Plant dicer-like proteins: double-stranded RNA-cleaving enzymes for small RNA biogenesis

Abstract

Dicer, a double-stranded RNA (dsRNA)-specific endoribonuclease, plays an essential role in triggering both transcriptional and post-transcriptional gene silencing in eukaryotes by cleaving dsRNAs or single-stranded RNAs bearing stem-loop structures such as microRNA precursor transcripts into 21- to 24-nt small RNAs. Unlike animals, plants have evolved to utilize at least four Dicer-like (DCL) proteins. Extensive genetic studies have revealed that each DCL protein participates in a specific gene silencing pathway, with some redundancy. However, a mechanistic understanding of how the specific action of each DCL protein is regulated in its respective pathway is still in its infancy due to the limited number of biochemical studies on plant DCL proteins. In this review, we summarize and discuss the biochemical properties of plant DCL proteins revealed by studies using highly purified recombinant proteins, crude extracts, and immunoprecipitates. With help from co-factor proteins and an ATPase/DExH-box RNA-helicase domain, the microRNA-producing enzyme DCL1 recognizes bulges and terminal loop structures in its substrate transcripts to ensure accurate and efficient processing. DCL4 prefers long dsRNA substrates and requires the dsRNA-binding protein DRB4 for its activity. The short-dsRNA preference of DCL3 is well suited for short-RNA transcription and subsequent dsRNA formation by coupling between a plant-specific DNA-dependent RNA-polymerase IV and RNA-dependent RNA-polymerase 2 in the transcriptional gene silencing pathway. Inorganic phosphate also seems to play a role in differential regulation of DCL3 and DCL4 activities. Further development of biochemical approaches will be necessary for better understanding of how plant DCL proteins are fine-tuned in each small RNA biogenesis pathway under various physiological conditions.

Global characterization of the Dicer-like protein DrnB roles in miRNA biogenesis in the social amoeba Dictyostelium discoideum

“…This strongly suggests that DrnB is dedicated to miRNA production and that the other Dicer-like protein DrnA, present in D. discoideum [48], is required for generating siRNAs. Similar division of functionality between Dicer/Dicer-like proteins is also seen in other organisms such as plants and the two Dicers present in Drosophila where Dicer-1 and Dicer-2 are devoted to miRNA and siRNA maturation, respectively [6,57,58]. However, in the single cell alga C. reinhardtii , one of the three Dicer homologs, DCL-3 is required for both miRNA and siRNA maturation, a situation similar to mammals and worms where a single Dicer protein produce both classes of small RNAs [31,57].…”

The ‘how’ and ‘where’ of plant microRNAs

“…pri-miRNAs are processed by the dicing complex, which contains DICER-LIKE1 (DCL1), HYPONASTIC LEAVES1 (HYL1) and SERRATE (SE) as core components, to yield mature miRNA/miRNA* duplexes (Park et al ., 2002; Reinhart et al ., 2002; Kurihara & Watanabe, 2004; reviewed in Fukudome & Fukuhara, 2017) (Fig. 1).…”

Dicer's Helicase Domain: A Meeting Place for Regulatory Proteins

“…Indeed, so far, a vertebrate Dicer has never been observed to require ATP. In contrast, there are multiple examples of invertebrate Dicers, those of plants, and that of Schizosaccharomyces pombe that show an ATP requirement in vitro (Liu et al 2003;Colmenares et al 2007;Welker et al 2011;Fukudome and Fukuhara 2017). Further, a mutation in the helicase Walker A motif of D. melanogaster Dicer-2 (dmDcr-2), a motif critical for ATP binding and hydrolysis, eliminates processing of siRNAs, establishing an ATP dependence in vivo (Lee et al 2004).…”