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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)

Using the S1 nuclease mapping technique, we demonstrated that the majority of Moloney murine leukemia RNA molecules, isolated either from the nucleus or cytoplasm of infected mouse cells, share a uniform 3' end located at the border of the R and U-5 regions of the long terminal repeat. When the long terminal repeat sequences were inserted in the pSV plasmid downstream of the simian virus 40 late promoter, the 3' end of the viral RNA was also generated close to the R region of the long terminal repeat. These results demonstrate that the long terminal repeat signals the generation of an authentic 3' end when situated downstream of an actively transcribed region.
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PMID:Generation of a uniform 3' end RNA of murine leukemia virus. 298 58

We examined the mos-specific intracellular RNA species in 6m2 cells, an NRK cell line nonproductively infected with the ts110 mutant of Moloney murine sarcoma virus. These cells present a normal phenotype at 39 degrees C and a transformed phenotype at 28 or 33 degrees C, expressing two viral proteins, termed P85gag-mos and P58gag, at 28 to 33 degrees C, whereas only P58gag is expressed at 39 degrees C. It has been previously shown that 6m2 cells contain two virus-specific RNA species, a 4.0-kilobase (kb) RNA coding for P58gag and a 3.5-kb RNA coding for P85gag-mos. Using both Northern blot and S1 nuclease analyses, we show here that the 3.5-kb RNA is the predominant viral RNA species in 6m2 cells grown at 28 degrees C, whereas only the 4.0-kb RNA is detected at 39 degrees C. During temperature shift experiments, the 3.5-kb RNA species disappears after a shift from 28 to 39 degrees C and is detected again after a shift back from 39 to 28 degrees C. By Southern blot analysis, we have detected only one ts110 proviral DNA in the 6m2 genome. This observation, as well as previously published heteroduplex and S1 nuclease analyses which showed that the 3.5-kb RNA species lacks about 430 bases found at the gag gene-mos gene junction in the 4.0-kb RNA, suggests that the 3.5-kb RNA is a splicing product of the 4.0-kb RNA. The absence of the 3.5-kb RNA when 6m2 cells are grown at 39 degrees C indicates that the splicing reaction is thermosensitive. The splicing defect of the ts110 Moloney murine sarcoma virus viral RNA in 6m2 cells cannot be complemented by acute Moloney murine leukemia virus superinfection, since no 3.5-kb ts110 RNA was detected in acutely superinfected 6m2 cells maintained at 39 degrees C. The spliced Moloney murine leukemia virus env mRNA, however, is found in acutely infected cells maintained at 39 degrees C, suggesting that the lack of ts110 viral RNA splicing at 39 degrees C is not due to an obvious host defect. In sharp contrast, however, 6m2 cells chronically superinfected with Moloney murine leukemia virus produce a 3.5-kb RNA species at 39 degrees C as well as at 28 degrees C and contain proviral DNAs corresponding to the two viral RNA species.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Temperature-sensitive viral RNA expression in Moloney murine sarcoma virus ts110-infected cells. 298 39

DNA-mediated gene transfer techniques have been used to study the effectiveness of a novel construction involving the feline leukemia virus long terminal repeat (FeLV LTR) for expressing the mouse H-2 Ld gene in mouse and human cells. In this construction, the transcription initiation (promoter) and termination (polyadenylation) functions of the FeLV LTR have been split by insertion of a promoterless H-2 gene between them. An S1 nuclease assay has been developed that makes it possible to measure accumulated LdRNA against a background of endogenous major histocompatibility antigen RNAs in mouse and human cells. In mouse cells, the H-2 Ld gene was expressed at approximately equal levels (measured as accumulated RNA) when driven either by its own promoter or by the FeLV LTR construction. In human cells, expression at the RNA level was highest when driven by the FeLV LTR. We conclude that the FeLV LTR construction is useful for expressing foreign genes in human cells.
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PMID:The promoter of the long terminal repeat of feline leukemia virus is effective for expression of a mouse H-2 histocompatibility gene in mouse and human cells. 298 2

