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)

The cDNA that accommodates the most distal 5'-end region of cholecystokinin mRNA was isolated from an internally primed cDNA library. Using primer extension and S1 nuclease protection analyses, we demonstrated multiple RNA molecules generated from the rat cholecystokinin gene, a single-copy sequence. The longest RNA is transcribed at position--225 upstream relative to the translation start site. The major transcription, more than 95% of the total cholecystokinin mRNA in rat brain, occurred at--59 and its promoter activity was determined by in vitro RNA synthesis in a HeLa cell extract. Deletion to--105 demonstrated an approximately 60% decrease in transcriptional level compared with the full promoter activity. At least the upstream region between--254 and--105 is necessary for transcription initiated at--59 of the cholecystokinin gene by the cell-free system.
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PMID:Transcription of the rat cholecystokinin gene is initiated at multiple sites: verification by an in vitro transcription system. 172 37

Cholecystokinin (CCK), the long known gut hormone has recently been found in the brain and suggested to be a neurotransmitter or neuromodulator. In order to clarify the structure of the CCK precursor, we cloned the cDNA to mRNA of rat brain. We determined the nucleotide sequence of the inserted cDNA and deduced the amino acid sequence and proteolytic cleavage sites of the CCK precursor. The availability of CCK cDNA probes allowed us to examine the levels of CCK mRNA in the brain and intestine. CCK mRNA was barely detectable in the fetal brain, but started to increase postnatally and attained a plateau after 20-30 days. The level of CCK mRNA was the highest in the frontal cortex, followed by those of the hippocampus and striatum. The cerebellum contained only negligible CCK mRNA. Furthermore, we succeeded in cloning the CCK gene and revealed its structure which contained three exons interrupted by two introns. The size of this gene spanned about 7 kbp. The transcription initiation sites were determined by using S1 nuclease mapping and a primer extension procedure, showing evidence for the existence of the major and minor start sites. To analyze the mechanism for CCK gene expression in the neuron and paraneuron, we tried to examine the transcription of the DNA fragments from CCK gene by an in vitro cell free system. These experiments confirmed the existence of two transcription start sites. From experiments with the deleted mutants of DNA fragments, we could deduce the promoter sequence.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Expression of the cholecystokinin gene in the neuron and paraneuron. 251 Aug 4

Cholecystokinin (CCK) is a neuropeptide which is present in brain and intestine and which stimulates gall bladder contraction and pancreatic secretion. Additional studies have demonstrated an appetite-suppressing effect of CCK in vivo. These data have aroused speculation that the physiology of this hormone could be relevant in the pathogenesis of the mouse obesity mutations ob on chromosome 6 and db on chromosome 4. In order to determine whether abnormalities of this hormone could be the primary defect in these obesity mutations, we have used three separate approaches to map the mouse Cck gene to distal chromosome 9, where it is part of a syntenic group between mouse chromosome 9 and human chromosome 3. These data therefore exclude cholecystokinin as the etiologic factor in the pathogenesis of any of the known mouse obesity syndromes. In order to exclude the possibility that there are differences in mutant animals in the level of CCK RNA, we have used an S1 nuclease protection assay as well as a novel radioimmunoassay that detects the CCK precursor, to show that there are no gross differences in CCK mRNA or protein precursor levels between ob/ob and wild-type animals.
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PMID:Level of expression and chromosome mapping of the mouse cholecystokinin gene: implications for murine models of genetic obesity. 257 82

Using previously cloned cDNAs to pig brain prepro-cholecystokinin mRNA and slot blot and S1 nuclease protection assays, the relative cholecystokinin mRNA levels in different regions of the pig brain were measured. The relative amounts of cholecystokinin mRNA generally correlated well with the levels of cholecystokinin-immunoreactive peptides in the various regions tested. One clear exception was noted in the cerebellum; in this region, levels of cholecystokinin mRNA were about 20% of the levels in brain cortex (or second highest level in all areas tested) whereas the mature forms of cholecystokinin peptides (cholecystokinin 58, cholecystokinin 8) were undetectable (less than 3 pmol/g). In vitro translation of cerebellar and cortical cholecystokinin mRNA indicated that there was no difference in the efficiency with which these two RNAs were translated into immunoreactive prepro-cholecystokinin. DNA sequence analysis confirmed that a cloned full-length cerebellar cholecystokinin cDNA was indistinguishable from its cortical counterpart and, therefore, must encode an identical prepro-cholecystokinin. We conclude that there are pronounced regional differences in cholecystokinin expression in pig brain. The apparent discrepancy between levels of immunoreactive cholecystokinin peptides and cholecystokinin mRNA in the cerebellum could be explained by a high turnover rate for the peptides, differential processing of the peptides, or tissue-specific inhibition of cholecystokinin mRNA translation.
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PMID:Cholecystokinin mRNA in porcine cerebellum. 366 32

Cholecystokinin (CCK) is a neuropeptide found in brain and intestine. In this report, we have isolated a cDNA clone that encodes CCK from a mouse brain cDNA library. This cDNA clone has extensive homology to CCK precursors that have been sequenced previously. Southern blots of genomic DNA probed with this cDNA clone revealed single bands for each of eight different restriction enzymes, all of which could be accounted for by a single genomic clone, suggesting that the CCK gene is present as a single-copy gene in mice. RNA blots, primer extensions, and S1 nuclease protection assays have suggested that the same RNA start site is utilized in brain and in gut. Finally, we have shown, by using RNA blots and a radioimmunoassay specific for CCK, that CCK is expressed at maximum adult levels in intestine at birth but that adult concentrations of CCK and its mRNA are not reached in brain until much later in development.
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PMID:Differential expression of the mouse cholecystokinin gene during brain and gut development. 386 83

The cholecystokinin-B and gastrin receptor is encoded by a single gene composed of five exons and spanning over 10 kilobases on human chromosome 11p 15.5-->15.4. Exon 4 has two possible alternative splicing donor sites that seem to be conserved in other species such as the canine, rat, Mastomys, and mouse. They could generate two receptor isoforms (short- and long-form), which differ in their putative third cytoplasmic domain of the serpentine G-protein-coupled receptors. In the present study, we examined whether an alternative splicing is operated in a tissue-specific manner and whether two receptor isoforms have functional differences. RNase-protection assay and S1 nuclease mapping demonstrated the preferential expression of the short-form in the human brain as well as the digestive organs, stomach and pancreas. The two putative isoforms of the cholecystokinin-B/gastrin receptor expressed in mouse fibroblasts showed the same characteristics in their ligand-bindings, the major signal transduction such as phosphoinositides production, cytoplasmic Ca2+ increase, tyrosine phosphorylation of focal adhesion kinase, activation of mitogen-activated protein kinase, and the induction of early-responsive genes such as c-fos, c-myc, and c-jun. Moreover, the ligand-dependent trophic effect was seen in both receptor isoforms. Taken together with the absence of tissue-specific expression of two receptor isoforms, these results suggest a species-specific dominant splice donor site in exon 4 of the human receptor gene.
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PMID:Functional characterization of two cholecystokinin-B/gastrin receptor isoforms: a preferential splice donor site in the human receptor gene. 784 14