The influence of the nucleic acid secondary structure on the fast (1 h) formation of bright red emissive silver nanoclusters (AgNCs) in a DNA sequence (DNA-12nt-RED-160), designed for the detection of a microRNA sequence (RNA-miR160), was investigated. The findings show that especially the propensity for mismatch self-dimer formation of the DNA probes can be a good indicator for the creation and stabilization of red emissive AgNCs. Also, the role of the thermal stability of the secondary DNA structures (mismatch self-dimer and hairpin monomers) and the observed AgNC red emission intensity were investigated. These findings can form the basis for a rationale to design new red emissive AgNC-based probes. As an example, a bright red emissive AgNC-based DNA probe was designed for RNA-miR172 detection. The latter opens the possibility to create a variety of AgNC-based DNA probes for the specific detection of plant and animal miRNAs.

Constitutive photomorphogenic 1 (COP1) is a RING-finger E3 ligase that plays a central role in photomorphogenesis by destabilizing many light-regulated transcription factors and photoreceptors. Here, we reveal a novel function for COP1 E3 ligase in controlling global miRNA biogenesis in Arabidopsis thaliana. In cop1 mutants, the level of miRNAs is dramatically reduced because of the diminution of HYPONASTIC LEAVES 1 (HYL1), an RNA-binding protein required for precise miRNA processing. HYL1 is destabilized by an unidentified protease, which we tentatively call protease X, that specifically cleaves the N-terminal region from HYL1, thus neutralizing its function. Our results further show that the cytoplasmic partitioning of COP1 under light is essential to protect HYL1 against protease X. Taken together, we suggest a novel regulatory network involving HYL1, protease X, COP1 and light signalling that is indispensable for miRNA biogenesis in Arabidopsis thaliana.

MicroRNAs (miRNAs) are small regulatory RNAs that have important regulatory roles in numerous developmental and metabolic processes in most eukaryotes. In Arabidopsis, DICER-LIKE1 (DCL1), HYPONASTIC LEAVES 1, SERRATE, HUA ENHANCER1 and HASTY are involved in processing of primary miRNAs (pri-miRNAs) to yield precursor miRNAs (pre-miRNAs) and eventually miRNAs. In addition to these components, mRNA cap-binding proteins, CBP80/ABA HYPERSENSITIVE1 and CBP20, also participate in miRNA biogenesis. Here, we show that STABILIZED1 (STA1), an Arabidopsis pre-mRNA processing factor 6 homolog, is also involved in the biogenesis of miRNAs. Similar to other miRNA biogenesis-defective mutants, sta1-1 accumulated significantly lower levels of mature miRNAs and concurrently higher levels of pri-miRNAs than wild type. The dramatic reductions of mature miRNAs were associated with the accumulation of their target gene transcripts and developmental defects. Furthermore, sta1-1 impaired splicing of intron containing pri-miRNAs and decreased transcript levels of DCL1. These results suggest that STA1 is involved in miRNA biogenesis directly by functioning in pri-miRNA splicing and indirectly by modulating the DCL1 transcript level.