The role of non-coding DNA sequences in transcription and processing of a yeast tRNA

Raymond, Gregory J.; Johnson, Jerry D.

doi:10.1093/nar/11.17.5969

Cited by 75 publications

(56 citation statements)

References 53 publications

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“…As has been shown previously (8,25,29), large intron alterations-base changes, insertions, and deletions-in S. cerevisiae tRNA precursors do not affect their absolute ability to be spliced. These observations have led to the following idea: if its sequence is not deleterious to the cerevisiae, this deletion mutant is efficiently transcribed and end-processed but is not spliced.…”

supporting

confidence: 71%

“…As has been shown previously (8,25,29) higher-order structure of the precursor, an intron plays no direct role in the maturation of a precursor tRNA. In short, the intron is passive in its own removal.…”

mentioning

confidence: 71%

“…While the splice junction sequences of the Alu mutant are not unique, the G-G 5' and C-C 3' Stu mutant junctions are not found among the natural pre-tRNAs (20). Raymond and Johnson (25) have reported that a mutant with a similar 3' splice junction was refractory to the splicing enzymes. Therefore, these data suggest that the splice junction-proximal primary sequence may prove to be critical for endonuclease recognition and cleavage.…”

Section: Methodsmentioning

confidence: 98%

“…6,1986 (20). An unspliced mutant of pre-tRNA3eu has only a 2-nucleotide 3' junction loop (25). Furthermore, by precise deletion at the 3' splice junction of pre-tRNAPrO, Knapp and co-workers have shown that 2 nucleotides is the minimum loop size for total endonuclease cleavage (G. Knapp, personal communication).…”

Section: Methodsmentioning

confidence: 99%

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Intron mutations affect splicing of Saccharomyces cerevisiae SUP53 precursor tRNA.

Strobel¹,

Abelson²

1986

Mol. Cell. Biol.

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The Saccharomyces cerevisiae amber suppressor tRNA gene SUP53 (a tRNALeu allele) was used to investigate the role of intron structure and sequence on precursor tRNA splicing in vivo and in vitro. This gene encodes a pre-tRNA which contains a 32-base intervening sequence. Two types of SUP53 intron mutants were constructed: ones with an internal deletion of the natural SUP53 intron and ones with a novel intron. These mutant genes were transcribed in vitro, and the end-processed transcripts were analyzed for their ability to serve as substrates for the partially purified S. cerevisiae tRNA endonuclease and ligase. The in vitro phenotype of these mutant RNAs was correlated with the in vivo suppressor tRNA function of these SUP53 alleles after integration of the genes into the yeast genome. Analysis of these mutant pre-tRNAs, which exhibited no perturbation of the mature domain, clearly showed that intron structure and sequence can have profound effects on pre-tRNA splicing. All of the mutant RNAs, which were inefficiently spliced or unspliced, evidenced cleavage only at the 5' splice junction. Base changes in the intron proximal to the 3' splice junction could partially rescue the splicing defect. The implications of these data for tRNA endonuclease-substrate interactions are discussed.Numerous studies have investigated yeast intragenic mutations which affect the production of a mature tRNA. These mutations map almost exclusively to the mature domain of the precursor tRNA (2, 3, 11-13, 16-18, 21, 35). In general, these mutations, which affect transcription, end-processing, and splicing of the precursor, perturb conserved base-pairing or alter conserved nucleotides in the mature domain. Notable by their paucity are mutations located in the intron. By genetic selection for an antisuppressor phenotype, only three intron mutants have been found.In the Saccharomyces cerevisiae SUP4 tRNATYr, a base change creates an RNA polymerase III transcription termination site in the intron (11). Willis et al. (35) found two intron alterations in the Schizosaccharomyces pombe sup9 tRNAser: a base insertion which has little effect on RNA maturation or suppressor tRNA function and a base change near the 3' intron-exon junction. The latter mutation significantly decreases the efficiency of end-processing and splicing. In each case, the mutation(s) enhances the formation of alternate base-pairing between the mature domain and intron sequences, which perturbs the mature domain structure. Therefore, these intron mutants probably exert their effect on RNA processing by their alteration of total precursor tRNA conformation.Mutations, selected for loss of suppressor tRNA function, which affect primarily splicing are found in the D stem-loop, the anticodon stem-loop, and the variable stem-loop (extra arm). As has been shown previously (8,25,29) higher-order structure of the precursor, an intron plays no direct role in the maturation of a precursor tRNA. In short, the intron is passive in its own removal.However, there is a single example of a...

