A functional interaction between the carboxy-terminal domain of RNA polymerase II and pre-mRNA splicing.
In the preceding study we found that Sm snRNPs and SerArg (SR) family proteins co-immunoprecipitate with Pol II molecules containing a hyperphosphorylated CTD (Kim et al., 1997). The association between Pol IIo and splicing factors is maintained in the absence of pre-mRNA, and the polymerase need not be transcriptionally engaged (Kim et al., 1997). The latter findings led us to hypothesize that a phosphorylated form of the CTD interacts with pre-mRNA splicing components in vivo. To test this idea, a nested set of CTD-derived proteins was assayed for the ability to alter the nuclear distribution of splicing factors, and to interfere with splicing in vivo. Proteins containing heptapeptides 1-52 (CTD52), 1-32 (CTD32), 1-26 (CTD26), 1-13 (CTD13), 1-6 (CTD6), 1-3 (CTD3), or 1 (CTD1) were expressed in mammalian cells. The CTD-derived proteins become phosphorylated in vivo, and accumulate in the nucleus even though they lack a conventional nuclear localization signal. CTD52 induces a selective reorganization of splicing factors from discrete nuclear domains to the diffuse nucleoplasm, and significantly, it blocks the accumulation of spliced, but not unspliced, human beta-globin transcripts. The extent of splicing factor disruption, and the degree of inhibition of splicing, are proportional to the number of heptapeptides added to the protein. The above results indicate a functional interaction between Pol II's CTD and pre-mRNA splicing.
Du
L
L
Department of Pathology, Yale University School of Medicine, New Haven, Connecticut 06510, USA.
Warren
S L
SL
D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D005347:Fibroblasts; D005914:Globins / Q000235:genetics; D000882:Haplorhini; D006367:HeLa Cells; D006801:Humans; D007668:Kidney; D008969:Molecular Sequence Data; D010766:Phosphorylation; D012319:RNA Polymerase II / Q000235:genetics / Q000378:metabolism*; D012322:RNA Precursors / Q000235:genetics / Q000378:metabolism*; D012326:RNA Splicing / Q000502:physiology*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D017411:Ribonucleoproteins, Small Nuclear / Q000378:metabolism
D012322:RNA Precursors; D011993:Recombinant Fusion Proteins; D017411:Ribonucleoproteins, Small Nuclear; D005914:Globins; D012319:RNA Polymerase II