Ribozyme cleavage reveals connections between mRNA release from the site of transcription and pre-mRNA processing.
We report a functional connection between splicing and transcript release from the DNA. A Pol II CTD mutant inhibited not only splicing but also RNA release from the site of transcription. A ribozyme situated downstream of the gene restored accurate splicing inhibited by the CTD mutant or a mutant poly(A) site, suggesting that cleavage liberates RNA from a niche that is inaccessible to splicing factors. Although ribozyme cleavage enhanced splicing, 3' end processing was impaired, indicating that an intact RNA chain linking the poly(A) site to Pol II is required for optimal processing. Surprisingly, poly(A)(-) beta-globin mRNA with a ribozyme-generated 3' end was exported to the cytoplasm. Ribozyme cleavage can therefore substitute for normal 3' end processing in stimulating splicing and mRNA export. We propose that mRNA biogenesis is coordinated by preventing splicing near the 3' end until the transcript is released by poly(A) site cleavage.
Bird
Gregory
G
Department of Biochemistry and Molecular Genetics, University of Colorado School of Medicine, UCHSC at Fitzsimons, Aurora, 80045, USA.
Fong
Nova
N
Gatlin
Jesse C
JC
Farabaugh
Susan
S
Bentley
David L
DL
D002460:Cell Line; D004247:DNA / Q000378:metabolism; D006801:Humans; D008957:Models, Genetic; D009154:Mutation; D009865:Oocytes / Q000378:metabolism; D011061:Poly A / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D026721:RNA 3' End Processing / Q000502:physiology; D012319:RNA Polymerase II / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012322:RNA Precursors / Q000378:metabolism*; D012326:RNA Splicing / Q000235:genetics / Q000502:physiology*; D016337:RNA, Catalytic / Q000378:metabolism*; D012333:RNA, Messenger / Q000096:biosynthesis / Q000378:metabolism*; D014158:Transcription, Genetic
D012322:RNA Precursors; D016337:RNA, Catalytic; D012333:RNA, Messenger; D011061:Poly A; D004247:DNA; D012319:RNA Polymerase II