The Significance of the DUF283 Domain for the Activity of Human Ribonuclease Dicer

Abstract

Dicers are multidomain proteins, usually comprising an amino-terminal putative helicase domain, a DUF283 domain (domain of unknown function), a PAZ domain, two RNase III domains (RNase IIIa and RNase IIIb) and a dsRNA-binding domain. Dicer homologs play an important role in the biogenesis of small regulatory RNAs by cleaving single-stranded precursors adopting stem-loop structures (pre-miRNAs) and double-strand RNAs into short RNA duplexes containing functional microRNAs or small interfering RNAs, respectively. Growing evidence shows that apart from the canonical role, Dicer proteins can serve a number of other functions. For example, results of our previous studies showed that human Dicer (hDicer), presumably through its DUF283 domain, can facilitate hybridization between two complementary RNAs, thus, acting as a nucleic acid annealer. Here, to test this assumption, we prepared a hDicer deletion variant lacking the amino acid residues 625-752 corresponding to the DUF283 domain. The respective 128-amino acid fragment of hDicer was earlier demonstrated to accelerate base-pairing between two complementary RNAs in vitro. We show that the ΔDUF(625-752) hDicer variant loses the potential to facilitate RNA-RNA base pairing, which strongly proves our hypothesis about the importance of the DUF283 domain for the RNA-RNA annealing activity of hDicer. Interestingly, the in vitro biochemical characterization of the obtained deletion variant reveals that it displays different RNA cleavage properties depending on the pre-miRNA substrate.

RNAi-Based Antiviral Innate Immunity in Plants

“…In general, the helicase domain utilizes ATP hydrolysis to facilitate the unwinding of dsRNA [19,21]. The DUF283 domain was recently described to facilitate RNA-RNA base pairing and RNA-binding [22,23]. The PAZ and RNase III domains are vital for dsRNA cleavage, the PAZ domain recognizes the terminus of the dsRNA and RNase III domains and cuts one of the strands of dsRNA, and the distance between the PAZ domain and RNase III domains is determined by the length of the products [22,24].…”

“…The DUF283 domain was recently described to facilitate RNA-RNA base pairing and RNA-binding [22,23]. The PAZ and RNase III domains are vital for dsRNA cleavage, the PAZ domain recognizes the terminus of the dsRNA and RNase III domains and cuts one of the strands of dsRNA, and the distance between the PAZ domain and RNase III domains is determined by the length of the products [22,24]. The dsRBD domain facilitates dsRNA binding and also serves as a nonclassical nuclear localization signal [23].…”

RNase III, Ribosome Biogenesis and Beyond

“…It carries two RIIIDs and multiple additional domains including a DExD/H helicase domain involved in the binding to the terminal loop of targeted pre-miRNAs [102][103][104]. An uncharacterized DUF283 domain was recently shown to contribute to RNA-RNA annealing in human Dicer [105]. The platform (PF) domain serves as a structural measurement for the size of the cleavage product [106] and a PAZ domain anchors the 3 2-nts overhanging end of the dsRNA substrate [107].…”

Research Progress in RNA-Binding Proteins

“…Even before transcription ends, RNA molecules enter the processing line, which is managed by an array of protein and ribonucleoprotein biogenesis factors that help to fold, modify, and cleave precursor transcripts, and hand the resulting mature RNAs over to the downstream proteins for transport and the realisation of their molecular functions. Among these factors, ribonucleases hold a special place, and three papers in this Special Issue are dedicated to this class of enzymes [6][7][8].…”

“…One such facet is the mysterious DUF283 domain, a domain of unknown function, as its name indicates, which had been found to possess ssRNA-binding and nucleic acid-annealing activities [11,12]. In a follow-up study published in this Special Issue, Szczepanska et al hypothesised that this part of the protein might be responsible for the elusive RNA basepairing ability of human Dicer [6]. They designed and purified human Dicer variants that lacked the DUF283 domain and assayed their activities in vitro and in vivo.…”