly synthesized tail-anchored proteins are transiently associated with a defined subset of cellular components. Among these, we identify several ER proteins, including subunits of the Sec61 translocon, Sec62p, Sec63p, and the 25-kDa subunit of the signal peptidase complex. When we analyze the cotranslational membrane insertion of a comparable signal-anchored protein we find the nascent polypeptide associated with a similar set of ER components. We conclude that the pathways for the integration of tail-anchored and signal-anchored membrane proteins at the ER exhibit a substantial degree of overlap, and we propose that this reflects similarities between co- and post-translational membrane insertion.

Cell Membrane↗

PubMed1994

Rhs elements of Escherichia coli: a family of genetic composites each encoding a large mosaic protein.

The Rhs family comprises a set of composite elements found in the chromosomes of many natural Escherichia coli strains. Five Rhs elements occur in strain K-12. The most prominent Rhs component is a giant core open reading frame (core ORF) whose features are suggestive of a cell surface ligand-binding protein. This hypothetical protein contains a peptide motif, xxGxxxRYxYDxxGRL(I or T)xxxx, that is repeated 28 times. A similar repeated motif is found in a Bacillus subtilis wall-associated protein. The Rhs core ORFs consist of two distinct parts: a large N-terminal core that is conserved in all Rhs elements, and a smaller C-terminus that is highly variable. Distinctive G+C contents of Rhs components indicate that the elements have a recent origin outside the E. coli species, and that they are composites assembled from segments with very different evolutionary histories. The Rhs cores fall into three sub-families that are mutually more than 20% divergent. Downstream of the core ORF is a second, much shorter ORF. Like the adjacent core extension, these are highly variable. In most examples, the hypothetical product of this ORF has a candidate signal sequence for transport across the cytoplasmic membrane. Another Rhs component, the 1.3 kb H-rpt, has features typical of insertion sequences. Structures homologous to H-rpt have been detected in other bacterial genera, such as Vibrio and Salmonella, where they are associated with loci that determine O-antigen variation.

Amino Acid Sequence↗

PubMed1996

Genomic complexity and plasticity of Burkholderia cepacia.

Burkholderia cepacia has attracted attention because of its extraordinary degradative abilities and its potential as a pathogen for plants and for humans. This bacterium was formerly considered to belong to the genus Pseudomonas in the gamma-subclass of the Proteobacteria, but recently has been assigned to the beta-subclass is based on rrn gene sequence analyses and other key phenotypic characteristics. The B. cepacia genome is comprised of multiple chromosomes and is rich in insertion sequences. These two features may have played a key role in the evolution of novel degradative functions and the unusual adaptability of this bacterium.

Biodegradation, Environmental↗

PubMed2002

Cloning, expression, and chromosomal localization of the mouse gene (Scgb3a1, alias Ugrp2) that encodes a member of the novel uteroglobin-related protein gene family.

The mouse UGRP gene family consists of two genes, Ugrp1 and Ugrp2. In this study, the genomic structure and expression patterns of Ugrp2 and its alternative spliced form were characterized. The authentic Ugrp2 gene has three exons and two introns, similar to the Ugrp1 gene, which produces a secreted protein. The Ugrp2 variant uses a sequence located between authentic exons 1 and 2, resulting in a cytoplasmic form due to a termination codon within the inserted sequence. Both mouse and human UGRP2 mRNAs are expressed in lung. In the case of human, the mRNA is expressed at the highest level in trachea, followed by salivary gland at a level similar to lung. Weak expression was also found in fetal lung and mammary gland. Ugrp2 was mapped by fluorescence in situ hybridization to mouse chromosome 11A5-B1 and human chromosome 5q35. These regions are known to be homologous. Interspecific mouse backcross mapping was also performed to obtain further detailed localization of mouse Ugrp1 and Ugrp2.

Alternative Splicing↗

PubMed1989

[Fast and slow genome rearrangements: molecular and genetic mechanisms of changes in virulence].

Different mechanisms of virulence changes at the molecular and populational levels are discussed. The methodology and mechanisms of Streptococcus pyogenes variability are analysed. The views of the adaptive role played by insertion sequences are developed, and the design of an epidemiologico-genetic experiment is proposed, which consists in obtaining highly virulent insertion mutants and testing their properties.

Enterobacteriaceae↗

PubMed1997

In vitro optimization of truncated stem-loop II variants of the hammerhead ribozyme for cleavage in low concentrations of magnesium under non-turnover conditions.

By truncating helix II to two base pairs in a hammerhead ribozyme having long flanking sequences (greater than 30 bases), the rate of cleavage in 1 mM magnesium can be increased roughly 100-fold. Replacing most of the nucleotides in a typical stem-loop II with 1-4 randomized nucleotides gave an RNA library that, even before selection, was more active in 1 mM magnesium than the parent ribozyme, but considerably less active than the truncated stem-loop II ribozyme. A novel, multiround selection for intermolecular cleavage was exploited to optimize this library for cleavage in low concentrations of magnesium. After three rounds of selection at sequentially lower concentrations of magnesium, the library cleaved substrate RNA 20-fold faster than the initial pool and was cloned. This pool was heavily enriched for one particular sequence (5'-CGUG-3') that represented 16 of 52 isolates (the next most common sequence was represented only six times). This sequence also represented the most active sequence, exceeding the activity of the short helix II variant under the conditions of the selection, thereby demonstrating the effectiveness of the selection technique. Analysis of the cleavage rates of RNAs made from eight isolates having different four-base insert sequences allowed assignment of highly preferred bases at each position in the insert. Analysis of pool clones having insert of differing lengths showed that, in general, activity decreased as the length of the insert decreased from 4 to 1. This supports the suggested role of stem-loop II in stabilizing the non-Watson-Crick interactions between the conserved bases of the catalytic core.

