Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.30.2 (endonuclease)
 18,621 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Previous studies have demonstrated that the mouse c-Harvey ras proto-oncogene (c-Ha-ras) promoter sequences are GC rich and contain several potential transcription factor SP1 binding sites. We investigated the endonuclease hypersensitivity of this region in nuclei in vitro and whole mouse tissues in vivo and identified a very strong, ubiquitous hypersensitive site covering the proximal promoter sequences. Footprint protection studies using nuclear extracts from various cell types including fibroblasts, erythroid cells, and both normal and transformed epithelial cells revealed a consistent protein-binding pattern. Five protein binding sites were observed, four of which correlated with potential SP1 binding sites. Competition experiments using an oligonucleotide corresponding to a consensus SP1 binding site confirmed that these sequences were indeed bound by the SP1 (or SP1-like) trans-acting factor. In addition, no differences were observed between the footprint patterns obtained using extracts from cells of different lineages or between normal and transformed epithelial cells carrying activated ras genes. The controlling elements responsible for differential c-Ha-ras transcription between cell types or at different stages of carcinogenesis therefore probably lie in other regions of the gene.
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PMID:Structural analysis of the mouse c-Ha-ras gene promoter. 204 51

To understand the basis of osteonectin (SPARC) transcriptional regulation, we have isolated a bovine genomic clone (lambda Og15) encoding exon 1 and 15 kilobase pairs (kb) of flanking DNA. Direct RNA sequencing of the 5' end of the osteonectin message showed it contained a sequence identical to that of a 2.4-kb EcoRI-BamHI fragment located midway in the clone lambda Og15. The results indicate exon 1 is located 10 kb away from exon 2 in the bovine genome. The DNA sequence unit CCTG is repeated five times in exon 1 which is composed exclusively of untranslated sequence. Sequence analysis of the 5'-flanking DNA revealed the presence of many regulatory motifs including a "GC" box with four overlapping SP1 consensus sequences. Immediately downstream from the GC box is a 72-base pair purine-rich stretch composed primarily of direct repeats of the sequence motifs GGGGA and GGA (GAGA box). Digestion of the flanking DNA in vitro with S1 endonuclease showed a site for the enzyme at position -55 which is just 3' to the GAGA box. Chimeric chloramphenicol acetyltransferase constructs were prepared containing the S1-sensitive site and showed substantial transcriptional activity in UMR-106 and fetal and adult human bone cells which are known to be high producers of the protein. The results indicate a potential regulatory activity of the S1 site in osteonectin gene activation.
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PMID:Osteonectin promoter. DNA sequence analysis and S1 endonuclease site potentially associated with transcriptional control in bone cells. 253 44

We have investigated protein-DNA interactions in the proximal promoter of the human amyloid precursor protein (APP) gene in temporal lobe neocortical nuclei isolated from control and Alzheimer disease (AD) affected brains. We report that the human APP 5' promoter sequence from -203 to +55 bp, which has been previously reported to contain essential regulatory elements for APP gene transcription, lies in a deoxyribonuclease I, micrococcal nuclease- and restriction endonuclease-sensitive, G+C-rich nucleosome-free gap flanked both 5' and 3' by typical nucleosome structures. As analyzed by electrophoretic mobility shift assay, this extended internucleosomal linker DNA is heavily occupied by nuclear protein factors, and interacts differentially with nuclear protein extracts obtained from HeLa and human brain neocortical nuclei. This suggests that the chromatin conformation of the APP gene promoter may vary in different cell types, and may correlate with differences in APP gene expression. Human recombinant transcription factors AP1, SP1 and TFIID (but not AP2 or brain histones H1, H2B and H4) interact with the -203 to +55 bp of the human APP promoter sequence. Only minor differences were observed in the chromatin structure of the immediate APP promoter between non-AD and AD affected neocortical nuclei, suggesting either that post-transcriptional processes, or that regulatory elements lying elsewhere in the APP gene may be important in the aberrant accumulation of the APP gene product.
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PMID:Protein-DNA interactions in the promoter region of the amyloid precursor protein (APP) gene in human neocortex. 801 72

