Two key arginine residues in the coat protein of Bamboo mosaic virus differentially affect the accumulation of viral genomic and subgenomic RNAs

Abstract

The interactions between viral RNAs and coat proteins (CPs) are critical for the efficient completion of infection cycles of RNA viruses. However, the specificity of the interactions between CPs and genomic or subgenomic RNAs remains poorly understood. In this study, Bamboo mosaic virus (BaMV) was used to analyse such interactions. Using reversible formaldehyde cross-linking and mass spectrometry, two regions in CP, each containing a basic amino acid (R99 and R227, respectively), were identified to bind directly to the 5' untranslated region of BaMV genomic RNA. Analyses of the alanine mutations of R99 and R227 revealed that the secondary structures of CP were not affected significantly, whereas the accumulation of BaMV genomic, but not subgenomic, RNA was severely decreased at 24 h post-inoculation in the inoculated protoplasts. In the absence of CP, the accumulation levels of genomic and subgenomic RNAs were decreased to 1.1%-1.5% and 33%-40% of that of the wild-type (wt), respectively, in inoculated leaves at 5 days post-inoculation (dpi). In contrast, in the presence of mutant CPs, the genomic RNAs remained about 1% of that of wt, whereas the subgenomic RNAs accumulated to at least 87%, suggesting that CP might increase the accumulation of subgenomic RNAs. The mutations also restricted viral movement and virion formation in Nicotiana benthamiana leaves at 5 dpi. These results demonstrate that R99 and R227 of CP play crucial roles in the accumulation, movement and virion formation of BaMV RNAs, and indicate that genomic and subgenomic RNAs interact differently with BaMV CP.

The molecular basis for flexibility in the flexible filamentous plant viruses

“…While the resolution of the data does not permit us to draw conclusions on the nature of the protein–RNA interactions, the model suggests that residues Arg99, Lys132, Lys157, Lys202, and Lys213 all potentially make protein–RNA contacts. Among them, Arg99 was found 20 to be part of the potential RNA binding motifs in BaMV CP.…”

Autophagy is involved in assisting the replication of Bamboo mosaic virus in Nicotiana benthamiana

“…Bamboo mosaic virus (BaMV) is a single-stranded positive-sense RNA virus belonging to Potexvirus of the family Alphaflexiviridae. The genome has five ORFs: ORF1 encodes the protein for viral RNA replication (Li et al , 1998, 2001a, 2001b; Huang et al , 2004; Meng and Lee, 2017); ORF2–ORF4 encode the proteins for cell-to-cell and systemic movement (Lin et al , 2004, 2006); and ORF5 encodes the capsid protein for encapsidation, symptom development, and viral movement (Lan et al , 2010; Hung et al , 2014a, 2014b). Host factors participating in the infection cycle of BaMV in N. benthamiana have been identified by using ultraviolet crosslinking (Lin et al , 2007; Prasanth et al , 2011; Huang et al , 2012), yeast two-hybrid (Lee et al , 2011), and cDNA-amplified fragment length polymorphism (Cheng et al , 2010) techniques.…”

Nuclear-Encoded Plastidal Carbonic Anhydrase Is Involved in Replication of Bamboo mosaic virus RNA in Nicotiana benthamiana

“…The movement of BaMV requires these three TGB proteins ( Lin et al, 2004 , 2006 ; Chou et al, 2013 ). ORF5, encoding a 25-kDa polypeptide viral capsid protein (CP) is required for cell-to-cell movement, symptom development, and virion assembly ( Lan et al, 2010 ; Hung et al, 2014a , b ). The 3′ untranslated region (UTR) plays roles in minus-strand RNA initiation, polyadenylation, and long-distance movement ( Chen et al, 2017 ).…”