The primary transcription product of a silkworm alanine tRNA gene: Identification of in vitro sites of initiation, termination and processing

Hagenbüchle, Otto; Larson, Drena D.; Hall, Gayann I.; Sprague, Karen U.

doi:10.1016/0092-8674(79)90234-4

Cited by 123 publications

(60 citation statements)

References 35 publications

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“…3 that the initiation of transcription of these tRNA genes occurs very near the 5'-terminus of the mature tRNA sequence. In contrast to some E. coli precursor tRNAs, the eukaryotic tRNA precursors analyzed to date have only short leader sequences (8,9,11).…”

Section: Discussionmentioning

confidence: 99%

“…The use of nuclear extracts from Xenopus oocytes has allowed in vitro transcription of Xenopus 5S RNA (5) or Drosophila, yeast, and Bombyx tRNA genes (6-9). Such in vitro experiments have defined initiation and termination sites for RNA transcription in Xenopus 5S RNA (10), Drosophila tRNAArg (11), and Bombyx tRNAAla (8,9).Comparison of the DNA sequences of several Xenopus 5S RNA genes revealed a conserved nucleotide sequence in the 5'-flanking region of the mature 5S RNA coding sequence (12). However, the results of RNA transcription experiments using 5S genes with shortened 5'-leader regions suggested that these conserved sequences are not important for initiation of 5S gene transcription (13).…”

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“…Injection of cloned genes for Xenopus 5S RNA (2), Xenopus initiator tRNA (3), and yeast tRNA (4) into the Xenopus oocyte nucleus results in the correct transcription of these genes into the mature RNA. The use of nuclear extracts from Xenopus oocytes has allowed in vitro transcription of Xenopus 5S RNA (5) or Drosophila, yeast, and Bombyx tRNA genes (6)(7)(8)(9). Such in vitro experiments have defined initiation and termination sites for RNA transcription in Xenopus 5S RNA (10), Drosophila tRNAArg (11), and Bombyx tRNAAla (8,9).…”

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confidence: 99%

“…Moreover, a tRNA gene that has only 10 nucleotides upstream of the RNA initiation site transcribes well (8,9). However, because the protein initiation factors required for 5S DNA transcription are different from those for tRNA genes (20), the DNA regions controlling transcription may not be the same for 5S and tRNA genes.…”

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Two control regions for eukaryotic tRNA gene transcription.

DeFranco¹,

Schmidt²,

Söll³

1980

Proc. Natl. Acad. Sci. U.S.A.

125

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Two Drosophila tRNALYS genes with identical coding sequences were shown to transcribe with very different efficiencies in nuclear extracts from Xenopus oocytes. The use of recombinant plasmids in which the 5'-flanking sequences of these genes were either "switched" or replaced by defined pBR322 sequences revealed two control regions for tRNA gene transcription. An internal control region comprising the mature tRNA coding sequence (and possibly its 3'-flanking sequences) is sufficient for transcription initiation, and an external control region comprising the 5'-flanking sequences represses this transcription. All transcripts have short leader sequences. Altered precursor tRNAs transcribed from truncated tRNALYS genes (missing a single base pair in the acceptor stem) are not processed well in vitro.Attempts to elucidate the signals controlling eukaryotic gene transcription by RNA polymerase III (1) have been greatly facilitated by the availability of genes of known DNA sequence that can be transcribed faithfully. Injection of cloned genes for Xenopus 5S RNA (2), Xenopus initiator tRNA (3), and yeast tRNA (4) into the Xenopus oocyte nucleus results in the correct transcription of these genes into the mature RNA. The use of nuclear extracts from Xenopus oocytes has allowed in vitro transcription of Xenopus 5S RNA (5) or Drosophila, yeast, and Bombyx tRNA genes (6-9). Such in vitro experiments have defined initiation and termination sites for RNA transcription in Xenopus 5S RNA (10), Drosophila tRNAArg (11), and Bombyx tRNAAla (8,9).Comparison of the DNA sequences of several Xenopus 5S RNA genes revealed a conserved nucleotide sequence in the 5'-flanking region of the mature 5S RNA coding sequence (12). However, the results of RNA transcription experiments using 5S genes with shortened 5'-leader regions suggested that these conserved sequences are not important for initiation of 5S gene transcription (13). Experiments with enzymatically truncated genes had an unexpected result: the sole requirement for proper transcription of 5S or 5S-size RNA is the maintenance of the gene segment between nucleotides +50 and +80 of the mature 5S RNA coding sequence (14,15).Sequence analysis of various eukaryotic tRNA genes (7, 16-19) revealed no obvious common "promoter-like" sequence in the 5'-flanking regions. Moreover, a tRNA gene that has only 10 nucleotides upstream of the RNA initiation site transcribes well (8, 9). However, because the protein initiation factors required for 5S DNA transcription are different from those for tRNA genes (20), the DNA regions controlling transcription may not be the same for 5S and tRNA genes. This is also suggested by recent experiments showing that both the anterior and posterior portions of a Xenopus tRNAMet gene unit are required for its proper transcription (21).Analysis of Drosophila tRNA genes gave the nucleotide sequences of four genes coding for tRNA2LYs (ref 19; this report).A sequence comparison (22) of the 5'-flanking regions of these genes revealed a highly conserved undec...

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Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

See 3 more Smart Citations

Two control regions for eukaryotic tRNA gene transcription.

DeFranco¹,

Schmidt²,

Söll³

1980

Proc. Natl. Acad. Sci. U.S.A.

125

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show abstract

t RNA Biosynthesis

Florentz¹,

Martín²

2002

Encyclopedia of Molecular Biology

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Genetic analysis of the processing of a spliced tRNA.

Nishikura¹,

Kurjan²,

Hall³

et al. 1982

The EMBO Journal

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We analyzed the effect of 18 single nucleotide changes on the processing of the transcripts produced by cloned yeast tRNATYr genes after microinjection into the nucleus of living Xenopus oocytes. The processing step most easily blocked by mutation is the early maturation of the 5' and 3' termini of the tRNATYr primary transcript, involving removal of 5' -leader and 3' -trailer sequences and CCA addition. The enzymes seem to recognize the whole tRNA cloverleaf structure since mutations in all regions of the molecule can stop processing. Mutations that affect splicing of the 92-nudeotide precursor (which has mature ends but still contains the intervening sequence, and is the normal substrate for the splicing enzymes), are located in the vicinity of the intervening sequence. Base modification enzymes that add pseudouridine, 1-methyladenosine and 5-methylcytosine appear rather insensitive to changes in secondary and tertiary structure of early transcripts in the 16 mutants examined. These enzymes may recognize only limited regions of the precursor RNA. RNA polymerase III behaves as if able to count the number of Us added before termination; and aberrant termination products in two mutants suggest that the secondary structure of the nascent transcript can be very important in eukaryotic transcription termination.

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The primary transcription product of a silkworm alanine tRNA gene: Identification of in vitro sites of initiation, termination and processing

Cited by 123 publications

References 35 publications

Two control regions for eukaryotic tRNA gene transcription.

Two control regions for eukaryotic tRNA gene transcription.

t RNA Biosynthesis

Genetic analysis of the processing of a spliced tRNA.

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