Recognition of prokaryotic transcription terminators by spinach chloroplast RNA polymerase

Chen, Liang-Jwu; Orozco, Emil M.

doi:10.1093/nar/16.17.8411

Cited by 65 publications

(64 citation statements)

References 32 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Plasmid DNA constructions The plasmid pTZ19-PLSTa contains the 5'-end of the spinach rbcL gene promoter (PLS) and the terminator from the threonine attenuator region (Ta) as previously described (1). The terminator variants were originally from the pTZ-19thr variants and pTZ-19T series as described by Jeng et al (13).…”

Section: Methodsmentioning

confidence: 99%

“…Besides (13) and YNANN mutants into the BamHI site of plasmid pTZ19-PLSTa (1). The sizes for transcripts initiating at the 'PLS' promoter and terminated at the terminator variants (227 nt) or at the wild-type terminator (335 nt) were indicated.…”

Section: Methodsmentioning

confidence: 99%

“…Such structures were initially presumed to be transcription terminators, analogous to bacterial factor-independent terminators. However, when they were analyzed in a homologous in vitro transcription system, no significant termination activity was observed (1,2). It was subsequently concluded that the 3' ends of the rbcL, atpB, psbA, petD and rpoA transcripts from spinach chloroplast are primarily the result of RNA processing and not transcription termination (2,3).…”

Section: Introductionmentioning

confidence: 99%

“…In contrast, a recent report indicates that the stem-loop structure of the 3' ends of the Chlamydomonas chloroplast rbcL and psaB genes appear to have significant termination activity in vivo (4). In addition, the prokaryotic factor-independent terminators that contain a typical d(G+C)-rich region of dyad symmetry and are followed by a poly(T) tract in the sense strand, such as the terminator for the threonine attenuator, thra from Escherichia coli, efficiently terminate transcription in vitro by spinach chloroplast RNA polymerase (1,5). This indicates that the transcription termination process in chloroplasts may have at least some features in common with the mechanism used in prokaryotes and that the stem-loop-dependent termination is probably sometimes functional in chloroplasts.…”

Section: Introductionmentioning

confidence: 99%

See 3 more Smart Citations

Transcription termination at the Escherichia coli thra terminator by spinach chloroplast RNA polymerase in vitro is influenced by downstream DNA sequences

Chen¹,

Liang²,

Jeng³

et al. 1995

Nucl Acids Res

Self Cite

View full text Add to dashboard Cite

We have investigated the mechanism of transcription termination in vitro by spinach chloroplast RNA polymerase using templates encoding variants of the transcription-termination structure (attenuator) of the regulatory region of the threonine (thi) operon of Escherichia coli. Fourteen sequence variants located within its d(G+C) stem-loop and d(A+T)-rich regions were studied. We found that the helix integrity in the stem-loop structure is necessary for termination but that its stability is not directly correlated with termination efficiency. The sequence of the G+C stem-loop itself also influences termination. Moreover, the dA template stretch at the 3' end of the terminator plays a major role in termination efficiency, but base pairing between the A and U tract of the transcript does not From the studies using deletion variants and a series of mutants that alter the sequences immediately downstream from the transcription termination site, we found that termination of transcription by spinach chloroplast RNA polymerase was also modulated by downstream DNA sequences in a sequence-specific manner. The second base immediately following the poly(T) tract is crucial for determining the termination efficiency by chloroplast RNA polymerase, but not of the T7 or E.coli enzymes.

show abstract

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Transcription termination at the Escherichia coli thra terminator by spinach chloroplast RNA polymerase in vitro is influenced by downstream DNA sequences

Chen¹,

Liang²,

Jeng³

et al. 1995

Nucl Acids Res

Self Cite

View full text Add to dashboard Cite

show abstract

“…23 The produced RNA was reverse transcribed in the presence of 500 µM dATP, dGTP and dTTP. For dCTP three different solutions were used.…”

Section: Samples and Preparationmentioning

confidence: 99%

Measuring, in Solution, Multiple-Fluorophore Labeling by Combining Fluorescence Correlation Spectroscopy and Photobleaching

et al. 2010

View full text Add to dashboard Cite

Determining the number of fluorescent entities that are coupled to a given molecule (DNA, protein, etc.) is a key point of numerous biological studies, especially those based on a single molecule approach. Reliable methods are important, in this context, not only to characterize the labeling process, but also to quantify interactions, for instance within molecular complexes. We combined Fluorescence Correlation Spectroscopy (FCS) and photobleaching experiments to measure the effective number of molecules and the molecular brightness as a function of the total fluorescence count rate on solutions of cDNA (containing a few percent of C bases labeled with Alexa Fluor 647). Here, photobleaching is used as a control parameter to vary the experimental outputs (brightness and number of molecules).Assuming a Poissonian distribution of the number of fluorescent labels per cDNA, the FCSphotobleaching data could be easily fit to yield the mean number of fluorescent labels per cDNA strand ( 2). This number could not be determined solely on the basis of the cDNA brightness, because of both the statistical distribution of the number of fluorescent labels and their unknown brightness when incorporated in cDNA. The statistical distribution of the number of fluorophores labeling cDNA was confirmed by analyzing the photon count distribution (with the cumulant method), which showed clearly that the brightness of cDNA strands varies from one molecule to the other. We also performed complementary continuous photobleaching experiments and found that the photobleaching decay rate of Alexa Fluor 647 in the excited state decreases by about 30% when incorporated into cDNA, while its non radiative decay rate is increased such that the brightness of individual Alexa labels is decreased by 25% compared to free Alexa dyes.

show abstract

Light-induced switch in barley psbD-psbC promoter utilization: a novel mechanism regulating chloroplast gene expression.

1990

View full text Add to dashboard Cite

The synthesis of reaction center protein D2 and mRNAs which encode this protein are differentially maintained at high levels in mature barley chloroplasts. To understand the differential maintenance of psbD mRNA abundance, we have studied the transcription and the RNAs produced from the psbD‐psbC operon in plastids of light and dark‐grown barley seedlings. Ten psbD‐psbC RNAs synthesized in dark‐grown barley share four different 5′‐ends, two of which arise by transcription initiation, and one of which is generated by 5′‐processing of longer psbD‐psbC transcripts. Illumination of dark‐grown barley causes the decline of these ten transcripts, and the accumulation of two different psbD‐psbC RNAs. Capping assays, in vitro transcription and RNA processing experiments and treatment of plants with tagetitoxin (a selective inhibitor of chloroplast transcription), indicate that the light‐induced transcripts arise by transcription initiation. Run‐on transcription and RNA quantitation experiments provide evidence that both light‐induced transcription and RNA stability play roles in the accumulation of the light‐induced RNAs. These data document a novel mechanism for regulating plastid gene expression involving a light‐induced switch in psbD‐psbC promoter utilization.

show abstract

Recognition of prokaryotic transcription terminators by spinach chloroplast RNA polymerase

Cited by 65 publications

References 32 publications

Transcription termination at the Escherichia coli thra terminator by spinach chloroplast RNA polymerase in vitro is influenced by downstream DNA sequences

Transcription termination at the Escherichia coli thra terminator by spinach chloroplast RNA polymerase in vitro is influenced by downstream DNA sequences

Measuring, in Solution, Multiple-Fluorophore Labeling by Combining Fluorescence Correlation Spectroscopy and Photobleaching

Light-induced switch in barley psbD-psbC promoter utilization: a novel mechanism regulating chloroplast gene expression.

Contact Info

Product

Resources

About