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supporting

confidence: 71%

mentioning

confidence: 71%

Section: Methodsmentioning

confidence: 98%

Section: Methodsmentioning

confidence: 99%

See 2 more Smart Citations

Intron mutations affect splicing of Saccharomyces cerevisiae SUP53 precursor tRNA.

Strobel¹,

Abelson²

1986

Mol. Cell. Biol.

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“…However, since the TR sequences of the rGH gene and ID cDNA clones are essentially the same, the basis for the striking difference in their Pol III activity is most likely due to either position effects and/or the presence of inhibitory or stimulatory flanking sequences. Although such influences have been reported, they are difficult to predict in cell-free studies (39)(40)(41). If the flanking DNA in some other way regulated the activity of the TR RNA polymerase III promotor, this could provide a mechanism whereby the activity of "identifier" or TR sequences is regulated by being dependent on the site of insertion.…”

Section: Cell-free Transcriptionmentioning

confidence: 99%

Transcription of two classes of rat growth hormone gene-associated repetitive DNA: differences in activity and effects of tandem repeat structure

et al. 1984

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The rat growth hormone (rGH) gene contains two classes of repetitive DNA arranged as clusters within intron B and the 3' flanking region. The major family is equivalent to the CHO type 2 DNA. The second ("truncated repeat", TR) is a truncated version of the first and occurs in certain neural-specific transcripts and genes ("identifier" elements, ID). Here we report, using the HeLa cell-free transcription assay, that RNA polymerase III (Pol III) efficiently initiates at internal promoters within a tandem array of rGH gene repetitive DNA monomers and results in a novel organization of overlapping Class III transcription units. Transcription competition studies revealed that the rat type 2 structures share Pol III transcription factors with a tRNA gene, a human Alu repeat, and a mutant VAl gene. Also, the rGH type 2 but not the TR DNA efficiently promotes Pol III initiation, yet other TR members, which differ only in flanking DNA, are transcribed. Thus, the rGH gene is strikingly enriched with 10 repetitive DNA monomers; multimeric type 2 elements are actively transcribed; rGH-TR sequences are expressed only as part of larger transcripts promoted by type 2 DNA; and, type 2 DNA uses tRNA gene transcription factors. These studies show that flanking sequences, promoter organization and factor competition may all affect rat repetitive DNA expression. INTRODUCTIONThe major component of mammalian short-length, middle-repetitive DNA is

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Differential binding of a S. cerevisiae RNA polymerase III transcription factor to two promoter segments of a tRNA gene.

Stillman¹,

Geiduschek²

1984

The EMBO Journal

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A Saccharomyces cerevisiae protein fraction which binds specifically to the internal promoter regions of genes that are transcribed by RNA polymerase III is shown to function as a transcription factor. We postulate that the stable DNA binding of the factor confers stability on polymerase III transcription complexes. Analysis of the binding by DNase ‘foot‐printing’ distinguishes three segments of the S. cerevisiae tRNALeu3 gene: a region surrounding the so‐called A block of the internal promoter, a region surrounding the B block and an intermediate segment. Binding to the A and B block regions is connected, but the B block region exerts a dominant effect.

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The role of non-coding DNA sequences in transcription and processing of a yeast tRNA

Cited by 75 publications

References 53 publications

Intron mutations affect splicing of Saccharomyces cerevisiae SUP53 precursor tRNA.

Intron mutations affect splicing of Saccharomyces cerevisiae SUP53 precursor tRNA.

Transcription of two classes of rat growth hormone gene-associated repetitive DNA: differences in activity and effects of tandem repeat structure

Differential binding of a S. cerevisiae RNA polymerase III transcription factor to two promoter segments of a tRNA gene.

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