Assembly and disassembly of the Drosophila RNA polymerase II complex during transcription

Initiation of mRNA synthesis by RNA polymerase II occurs by a multistep pathway that involves the polymerase along with several auxiliary factors. Here I describe an analysis of the kinetics and mechanism of action of Drosophila RNA polymerase II by using the detergent Sarkosyl to inhibit different...

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Bibliographic Details
Published in:The Journal of biological chemistry 1990-02, Vol.265 (5), p.2624-2631
Main Author: Kadonaga, J.T
Format: Article
Language:English
Subjects:
Animals
Base Sequence
Biological and medical sciences
Detergents - pharmacology
Diptera
DNA
DNA - genetics
DNA-directed RNA polymerase
Drosophila
Drosophila - enzymology
Drosophilidae
Embryo, Nonmammalian
Fundamental and applied biological sciences. Psychology
kinetics
Molecular and cellular biology
Molecular genetics
Molecular Sequence Data
Oligonucleotide Probes
Plasmids
Promoter Regions, Genetic
RNA Polymerase II - metabolism
Sarcosine - analogs & derivatives
Sarcosine - pharmacology
synthesis
Templates, Genetic
Transcription, Genetic - drug effects
Transcription. Transcription factor. Splicing. Rna processing
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Summary:Initiation of mRNA synthesis by RNA polymerase II occurs by a multistep pathway that involves the polymerase along with several auxiliary factors. Here I describe an analysis of the kinetics and mechanism of action of Drosophila RNA polymerase II by using the detergent Sarkosyl to inhibit different steps in the pathway leading to transcription initiation. The template DNA and Drosophila transcription factors assemble into a partially Sarkosyl-resistant initiation complex with a kinetically first order time course characterized by a t1/2 of about 3 min, and subsequent conversion of the initiation complex into the elongating complex occurs rapidly (less than 5 s). During the formation of the first two phosphodiester bonds, there is a stepwise transition from the initiation complex to the 0.25% Sarkosyl-resistant elongating polymerase. In contrast to the mammalian RNA polymerase II systems, the Drosophila extracts possess the ability to carry out many rounds (approximately 10) of transcription in vitro. Finally, the transcription factors are committed to the template DNA as an initiation complex, but then no longer appear to remain committed to the DNA after initiation of transcription. This finding suggests that there is complete assembly and disassembly of the transcription complex during each round of transcription by RNA polymerase II.
ISSN:0021-9258
1083-351X
DOI:10.1016/s0021-9258(19)39847-3