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At least 271 records · Page 15Linked to original sources
PubMed2002

Identification of a variant antioxidant response element in the promoter of the human glutamate-cysteine ligase modifier subunit gene. Revision of the ARE consensus sequence.

Constitutive and inducible expression of the gene encoding the modulator subunit of human glutamate-cysteine ligase (GCLM) is regulated by either of two regions of the promoter; an antioxidant response element (ARE) at -302:-291 and a 44-bp fragment (-346:-303) upstream of the ARE. This second region includes a consensus AP-1 site previously considered responsible for the enhancer activity of the upstream fragment. Deletion of a 165-bp fragment (-348:-183) including the ARE and upstream 44-bp fragment totally ablated t-butyl hydroquinone (tBHQ) inducibility of a GCLM promoter/luciferase transgene. Mutation analyses confirmed that both the ARE and the -346:-303 fragment could support induction following tBHQ exposure but demonstrated that induction in the latter case did not involve the AP-1 site at -341:-335. A region sharing significant homology with the consensus ARE sequence except for a single nucleotide mismatch at -330 (5'-TTACnnnGCA-3' versus 5'-TGACnnnGCA-3') was identified at the 5'-end of the 44-bp fragment immediately adjacent to the AP-1 site. A G in this position has been considered an invariant requirement of functional ARE sequences. Mutation of T(-330) to A (a substitution known to ablate ARE function) or C eliminated basal and inducible expression. Substitution of a G at -330 enhanced basal expression relative to the wild-type sequence, but induction following tBHQ exposure was comparable, indicating that either sequence (5'-TTACnnnGCA-3' versus 5'-TGACnnnGCA-3') may function as an ARE, although the former sequence is less effective at directing basal expression. This possibility was confirmed by similar mutational analyses of the core sequence of hNQO1, a prototypic ARE. Electrophoretic mobility shift competition assays revealed that the 5'-TTACnnnGCA-3' sequence could compete with the hNQO1 ARE for protein binding but was less effective than a similar probe containing the 5'-TGACnnnGCA-3' motif. Probes including the T(-330)A or T(-330)C mutations were ineffective. These results reveal that the GCLM promoter includes two functional AREs, one having a variant sequence. The results indicate that the consensus ARE sequence should be revised to 5'-RTKAYnnnGCR-3'.

Amino Acid Sequence↗
PubMed2001

Expression of nuclear transcription factor interferon consensus sequence binding protein in chronic myeloid leukemia correlates with pretreatment risk features and cytogenetic response to interferon-alpha.

Recently, it was shown that interferon consensus sequence binding protein (ICSBP), a member of the interferon regulatory factor (IRF) family, has a potential role in chronic myeloid leukemia (CML). Deletion of ICSBP gene in mice leads to a CML-like syndrome and samples from CML patients exhibited impaired ICSBP expression. The present study found that ICSBP expression correlated with risk features determined by Sokal score in untreated CML (P =.007 for high versus low risk). In addition, analyzing ICSBP expression during interferon-alpha (IFN-alpha) therapy in "good" (n = 27) versus "poor" (n = 15) cytogenetic responders, high ICSBP levels were only observed in "good" responders (P =.0002). Together, these data suggest that ICSBP levels are related to initial presentation of CML and the therapeutic response of CML to IFN-alpha, indicating an important role of ICSBP in CML. (Blood. 2001;97:3648-3650)

Cytogenetic Analysis↗
PubMed1995

Enterobacterial repetitive intergenic consensus sequences and the PCR to generate fingerprints of genomic DNAs from Vibrio cholerae O1, O139, and non-O1 strains.

Enterobacterial repetitive intergenic consensus (ERIC) sequence polymorphism was studied in Vibrio Cholerae strains isolated before and after the cholera epidemic in Brazil (in 1991), along with epidemic strains from Peru, Mexico, and India, by PCR. A total of 17 fingerprint patterns (FPs) were detected in the V. cholerae strains examined; 96.7% of the toxigenic V. cholerae O1 strains and 100% of the O139 serogroup strains were found to belong to the same FP group comprising four fragments (FP1). The nontoxigenic V. cholerae O1 also yielded four fragments but constituted a different FP group (FP2). A total of 15 different patterns were observed among the V. cholerae non-O1 strains. Two patterns were observed most frequently for V. cholerae non-01 strains, 25% of which have FP3, with five fragments, and 16.7% of which have FP4, with two fragments. Three fragments, 1.75, 0.79, and 0.5 kb, were found to be common to both toxigenic and nontoxigenic V. cholerae O1 strains as well as to group FP3, containing V. cholerae non-O1 strains. Two fragments of group FP3, 1.3 and 1.0 kb, were present in FP1 and FP2 respectively. The 0.5-kb fragment was common to all strains and serogroups of V. cholerae analyzed. It is concluded from the results of this study, based on DNA FPs of environmental isolates, that it is possible to detect an emerging virulent strain in a cholera-endemic region. ERIC-PCR constitutes a powerful tool for determination of the virulence potential of V. cholerae O1 strains isolated in surveillance programs and for molecular epidemiological investigations.

