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)

We have isolated a 34-kilobase pair genomic thrombospondin clone and determined the sequence of 2033 base pairs of the 5'-flanking region, the first entirely untranslated exon and the first intron. A number of interesting regulatory motifs were identified. The transcription start site, determined by primer extension and S1 nuclease analysis, was shown to be 20-30 bases 3' of a TATA-like sequence, TTTAAAA. We have also shown that a thrombospondin promoter-bovine growth hormone fusion gene is transcribed in transiently transfected cells. We conclude that the genomic clone contains the transcriptional signals of the thrombospondin gene and will therefore be useful in the analysis of cis-acting elements that regulate the expression of this gene.
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PMID:Structural analysis and expression of the human thrombospondin gene promoter. 284 1

Transgenic mice carrying the H-2K/human growth hormone (hGH) fusion gene were produced by microinjecting into the pronucleus of fertilized eggs DNA molecules containing 2 kb of the 5' flanking sequences (including promoter) of the class I H-2Kb gene joined to the coding sequences of the hGH gene. Thirteen transgenic mice were obtained which all contained detectable levels of hGH hormone in their blood. Nine grew larger than their control litter-mates. Endogenous H-2Kb and exogenous hGH mRNA levels were analysed by S1 nuclease digestion experiments. hGH transcripts were found in all the tissues examined and the pattern of expression paralleled that of endogenous H-2K gene expression, being high in liver and lymphoid organs and low in muscle and brain. Thus 2 kb of the 5' promoter/regulatory region of the H-2K gene are sufficient to ensure regulated expression of hGH in transgenic mice. This promoter may therefore be of use to target the expression of different exogenous genes in most tissues of transgenic mice and to study the biological role of the corresponding proteins in different cellular environments.
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PMID:Studies on the expression of an H-2K/human growth hormone fusion gene in giant transgenic mice. 301 67

Stably transfected cell lines containing the normal human growth hormone (hGH-N) and human growth hormone-variant (hGH-V) genes have been established in order to study the expression of these two highly homologous genes. Each gene was inserted into a bovine papillomavirus shuttle vector under the transcriptional control of the mouse metallothionein gene promoter and the resultant recombinants were transfected into mouse C127 cells. The transfected cells containing the hGH-N gene secrete two hGH proteins, 91% migrating at 22 kD and 9% migrating at 20 kD, the same relative proportions synthesized in vivo by the human pituitary. S1 nuclease analysis of mRNA from these cells confirms that 20 kD hGH is encoded by an alternatively spliced product of the primary hGH-N gene transcript in which the normal exon 3 splice-acceptor site is bypassed for a secondary site 15 codons within exon 3. Although the hGH-V gene is identical to the hGH-N gene for at least 15 nucleotides on either side of the normal and alternative exon 3 AG splice-acceptor sites, hGH-V synthesizes only a 22-kD protein. Reciprocal exchange of exon 3 and its flanking intron sequences between the hGH-N gene and the hGH-V gene, eliminates the synthesis of the 20-kD protein in both resultant chimeric genes. These results directly demonstrate that both the major 22-kD and the minor 20-kD forms of pituitary hGH are encoded by the alternative splicing products of a single hGH-N gene transcript. This alternative splicing is neither species nor tissue-specific and appears to be regulated by at least two separate regions remote from the AG splice-acceptor site.
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PMID:Human growth hormone gene and the highly homologous growth hormone variant gene display different splicing patterns. 339 9