Base Sequence↗

PubMed1994

A 5' splice site mutation affecting the pre-mRNA splicing of two upstream exons in the collagen COL1A1 gene. Exon 8 skipping and altered definition of exon 7 generates truncated pro alpha 1(I) chains with a non-collagenous insertion destabilizing the triple helix.

A heterozygous de novo G to A point mutation in intron 8 at the +5 position of the splice donor site of the gene for the pro alpha 1(I) chain of type I procollagen, COL1A1, was defined in a patient with type IV osteogenesis imperfecta. The splice donor site mutation resulted not only in the skipping of the upstream exon 8 but also unexpectedly had the secondary effect of activating a cryptic splice site in the next upstream intron, intron 7, leading to re-definition of the 3' limit of exon 7. These pre-mRNA splicing aberrations cause the deletion of exon 8 sequences from the mature mRNA and the inclusion of 96 bp of intron 7 sequence. Since the mis-splicing of the mutant allele product resulted in the maintenance of the correct codon reading frame, the resultant pro alpha 1(I) chain contained a short non-collagenous 32-amino-acid sequence insertion within the repetitive Gly-Xaa-Yaa collagen sequence motif. At the protein level, the mutant alpha 1(I) chain was revealed by digestion with pepsin, which cleaved the mutant procollagen within the protease-sensitive non-collagenous insertion, producing a truncated alpha 1(I). This protease sensitivity demonstrated the structural distortion to the helical structure caused by the insertion. In long-term culture with ascorbic acid, which stimulates the formation of a mature crosslinked collagen matrix, and in tissues, there was no evidence of the mutant chain, suggesting that during matrix formation the mutant chain was unable to stably incorporated into the matrix and was degraded proteolytically.

Amino Acid Sequence↗

PubMed2003

Revised Escherichia coli selenocysteine insertion requirements determined by in vivo screening of combinatorial libraries of SECIS variants.

To investigate the stringency of the Escherichia coli selenocysteine insertion sequence (SECIS) requirements, libraries of SECIS variants were screened via a novel method in which suppression of the selenocysteine (Sec) opal codon was coupled to bacteriophage plaque formation. The SECIS variant libraries were designed with a mostly paired lower stem, so that randomization could be focused on the upper stem and loop regions. We identified 19 functional non-native SECIS sequences that violated the expected pairing requirements for the SECIS upper stem. All of the SECIS variants were shown to permit Sec insertion in phage (by chemical modification of the Sec residue) and fused to lacZalpha (by beta-galactosidase assay). The diminished pairing of the upper stem appears to be mitigated by the overall stem stability; a given upper stem variant has significantly higher readthrough in the context of a paired, rather than unpaired, lower stem. These results suggest an unexpected downstream sequence flexibility in prokaryotic selenoprotein expression.

Bacteriophages↗

PubMed1990

Construction and hormone regulation of a novel retroviral vector.

We report the analysis of a self-inactivating retroviral vector, constructed to allow inducible gene expression of inserted sequences from the mouse mammary tumour virus hormonal response element. The cloning strategy has been designed to allow for ease of insertion of the genes of interest. The vector contains the aph gene, allowing geneticin-resistance selection in mammalian cells. We have characterised dexamethasone (Dex)-induced increase in gene expression using the reporter gene encoding chloramphenicol acetyltransferase (CAT) inserted into the retroviral vector. We observe low basal levels of CAT activity in infected cells which is increased up to 50-fold by induction with Dex. The induction of pooled clones is 13.3-fold. Variation in Dex-induced CAT activity is observed in independently infected clones, which is not explained by proviral copy number.

Animals↗

PubMed2001

The complete genome of the crenarchaeon Sulfolobus solfataricus P2.

The genome of the crenarchaeon Sulfolobus solfataricus P2 contains 2,992,245 bp on a single chromosome and encodes 2,977 proteins and many RNAs. One-third of the encoded proteins have no detectable homologs in other sequenced genomes. Moreover, 40% appear to be archaeal-specific, and only 12% and 2.3% are shared exclusively with bacteria and eukarya, respectively. The genome shows a high level of plasticity with 200 diverse insertion sequence elements, many putative nonautonomous mobile elements, and evidence of integrase-mediated insertion events. There are also long clusters of regularly spaced tandem repeats. Different transfer systems are used for the uptake of inorganic and organic solutes, and a wealth of intracellular and extracellular proteases, sugar, and sulfur metabolizing enzymes are encoded, as well as enzymes of the central metabolic pathways and motility proteins. The major metabolic electron carrier is not NADH as in bacteria and eukarya but probably ferredoxin. The essential components required for DNA replication, DNA repair and recombination, the cell cycle, transcriptional initiation and translation, but not DNA folding, show a strong eukaryal character with many archaeal-specific features. The results illustrate major differences between crenarchaea and euryarchaea, especially for their DNA replication mechanism and cell cycle processes and their translational apparatus.

Cell Cycle Proteins↗

PubMed2006

Evidence for the acquisition of multi-drug resistance in an HIV-1 clinical isolate via human sequence transduction.

Insertions in HIV-1 reverse transcriptase's fingers subdomain can enhance chain terminator excision and confer resistance to multiple nucleoside analogs. Inserts that resemble flanking sequences likely arise by local sequence duplication. However, a remarkable variety of non-repeat fingers insertions have been observed. Here, molecular epidemiology, sequence analyses and mechanistic modeling were employed to show that one Japanese isolate's RT fingers insert likely resulted from non-homologous recombination between virus and host sequences and the transductive copying of 37 nucleotides from human chromosome 17. These findings provide evidence that human sequence transduction can, at least rarely, contribute to genetic and phenotypic variation in pandemic HIV.

Anti-HIV Agents↗