Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.1.30.1 (S1 nuclease)
3,660 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

amfC plays a regulatory role in aerial mycelium formation in Streptomyces griseus and is distributed widely among Streptomyces species. Disruption of the chromosomal amfC gene in Streptomyces coelicolor A3(2) severely reduced formation of aerial hyphae, indicating that amfC is important in morphological development. In addition, the disruption caused S. coelicolor A3(2) M130 to produce much less actinorhodin, and to produce undecylprodigiosin at a later stage of growth, indicating that amfC also regulates secondary metabolism. S1 nuclease mapping showed that transcription of actII-ORF4, the pathway-specific transcriptional activator in the act gene cluster, was greatly reduced in the amfC disruptants. The defect in secondary metabolite formation was suppressed or overcome by a mutation in sre-1. Consequently, an amfC-disrupted strain derived from S. coelicolor A3(2) M145, an actinorhodin-overproducing strain due to the sre-1 mutation, still produced a large amount of actinorhodin. Extra copies of amfC in strains M130 and M145 did not change spore-chain morphology or secondary metabolite formation. However, the spores in these strains remained white even after prolonged incubation. Since only spore pigmentation was affected, all known whi genes, except whiE, responsible for the polyketide spore pigment formation, were assumed to function normally. S1 nuclease mapping showed that transcription of whiEP1, one of the promoters in the whiE locus, was reduced in S. coelicolor A3(2) containing extra copies of amfC. Introducing amfC into several other Streptomyces species, such as Streptomyces lividans, Streptomyces lavendulae and Streptomyces lipmanii, also abolished spore pigment formation. An increase in the amount of AmfC appeared to disturb the maturation of spores.
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PMID:Involvement of amfC in physiological and morphological development in Streptomyces coelicolor A3(2). 1051 80

In Streptomyces griseus, A-factor (2-isocapryloyl-3R-hydroxymethyl-gamma-butyrolactone) at an extremely low concentration triggers streptomycin biosynthesis and cell differentiation by binding a repressor-type receptor protein (ArpA) and dissociating it from DNA. An A-factor-responsive transcriptional activator (AdpA) able to bind the promoter of strR, a pathway-specific regulatory gene responsible for transcription of other streptomycin biosynthetic genes, was purified to homogeneity and adpA was cloned by PCR on the basis of amino acid sequences of purified AdpA. adpA encoding a 405-amino-acid protein containing a helix-turn-helix DNA-binding motif at the central region showed sequence similarity to transcriptional regulators in the AraC/XylS family. The -35 and -10 regions of the adpA promoter were found to be a target of ArpA; ArpA bound the promoter region in the absence of A-factor and exogenous addition of A-factor to the DNA-ArpA complex immediately released ArpA from the DNA. Consistent with this, S1 nuclease mapping showed that adpA was transcribed only in the presence of A-factor and strR was transcribed only in the presence of intact adpA. Furthermore, adpA disruptants produced no streptomycin and overexpression of adpA caused the wild-type S. griseus strain to produce streptomycin at an earlier growth stage in a larger amount. On the basis of these findings, we propose here a model to demonstrate how A-factor triggers streptomycin biosynthesis at a late exponential growth stage.
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PMID:The A-factor regulatory cascade leading to streptomycin biosynthesis in Streptomyces griseus : identification of a target gene of the A-factor receptor. 1054 Feb 89

In Saccharomyces cerevisiae, the phospholipid biosynthetic genes are transcriptionally regulated in response to inositol and choline. This regulation requires the transcriptional activator proteins Ino4p and Ino2p, which form a heterodimer that binds to the UAS(INO) element. We have previously shown that the promoters of the INO4 and INO2 genes are among the weakest promoters characterized in yeast. Because little is known about the promoters of weakly expressed yeast genes, we report here the analysis of the constitutive INO4 promoter. Promoter deletion constructs scanning 1,000 bp upstream of the INO4 gene identified a small region (-58 to -46) that is absolutely required for expression. S1 nuclease mapping shows that this region contains the transcription start sites for the INO4 gene. An additional element (-114 to -86) modestly enhances INO4 promoter activity (fivefold). Thus, the region required for INO4 transcription is limited to 68 bp. These studies also found that INO4 gene expression is not autoregulated by Ino2p and Ino4p, despite the presence of a putative UAS(INO) element in the INO4 promoter. We further report that the INO4 steady-state transcript levels and Ino4p levels are regulated twofold in response to inositol and choline, suggesting a posttranscriptional mechanism of regulation.
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PMID:The promoter of the yeast INO4 regulatory gene: a model of the simplest yeast promoter. 1078 42