A plasmid containing 1.8 kilobase pairs of rat growth hormone (rGH) promoter and upstream flanking sequences fused to the bacterial gene for chloramphenicol acetyltransferase (CAT) was transiently introduced into pituitary, fibroblast, and kidney cell lines. Significant CAT activity was detectable only in the pituitary cell lines, demonstrating that this relatively large fragment directs strongly cell-type-specific expression. However, plasmids containing only 200-300 bases of rGH promoter and flanking sequences directed expression of CAT in all three cell types, suggesting that upstream sequences directly repress the activity of a minimal rGH promoter in nonpituitary cell types. S1 nuclease analysis showed that the RNA synthesis directed by one of the short rGH promoter fragments in fibroblasts initiated from the site used by the natural promoter in pituitary cells. Insertion of rGH upstream sequences in their natural orientation upstream of the mouse metallothionein I promoter caused a decrease in its activity in fibroblasts by a factor of 4, but there was a 2.5-fold increase in its activity in pituitary cells. Insertion of the rGH fragment upstream of the thymidine kinase promoter in either orientation lowered its activity in both fibroblasts and pituitary cells. Thus, the negatively acting rGH flanking sequences can act on a heterologous promoter and have at least some of the properties of positively acting enhancers.
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PMID:Repression mediates cell-type-specific expression of the rat growth hormone gene. 346 54

The characterization of a 20 kilodalton (20 kD) variant of rat growth hormone is reported. The 20 kD variant from rat pituitary gland extracts was identified on Western immunoblots of polyacrylamide gels. It was also shown that pituitary tissue maintained in culture secretes the 20 kD form. A rat growth hormone cDNA fragment was used as a probe in S1 nuclease mapping experiments of rat pituitary poly (A) mRNA to detect the presence of two growth hormone mRNAs in the rat pituitary gland. The protected mRNAs correspond to the predicted sizes that would encode the 22 kD and 20 kD forms of growth hormone. The site of variation between the mRNAs maps to a potential alternative 3' splice site in the 5' end of exon 3 of the coding sequence. The results support the hypothesis that the 20 kD variant in rat is the product of an mRNA alternatively spliced in exon 3, as is the case for the human growth hormone.
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PMID:Alternative splicing model for the synthesis and secretion of the 20 kilodalton form of rat growth hormone. 363 90

We have characterized the process by which the growth hormone (GH) gene is stimulated in rat pituitary tumor cells (GC or GH3) by the steroid hormone dexamethasone (Dex) and the thyroid hormone, L-triiodothyronine (T3). A primary transcriptional response is detected within 60 minutes of addition of T3 or Dex + T3 to GH-producing cells (GC or GH3). A fivefold transcriptional stimulation of GH nuclear RNA occurs in cells cultured with serum substitute medium and induced with Dex + T3, while T3 alone induces a modest two- to threefold stimulation. The absence of fetal calf serum from the cell culture medium does not decrease the level of transcriptional activity of the GH gene during hormone stimulation. Twenty-four hours after addition of Dex + T3 the cytoplasmic GH mRNA shows a 50-fold increase, as measured by S1 nuclease analysis. This large accumulation of cytoplasmic GH mRNA in contrast to the relatively small changes in GH gene activity is inconsistent with solely a transcriptional mechanism of hormone induction. We suggest that a change in specific GH mRNA stability also takes place in response to Dex + T3. In contrast to other reports, transcriptional stimulation of the GH gene by Dex is insignificant except in the presence of T3.
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PMID:Regulation of growth hormone messenger RNA synthesis by dexamethasone and triiodothyronine. Transcriptional rate and mRNA stability changes in pituitary tumor cells. 398 36

We examined whether the sequence extending 3' to the polyadenylylation site of the bovine growth hormone gene contains any signal that affects the polyadenylylation of the growth hormone mRNA. For this purpose, cloned copies of this gene, each containing a different length of growth hormone-specific sequence 3' to the wild-type polyadenylylation site, were used to transfect COS-1 cells. The polyadenylylation site on the mRNAs produced from the exogenously added growth hormone genes were analyzed with an S1 nuclease mapping procedure. We found that a gene containing 84 base pairs of its own 3' flanking sequence is capable of producing an accurately polyadenylylated mRNA. On the other hand, genes containing only 1, 10, or 13 base pairs of 3' flanking sequence were principally polyadenylylated at discrete sites either upstream or downstream from the wild-type position. Using a computer program, we examined whether secondary structures on the primary growth hormone transcript correlated with the site where the mRNA is polyadenylylated.
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PMID:Requirement for the 3' flanking region of the bovine growth hormone gene for accurate polyadenylylation. 614 35

