RNA editing and regulation of Drosophila 4f-rnp expression by sas-10 antisense readthrough mRNA transcripts.
We have previously described an example of extensively A-to-G edited cDNA derived from adult heads of the fruitfly Drosophila melanogaster. In that study, the source of the predicted antisense RNA pairing strand for template recognition by dADAR editase was not identified, and the biological significance of the observed hyperediting was not known. Here, we address each of these questions. 4f-rnp and sas-10 are closely adjacent X-linked genes located on opposite DNA strands that produce convergent transcripts. We show that developmentally regulated antisense sas-10 readthrough mRNA arises by activation of an upstream promoter P2 during the late embryo stage of fly development. The sas-10 readthrough transcripts pair with 4f-rnp mRNA to form double-stranded molecules, as indicated by A-to-G editing observed in both RNA strands. It would be predicted that perfect RNA duplexes would be targeted for modification/degradation by enzyme pathways that recognize double-stranded RNAs, leading to decline in 4f-rnp mRNA levels, and this is what we observe. The observation using quantitative RT-PCR that sas-10 readthrough and 4f-rnp transcript levels are inversely related suggests a role for the antisense RNA in posttranscriptional regulation of 4f-rnp gene expression during development. Potential molecular mechanisms that could lead to this result are discussed, one of which is targeted transcript degradation via the RNAi pathway. Insofar as the dADAR editase and RNAi pathways are known to be constitutive in this system, it is likely that control of antisense RNA transcription is the rate-limiting factor. The results provide insight into roles of naturally occurring antisense RNAs in regulation of eukaryotic gene expression.
Peters
Nick T
NT
Department of Zoology, Miami University, Oxford, Ohio 45056, USA.
Rohrbach
Justin A
JA
Zalewski
Brian A
BA
Byrkett
Colleen M
CM
Vaughn
Jack C
JC
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D029721:Drosophila Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism; D004331:Drosophila melanogaster / Q000196:embryology / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism; D018507:Gene Expression Regulation, Developmental; D017344:Genes, Insect; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000096:biosynthesis / Q000235:genetics*; D011401:Promoter Regions, Genetic; D017393:RNA Editing; D020871:RNA Stability; D016372:RNA, Antisense / Q000502:physiology*; D012330:RNA, Double-Stranded / Q000378:metabolism; D012333:RNA, Messenger / Q000096:biosynthesis / Q000378:metabolism*; D016601:RNA-Binding Proteins; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012261:Ribonucleoproteins / Q000235:genetics* / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D017386:Sequence Homology, Amino Acid; D019914:Taq Polymerase; D014158:Transcription, Genetic
D029721:Drosophila Proteins; D009687:Nuclear Proteins; D016372:RNA, Antisense; D012330:RNA, Double-Stranded; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; C100381:RNA-binding protein 4F, Drosophila; D012261:Ribonucleoproteins; D029701:Saccharomyces cerevisiae Proteins; C475181:Sas10 protein, Drosophila; C114039:Sas10 protein, S cerevisiae; D019914:Taq Polymerase