To define the mechanisms of 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced transcription of the ornithine decarboxylase (ODC) gene, we isolated a genomic clone (hODC41B) of ODC from a human leukocyte genomic DNA library. The restriction endonuclease map, in comparison with the previously published sequences of the human ODC gene, indicated that hODC41B contained a 15.7-kb sequence that extended from the sixth exon to about 10 kb upstream of the ODC gene. A 2.5-kb genomic fragment containing the 5' flanking region and the first exon was subcloned and sequenced. Sequence analysis revealed multiple putative promoter/enhancer elements (a TATA box, a CAAT box, 17 GC boxes, and a cAMP-responsive element) but no consensus AP-1 sequences (TGAGTCA) in the 2.5-kb 5' flanking region. However, three AP-1 sequences were located in introns 3, 5, and 11. We constructed a series of chimeric genes containing part of the first exon and increasingly longer 5' flanking sequences of the ODC gene fused to either bacterial chloramphenicol acetyltransferase (CAT) or luciferase reporter genes. TPA inducibility was determined by transient transfection and measurement of CAT or luciferase expression in HeLa cells. The induction of CAT activity by TPA decreased with decreasing lengths of the 5' flanking sequences up to nt -82. The TPA induction from the construct -72 ODC CAT was threefold to sevenfold, and the TPA inducibility of the same fragment was about ninefold to 30-fold with the luciferase reporter gene. Further deletion analysis revealed TPA-responsive sequences in ODC nt -42 to +54. Gel mobility shift assays using alpha-32P-end labeled ODC nt -42 to +60 revealed that nt -42 to +60 specifically bound HeLa cell nuclear proteins. HeLa cell nuclear protein binding to ODC nt -42 to +60 could not be completely competed by AP-1-, AP-2-, AP-3-, or SP1-responsive sequences.
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PMID:Non-AP-1 tumor promoter 12-O-tetradecanoylphorbol-13-acetate-responsive sequences in the human ornithine decarboxylase gene. 804 98

To identify the SV40 regulatory sequences responsible for the chromatin remodeling associated with early transcription, SV40 chromosomes containing potential remodeling sequences inserted adjacent to a reporter region were isolated at various times within the first 6 h of infection and analyzed by a combination of restriction endonuclease digestion and competitive PCR amplification. The sequences analyzed included the early domain, the enhancer, the late domain, the early phasing element, the AP-1 element, two tandem copies of the SP1 element, and the AP-4 element. From 30 min to 3 h postinfection only the enhancer, the AP-1 element, and the two tandem copies of the SP1 element caused a change in nuclease sensitivity consistent with chromatin remodeling. These results suggest that the changes in chromatin structure seen in the promoter during activation of early transcription are most likely a result of remodeling by the AP-1 and/or SP1.
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PMID:SP1 and AP-1 elements direct chromatin remodeling in SV40 chromosomes during the first 6 hours of infection. 1188 75

Quality control during mass production of Spodoptera exigua multiple nucleopolyhedrovirus (SeMNPV), and studies on environmental fate following the use of this virus as a biological pesticide, would be facilitated by a rapid method for the detection and identification of isolates. A molecular biology tool was developed that combined the polymerase chain reaction and restriction fragment length polymorphism (PCR-RFLP) to differentiate SeMNPV isolates. Oligonucleotide primers were designed to amplify five variable SeMNPV genomic regions (V01, V02, V03, V04, V05). Four wild-type SeMNPV strains isolated from the United States (US2) and Spain (SP1, SP2, and SP3), and a laboratory cloned genotype (US1A), were analyzed with 36 different primer-endonuclease combinations. BglII digestion of the variable region 1 (V01) amplicon was the only combination that differentiated each of the five virus isolates tested, although genetic heterogeneity limited the discriminatory power of the technique. Six novel SeMNPV isolates originating from greenhouse soils in southern Spain were successfully identified using this method. As judged by sequence analysis, the V01 region, which comprises the homologous region 1 (hr1), is the most variable genomic region among the genotypes present in the Spanish isolates. This method constitutes a useful tool for processing large number of environmental samples and could be used to address environmental biosafety concerns.
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PMID:Application of the PCR-RFLP method for the rapid differentiation of Spodoptera exigua nucleopolyhedrovirus genotypes. 1649 53