The sequence of one of the two major expressed human chorionic somatomammotropin genes (hCS-1) was determined. The hCS-1 gene and the human growth hormone gene (hGH-1) share 92% nucleotide sequence homology in their 5'- and 3'-flanking regions, introns and exons. This finding, in addition to the existence of multiple closely linked hGH and hCS genes, suggests that these genes are evolving by concerted mechanisms. S1 nuclease, hybridization, and primer extension analysis of placental poly(A+) RNA demonstrated the presence of two functional initiation sites within the hCS-1 and/or the hCS-2 gene(s). The majority (about 95%) of the transcripts initiate 30 nucleotides downstream from the TATAAA sequence, and about 5% of the transcripts initiate 30 nucleotides downstream from a CATAAA sequence located 55 nucleotides 5' to the TATAAA sequence. The presence of two functional promoter elements as well as direct repeated sequences flanking the TATAAA sequence and exon I of the hCS genes are consistent with the hypothesis (Cooke, N. E., and Baxter, J. D. (1982) Nature (Lond.) 297, 603-606) that an important regulatory element may have been inserted into the gene in a separate evolutionary event. An analysis of other direct repeats, internal homology, and homology between other growth hormone and chorionic somatomammotropin genes offers a more extensive conceptualization of how this gene family evolved.
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PMID:Analysis of a major human chorionic somatomammotropin gene. Evidence for two functional promoter elements. 620 92

The present communication describes the molecular cloning and DNA sequence determination of the rat growth hormone (rGH) gene. The rGH gene was cloned on an 11 kilobase EcoRI fragment of total rat DNA; it has four intervening sequences which correspond in position to those of the human growth hormone (hGH) gene. One of the intervening sequences in the rGH gene contains a possible transposable element: a 200 base pair direct repeat that is itself flanked by an exact 15 base pair direct repeat. The DNA sequence was used to estimate the location of the 5' end of the mature growth hormone mRNA. By S1 nuclease mapping it was located approximately 25 bases "downstream" from a TATAAA sequence presumed to play a role in initiation of transcription of the rGH gene.
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PMID:DNA sequence of the rat growth hormone gene: location of the 5' terminus of the growth hormone mRNA and identification of an internal transposon-like element. 627 24

A library of cloned bovine DNA fragments was constructed and screened for growth hormone sequences. The growth hormone gene was isolated from this library and its nucleotide sequence determined. The likely transcription initiation site was located using the S1 nuclease protection procedure. The bovine growth hormone gene contains approximately 1793 nucleotides and consists of five exons separated by four intervening sequences. The sequence TATAAA is found in the 5' flanking region and probably is involved in facilitating transcription initiation. Comparison of the bovine growth hormone gene to the known sequence of the rat and human genes reveals that the coding regions of the three genes are highly conserved. In general the intervening sequences are much less similar than the coding regions. Interestingly, all three growth hormone genes share a conserved (but nonidentical) 40 base pair region within the 5' flanking region. This conserved region may be an important sequence involved in the hormonal regulation of growth hormone gene transcription. Analysis of GH sequences present in total bovine DNA suggests that the bovine genome contains a gene similar to the cloned gene as well as a different, but related, gene. The functional significance of the two genes remains to be explored. Analysis of nuclear species of growth hormone mRNA has demonstrated the presence of RNAs of 2100, 1400 and 1000 nucleotides containing growth hormone sequences. These likely correspond to a polyadenylated primary transcript, a processing intermediate and mature growth hormone mRNA, respectively.
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PMID:Nucleotide sequence of the bovine growth hormone chromosomal gene. 635 99


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