Bacterial Toxins↗
PubMed1999

Interferon consensus sequence binding protein and interferon regulatory factor-4/Pip form a complex that represses the expression of the interferon-stimulated gene-15 in macrophages.

Interferon consensus sequence binding protein (ICSBP), a transcription factor of the interferon (IFN) regulatory factor (IRF) family, binds to the IFN-stimulated response element (ISRE) in the regulatory region of IFNs and IFN-stimulated genes (ISG). To identify target genes, which are deregulated by an ICSBP null-mutation in mice (ICSBP-/-), we have analyzed transcription of an ISRE-bearing gene, ISG15. We have found that although ISG15 expression is unchanged in B cells, it is upregulated in macrophages from ICSBP-/- mice. Three factors, ICSBP, IRF-2, and IRF-4/Pip interact with the ISRE in B cells, however only ICSBP and IRF-4/Pip were found to bind this sequence in macrophages of wild-type mice. Although IRF-4 was considered to be a lymphoid-specific factor, we provide evidence for its role in macrophage gene regulation. Our results suggest that the formation of cell-type-specific heteromeric complexes between individual IRFs plays a crucial role in regulating IFN responses.

Animals↗
PubMed1996

Enterobacterial intergenic consensus sequence polymerase chain reaction as a typing method for Burkholderia (Pseudomonas) cepacia.

OBJECTIVE: We developed a rapid polymerase chain reaction (PCR) method, which utilizes primers from an enterobacterial repetitive consensus sequence, to differentiate among strains of Burkholderia cepacia. We then applied this method to isolates from a device-associated pseudoinfection and also compared the discriminative typing potential with that of another PCR-based method. METHODS: A simplified lysis procedure was used to generate DNA substrate for amplification by a PCR method which utilized primer ERIC2 and a reverse complement of ERIC1R. The alternative PCR-based method employed primers which are directed to Bordetella pertussis repetitive sequences. Amplification products were visually assessed after conventional agarose gel electrophoresis and staining. RESULTS: The ERIC-based method appropriately clustered strains for the device-associated pseudo-infection. In addition, there was consistency in the definition of similar or dissimilar strains between the two PCR---based systems. CONCLUSIONS: The ERIC-based PCR typing method offers sufficient discriminatory power for it to be used for epidemiologic purposes.

Journal Article↗
PubMed2003

Interferon consensus sequence binding protein (ICSBP; IRF-8) antagonizes BCR/ABL and down-regulates bcl-2.

BCR/ABL is the causative genetic aberration in chronic myelogenous leukemia (CML). Mice lacking expression of the interferon (IFN) consensus sequence binding protein (ICSBP), an IFN gamma-inducible transcription factor of the interferon regulatory factor (IRF) family, develop a disease similar to human CML. Mounting evidence suggests a role for ICSBP in the pathogenesis of CML. However, the underlying mechanisms are largely unknown. By stable and conditional expression of ICSBP in wild-type and BCR/ABL-transformed 32D cells (32D/wt and 32D/BA), we found that ICSBP inhibited BCR/ABL-mediated leukemogenesis in vivo. Moreover, ICSBP also overrode BCR/ABL-mediated morphology changes, chemotherapy, and imatinib resistance, as well as BCR/ABL-induced repression of differentiation. Some of these ICSBP effects may be explained in part by an ICSBP-mediated repression of bcl-2, a major antiapoptotic target of BCR/ABL, on transcriptional and protein level. Using reporter gene assays and electrophoretic mobility shift assays we identified that the bcl-2 promoter activity was inhibited by ICSBP by way of a fragment containing 2 characteristic ICSBP-responsive elements. An inverse correlation between ICSBP and bcl-2 expression was confirmed in vivo. Collectively, our findings suggest that ICSBP antagonizes BCR/ABL by down-regulation of bcl-2 and implicates a central role for ICSBP in the pathogenesis of CML, as well as a therapeutic target to overcome drug resistance in bcl-2-dependent tumors.

