title: Single-molecule imaging of transcriptionally coupled and uncoupled splicing.
abstract: Introns are removed from pre-mRNAs during transcription while the pre-mRNA is still tethered to the gene locus via RNA polymerase. However, during alternative splicing, it is important that splicing be deferred until all of the exons and introns involved in the choice have been synthesized. We have developed an in situ RNA imaging method with single-molecule sensitivity to define the intracellular sites of splicing. Using this approach, we found that the normally tight coupling between transcription and splicing is broken in situations where the intron's polypyrimidine tract is sequestered within strong secondary structures. We also found that in two cases of alternative splicing, in which certain exons are skipped due to the activity of the RNA-binding proteins Sxl and PTB, splicing is uncoupled from transcription. This uncoupling occurs only on the perturbed introns, whereas the preceding and succeeding introns are removed cotranscriptionally. PAPERCLIP:
authors: Public Health Research Institute, New Jersey Medical School, University of Medicine and Dentistry of New Jersey, Newark, NJ 07103, USA.
authors: Diana Y
authors: DY
authors: Vargas
authors: Khyati
authors: K
authors: Shah
authors: Mona
authors: M
authors: Batish
authors: Michael
authors: M
authors: Levandoski
authors: Sourav
authors: S
authors: Sinha
authors: Salvatore A E
authors: SA
authors: Marras
authors: Paul
authors: P
authors: Schedl
authors: Sanjay
authors: S
authors: Tyagi
mesh_terms: D017398:Alternative Splicing; D000818:Animals; D001483:Base Sequence; D029721:Drosophila Proteins / Q000235:genetics; D004331:Drosophila melanogaster / Q000235:genetics*; D005091:Exons; D005786:Gene Expression Regulation; D016762:Genes, fos; D006367:HeLa Cells; D006801:Humans; D057054:Molecular Imaging / Q000379:methods*; D008969:Molecular Sequence Data; D009154:Mutation; D038941:Polypyrimidine Tract-Binding Protein / Q000378:metabolism; D012322:RNA Precursors / Q000737:chemistry / Q000378:metabolism; D012326:RNA Splicing; D016601:RNA-Binding Proteins / Q000235:genetics; D014158:Transcription, Genetic
chemical_list: D029721:Drosophila Proteins; D012322:RNA Precursors; D016601:RNA-Binding Proteins; C067777:Sxl protein, Drosophila; D038941:Polypyrimidine Tract-Binding Protein