Dissecting the multifunctional role of the N‐terminal domain of the Melon necrotic spot virus coat protein in RNA packaging, viral movement and interference with antiviral plant defence

Abstract

The coat protein (CP) of Melon necrotic spot virus (MNSV) is structurally composed of three major domains. The middle S-domain builds a robust protein shell around the viral genome, whereas the C-terminal protruding domain, or P-domain, is involved in the attachment of virions to the transmission vector. Here, we have shown that the N-terminal domain, or R-domain, and the arm region, which connects the R-domain and S-domain, are involved in different key steps of the viral cycle, such as cell-to-cell movement and the suppression of RNA silencing and pathogenesis through their RNA-binding capabilities. Deletion mutants revealed that the CP RNA-binding ability was abolished only after complete, but not partial, deletion of the R-domain and the arm region. However, a comparison of the apparent dissociation constants for the CP RNA-binding reaction of several partial deletion mutants showed that the arm region played a more relevant role than the R-domain in in vitro RNA binding. Similar results were obtained in in vivo assays, although, in this case, full-length CPs were required to encapsidate full-length genomes. We also found that the R-domain carboxyl portion and the arm region were essential for efficient cell-to-cell movement, for enhancement of Potato virus X pathogenicity, for suppression of systemic RNA silencing and for binding of small RNAs. Therefore, unlike other carmovirus CPs, the R-domain and the arm region of MNSV CP have acquired, in addition to other essential functions such as genome binding and encapsidation functions, the ability to suppress RNA silencing by preventing systemic small RNA transport.

Hsc70-2 is required for Beet black scorch virus infection through interaction with replication and capsid proteins

“…A growing body of evidence shows that viral CP is a multi-functional protein with roles outside of virus packaging 15 , 44 . To investigate what role of BBSV CP in virus replication, three different viral clones were constructed as depicted in Fig.…”

Cucurbit chlorotic yellows virus p22 suppressor of RNA silencing binds single-, double-stranded long and short interfering RNA molecules in vitro

“…In these experiments, Melon necrotic spot virus (MNSV) coat protein (CP) was used as a positive control for its previously documented ability to bind ssRNA in a non-specific manner (Fig. 1A; Serra-Soriano et al, 2017). When a 390 nt long ssRNA derived from the MNSV 3'-untranslated region (UTR) was used as a probe (Navarro et al, 2006), an alteration on the RNA migration pattern could be observed in the presence relatively large amounts of CCYV p22: an initial RNA shift was detected when 2 g of protein were used and, when 4 g or more protein was introduced, a high molecular weight RNA:protein complex could be observed in the upper part of the gel (Fig.…”

An Update on the Intracellular and Intercellular Trafficking of Carmoviruses

“…In both cases, the normal extent of local movement was recovered when other well-established viral suppressors were provided in trans ( Genovés et al, 2006 ; Powers et al, 2008 ; Shi et al, 2009 ). Therefore, the apparent effect of these CPs on cell-to-cell movement was most likely associated with their ability to suppress RNA silencing ( Qu et al, 2003 ; Thomas et al, 2003 ; Genovés et al, 2006 ; Serra-Soriano et al, 2017 ). Besides, the TCV CP was dispensable for cell-to-cell movement in Arabidopsis thaliana but it was essential to facilitate the entry of virions into the vasculature and cause systemic infection ( Cohen et al, 2000a ).…”