The stability of E. coli gene transcripts is dependent on determinants localized to specific mRNA segments

Belasco, Joel G.; Nilsson, Gisela; Gabain, Alexander von; Cohen, Stanley N.

doi:10.1016/0092-8674(86)90741-5

Cited by 181 publications

(117 citation statements)

References 28 publications

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“…The data presented above indicate that the half-lives of E. coli transcripts extend over a wide range. Earlier investigations have shown that discrete determinants of mRNA decay exist and can be localized to particular mRNA sites rather than being dispersed widely and nonspecifically throughout the transcript (30). Consistent with such evidence for the site specificity of RNA decay determinants, our global analysis of RNA decay showed no correlation between ORF or operon length and half-life (Fig.…”

Section: Global Analysis Of Mrna Half-life In Cells Growing At Differsupporting

confidence: 73%

Global analysis of mRNA decay and abundance in Escherichia coli at single-gene resolution using two-color fluorescent DNA microarrays

Bernstein

Khodursky

Lin

et al. 2002

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

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773

711

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Much of the information available about factors that affect mRNA decay in Escherichia coli, and by inference in other bacteria, has been gleaned from study of less than 25 of the Ϸ4,300 predicted E. coli messages. To investigate these factors more broadly, we examined the half-lives and steady-state abundance of known and predicted E. coli mRNAs at single-gene resolution by using two-color fluorescent DNA microarrays. An rRNA-based strategy for normalization of microarray data was developed to permit quantitation of mRNA decay after transcriptional arrest by rifampicin. We found that globally, mRNA half-lives were similar in nutrient-rich media and defined media in which the generation time was approximately tripled. A wide range of stabilities was observed for individual mRNAs of E. coli, although Ϸ80% of all mRNAs had half-lives between 3 and 8 min. Genes having biologically related metabolic functions were commonly observed to have similar stabilities. Whereas the half-lives of a limited number of mRNAs correlated positively with their abundance, we found that overall, increased mRNA stability is not predictive of increased abundance. Neither the density of putative sites of cleavage by RNase E, which is believed to initiate mRNA decay in E. coli, nor the free energy of folding of 5 or 3 untranslated region sequences was predictive of mRNA half-life. Our results identify previously unsuspected features of mRNA decay at a global level and also indicate that generalizations about decay derived from the study of individual gene transcripts may have limited applicability.RNA degradation ͉ RNA half-life ͉ transcript abundance T he rate of mRNA degradation plays a central role in the metabolism of nucleic acids in both prokaryotic and eukaryotic cells (for reviews see refs. 1-5). mRNA decay has been studied in a range of organisms, and much has been learned about the substrate features and ribonucleolytic enzymes that influence mRNA stability. In the case of Escherichia coli, experimenters have demonstrated that extensive variation exists in the rate of decay of individual mRNAs and that specific features of the mRNA sequence as well as transcript secondary structure can be important determinants of such variation (1, 3). Additionally, although the half-lives of some RNAs depend on the physiological state of the cell, as influenced by genetic and environmental factors, the half-lives of other RNAs seem to be independent of cell physiology (6, 7).Much of the information available about RNA decay has been derived from study of RNAs encoded by a very limited number of genes. In fact, we can find published reports of RNA half-life for less than 25 of the 4,288 predicted ORFs in the E. coli genome. Current models of mRNA decay are based on this relatively limited experimental sampling of message turnover. To gain a broader understanding of the fate of mRNAs in E. coli we adapted DNA microarray methodology to measure the decay of each chromosomally encoded mRNA simultaneously.DNA microarrays containing sequences derived ...

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Section: Global Analysis Of Mrna Half-life In Cells Growing At Differsupporting

confidence: 73%

Global analysis of mRNA decay and abundance in Escherichia coli at single-gene resolution using two-color fluorescent DNA microarrays

Bernstein

Khodursky

Lin

et al. 2002

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

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773

711

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“…Similar RNA structures stabilize transcripts for ompA (Belasco et al, 1986;Bechhofer, 1993), papA, pufBA, and T4 phage gene 32 (Emory et al, 1992;Bechhofer, 1993). In E. coli, for example, mRNA half-lives range from a few seconds to 1 h, with an average lifetime of 2 to 4 min (Emory and Belasco, 1990).…”

