Highlights d The dsRNA binding protein SGS3 is required for microRNAmediated ribosome stalling d SGS3 interacts directly with the 3 0 end of the AGO-loaded microRNA on the target d SGS3-microRNA-AGO can pause ribosomes on cleavable and non-cleavable sites d Ribosome pausing correlates positively with production of secondary siRNAs

Secondary small interfering RNA (siRNA) production, triggered by primary small RNA targeting, is critical for proper development and antiviral defense in many organisms. RNA-dependent RNA polymerase (RDR) is a key factor in this pathway. However, how RDR specifically converts the targets of primary small RNAs into double-stranded RNA (dsRNA) intermediates remains unclear. Here, we develop an in vitro system that allows for dissection of the molecular mechanisms underlying the production of trans-acting siRNAs, a class of plant secondary siRNAs that play roles in organ development and stress responses. We find that a combination of the dsRNA-binding protein, SUPPRESSOR OF GENE SILENCING3; the putative nuclear RNA export factor, SILENCING DEFECTIVE5, primary small RNA, and Argonaute is required for physical recruitment of RDR6 to target RNAs. dsRNA synthesis by RDR6 is greatly enhanced by the removal of the poly(A) tail, which can be achieved by the cleavage at a second small RNA-binding site bearing appropriate mismatches. Importantly, when the complementarity of the base pairing at the second target site is too strong, the small RNA–Argonaute complex remains at the cleavage site, thereby blocking the initiation of dsRNA synthesis by RDR6. Our data highlight the light and dark sides of double small RNA targeting in the secondary siRNA biogenesis.

The path of ribosomes on mRNAs can be impeded by various obstacles. One such example is halting of ribosome movement by microRNAs, though the exact mechanism and physiological role remain unclear. Here, we find that ribosome stalling caused by the Argonaute-miRNA-SGS3 complex regulates production of secondary siRNA biogenesis in plants. We show that the double-stranded RNA-binding protein, SGS3, directly interacts with the 3′ end of the microRNA-Argonaute complex, resulting in ribosome stalling. Strikingly, microRNA-mediated ribosome stalling enhances production of secondary small interfering RNAs (siRNAs) from target mRNAs. Our results uncover a previously uncharacterized role for paused ribosomes in regulation of small RNA function that may have broad biological implications across the plant kingdom.

While both active and granular matter have been extensively studied, here we investigate what happens when we mix the two of them, in a model system combining microparticles and cell assemblies. On a substrate covered with polystyrene or silica microparticles, we notice two regimes in the spreading of a cell aggregate: light particles are pushed by the cells and form a ring, which bonds to the substrate by adhesion forces that oppose spreading, while for heavy particles, the cell monolayer spreads above the particle bed. In both cases, cell activity is transmitted to inert beads, leading to the formation of cell-microparticle aggregates, which flicker and diffuse. We then study the formation and the spreading of hybrid aggregates of microparticles and living cells and observe phase separations and jamming transitions. Our study may have implications on processes such as cancer metastasis and development, and may guide cancer therapies based on inert particles.