Respective contributions of Arabidopsis DCL2 and DCL4 to RNA silencing.
Dicer proteins are central to the different mechanisms involving RNA interference. Plants have evolved multiple DICER-LIKE (DCL) copies, thus enabling functional diversification. In Arabidopsis, DCL2 and DCL4 process double-stranded RNA into 22 and 21 nucleotide small interfering (si)RNAs, respectively, and have overlapping functions with regards to virus and transgene silencing. Nonetheless, some studies have reported that dcl2 or dcl4 single mutations are sometimes sufficient to hinder silencing. To better dissect the role of DCL2 and DCL4, we analyzed silencing kinetics and efficiencies using different transgenic systems in single and double mutant backgrounds. The results indicate that DCL2 stimulates transitivity and secondary siRNA production through DCL4 while being sufficient for silencing on its own. Notably, silencing of 35S-driven transgenes functions more efficiently in dcl4 mutants, indicating that DCL4 mostly obscures DCL2 in wild-type plants. Nonetheless, in a dcl4 mutant compromised in phloem-originating silencing, ectopically expressed DCL2 allows restoration of silencing, suggesting that DCL2 is not, or poorly, expressed in phloem. Remarkably, this ectopic DCL2 contribution to phloem-originating silencing is dependent on the activity of RNA-DEPENDENT RNA POLYMERASE6. These results indicate that, despite differences in the silencing activity of their small RNA products, DCL2 and DCL4 mostly act redundantly yet hierarchically when present simultaneously.
Institut Jean-Pierre Bourgin, UMR1318, INRA, 78026, Versailles Cedex, France.
Jean-Sébastien
JS
Parent
Nathalie
N
Bouteiller
Taline
T
Elmayan
Hervé
H
Vaucheret
D017360:Arabidopsis / Q000235:genetics / Q000378:metabolism*; D029681:Arabidopsis Proteins / Q000235:genetics / Q000378:metabolism*; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism*; D020868:Gene Silencing / Q000502:physiology; D034622:RNA Interference / Q000502:physiology; D034741:RNA, Small Interfering / Q000235:genetics / Q000502:physiology; D043244:Ribonuclease III / Q000235:genetics / Q000378:metabolism*
D029681:Arabidopsis Proteins; D018797:Cell Cycle Proteins; D034741:RNA, Small Interfering; C504982:DCL4 protein, Arabidopsis; C511919:DCL2 protein, Arabidopsis; D043244:Ribonuclease III
Arabidopsis thaliana; DICER-LIKE proteins; RNA interference; plant genetics; small RNA; transgene silencing