Section: Discussionmentioning

confidence: 99%

Transcriptional and Posttranscriptional Control of mRNA fromlrtA, a Light-Repressed Transcript inSynechococcus sp. PCC 70021

Samartzidou

Widger

1998

Plant Physiology

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Transcription regulation and transcript stability of a lightrepressed transcript, lrtA, from the cyanobacterium Synechococcus sp. PCC 7002 were studied using ribonuclease protection assays. The transcript for lrtA was not detected in continuously illuminated cells, yet transcript levels increased when cells were placed in the dark. A lag of 20 to 30 min was seen in the accumulation of this transcript after the cells were placed in the dark. Transcript synthesis continued in the dark for 3 h and the transcript levels remained elevated for at least 7 h. The addition of 10 M rifampicin to illuminated cells before dark adaptation inhibited the transcription of lrtA in the dark. Upon the addition of rifampicin to 3-h dark-adapted cells, lrtA transcript levels remained constant for 30 min and persisted for 3 h. A 3-h half-life was estimated in the dark, whereas a 4-min half-life was observed in the light. Extensive secondary structure was predicted for this transcript within the 5 untranslated region, which is also present in the 5 untranslated region of lrtA from a different cyanobacterium, Synechocystis sp. PCC 6803. Evidence suggests that lrtA transcript stability is not the result of differences in ribonuclease activity from dark to light. Small amounts of lrtA transcript were detected in illuminated cells upon the addition of 25 g mL ؊1 chloramphenicol. The addition of chloramphenicol to dark-adapted cells before illumination allowed detection of the lrtA transcript for longer times in the light relative to controls without chloramphenicol. These results suggest that lrtA mRNA processing in the light is different from that in the dark and that protein synthesis is required for light repression of the lrtA transcript.

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“…This m e c h a n i s m has been demonstrated for chloroplast and Escherichia coli mRNAs; that is, s t e m -l o o p structures present in the 3'-UTRs of certain mRNAs are responsible for the relative stability of those messages (Burton et al 1983;Belasco et al 1986;Stem and Gruissem 1987). To test whether a stable secondary structure was sufficient to increase gene expression, a reporter m R N A was constructed with the origin of assembly sequence {OAS; Butler 19841 of TMV at its 3' terminus.…”

Section: The Presence Of Higher-order Structure At the 3' Terminus Domentioning

confidence: 99%

RNA pseudoknot domain of tobacco mosaic virus can functionally substitute for a poly(A) tail in plant and animal cells.

Gallie¹,

Walbot²

1990

Genes Dev.

144

119

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The genomes of many RNA viruses terminate in a tertiary structure similar to the L-conformation of tRNAs and this structure is recognized by many tRNA-specific enzymes such as aminoacyl-tRNA synthetase. Virtually the entire 3'-untranslated region (UTR) of tobacco mosaic virus (TMV) RNA is involved in an extended tertiary structure containing, in addition to a tRNA-like structure, a pseudoknot domain that lies immediately upstream. Although the functions of these structures are not well understood, they are essential to the virus. We demonstrate that the addition of the 204-base TMV 3'-untranslated region to foreign mRNA constructs can increase gene expression up to 100-fold compared to nonadenylated mRNA. The 3'-UTR of TMV was equal to or greater than a polyadenylated tail in enhancing gene expression in electroporated dicot and monocot protoplasts. The TMV 3'-UTR is functionally similar to a polyadenylated tail in that it increases mRNA stability and translation and must be positioned at the 3' terminus to function efficiently. Similar effects on expression were observed in Chinese hamster ovary cells, demonstrating that the sequence functions in a wide range of eukaryotes. When the extended tertiary structure was dissected, the upstream pseudoknot domain was found to be largely responsible for increasing expression. The inclusion of the tRNA-like structure, however, was important for full regulation.

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The stability of E. coli gene transcripts is dependent on determinants localized to specific mRNA segments

Cited by 181 publications

References 28 publications

Global analysis of mRNA decay and abundance in Escherichia coli at single-gene resolution using two-color fluorescent DNA microarrays

Global analysis of mRNA decay and abundance in Escherichia coli at single-gene resolution using two-color fluorescent DNA microarrays

Transcriptional and Posttranscriptional Control of mRNA fromlrtA, a Light-Repressed Transcript inSynechococcus sp. PCC 70021

RNA pseudoknot domain of tobacco mosaic virus can functionally substitute for a poly(A) tail in plant and animal cells.

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