Structural homology between bamboo mosaic virus and its satellite RNAs in the 5'untranslated region

Abstract

A structural element was identified in the 59-proximal sequence of the bamboo mosaic virus (BaMV) RNA. Mutational analysis of the hairpin showed that disruptions of the secondary structure or substitutions of the loop sequences resulted in reduced accumulation of BaMV genomic RNA. Phylogenetic analysis further suggested the presence of structural homologues of this hairpin in all other potexviruses. In addition, remarkable structural homology was discovered between the BaMV hairpin and a stem-loop in the 59untranslated region of satellite RNAs responsible for attenuation of BaMV in co-infected plants. The role of this homology in the helpersatellite interaction is discussed. Bamboo mosaic virus (BaMV) is a plant virus of the genusPotexvirus with a positive-sense RNA genome of 6366 nt, comprising five open reading frames (ORFs) . Infections by BaMV are frequently associated with the presence of satellite (sat) RNAs, some of which may reduce accumulation of BaMV RNA and attenuate BaMVinduced symptoms in co-infected plants (Hsu et al., 1998).The 140 nt 39untranslated region (UTR) of BaMV genomic RNA (gRNA) is composed of a pseudoknot structure involving the poly-A tail, a stem-loop structure harbouring a conserved hexamer motif and a cloverleaf-like structure formed by three adjacent hairpins Tsai et al., 1999). These structural elements have been implicated in the initiation of minus-strand synthesis by the viral RNAdependent RNA polymerase (Huang et al., 2001;Cheng et al., 2002). However, structural elements in the 59UTR of BaMV gRNA that potentially play roles in replication and/or translation have not been investigated in detail, although two stem-loop structures in the corresponding region of the negative-strand RNA have been suggested to be involved in positive-strand RNA synthesis (Lin et al., 2005).Using Mfold (Zuker, 2003), a large stem-loop structure was predicted in the 59 117 nt of BaMV positive-strand RNA (Fig. 1a). To verify the secondary structure of the predicted hairpin, in particular the upper part (nt 53-99), a transcript of approximately 1000 nt was synthesized by in vitro transcription of a DNA template obtained by PCR amplification of a full-length cDNA of BaMV-S (Lin et al., 2004). Purified transcripts were probed with enzymes and chemicals, and analysed by denaturing gel-electrophoresis using an RNA sequencing ladder as reference, as described previously (Chen et al., 2007b). Fig. 1(b) shows that nt C80, C77, A71, A70, C69, A68, C52, A51, A50 and A41 are strongly reactive to dimethyl sulfate (DMS) while A42, A57, A58, C86 and A90 are weakly reactive. Interestingly, the absence of strong modification on C63, C64, C89 and A90 (Fig. 1b) could indicate that the C : C and the C : A mismatches in stem-II (Fig. 1a) potentially form non-Watson-Crick base pairs, as had been found in the turnip yellow mosaic virus (TYMV) 59 hairpins (Hellendoorn et al., 1997;Bink et al., 2002). Nuclease S1 recognized nt A68-U73 in internal loop (IL)-I and U78 in the apical loop (AL). To some extent, U60 and U62 in st...

Identification of Tobacco Proteins Associated with the Stem-Loop 1 RNAs of Potato virus X

“…Previous studies have demonstrated that the sequences of upper region and C:C mismatch in SL structures of PVX and Bamboo mosaic virus (BaMV) play important roles in accumulation of genomic RNA (Chen et al, 2010;Miller et al, 1998). It has been shown that SL1(+) RNA binds to PVX CP and several tobacco proteins including the N. tabacum WRKY1 transcription factor (NtWRKY1) (Kim et al, 2002;Part and Kim, 2011).…”

A nationwide survey and genetic analysis of rice stripe virus in South Korea during 2012–2013

“…In all potexviruses, homologous stemloop structures were predicted by Mfold in the 5 0 untranslated region and can be clustered into four types (Chen et al 2010). A conserved hairpin structure is also predicted in the IR of RSV RNA3.…”

Host Factors in the Infection Cycle of Bamboo mosaic virus

“…Furthermore, a satellite RNA (satBaMV) was identified to associate with BaMV and could be amplified by BaMV replicase (Lin and Hsu, 1994; Lin et al, 2006). The tertiary structures of the 5′ and 3′ UTRs of satBaMV were revealed to be similar to those of BaMV (Huang et al, 2009; Chen et al, 2010). …”