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Molecular and Cellular Biology, August 2003, p. 5768-5779, Vol. 23, No. 16
0270-7306/03/$08.00+0     DOI: 10.1128/MCB.23.16.5768-5779.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Cotranscriptional Recruitment of the U1 snRNP to Intron-Containing Genes in Yeast

Kimberly M. Kotovic, Daniel Lockshon,{dagger} Lamia Boric,{ddagger} and Karla M. Neugebauer*

Max Planck Institute of Molecular Cell Biology and Genetics, 01307 Dresden, Germany, and Department of Neurology, University of Washington School of Medicine, Seattle, Washington 98195

Received 13 March 2003/ Returned for modification 21 April 2003/ Accepted 14 May 2003

Evidence that pre-mRNA processing events are temporally and, in some cases, mechanistically coupled to transcription has led to the proposal that RNA polymerase II (Pol II) recruits pre-mRNA splicing factors to active genes. Here we address two key questions raised by this proposal: (i) whether the U1 snRNP, which binds to the 5' splice site of each intron, is recruited cotranscriptionally in vivo and, (ii) if so, where along the length of active genes the U1 snRNP is concentrated. Using chromatin immunoprecipitation (ChIP) in yeast, we show that elevated levels of the U1 snRNP were specifically detected in gene regions containing introns and downstream of introns but not along the length of intronless genes. In contrast to capping enzymes, which bind directly to Pol II, the U1 snRNP was poorly detected in promoter regions, except in genes harboring promoter-proximal introns. Detection of the U1 snRNP was dependent on RNA synthesis and was abolished by intron removal. Microarray analysis revealed that intron-containing genes were preferentially selected by ChIP with the U1 snRNP. Thus, U1 snRNP accumulation at genes correlated with the presence and position of introns, indicating that introns are necessary for cotranscriptional U1 snRNP recruitment and/or retention.

* Corresponding author. Mailing address: Max Planck Institute for Molecular Cell Biology and Genetics, Pfotenhauerstrasse 108, 01307 Dresden, Germany. Phone: 49(0)351-210-2589. Fax: 49(0)351-210-1209. E-mail: neugebauer{at}mpi-cbg.de.

{dagger} Present address: Department of Biochemistry, University of Washington, Seattle, WA 98195.

{ddagger} Present address: Oregon Health and Science University, Portland, OR 97239.

Molecular and Cellular Biology, August 2003, p. 5768-5779, Vol. 23, No. 16
0022-538X/03/$08.00+0     DOI: 10.1128/MCB.23.16.5768-5779.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



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