Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.1.30.1 (S1 nuclease)
 3,660 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We previously isolated cDNA clones coding for the entire murine laminin B2 chain. In this report, we have screened a mouse genomic library with a 5' portion of the laminin B2 chain cDNA and isolated two genomic clones which contain the first and second exons. The transcription initiation site was determined by primer extension and S1 nuclease mapping. Exon 1 contained 641 bp (base pairs) including 229 bp of 5'-untranslated segment, sequences coding for the signal peptide and the N-terminal portion of the protein, while exon 2 contained 305 bp. Nucleotide sequencing of 830 bp of the 5'-flanking region of the gene showed several interesting features including the presence of 9 "GC" boxes, a stretch of 9 nearly identical repeats of 11 nucleotides between -200 and -450, and a sequence which is similar to the cAMP consensus sequence. There was no TATA box or CAAT box. A recombinant plasmid containing the 830 bp promoter segment coupled to the chloramphenicol acetyltransferase gene was constructed and transfected into various cells. Differentiated F9 cells transfected with this construct showed twice as much chloramphenicol acetyltransferase activity as the undifferentiated cells. The 830-bp B2 laminin promoter was also active in NIH-3T3 cells which produce little laminin, but was not active in human HT-1080 cells. These results indicate that this structurally unique promoter contains DNA sequences that help regulate the gene during differentiation.
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PMID:The laminin B2 chain promoter contains unique repeat sequences and is active in transient transfection. 283 21

A gene-specific transcription assay was developed that is based on pulse-labeled incorporation of [3H]uridine into nuclear RNA. Transcription is quantified by scintillation counting of [3H]uridine incorporated into nuclear RNA that is protected from S1 nuclease digestion by hybridization with cold gene probes. This assay was dependent upon partial degradation of nuclear RNA and optimization of hybridization and nuclease digestion conditions. To validate this assay, transcription of beta-actin and c-myc genes was measured in two different human cell lines using the incorporation assay in parallel with the nuclear run-off assay. Transcription kinetics of the beta-actin and c-myc genes in serum-stimulated fibrosarcoma HT-1080 cells determined by [3H]uridine incorporation were comparable to that determined by the nuclear run-off method. For beta-actin, there was an approximate 2-fold increase in transcription rate within two hours of stimulation that declined to basal levels by 20 h. The c-myc gene response followed a similar kinetics as for the beta-actin gene except that maximal enhancement was greater at 6-9-fold. The relative transcriptional activities of the beta-actin gene to that of the c-myc gene were virtually identical using the two assay methods. Comparable transcription results using both methods were also observed when beta-actin and c-myc gene transcription were measured in log-phase HL-60 leukemia cells.
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PMID:Measurement of gene-specific transcription by nuclease protection of pulse-labeled nuclear RNA. 769 98