Functional characterization of the RNase III gene of rock bream iridovirus

Abstract

All of the fully sequenced iridoviruses have an ORF resembling a putative RNase III gene. However, to the best of our knowledge, functional characterization of the iridovirus-encoded RNase III has not been done. In the present study, we have characterized the putative RNase III of rock bream iridovirus (RBIV), the major cause of mass mortality of cultured rock bream Oplegnathus fasciatus in Korea. RBIV RNase III has a single N-terminal endonuclease domain followed by a C-terminal double-stranded RNA (dsRNA) binding domain. The true presence of the predicted ORF encoding RNase III in RBIV was confirmed by temporal transcription analysis of the ORF in RBIV-infected grunt fin (GF) cells. Comparing the catalytic activity to that of previously reported RNase III proteins, including Escherichia coli RNase III, the present RBIV RNase III had different features in that: (1) the dsRNA substrate was cleaved by the RBIV RNase III at high concentrations of Mg(2+) (5-20 mM) at low salt concentration (50 mM), but the enzyme activity was completely inhibited at 200 mM NaCl (within physiological ranges) irrespective of Mg(2+) concentrations (0.5-20 mM); (2) the substrate dsRNA was cleaved at low concentrations of Mn(2+) (0.5-1 mM) at low salt concentration (50 mM) and was cleaved by increasing Mn(2+) (5-20 mM) at 200 mM salt. These features of RBIV RNase III are similar to E. coli RNase III devoid of the C-terminal dsRBD region. The exact role of the RNase III in RBIV replication is not known, and further studies are needed to elucidate whether the RNase III is involved in the suppression of host RNA interference, which attacks viral mRNAs, or in the processing of viral RNAs for effective replication.

An Ascovirus-Encoded RNase III Autoregulates Its Expression and Suppresses RNA Interference-Mediated Gene Silencing

“…These are closely related to RNase III from iridoviruses. Recently, it was shown that the RNase III encoded by rock bream iridovirus has activity against dsRNA (33). Our in vitro cleavage assays with the purified protein confirmed endoND activity of HvAV-3e RNase III against dsRNA, which is a characteristic feature of all RNases (11,28).…”

“…Thus far, well-studied examples of viral RNase III proteins among RNA viruses is from sweet potato chlorotic stunt virus (SPCSV) (20), while in DNA viruses are from chlorella virus (PBCV-1) (34) and rock bream iridovirus (33). Genome annotations of other DNA viruses belonging to Ascoviridae and Iridoviridae indicate the presence of open reading frames (ORFs) that putatively encode RNase III proteins.…”

Genomic and Proteomic Analysis of Invertebrate Iridovirus Type 9

“…The presence of an RNase III gene (orf034L) that has also been identified in IIV-3, IIV-6, and vertebrate IV (45), and a dsRNA binding protein (060R; IIV-6 340R), supports a role for noncoding RNA in IV replication. RNase III was identified in purified virus particles and infected cells by our proteomic experiments (Table 2; see also Tables S1 and S3 in the supplemental material) and was previously identified in particles of SGIV (31), confirming that this protein is produced and, hence, likely to have a role in IV replication.…”

Functional characterisation and expression analysis of recombinant serum amyloid P isoform 1 (RbSAP1) from rock bream (Oplegnathus fasciatus)

“…However, an alarming increase in the prevalence and virulence of pathogenic infections in rock bream mariculture has caused considerable economic loss in Korea [1]. Therefore, research on immunity-related genes in rock bream is necessary for disease prevention and treatment.…”