Animals↗
PubMed1995

Structural analysis of the N- and C-termini in a peptide with consensus sequence.

We present a structural analysis of a peptide, the sequence of which includes amino acids that show preferences for specific positions near the N- and C-termini in protein helices. This peptide has the sequence ac-YMSEDELKAAEAAFKRHGVP-amide, which includes a strong version of an N-terminal Harper-Rose capping box structure as well as a Gly located close to the C-terminus designed to elucidate its role in C-terminal capping. The sequence of five residues at the middle is inserted to separate effects at the two ends via a helix-stabilizing linker. Application of a simulated annealing procedure using interproton distance constraints derived from 1H NOESY experiments in water reveals the presence of a C-terminal structure in this model. The C-terminus forms a folded back structure in a significant fraction of structures generated by the annealing, in most of which Gly assumes an alpha L conformation. This structure occurs within a highly flexible region of the molecule and hence is occupied only a fraction of the time.

Amino Acid Sequence↗
PubMed1999

The complete consensus sequence of coxsackievirus B6 and generation of infectious clones by long RT-PCR.

The full length sequence for the human pathogen coxsackievirus B6 (CVB6, Schmitt strain) has been determined. We used long RT-PCR to generate full length DNA amplicon of CVB6, and then directly sequenced the amplicons. One-step cloning of the full length amplicon enabled us to obtain an infectious clone of CVB6. RNA generated from CVB6 amplicon DNA or CVB6 clones, by transcription with T7 RNA polymerase, was demonstrated to be infectious upon transfection into HeLa cells in vitro. The CVB6 genome is characteristic of enteroviruses, with a 5'-non-translated region (743 nucleotides) followed by an open reading frame (encoding a 2184 amino acid polyprotein) and a 3'-non-translated region (100 nucleotides) and polyadenylated tail. The predicted amino acid sequence of CVB6 clustered with the other CVB serotypes and swine vesicular disease virus (SVDV).

3' Untranslated Regions↗
PubMed1987

A new natural hGH variant--17.5 kd--produced by alternative splicing. An additional consensus sequence which might play a role in branchpoint selection.

From a human pituitary cDNA library, we have cloned 3 distinct human growth hormone (hGH) cDNAs, coding respectively for the 22 K hGH, the 20 K variant, and a yet unknown 17.5 K variant. S1 mapping analysis using human pituitary RNA confirms the existence of at least four distinct hGH mRNAs originating from alternative acceptor sites at the second intron of the primary transcript. We have analysed the hGH gene sequence to explain the high frequency of alternative splicings which occur only at this location. In this study we propose CTTGNNPyPyPy as an additional consensus sequence guiding the selection of the branched nucleotide.

Base Sequence↗
PubMed1994

The leader peptide of colicin V shares consensus sequences with leader peptides that are common among peptide bacteriocins produced by gram-positive bacteria.

Colicin V is a ribosomally synthesized antimicrobial peptide produced by Escherichia coli. Four recently characterized genes, arranged in two convergent operons on the plasmid pCoIV-K30, are required for colicin V synthesis, export and immunity. We report the purification and N-terminal amino acid sequencing of the colicin V protein. Our results demonstrate that the colicin V primary translation product, which consists of 103 amino acids, is proteolytically processed. A leader peptide, consisting of 15 amino acid residues, is removed from the N-terminus during maturation of colicin V. This leader peptide is not related to the N-terminal signal sequences which direct proteins across the cytoplasmic membrane via the Sec pathway. The molecular mass of colicin V, obtained by mass spectrometry analysis, showed that the peptide consists of only unmodified amino acids. The deduced amino acid sequence of the leader peptide was highly homologous to the N-terminal extensions found in non-lantibiotic, peptide bacteriocins produced by Gram-positive bacteria. These findings strongly indicate that colicin V belongs to a family of small peptide bacteriocins that have been found previously only among the Gram-positive lactic acid bacteria.

Amino Acid Sequence↗
PubMed1993

Rotavirus protein NSP3 (NS34) is bound to the 3' end consensus sequence of viral mRNAs in infected cells.