The human T-cell leukemia viruses (HTLV) are replication-competent retroviruses whose genomes contain gag, pol, and env genes as well as a fourth gene, termed x, which is believed to be the transforming gene of HTLV. The product of the x gene is now shown to be encoded by a 2.1-kilobase messenger RNA derived by splicing of at least two introns. By means of S1 nuclease mapping of this RNA and nucleic acid sequence analysis of a complementary DNA clone, the complete primary structure of the x-gene product has been determined. It is encoded by sequences containing the env initiation codon and one nucleotide of the next codon spliced to the major open reading frame of the HTLV-I and HTLV-II x gene.
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PMID:HTLV x-gene product: requirement for the env methionine initiation codon. 299 32

Chromosomal rearrangements involving the c-myc oncogene are a prevalent feature of plasmacytomas that arise after inoculating BALB/c mice with pristane and Abelson murine leukaemia virus (A-MuLV). With this observation in mind, we decided to determine if any genetic alterations of the c-myc locus could be observed in cells of a different type, when transformed in vitro by A-MuLV. Here we have analysed three independent A-MuLV-transformed NIH 3T3 lines (ANN-I, 54c12 and N25), and found that the c-myc locus is amplified 8-19-fold in each transformant. Quantitative S1 nuclease mapping performed on ANN-I and 54c12 RNAs demonstrated that: (1) c-myc messenger RNAs accumulated to double the levels found in NIH 3T3 cells; and (2) a shift in the use of the two normal c-myc transcription initiation sites (P1 and P2) occurred in favour of the 3' site, P2. Analysis of c-myc chromatin by DNase I treatment of 54c12 nuclei revealed that most, if not all, of the c-myc gene copies were transcriptionally competent. We present alternative ideas to explain why amplification of the c-myc gene occurs repeatedly in A-MuLV-transformed fibroblasts. Finally, we discuss our results in relation to the hypothesis linking the phenomenon of tumour progression with the amplification of oncogenes.
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PMID:Amplification and altered expression of the c-myc oncogene in A-MuLV-transformed fibroblasts. 299 29

The long terminal repeat (LTR) from proviral DNA of Moloney murine leukemia virus (Mo-MLV) was cloned on a derivative of pBR322, and after introducing superhelical torsions into the resulting recombinant, the sites of conformational transition were investigated by the nuclease S1-digestion method. With an increase in the negative linking differences, fourteen dominant cutting sites were identified, of which two were mapped inside the LTR and one at the 3' end of the LTR. By searching the sequence data, all these sites were localized in the regions having either palindromic sequences or AT-rich sequences. Free energy calculation for the local secondary structure on one strand indicated that nuclease S1 attacked the palindromic sequence regions which could form relatively stable hairpin structures. Under the conditions used, no correlation was found between the S1-sensitive sites and the potential Z-DNA-forming regions, including those within the enhancer sequence.
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PMID:Nuclease S1-sensitive sites on superhelical DNA molecules carrying the LTR region of Moloney murine leukemia virus. 303 62

The c-myc gene is rearranged in a subset of feline T-cell lymphosarcomas. Detailed mapping of c-myc rearrangements showed that some result from feline leukaemia virus (FeLV) proviral integration within or upstream of c-myc, but one case involves a complex 3' alteration and amplification which is apparently not directly virus-induced. S1 nuclease mapping of RNA from normal cells using c-myc probes revealed two presumptive 5' ends, each corresponding to a promoter-like sequence (P1 and P2), and a major 3' discontinuity which mapped to the 3'-most of two possible polyadenylation signals. Analysis of RNA from a series of tumours revealed different modes of c-myc expression. All tumours produced P1 and P2 transcripts with apparently normal structure except for one case where an insertion in intron 1 displaced exon 1 sequences. The abundance ratio of P1/P2 transcripts varied considerably and was high in tumours which carry a rearrangement adjacent to c-myc, but some other T-cell tumours with no apparent myc alteration displayed an equally high ratio. However, a consistent feature was the lack of detectable RNA from normal c-myc alleles in tumours which express a rearranged c-myc allele or a transduced FeLV v-myc gene. We suggest that this may prove to be a useful indicator of the presence of an oncogenically active myc gene, whether this is a rearranged c-myc or transduced v-myc sequence.
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PMID:Altered structure and expression of c-myc in feline T-cell tumours. 303 89

