Regulation of AUXIN RESPONSE FACTOR3 by TAS3 ta-siRNA Affects Developmental Timing and Patterning in Arabidopsis

MicroRNAs (miRNAs) and trans-acting siRNAs (ta-siRNAs) in plants form through distinct pathways, although they function as negative regulators of mRNA targets by similar mechanisms [1-7]. Three ta-siRNA gene families (TAS1, TAS2, and TAS3) are known in Arabidopsis thaliana. Biogenesis of TAS3 ta-siRNAs, which target mRNAs encoding several AUXIN RESPONSE FACTORs (including ARF3/ETTIN and ARF4 [1, 8]) involves miR390-guided processing of primary transcripts, conversion of a precursor to dsRNA through RNA-DEPENDENT RNA POLYMERASE6 (RDR6) activity, and sequential DICER-LIKE4 (DCL4)-mediated cleavage events. We show that the juvenile-to-adult phase transition is normally suppressed by TAS3 ta-siRNAs, in an ARGONAUTE7-dependent manner, through negative regulation of ARF3 mRNA. Expression of a nontargeted ARF3 mutant (ARF3mut) in a wild-type background reproduced the phase-change phenotypes detected in rdr6-15 and dcl4-2 mutants, which lose all ta-siRNAs. Expression of either ARF3 or ARF3mut in rdr6-15 plants, in which both endogenous and transgenic copies of ARF3 were derepressed, resulted in further acceleration of phase change and severe morphological and patterning defects of leaves and floral organs. In light of the functions of ARF3 and ARF4 in organ asymmetry, these data reveal multiple roles for TAS3 ta-siRNA-mediated regulation of ARF genes in developmental timing and patterning.