Recent research in various areas has appreciably expanded our knowledge of streptokinase, a plasminogen activator produced by all human group A (GAS), group C (GCS), and group G (GGS) streptococci. Several molecular genetic approaches are described here to study the expression of the streptokinase gene, skn. Southern hybridization analysis demonstrated homology of synteny of ska, skc, and skg in the genomes of the above serogroups. S1 nuclease mapping, the use of transcriptional fusions to beta-galactosidase and luciferase reporter genes, in conjunction with site-directed mutagenesis, led to the localization of the core promoter region of skc and the identification of a cis-active upstream region required for full promoter activity. Circular permutation analysis of the promoter upstream region identified an intrinsic DNA bending locus as the pivotal DNA element stimulating the activity of the core promoter. The detection of skn allele-specific expression phenotypes, which proved not to be due to different skn mRNA half-lives, prompted allele swap experiments, showing that promoter activity is dictated by the host genetic background, rather than the sequence of the regulatory region. These findings suggest the involvement in skn expression of an as yet unidentified transcriptional activator that contacts the bent DNA region. Transcription termination of skc is directed by a bidirectional terminator whose structural requirements for termination efficiency were determined with base substitution mutants fused to a chloramphenicol acetyl transferase reporter. Finally, mutagenic plasmids are described for insertion-duplication and allele replacement mutagenesis of the skn locus.
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PMID:Expression and regulation of the streptokinase gene. 1081 72

A-factor (2-isocapryloyl-3R-hydroxymethyl-gamma-butyrolactone) at an extremely low concentration triggers streptomycin production and aerial mycelium formation in Streptomyces griseus. A-factor induces the expression of an A-factor-dependent transcriptional activator, AdpA, essential for both morphological and physiological differentiation by binding to the A-factor receptor protein ArpA, which has bound and repressed the adpA promoter, and dissociating it from the promoter. Nine DNA fragments that were specifically recognized and bound by histidine-tagged AdpA were isolated by cycles of a gel mobility shift-PCR method. One of them was located in front of a gene encoding an extracytoplasmic function sigma factor belonging to a subgroup of the primary sigma(70) family. The cloned gene was named AdpA-dependent sigma factor gene (adsA), and the gene product was named sigma(AdsA). Transcription of adsA depended on A-factor and AdpA, since adsA was transcribed at a very low and constant level in an A-factor-deficient mutant strain or in an adpA-disrupted strain. Consistent with this, transcription of adsA was greatly enhanced at or near the timing of aerial hyphae formation, as determined by low-resolution S1 nuclease mapping. High-resolution S1 mapping determined the transcriptional start point 82 nucleotides upstream of the translational start codon. DNase I footprinting showed that AdpA bound both strands symmetrically between the transcriptional start point and the translational start codon; AdpA protected the antisense strand from positions +7 to +41 with respect to the transcriptional start point and the sense strand from positions +12 to +46. A weak palindrome was found in the AdpA-binding site. The unusual position bound by AdpA as a transcriptional activator, in relation to the promoter, suggested the presence of a mechanism by which AdpA activates transcription of adsA in some unknown way. Disruption of the chromosomal adsA gene resulted in loss of aerial hyphae formation but not streptomycin or yellow pigment production, indicating that sigma(AdsA) is involved only in morphological development and not in secondary metabolic function. The presence of a single copy in each of the Streptomyces species examined by Southern hybridization suggests a common role in morphogenesis in this genus.
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PMID:An A-factor-dependent extracytoplasmic function sigma factor (sigma(AdsA)) that is essential for morphological development in Streptomyces griseus. 1091 94

