Boundary Formation is an Inherent Property of Mobile Small RNA Gradients

Small RNAs recently emerged as a new class of mobile instructive signals in development. Here, we investigate their mechanism of action and show that the gradients formed by mobile small RNAs generate sharply defined domains of target gene expression. By modulating the source of artificial miRNAs we show that boundary formation is an inherent property of the small RNA gradient itself. The threshold-based readout of such gradients is highly sensitive to small RNA levels at the source, allowing plasticity in the positioning of a target gene expression boundary. In addition to generating sharp expression domains of their immediate targets, the readouts of opposing small RNA gradients enable formation of stable and uniformly positioned developmental boundaries. These novel patterning properties of small RNAs are reminiscent of those of morphogens in animal systems. However, their exceptionally high specificity, direct mode of action, and the fully intrinsic nature of their gradients, distinguish mobile small RNAs from classical morphogens. Our findings present mobile small RNAs and their targets as highly portable and evolutionarily-tractable regulatory modules through which to create pattern in development and beyond. Overall design: Examination of artificial miRNA (miRGFP) expression and GFP-derived secondary siRNA production, in p35S:miRGFP and control p35S:3xNLS-GFP (no miRGFP) Arabidopsis seedlings, in an rdr6 null mutant background impaired for siRNA production.