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Molecular and Cellular Biology, October 2002, p. 6706-6718, Vol. 22, No. 19
0270-7306/02/$04.00+0     DOI: 10.1128/MCB.22.19.6706-6718.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

In Vivo Kinetics of mRNA Splicing and Transport in Mammalian Cells

A. Audibert,{dagger} D. Weil,* and F. Dautry

CNRS-UPR 1983, Institut André Lwoff, 94801 Villejuif Cedex, France

Received 15 March 2002/ Returned for modification 14 May 2002/ Accepted 20 June 2002

The kinetics of pre-mRNA processing in living cells is poorly known, preventing a detailed analysis of the regulation of these reactions. Using tetracycline-regulated promoters we performed, during a transcriptional induction, a complete analysis of the maturation of two cellular mRNAs, those for LT-{alpha} and ß-globin. In both cases, splicing was appropriately described by first-order reactions with corresponding half-lives ranging between 0.4 and 7.5 min, depending on the intron. Transport also behaved as a first-order reaction during the early phase of ß-globin expression, with a nuclear dwelling time of 4 min. At a later time, analysis was prevented by the progressive accumulation within the nucleus of mature mRNA not directly involved in export. Our results further establish for these genes that (i) splicing components are never limiting, even when expression is induced in naive cells, (ii) there is no significant RNA degradation during splicing and transport, and (iii) precursor-to-product ratios at steady state can be used for the determination of splicing rates. Finally, the comparison between the kinetics of splicing during transcriptional induction and during transcriptional shutoff reveals a novel coupling between transcription and splicing.

* Corresponding author. Mailing address: CNRS-UPR 1983, Institut André Lwoff, 7 rue Guy Moquet, 94801 Villejuif Cedex, France. Phone: 33 1 4958 3380. Fax: 33 1 4958 3578. E-mail: weil{at}infobiogen.fr.

{dagger} Present address: UMR 7622, Université Pierre et Marie Curie, 75005 Paris, France.

Molecular and Cellular Biology, October 2002, p. 6706-6718, Vol. 22, No. 19
0022-538X/02/$04.00+0     DOI: 10.1128/MCB.22.19.6706-6718.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.



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