The platelet-derived growth factor (PDGF)-A promoter is regulated by a number of GC-rich regulatory elements that possess non-B-form DNA structures. Screening of a HeLa cDNA expression library with the C-rich strand of a PDGF-A silencer sequence (5'-S1 nuclease-hypersensitive site (SHS)) yielded three cDNA clones encoding NM23-H1, a protein implicated as a suppressor of metastasis in melanoma and breast carcinoma. Recombinant human NM23-H1 cleaved within the 3'-portions of both 5'-SHS strands in either single-stranded or duplex forms. In contrast, NM23-H2, known as a transcriptional activator with a DNA cleavage function, cleaved within the 5'-portions of both strands, revealing that NM23-H1 and NM23-H2 cleave at distinct sites of the 5'-SHS and by different mechanisms. NM23-H1 and NM23-H2 also cleaved within the PDGF-A basal promoter region, again exhibiting preferences for cleavage within the 5'- and 3'-portions of the element, respectively. Transient transfection analyses in HepG2 cells revealed that both NM23-H1 and -H2 repressed transcriptional activity driven by the PDGF-A basal promoter (-82 to +8). Activity of the negative regulatory region (-1853 to -883), which contains the 5'-SHS, was also inhibited modestly by NM23-H1 and NM23-H2. These studies demonstrate for the first time that NM23-H1 interacts both structurally and functionally with DNA. They also indicate a role for NM23 proteins in repressing transcription of a growth factor oncogene, providing a possible molecular mechanism to explain their metastasis-suppressing effects.
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PMID:NM23-H1 and NM23-H2 repress transcriptional activities of nuclease-hypersensitive elements in the platelet-derived growth factor-A promoter. 1169 15

AfsR is a pleiotropic, global regulator that controls the production of actinorhodin, undecylprodigiosin and calcium-dependent antibiotic in Streptomyces coelicolor A3(2). AfsR, with 993 amino acids, is phosphorylated on serine and threonine residues by a protein serine/threonine kinase AfsK and contains an OmpR-like DNA-binding fold at its N-terminal portion and A- and B-type nucleotide-binding motifs in the middle of the protein. The DNA-binding domain, in-dependently of the nucleotide-binding domain, contributed the binding of AfsR to the upstream region of afsS that locates immediately 3' to afsR and encodes a 63-amino-acid protein. No transcription of afsS in the DeltaafsR background and restoration of afsS transcription by afsR on a plasmid in the same genetic background indicated that afsR served as a transcriptional activator for afsS. Interestingly, the AfsR binding site overlapped the promoter of afsS, as determined by DNase I protection assay and high-resolution S1 nuclease mapping. The nucleotide-binding domain contributed distinct ATPase and GTPase activity. The phosphorylation of AfsR by AfsK greatly enhanced the DNA-binding activity and modulated the ATPase activity. The DNA-binding ability of AfsR was independent of the ATPase activity. However, the ATPase activity was essential for transcriptional activation of afsS, probably because the energy available from ATP hydrolysis is required for the isomerization of the closed complex between AfsR and RNA polymerase to a transcriptionally competent open complex. Thus, AfsR turns out to be a unique transcriptional factor, in that it is modular, in which DNA-binding and ATPase activities are physically separable, and the two functions are modulated by phosphorylation on serine and threonine residues.
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PMID:afsS is a target of AfsR, a transcriptional factor with ATPase activity that globally controls secondary metabolism in Streptomyces coelicolor A3(2). 1195 95

