1. SR Protein Family Members Display Diverse Activities in the Formation of Nascent and Mature mRNPs In Vivo
- Author
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Aparna K. Sapra, Jarod Rollins, Minna-Liisa Änkö, Mike Lorenz, Eva Maria Weiland, Ina Poser, Karla M. Neugebauer, Inna Grishina, and Marta Pabis
- Subjects
Chromatin Immunoprecipitation ,Chromosomes, Artificial, Bacterial ,Transcription, Genetic ,RNase P ,RNA Splicing ,Recombinant Fusion Proteins ,Green Fluorescent Proteins ,RNA polymerase II ,SR protein ,Transcription (biology) ,Fluorescence Resonance Energy Transfer ,Humans ,Promoter Regions, Genetic ,Molecular Biology ,Gene ,Messenger RNA ,biology ,Genes, fos ,Nuclear Proteins ,RNA-Binding Proteins ,RNA ,Cell Biology ,Molecular biology ,Cell biology ,Ribonucleoproteins ,RNA splicing ,biology.protein ,HeLa Cells - Abstract
The SR proteins are a family of pre-mRNA splicing factors with additional roles in gene regulation. To investigate individual family members in vivo, we generated a comprehensive panel of stable cell lines expressing GFP-tagged SR proteins under endogenous promoter control. Recruitment of SR proteins to nascent FOS RNA was transcription dependent and RNase sensitive, with unique patterns of accumulation along the gene specified by the RNA recognition motifs (RRMs). In addition, all SR protein interactions with Pol II were RNA dependent, indicating that SR proteins are not preassembled with Pol II. SR protein interactions with RNA were confirmed in situ by FRET/FLIM. Interestingly, SC35-GFP also exhibited FRET with DNA and failed to associate with cytoplasmic mRNAs, whereas all other SR proteins underwent nucleocytoplasmic shuttling and associated with specific nuclear and cytoplasmic mRNAs. Because different constellations of SR proteins bound nascent, nuclear, and cytoplasmic mRNAs, mRNP remodeling must occur throughout an mRNA's lifetime.
- Published
- 2009