Mapping the RNA-binding domain on the Apple chlorotic leaf spot virus movement protein

Abstract

The RNA-binding properties of the cell-to-cell movement protein (MP) of Apple chlorotic leaf spot virus were analysed. MP was expressed in Escherichia coli and was used in UV-crosslinking analysis, using a digoxigenin-UTP-labelled RNA probe and gel-retardation analysis. The analyses demonstrated that MP bound cooperatively to single-stranded RNA (ssRNA). When analysed for NaCl dependence of the RNA-binding activity, the majority of the MP could bind ssRNA even in binding buffer with 1 M NaCl. Furthermore, competition binding experiments showed that the MP bound preferentially to ssRNA and single-stranded DNA without sequence specificity. MP deletion mutants were used to identify the RNA-binding domain by UV-crosslinking analysis. Amino acid residues 82-126 and 127-287 potentially contain two independently active, single-stranded nucleic acid-binding domains.

Cucumber Mosaic VirusMovement Protein Severs Actin Filaments to Increase the Plasmodesmal Size Exclusion Limit in Tobacco

“…The absence of any intermediate retarded bands indicated that CMV MP bound in a cooperative manner to CMV RNA3, which is consistent with the results reported by Li and Palukaitis (1996). The retarded probes barely entered the gel, and this may be due to a highly cooperative binding between MP and the labeled probe (Nurkiyanova et al, 2001;Isogai and Yoshikawa, 2005). A competition binding assay with unlabeled CMV RNA3 competitor further confirmed that the purified CMV MP was able to bind vRNA in vitro (see Supplemental Figure 2A online).…”

Subcellular localization and expression of bamboo mosaic virus satellite RNA-encoded protein

“…The N-terminal arginine-rich motif of P20 potentiates these activities as well as intracellular targeting (Palani et al , 2006). In this regard, P20 shares common biological properties with other viral MPs (Carvalho & Lazarowitz, 2004; Hsu et al , 2004; Isogai & Yoshikawa, 2005). In BaMV and satBaMV co-infected N. benthamiana leaves, accumulation of P20 and a serologically related polypeptide of a low molecular mass of 16 kDa (P16) has been detected (Palani et al , 2006).…”

Host and viral RNA-binding proteins involved in membrane targeting, replication and intercellular movement of plant RNA virus genomes

“…After the first report of TMV MP binding to nucleic acids, the binding of other MPs to nucleic acids has been reported for 26 other viruses, including (+)RNA viruses, (-) RNA viruses, and DNA viruses (reviewed by Waigmann et al, 2004). In the past decade, at least nine MPs have been added to the list of nucleic acid-binding MPs: Cowpea mosaic virus (CPMV; Carvalho et al, 2004); Prunus necrotic ringspot virus (PNRSV; Herranz and Pallás, 2004); Rice yellow stunt virus (Huang et al, 2005); Apple chlorotic leaf spot virus (Isogai and Yoshikawa, 2005); Rice dwarf virus (Ji et al, 2011); Parietaria mottle virus (Martínez et al, 2014); Pelargonium flower break virus (Martínez-Turiño and Hernández, 2011); Melon necrotic spot virus (Navarro et al, 2006); Rice stripe virus (Xiong et al, 2008). Most MPs can bind single-stranded nucleic acids, and only three viral MPs (one geminivirus and two hordeiviruses) have been shown to bind double-stranded nucleic acids (Bleykasten et al, 1996; Donald et al, 1997; Rojas et al, 1998).…”