A multiple-gene locus for polyamine uptake and utilization was discovered in Pseudomonas aeruginosa PAO1. This locus contained nine genes designated spuABCDEFGHI (spu for spermidine and putrescine utilization). The physiological functions of the spu genes in utilization of two polyamines (putrescine and spermidine) were analyzed by using Tn5 transposon-mediated spu knockout mutants. Growth and uptake experiments support that the spuDEFGH genes specify components of a major ABC-type transport system for spermidine uptake, and enzymatic measurements indicated that spuC encodes putrescine aminotransferase with pyruvate as the amino group receptor. Although spuA and spuB mutants showed an apparent defect in spermidine utilization, the biochemical functions of the gene products have yet to be elucidated. Assays of lacZ fusions demonstrated the presence of agmatine-, putrescine-, and spermidine-inducible promoters for the spuABCDEFGH operon and the divergently transcribed spuI gene of unknown function. Since the observed induction effect of agmatine was abolished in an aguA mutant where conversion of agmatine into putrescine was blocked, putrescine or spermidine, but not agmatine, serves as the inducer molecule of the spuA-spuI divergent promoters. S1 nuclease mappings confirmed further the induction effects of the polyamines on transcription of the divergent promoters and localized the transcription initiation sites. Gel retardation assays with extracts from the cells grown on putrescine or spermidine demonstrated the presence of a polyamine-responsive regulatory protein interacting with the divergent promoter region. Finally, the absence of the putrescine-inducible spuA expression and putrescine aminotransferase (spuC) formation in the cbrB mutant indicated that the spu operons are regulated by the global CbrAB two-component system perhaps via the putative polyamine-responsive transcriptional activator.
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PMID:Functional analysis and regulation of the divergent spuABCDEFGH-spuI operons for polyamine uptake and utilization in Pseudomonas aeruginosa PAO1. 1208 45

Nitrogen fixation in Azotobacter vinelandii is regulated by the nifLA operon. NifA activates the transcription of nif genes, while NifL antagonizes the transcriptional activator NifA in response to fixed nitrogen and molecular oxygen levels. However, transcriptional regulation of the nifLA operon of A. vinelandii itself is not fully understood. Using the S1 nuclease assay, we mapped the transcription start site of the nifLA operon, showing it to be similar to the sigma54-dependent promoters. We also identified a positive cis-acting regulatory element (+134 to +790) of the nifLA operon within the coding region of the nifL gene of A. vinelandii. Deletion of this element results in complete loss of promoter activity. Several protein factors bind to this region, and the specific binding sites have been mapped by DNase I foot printing. Two of these sites, namely dR1 (+134 to +204) and dR2 (+745 to +765), are involved in regulating the nifLA promoter activity. The absence of NtrC-like binding sites in the upstream region of the nifLA operon in A. vinelandii makes the identification of these downstream elements a highly significant finding. The interaction of the promoter with the proteins binding to the dR2 region spanning +745 to +765 appears to be dependent on the face of the helix as introduction of 4 bases just before this region completely disrupts promoter activity. Thus, the positive regulatory element present within the BglII-BglII fragment may play, in part; an important role in nifLA regulation in A. vinelandii.
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PMID:Identification of a positive transcription regulatory element within the coding region of the nifLA operon in Azotobacter vinelandii. 1600 Jul 81

AtrA, a transcriptional activator for actII-ORF4, encoding the pathway-specific transcriptional activator of the actinorhodin biosynthetic gene cluster in Streptomyces coelicolor A3(2), has been shown to bind the region upstream from the promoter of strR, encoding the pathway-specific transcriptional activator of the streptomycin biosynthetic gene cluster in Streptomyces griseus [Uguru et al. (2005) Mol Microbiol 58, 131-150]. The atrA orthologue (atrA-g) in S. griseus was constitutively transcribed throughout growth from a promoter located about 250 nt upstream of the translational start codon, as determined by S1 nuclease mapping. DNase I footprinting showed that histidine-tagged AtrA-g bound an inverted repeat located upstream of strR at positions -117 to -142 relative to the transcriptional start point of strR as +1. This AtrA-g-binding site was between two AdpA-binding sites at approximately nucleotide positions -270 and -50. AdpA is a central transcriptional activator in the A-factor regulatory cascade and essential for the transcription of strR. AtrA-g and AdpA simultaneously bound the respective binding sites. In contrast to AdpA, AtrA-g was non-essential for strR transcription; an atrA-g-disrupted strain produced streptomycin on routine agar media to the same extent as the wild-type strain. However, the atrA-g-disrupted strain tended to produce a smaller amount of streptomycin than the wild-type strain under some conditions, for example, on Bennett agar containing 1 % maltose and on a minimal medium. Therefore, AtrA-g had a conditionally positive effect on streptomycin production, as a tuner, probably by enhancing the AdpA-dependent transcriptional activation of strR in a still unknown manner.
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PMID:Conditionally positive effect of the TetR-family transcriptional regulator AtrA on streptomycin production by Streptomyces griseus. 1831 36


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