In VitroStudies of Transcript Initiation byEscherichia coliRNA Polymerase. 1. RNA Chain Initiation, Abortive Initiation, and Promoter Escape at Three Bacteriophage Promoters†

Biochemistry ◽  
2003 ◽  
Vol 42 (13) ◽  
pp. 3777-3786 ◽  
Author(s):  
Lilian M. Hsu ◽  
Nam V. Vo ◽  
Caroline M. Kane ◽  
Michael J. Chamberlin
Keyword(s):  
Chain Initiation ◽  
Promoter Escape ◽  
Transcript Initiation ◽  
Abortive Initiation

scholarly journals Probing the Transcription Mechanisms of Reovirus Cores with Molecules That Alter RNA Duplex Stability

2009 ◽  
Vol 83 (11) ◽  
pp. 5659-5670 ◽  
Author(s):  
Alexander A. Demidenko ◽  
Max L. Nibert
Keyword(s):  
Small Molecules ◽  
Dimethyl Sulfoxide ◽  
Active Fraction ◽  
Core Particle ◽  
Double Stranded Rna ◽  
Promoter Escape ◽  
Duplex Stability ◽  
Rna Duplexes ◽  
Dsrna Viruses ◽  
Transcript Initiation

ABSTRACT The mammalian reovirus (MRV) genome comprises 10 double-stranded RNA (dsRNA) segments, packaged along with transcriptase complexes inside each core particle. Effects of four small molecules on transcription by MRV cores were studied for this report, chosen for their known capacities to alter RNA duplex stability. Spermidine and spermine, which enhance duplex stability, inhibited transcription, whereas dimethyl sulfoxide and trimethylglycine, which attenuate duplex stability, stimulated transcription. Different mechanisms were identified for inhibition or activation by these molecules. With spermidine, one round of transcription occurred normally, but subsequent rounds were inhibited. Thus, inhibition occurred at the transition between the end of elongation in one round and initiation in the next round of transcription. Dimethyl sulfoxide or trimethylglycine, on the other hand, had no effect on transcription by a constitutively active fraction of cores in each preparation but activated transcription in another fraction that was otherwise silent for the production of elongated transcripts. Activation of this other fraction occurred at the transition between transcript initiation and elongation, i.e., at promoter escape. These results suggest that the relative stability of RNA duplexes is most important for certain steps in the particle-associated transcription cycles of dsRNA viruses and that small molecules are useful tools for probing these and probably other steps.

scholarly journals Direct Observation of Abortive Initiation and Promoter Escape within Single Immobilized Transcription Complexes

2006 ◽  
Vol 90 (4) ◽  
pp. 1419-1431 ◽  
Author(s):  
Emmanuel Margeat ◽  
Achillefs N. Kapanidis ◽  
Philip Tinnefeld ◽  
You Wang ◽  
Jayanta Mukhopadhyay ◽  
...  
Keyword(s):  
Direct Observation ◽  
Promoter Escape ◽  
Transcription Complexes ◽  
Abortive Initiation

scholarly journals Distinct Roles for the Helicases of TFIIH in Transcript Initiation and Promoter Escape

2000 ◽  
Vol 275 (4) ◽  
pp. 2532-2538 ◽  
Author(s):  
John Bradsher ◽  
Frederic Coin ◽  
Jean-Marc Egly
Keyword(s):  
Promoter Escape ◽  
Transcript Initiation

scholarly journals Strong Natural Pausing by RNA Polymerase II within 10 Bases of Transcription Start May Result in Repeated Slippage and Reextension of the Nascent RNA

2002 ◽  
Vol 22 (1) ◽  
pp. 30-40 ◽  
Author(s):  
Mahadeb Pal ◽  
Donal S. Luse
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Rna Synthesis ◽  
Early Stage ◽  
Transcription Start ◽  
Promoter Escape ◽  
Transcript Length ◽  
Template Sequence ◽  
Transcription Complexes ◽  
Transcript Initiation

ABSTRACT We find that immediately following transcript initiation, RNA polymerase II pauses at several locations even in the presence of relatively high (200 μM) levels of nucleoside triphosphates. Strong pauses with half-lives of >30 s were observed at +7, +18/19, and about +25 on the template used in these experiments. We show that the strong pause at +7, after the synthesis of 5′-ACUCUCU, leads to repeated cycles of upstream slippage of the RNA-DNA hybrid followed by re-pairing with the DNA and continued RNA synthesis. The resulting transcripts are 2, 4, and 6 bases longer than predicted by the template sequence. Slippage is efficient when transcription is primed with the +1/+2 (ApC) dinucleotide, and it occurs at even higher levels with the +2/+3 primer (CpU). Slippage can occur at high levels with ATP initiation, but priming with CpA (−1/+1) supports very little slippage. This latter result is not simply an effect of transcript length at the point of pausing. Slippage can also occur with a second template on which the polymerase can be paused after synthesizing ACUCU. Slippage is not reduced by an ATP analog that blocks promoter escape, but it is inhibited by substitution of 5Br-U for U in the RNA. Our results reveal an unexpected flexibility of RNA polymerase II ternary complexes during the very early stage of transcription, and they suggest that initiation at different locations within the same promoter gives rise to transcription complexes with different properties.

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