A Drosophila metabolic gene transcript is alternatively processed

Henikoff, Steven; Sloan, James S.; Kelly, James D.

doi:10.1016/0092-8674(83)90374-4

Cited by 68 publications

(33 citation statements)

References 31 publications

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“…A segment of D. melanogaster DNA has previously been shown to complement an S. cerevisiae adenine-8 mutation (15). Transcription of this Drosophila segment has been mapped both on a plasmid in S. cerevisiae (12) and in the Drosophila genome (14). Although sites for the initiation of transcription differ considerably, the 3'-end locations of polyadenylic acid on the mRNAs isolated from the two organisms nearly coincide.…”

Section: Resultsmentioning

confidence: 96%

Sequences responsible for transcription termination on a gene segment in Saccharomyces cerevisiae.

Henikoff¹,

Cohen²

1984

Mol. Cell. Biol.

156

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We have mapped a signal sequence for mRNA 3'-end formation in Saccharomyces cerevisiae by using a Drosophila melanogaster DNA segment that complements a yeast adenine-8 mutation. That the 3' end of the transcript in S. cerevisiae nearly coincides with that in D. melanogaster is consistent with the possibility that mRNA termini are similarly determined in both organisms. Deletion analysis reveals that the complete signal is no more than 21 base pairs long. Part of the signal is the sequence TTTTTATA, which is seen in the termination region of several yeast genes. TTTTTATA appears to be able to act autonomously as a partial termination signal. The efficiency of the complete signal is affected by substitution of sequences downstream from it. This modulation of the effect of a signal is consistent with termination in S. cerevisiae, resembling rhodependent termination in bacteria.Until recently, transcription termination in eucaryotes has been poorly understood, in part because of the rapid processing that occurs at the 3' ends of transcripts for proteincoding genes. Processing usually involves cleavage of a precursor followed by polyadenylation in higher eucaryotes (6, 21). Histone mRNA 3' ends which lack polyadenylic acid were once thought to result from transcription termination (3) but now appear to be generated by processing (2, 7). The biochemical elucidation of mRNA 3' processing should be possible with the recent development of a cell-free system (23). In contrast to processing, actual transcription termination, which can occur at a considerable distance downstream from the mRNA 3' end (6, 27), has been relatively refractory to analysis.In Saccharomyces cereisisiae, polyadenylic acid addition and transcription termination appear to be coupled, as essentially all transcripts from protein-coding genes are polyadenylated [poly(A)+] (29). Therefore, the strong possibility exists that the study of 3'-end formation of an mRNA in yeast cells will shed light on the actual transcription termination event. Termination is known to be important in the regulation of a variety of bacterial operons (reviewed in references 17 and 24) and appears to be involved in the regulation of at least one eucaryotic gene (9, 10, 28). We have been investigating the sequence determinants of transcription termination on a DNA segment in S. cerev,isiae. This segment is derived from Drosophila melanogaster and complements a yeast adenine-8 mutation (15). We showed previously that mRNA 3' ends actually occur 50 to 90 base pairs (bp) downstream from a control signal, the 3' boundary of which we mapped precisely (13). Here we show that this signal is no more than 21 bp long. We also present evidence suggesting that an 8-bp portion of this signal promotes partial termination. It

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Section: Resultsmentioning

confidence: 96%

Sequences responsible for transcription termination on a gene segment in Saccharomyces cerevisiae.

Henikoff¹,

Cohen²

1984

Mol. Cell. Biol.

156

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“…From published data on several other abundantly expressed genes (the rat calcitonin gene [23], the Drosophila actin genes [9], and the chicken ovalbumin gene [17]), we estimate that less than 1% of the RNA homologous to these genes is of precursor length. Similarly, in two published examples of Drosophila genes, no evidence of intron-containing RNAs can be detected in RNA blot analyses (13,22). In the case of the white gene (22), probes from within an intron fail to detectably hybridize any poly(A)+ RNAs containing the intron sequences.…”

Section: Methodsmentioning

confidence: 99%

High levels of intron-containing RNAs are associated with expression of the Drosophila DOPA decarboxylase gene.

Beall¹,

Hirsh²

1984

Mol. Cell. Biol.

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We have examined the structure and expression during embryonic development of the Drosophila DOPA decarboxylase gene, Ddc. The Ddc gene is transcribed to make at least five different size classes of RNA. These RNA species first appear late in embryogenesis, coincident with induction of Dde enzyme activity. The most abundant and smallest RNA appears to be Ddc mRNA. The sequences encoding this RNA are split by two intervening sequences. Each of the larger RNA species contains some or all of the intervening sequences. We have noted two unusual features of Ddc expression during embryogenesis. First, the intervening-sequencecontaining RNAs are present as 20% or more of the polyadenylated Dde RNA molecules, an exceptionally high proportion. Second, these RNAs do not disappear as rapidly as Ddc mRNA after Ddc enzyme activity reaches fully induced levels. These observations indicate slow rates of RNA processing relative to mRNA half-life and suggest that post-transcriptional steps participate in regulating Ddc expression. Although four of the five RNA species were detected at multiple developmental stages during which Dde is expressed, one was found uniquely during embryogenesis. This RNA differs from Ddc mRNA in length and in time of expression during embryogenesis but is transcribed in the same orientation and from the same genomic sequences as the Ddc primary transcript.

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“…The entire DNA sequence has been determined, both mRNAs have been mapped at the sequence level, and all three enzyme activities have been localized within the two encoded polypeptides (14,(28)(29)(30). A combined genetic and molecular analysis might lead to an understanding of the regulation and expression of a complex "housekeeping" gene in a developing organism.…”

Section: Resultsmentioning

confidence: 99%

Two Drosophila melanogaster mutations block successive steps of de novo purine synthesis.

Henikoff¹,

Nash²,

Hards³

et al. 1986

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

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Drosophila melanogaster purine auxotrophs ade2(1) and ade3(1) have been characterized biochemically. The ade2(1) strain is deficient in the fourth step of the de novo purine synthetic pathway catalyzed by phosphoribosylglycinamidine synthase (phosphoribosylformylglycinamide amidotransferase). The ade3(1) strain is deficient in the previous step catalyzed by phosphoribosylglycinamide formyltransferase (GART). The mutation responsible for the slightly leaky ade3(1) phenotype was characterized further. First, the mutant GART polypeptide was found to be of normal size and present at normal levels. Second, the GART-encoding region of the mutant was cloned, inserted into a yeast-Escherichia coli shuttle vector, and used to transform mutant yeast. Transformants showed very slight in vivo activity when compared to wild type, verifying that the mutation is in the GART coding sequence. Lastly, the region of the gene encoding GART activity from mutant and inbred parental strain flies was completely sequenced. A single base transition was found, leading to the substitution of a serine for a highly conserved glycine. These two mutations provide examples of blocks in the de novo purine synthetic pathway in a whole animal.

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A Drosophila metabolic gene transcript is alternatively processed

Cited by 68 publications

References 31 publications

Sequences responsible for transcription termination on a gene segment in Saccharomyces cerevisiae.

Sequences responsible for transcription termination on a gene segment in Saccharomyces cerevisiae.

High levels of intron-containing RNAs are associated with expression of the Drosophila DOPA decarboxylase gene.

Two Drosophila melanogaster mutations block successive steps of de novo purine synthesis.

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