Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.1.27.1 (RNase)
 16,360 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have isolated a cDNA clone (RBP-2) for the protein (RBP-J kappa) which binds to immunoglobulin recombination signals with 23-base pair spacers (Matsunami, N., Hamaguchi, Y., Yamamoto, Y., Kuze, K., Kangawa, K., Matsuo, H., Kawaichi, M., and Honjo, T. (1989) Nature 342, 934-937). During further screening of a cDNA library from the same mouse pre-B cell line (38B9), we have isolated a second cDNA clone (RBP-2N) which differs from RBP-2 in its 5' sequence. RNase protection assays indicated that the RBP-2N type mRNA was produced in 10-20 times the quantity as RBP-2 mRNA. To elucidate the relationship between these two mRNAs, we analyzed the genomic organization of the RBP-J kappa gene. Southern hybridization of mouse genomic DNA detected at least 7 EcoRI fragments hybridizing to an RBP-2 cDNA probe, suggesting a complex structure for the RBP-J kappa gene. Cloning of each EcoRI fragment revealed one functional RBP-J kappa gene and three related genes. The functional gene was composed of 11 exons and spanned at least 50 kilobase pairs. The sequence of exon 1 and its 5'-flanking region contained a GC-rich promoter-like region but no apparent TATA box. The initiation site of transcription was heterogeneous, and the two types of mRNA are produced from the same exon by transcription initiation at different sites and by different usage of splice signals. Two of the three related genes were processed pseudogenes with scattered stop codons. The other was also a processed gene with a sequence exactly the same as that of RBP-2, except that this gene lacked the sequence corresponding to the first exon of the functional gene.
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PMID:Genomic organization of mouse J kappa recombination signal binding protein (RBP-J kappa) gene. 174 Apr 50

Transgenic mice carrying a liver-specific promoter fused to a nuclear-targeted lacZ reporter gene were generated. Three separate lines of mice showed liver expression in the adult and no expression elsewhere in the animal. These results show that a 1.7 kb 5'-flanking region in the retinol-binding protein gene contains necessary and sufficient transcriptional signals for expression in adult livers. A fourth line (R197) did not express the transgene in the liver; instead, constitutive lacZ expression was seen during postimplantation stages of development from Day 9.5 onwards and appeared to be associated with segmented structures including somites, branchial arches, and hindbrain rhombomeres until late fetal stages. The beta-galactosidase positive cells in R197 were later seen to give rise to facial and flank musculature, and to other region-specific subpopulations of the jaws, neocortex, and brain stem. Northern blot analysis for the host retinol-binding protein RNA transcript did not show overlapping tissue expression with the reporter gene and suggests that transgenic phenotype seen in segmented embryonic structures of R197 and other extra-hepatic sites is from novel cis-acting transcriptional specificity. RNase protection assays of the R197 mRNA indicate that the lacZ sequences are appropriately transcribed downstream of the RBP canonical TATA box, even though the RBP promoter is itself silent. This result would suggest host flanking sequences with enhancer activity may have either activated the lacZ reporter gene or cooperated with the RBP promoter to create novel region-specific transcription. Breeding experiments have so far failed to produce offsprings that are homozygous for the transgene, and mating of transgenic F1 siblings routinely produce reduced litter sizes. Embryos that are homozygous for the transgene appear to be unable to survive beyond the egg cylinder stages of development. Thus, disruption of the host genome by insertional mutation appears to be manifest at an earlier stage than when position-effect expression of the transgene is first apparent. These experiments demonstrate that the component parts of a transgene may be subject to differential activation or suppression by host genomic flanking sequences and that even a strong, tissue-specific promoter may be overridden by host genes.
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PMID:Liver-specific and position-effect expression of a retinol-binding protein-lacZ fusion gene (RBP-lacZ) in transgenic mice. 206 Jul 5

