A potential link between transgene silencing and poly(A) tails.
Argonaute proteins function in gene silencing induced by double-stranded RNA (dsRNA) in various organisms. In Drosophila, the Argonaute proteins AGO1 and AGO2 have been implicated in post-transcriptional gene-silencing (PTGS)/RNA interference (RNAi). In this study, we found that AGO1 and AGO2 depletion caused the accumulation of multicopied enhanced green fluorescence protein (EGFP) transgene transcripts in Drosophila S2 cells. Depletion of AGO1, the essential factor for miRNA biogenesis, led to an increased transcriptional rate of the transgenes. In contrast, depletion of AGO2, the essential factor for siRNA-directed RNAi, resulted in EGFP mRNA stabilization with concomitant shortening of the EGFP mRNA poly(A) tail. Our findings suggest that AGO1 and AGO2 mediate multicopied transgene silencing by different mechanisms. Intriguingly, Dicer2 depletion phenocopies AGO2 depletion, with an increase in EGFP protein levels and shortening of the EGFP mRNA poly(A) tail. The possibility that AGO2 and Dicer2 involve, at least in part, poly(A) length maintenance of transgene mRNA suggests a potentially important link between transgene silencing and poly(A) tails.
Institute for Genome Research, University of Tokushima, Tokushima 770-8503, Japan. siomim@genome.tokushima-u.ac.jp
Mikiko C
MC
Siomi
Hiroko
H
Tsukumo
Akira
A
Ishizuka
Tomoko
T
Nagami
Haruhiko
H
Siomi
D000818:Animals; D060565:Argonaute Proteins; D015153:Blotting, Western; D002478:Cells, Cultured; D004330:Drosophila / Q000166:cytology / Q000235:genetics / Q000378:metabolism; D029721:Drosophila Proteins / Q000172:deficiency / Q000378:metabolism; D039642:Eukaryotic Initiation Factors; D018628:Gene Dosage; D020868:Gene Silencing; D049452:Green Fluorescent Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D007700:Kinetics; D035683:MicroRNAs / Q000096:biosynthesis; D011061:Poly A / Q000737:chemistry*; D011233:Precipitin Tests; D034622:RNA Interference; D012323:RNA Processing, Post-Transcriptional; D012326:RNA Splicing; D012333:RNA, Messenger / Q000737:chemistry / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism; D034743:RNA-Induced Silencing Complex / Q000172:deficiency / Q000378:metabolism; D014158:Transcription, Genetic; D019076:Transgenes
C489654:AGO1 protein, Drosophila; C433807:AGO2 protein, Drosophila; D060565:Argonaute Proteins; D029721:Drosophila Proteins; D039642:Eukaryotic Initiation Factors; D035683:MicroRNAs; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D034743:RNA-Induced Silencing Complex; D049452:Green Fluorescent Proteins; D011061:Poly A