Expression of most RNA polymerase II transcripts requires the coordinated execution of transcription, splicing, and 3′ processing. We have previously shown that upon transcriptional activation of a gene in vivo, pre-mRNA splicing factors are recruited from nuclear speckles, in which they are concentrated, to sites of transcription (Misteli, T., J.F. Cáceres, and D.L. Spector. 1997. Nature. 387:523–527). This recruitment process appears to spatially coordinate transcription and pre-mRNA splicing within the cell nucleus. Here we have investigated the molecular basis for recruitment by analyzing the recruitment properties of mutant splicing factors. We show that multiple protein domains are required for efficient recruitment of SR proteins from nuclear speckles to nascent RNA. The two types of modular domains found in the splicing factor SF2/ ASF exert distinct functions in this process. In living cells, the RS domain functions in the dissociation of the protein from speckles, and phosphorylation of serine residues in the RS domain is a prerequisite for this event. The RNA binding domains play a role in the association of splicing factors with the target RNA. These observations identify a novel in vivo role for the RS domain of SR proteins and suggest a model in which protein phosphorylation is instrumental for the recruitment of these proteins to active sites of transcription in vivo.
Serine Phosphorylation of SR Proteins Is Required for Their Recruitment to Sites of Transcription In Vivo
T. Misteli dedicates this paper to the memory of Thomas Kreis.
T. Misteli was supported by the Human Frontiers Science Program and the Roche Research Foundation. A.R. Krainer was supported by National Cancer Institute grant CA13106, M.F. Wilkinson was supported by National Institutes of Health (NIH) grant GM39586 and National Science Foundation grant MCB-9307963, and D.L. Spector was supported by NIH grant GM42694.
Address all correspondence to D.L. Spector, Cold Spring Harbor Laboratory, Cold Spring Harbor, NY 11724. Tel.: (516) 367-8456. Fax: (516) 367-8876. E-mail: [email protected]
Tom Misteli, Javier F. Cáceres, Jade Q. Clement, Adrian R. Krainer, Miles F. Wilkinson, David L. Spector; Serine Phosphorylation of SR Proteins Is Required for Their Recruitment to Sites of Transcription In Vivo . J Cell Biol 19 October 1998; 143 (2): 297–307. doi: https://doi.org/10.1083/jcb.143.2.297
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