Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.26.4 (RNase H)
 2,751 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The human 7SK ribonucleoprotein (RNP) has been analyzed to determine its RNA secondary structure and protein constituents. HeLa cell 7SK RNA alone and within its RNP have been probed by chemical modification and enzymatic cleavage, and sites of modification or cleavage have been mapped by primer extension. The resulting secondary structure suggests that structural determinants necessary for capping (a 5' stem followed by the sequence AUPuUPuC) and nuclear migration (the sequence AUPuUPuC) of 7SK RNA may be similar to those for U6 small nuclear RNA (snRNA). It also supports existence of a 3' stem structure which could serve to self-prime cDNA synthesis during pseudogene formation. Oligonucleotide-directed RNase H digestion indicated regions of 7SK RNA capable of base pairing with other nucleic acids. Antisense 2'-O-methyl RNA oligonucleotides were used to affinity select the 7SK RNP from an in vivo 35S-labeled cell sonic extract and identify eight associated proteins of 83, 48, 45, 43, 42, 21, 18, and 13 kDa. 7SK RNA has extensive sequence complementarity to U4 snRNA, within the U4/U6 base pairing domain, and also to U11 snRNA. The possibility that the 7SK RNP is an unrecognized component of the pre-mRNA processing machinery is discussed.
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PMID:Structural analyses of the 7SK ribonucleoprotein (RNP), the most abundant human small RNP of unknown function. 164 89

A direct method for mapping introns has been devised. The technique makes use of a radioactive synthetic RNA transcript of the gene and a complementary, single-stranded DNA copy of mRNA-derived sequences. Upon hybridization of the cDNA to RNA and cleavage with ribonuclease H, only exonic RNA sequences are degraded. The surviving RNA fragments are the introns. Electrophoretic analysis in denaturing agarose gels reveals the number and size of the introns. The order of the introns is determined separately using unlabeled RNA transcripts; surviving RNA fragments are transferred to a solid support and the blot is probed sequentially with a nested set of genomic RNA transcripts of the opposite strand. Using the human prothymosin alpha gene as an example, four introns were identified which from 5' to 3' were 2.6, 0.47, 0.47, and 0.28 kb in size. From mapping and sequencing experiments the sizes are 2.6, 0.465, 0.459, and 0.295 kb, respectively. Similarly, the presence of two 300-bp insertions in a human prothymosin alpha pseudogene was established; the inserts were later identified as 295-bp Alu repetitive elements.
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PMID:Mapping introns by exon excision. 255 29

The POMC gene is predominantly expressed in the pituitary gland; it is also expressed in various extrapituitary tissues. While POMC mRNAs of similar size (approximately equal to 1000 nucleotides) are present in the anterior and neurointermediate lobes of the pituitary, other POMC-expressing tissues contain POMC mRNAs of different sizes. Longer POMC mRNAs are observed in the hypothalamus. Using S1 nuclease mapping and mRNA deadenylation by RNase H, we have shown that these large hypothalamic POMC mRNAs have longer poly(A) tails than pituitary POMC transcripts but contain the same transcripted sequences. In contrast, the testes contain POMC transcripts which are smaller than pituitary POMC mRNA. RNase and S1 nuclease mapping analyses suggest that these short transcripts do not contain sequences transcribed from pituitary exons 1 and 2. Indeed, as revealed by primer-extension experiments, these transcripts appear to initiate within exon 3 sequences of the POMC gene. The heterogeneous 5'-ends of these short testicular transcripts map into the NH2-terminal portion of the precursor in the region encoding gamma MSH; if ever translated, these transcripts would produce a form of POMC that would be truncated at the NH2-terminus and therefore would be devoid of any signal peptide sequence. Interestingly, the sequence of the short testicular transcripts corresponds to that of the mouse POMC pseudogene, suggesting that this POMC pseudogene may have derived from genomic integration of testicular transcripts via a cDNA intermediate.
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PMID:Unusual proopiomelanocortin ribonucleic acids in extrapituitary tissues: intronless transcripts in testes and long poly(A) tails in hypothalamus. 285 1

We have analysed the reverse transcriptase (RT) activity of the human LINE retrotransposon and that of two retroviruses, using an in vivo assay within mammalian (murine and human) cells. The assay relies on transfection of the cells with expression vectors for the RT of the corresponding elements and PCR analysis of the DNA extracted 2-4 days post-transfection using primers bracketing the intronic domains of co-transfected reporter genes or of cellular genes. This assay revealed high levels of reverse-transcribed cDNA molecules, with the intron spliced out, with expression vectors for the LINE. Generation of cDNA molecules requires LINE ORF2, whereas ORF1 is dispensable. Deletion derivatives within the 3.8 kb LINE ORF2 allowed further delineation of the RT domain: > 0.7 kb at the 5'-end of the LINE ORF2 is dispensable for reverse transcription, consistent with this domain being an endonuclease-like domain, as well as 1 kb at the 3'-end, a putative RNase H domain. Conversely, the RT of the two retroviruses tested, Moloney murine leukemia virus and human immunodeficiency virus, failed to produce similar reverse transcripts. These experiments demonstrate a specific and high efficiency reverse transcription activity for the LINE RT, which applies to RNA with no sequence specificity, including those from cellular genes, and which might therefore be responsible for the endogenous activity that we previously detected within mammalian cells through the formation of pseudogene-like structures.
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PMID:Functional differences between the human LINE retrotransposon and retroviral reverse transcriptases for in vivo mRNA reverse transcription. 935 39