Abstract
MicroRNAs (miRNAs) are endogenous antisense regulators that trigger endonucleolytic mRNA cleavage, translational repression, and/or mRNA decay. miRNA-mediated gene regulation is important for numerous biological pathways, yet the underlying mechanisms are still under rigorous investigation. Here we identify human UPF1 (hUPF1) as a protein that contributes to RNA silencing. When hUPF1 is knocked down, miRNA targets are upregulated. The depletion of hUPF1 also increases the off-target messages of small interfering RNAs (siRNAs), which are imperfectly complementary to transfected siRNAs. Conversely, when overexpressed, wild-type hUPF1 downregulates miRNA targets. The helicase domain mutant of hUPF1 fails to suppress miRNA targets. hUPF1 interacts with human Argonaute 1 (hAGO1) and hAGO2 and colocalizes with hAGO1 and hAGO2 in processing bodies, which are known to be the sites for translational repression and mRNA destruction. We further find that the amounts of target messages bound to hAGO2 are reduced when hUPF1 is depleted. Our data thus suggest that hUPF1 may participate in RNA silencing by facilitating the binding of the RNA-induced silencing complex to the target and by accelerating the decay of the mRNA.
SUPPLEMENTAL MATERIAL
Supplemental material for this article may be found at http://mcb.asm.org/ .
ACKNOWLEDGMENTS
We specially thank Lynne Maquat and Yoon Ki Kim for hUPF1 cDNA and NMD reporters. We also thank Elisa Izaurralde and Matthias Hentze for critical comments, Zissimos Mourelatos for Argonaute cDNAs, Gideon Dreyfuss for anti-Y14 and anti-hnRNP A1 antibodies, Tom Tuschl for Argonaute cDNAs, J. Lykke-Andersen and Mikiko C. Siomi for antibodies, Marvin J. Fritzler for anti-GW182 antibody, Hyunsook Lee for anti-myc antibody, and Young-Kook Kim for discussion and technical help.
This work was supported by Creative Research Initiatives (R16-2007-073-01000-0). H.J., J.H., and K.-H.Y. were supported by the BK21 studentship. H.J. was supported by the Seoul Science Fellowship.