Coordination of editing and splicing of glutamate receptor pre-mRNA.
Adenosine deaminase that acts on RNA, ADAR, catalyzes the conversion of adenosine into inosine within double-stranded RNA. This type of editing has mainly been found in genes involved in neurotransmission. Site-specific A to I modifications often require intronic sequences to create the double-stranded structure necessary for editing. A system was developed to investigate if editing and splicing of pre-mRNA are coordinated. We have focused on a selectively edited site (R/G) in the glutamate receptor subunit B pre-mRNA. This editing site is situated in close proximity to a 5' splice site. To ensure efficient splicing, the editing site, together with its natural 5' splice site, was fused to a 3' splice site of the major late transcript from adenovirus. In vitro, on a premade transcript, ADAR2 editing and splicing were found to interfere with each other. The stable stem-loop required for ADAR2 editing had a negative effect on in vitro splicing, possibly by sequestering the 5' splice site. Further, RNA helicase A was shown to overcome the splicing inhibition caused by ADAR2. In vivo, allowing cotranscriptional processing, the same construct was found to efficiently edit and splice without interference, suggesting that the two RNA processing events are coordinated.
Bratt
Eva
E
Department of Molecular Biology and Functional Genomics, Stockholm University, 106 91 Stockholm, Sweden.
Ohman
Marie
M
D000243:Adenosine Deaminase / Q000378:metabolism; D000818:Animals; D001324:Autoantigens / Q000378:metabolism; D053487:DEAD-box RNA Helicases; D066298:In Vitro Techniques; D009363:Neoplasm Proteins; D017393:RNA Editing / Q000502:physiology*; D020365:RNA Helicases / Q000378:metabolism; D012319:RNA Polymerase II / Q000378:metabolism; D022821:RNA Splice Sites; D012326:RNA Splicing / Q000502:physiology*; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D051381:Rats; D017470:Receptors, Glutamate / Q000235:genetics*
D001324:Autoantigens; D009363:Neoplasm Proteins; D022821:RNA Splice Sites; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D017470:Receptors, Glutamate; D012319:RNA Polymerase II; C567955:ADARB1 protein, human; D000243:Adenosine Deaminase; C098487:DHX9 protein, human; D053487:DEAD-box RNA Helicases; D020365:RNA Helicases