Interaction between viral proteins and RNAs has been studied in rotavirus-infected cells. The use of UV cross-linking followed by immunoprecipitation and labeling with T4 polynucleotide kinase allowed us to detect interactions between RNA and nonstructural viral proteins. The RNAs linked to the nonstructural protein NSP3 have been identified as rotavirus mRNAs, and the sequences of the RNase T1-protected fragments have been established. These sequences correspond to the 3' end sequence common to all rotavirus group A genes. We also show that the last 3' nucleotide is cross-linked to the protein and that monomeric and multimeric forms of NSP3 are bound to rotavirus mRNA. The role of NSP3 in rotavirus replication is discussed in the light of our results and by comparison with other RNA-binding proteins of members of the Reoviridae family.

Base Sequence↗
PubMed1992

The use of consensus sequence for amplification and oriented cloning of human alpha interferon genes.

A clone from human cDNA library was amplified in polymerase chain reaction (PCR) by the synthetic oligonucleotides. The final construct after linker ligation and digestion with restriction endonucleases was suitable for oriented cloning into E. coli expression vector. The interferon (IFN) expression could be detected by SDS-PAGE electrophoresis. Western blotting and biological antiviral assay. This set of oligonucleotides can be used also for amplification of genomic DNA or cDNA libraries.

Base Sequence↗
PubMed1990

A 3' splice site consensus sequence mutation in the cystic fibrosis gene.

In the cystic fibrosis (CF) gene, recently cloned, a three base pair deletion (delta F508) has been identified in a majority of CF patients. This deletion has been found in 80% of CF chromosomes in families from north west Brittany. In order to identify new mutations we have selected 43 chromosomes negative for the three base pair deletion from these families and directly sequenced exon 11 after DNA amplification by the polymerase chain reaction. We have detected a base change (G----A) at the 3' end of the consensus sequence of intron ten (namely 1717-1). This mutation destroys a splice site in the cystic fibrosis gene which probably produces a mutant allele. This single nucleotide mutation has been reported on two other CF chromosomes.

Cystic Fibrosis↗
PubMed2004

Toll-like receptor 9 signaling activates NF-kappaB through IFN regulatory factor-8/IFN consensus sequence binding protein in dendritic cells.

Unmethylated CpG DNA binds to the Toll-like receptor 9 (TLR9) and activates NF-kappaB to induce cytokine genes in dendritic cells (DCs). IFN regulatory factor (IRF)-8/IFN consensus sequence binding protein is a transcription factor important for development and activation of DCs. We found that DCs from IRF-8(-/-) mice were unresponsive to CpG and failed to induce TNF-alpha and IL-6, targets of NF-kappaB. Revealing a signaling defect selective for CpG, these cytokines were robustly induced in IRF-8(-/-) DCs in response to LPS that signals through TLR4. IRF-8(-/-) DCs expressed TLR9, adaptor myeloid differentiation factor 88, and other signaling molecules, but CpG failed to activate NF-kappaB in -/- cells. This was due to the selective inability of -/- DCs to activate I-kappaB kinase alphabeta, the kinases required for NF-kappaB in response to CpG. IRF-8 reintroduction fully restored CpG activation of NF-kappaB and cytokine induction in -/- DCs. Together, TLR signals that activate NF-kappaB are diverse among different TLRs, and TLR9 signaling uniquely depends on IRF-8 in DCs.

Dendritic Cells↗
PubMed1989

35 kDa fragment of h-caldesmon conserves two consensus sequences of the tropomyosin-binding domain in troponin T.

Using a tropomyosin-coupled affinity column, we have demonstrated a direct association between the chymotryptic 35 kDa fragment of h-caldesmon, which is located at the C-terminal of the parent molecule, and gizzard tropomyosin. We have subsequently determined the nucleotide sequence of cDNA clones encoding the 35 kDa fragment from the cDNA library prepared from chick embryo gizzards, and have deduced the amino acid sequence. Calculating from the predicted sequence, the 35 kDa fragment is composed of 306 amino acid residues. In agreement with the tropomyosin-binding ability, the 35 kDa fragment conserves two consensus sequences of the tropomyosin-binding domain in troponin T. These results suggest that the 35 kDa fragment of h-caldesmon, at least in part, has a common property to the striated muscle troponin T.

Amino Acid Sequence↗