We have studied the pathways of regulation of cytokine and cell cycle control proteins during infection of human B lymphocytes by Epstein-Barr virus (EBV). Among 30 cytokine RNAs analyzed by the RNase protection assay, tumor necrosis factor alpha (TNF-alpha), granulocyte colony-stimulating factor, lymphotoxin (LT), and LTbeta were found to be regulated within 20 h of EBV infection of primary B cells. Similar results were obtained using the estrogen-regulated EBNA-2 cell line EREB2.5, in which RNAs for LT and TNF-alpha were induced within 6 h of activation of EBNA-2. Expression of Notch also caused an induction of TNF-alpha RNA. The induction of TNF-alpha RNA by EBNA-2 was indirect, and constitutive expression of either LMP-1 or c-myc proteins did not substitute for EBNA-2 in induction of TNF-alpha RNA. Cyclin D2 is also an indirect target of EBNA-2-mediated transactivation. EBNA-2 was found to activate the cyclin D2 promoter in a transient-transfection assay. A mutant of EBNA-2 that does not bind RBP-Jkappa retained some activity in this assay, and activation did not depend on the presence of B-cell-specific factors. Deletion analysis of the cyclin D2 promoter revealed that removal of sequences containing E-box c-myc consensus DNA binding sequences did not reduce EBNA-2-mediated activation of the cyclin D2 promoter in the transient-transfection assay. The results indicate that cytokines are an early target of EBNA-2 and that EBNA-2 can regulate cyclin D2 transcription in EBV-infected cells by mechanisms additional to the c-myc pathway.
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PMID:Direct and indirect regulation of cytokine and cell cycle proteins by EBNA-2 during Epstein-Barr virus infection. 1126 43

RNA interference (RNAi) is a biological process in which animal and plant cells destroy double-stranded RNA (dsRNA) and consequently the mRNA that shares sequence homology to the dsRNA. Although it is known that the enzyme Dicer is responsible for the digestion of dsRNA into approximately 22 bp fragments, the mechanism through which these fragments are associated with the RNA-induced silencing complex (RISC) is mostly unknown. To find protein components in RISC that interact with the approximately 22 bp fragment, we synthesized a (32)P- and photoaffinity moiety-labeled 22 bp dsRNA fragment and used it as bait to fish out protein(s) directly interacting with the dsRNA fragment. One of the proteins that we discovered by mass spectrometric analysis was TB-RBP/translin. Further analysis of this DNA/RNA binding protein showed that it possesses both ssRNase and dsRNase activities but not DNase activity. The protein processes long dsRNA mainly into approximately 25 bp fragments by binding to the open ends of dsRNA and cutting it with almost no turnover due to its high affinity toward the products. The activity requires physiological ionic strength. However, with single-stranded RNA as substrate, the digestion appeared to be more complete. Both ssRNase and dsRNase activities are inhibited by high levels of common RNase inhibitors. Interestingly, both activities can be enhanced greatly by EDTA.
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PMID:Testis brain ribonucleic acid-binding protein/translin possesses both single-stranded and double-stranded ribonuclease activities. 1549 Nov 49

RNA immunoprecipitation in tandem (RIPiT) is a method for enriching RNA footprints of a pair of proteins within an RNA:protein (RNP) complex. RIPiT employs two purification steps. First, immunoprecipitation of a tagged RNP subunit is followed by mild RNase digestion and subsequent non-denaturing affinity elution. A second immunoprecipitation of another RNP subunit allows for enrichment of a defined complex. Following a denaturing elution of RNAs and proteins, the RNA footprints are converted into high-throughput DNA sequencing libraries. Unlike the more popular ultraviolet (UV) crosslinking followed by immunoprecipitation (CLIP) approach to enrich RBP binding sites, RIPiT is UV-crosslinking independent. Hence RIPiT can be applied to numerous proteins present in the RNA interactome and beyond that are essential to RNA regulation but do not directly contact the RNA or UV-crosslink poorly to RNA. The two purification steps in RIPiT provide an additional advantage of identifying binding sites where a protein of interest acts in partnership with another cofactor. The double purification strategy also serves to enhance signal by limiting background. Here, we provide a step-wise procedure to perform RIPiT and to generate high-throughput sequencing libraries from isolated RNA footprints. We also outline RIPiT's advantages and applications and discuss some of its limitations.
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PMID:Identification of Footprints of RNA:Protein Complexes via RNA Immunoprecipitation in Tandem Followed by Sequencing (RIPiT-Seq). 3135 89