We have examined the S1 nuclease sensitivity of supercoiled plasmids harboring the Moloney Murine Leukemia Virus (MoMuLV) long terminal repeat (LTR). S1 sensitivity was found within the LTR enhancer direct repeats. Transformation of E. coli DH5 cells with a construct containing most of the MoMuLV LTR yielded the precise deletion of one direct repeat and loss of S1 sensitivity. The dependence of S1 sensitivity on the presence of both direct repeats, together with the exact excision of one direct repeat by E. coli, suggests the presence of slipped DNA within the enhancer. Such structures may represent targets for effector proteins which mediate vital functions during viral propagation.
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PMID:Slipped DNA structures within the enhancer region of the Moloney murine leukemia virus. 305 52

Adenovirus early region 1A (E1A), which gives rise to three overlapping transcripts, was inserted into a murine leukemia virus-derived vector, and recombinant viruses were used to prepare permanent cell lines of NIH 3T3 cells containing DNA copies of the individual 13S, 12S, and 9S mRNAs. Integrated proviral copies of the recombinant genomes were rescued as bacterial plasmids from each of the cell lines, and the DNA sequence of E1A was demonstrated to be a precise copy of the individual transcripts. The DNA copies were shown to be expressed as part of the full-length retroviral transcript by S1 nuclease analysis, and the synthesis of their encoded polypeptides was demonstrated by immunoprecipitation. Those cell lines expressing the polypeptide encoded by the 13S transcript were shown to contain that function required for regulating the accumulation of mRNAs from adenovirus early genes by their ability to complement the adenovirus type 5 E1A deletion mutant dl312. Cell lines expressing polypeptides encoded by the 13S, 12S, and 9S transcripts showed characteristic alterations in morphology. Two-dimensional gel electrophoresis of total cellular protein derived from the three cell lines demonstrated that each E1A gene product elicits specific alterations in the patterns of proteins expressed. Studies of the expression of two specific genes, those encoding fibronectin and collagen type 1, indicated that the observed alteration in levels of the two proteins results from a reduction in RNA levels induced by E1A functions.
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PMID:Individual adenovirus type 5 early region 1A gene products elicit distinct alterations of cellular morphology and gene expression. 405 56

Activation of transcription by the Moloney Murine Leukemia Virus (MLV) and Simian Virus (SV40) enhancers was compared by transfecting recombinants containing these enhancers in either mouse or human cell-lines, and analysing RNA 48 h later by quantitative S1 nuclease mapping. The enhancers share the following properties. They stimulate transcription in an orientation-independent manner from the same startsites on the natural heterologous conalbumin (+62 to -102) or SV40 early promoter elements as well as on substitute promoter elements. The enhancers are most efficient when they are located directly upstream from the conalbumin (+62 to -102) promoter element, but they still stimulate transcription when they are either immediately downstream from the promoter element, or further upstream. Increasing the distance by interposing DNA sequences between the enhancers and the conalbumin promoter fragment results in decreased activation. Both enhancers show some cell-line specificity for activation of transcription. However, in all cell-lines and constructions tested the MLV enhancer was always less efficient than the SV40 enhancer. These results suggest that the MLV and SV40 enhancers stimulate transcription by similar mechanisms.
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PMID:The MLV and SV40 enhancers have a similar pattern of transcriptional activation. 609 7


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