p15 encoded by Garlic virus X is a pathogenicity factor and RNA silencing suppressor

Abstract

Garlic virus X (GarVX) encodes a 15 kDa cysteine-rich protein (CRP). To investigate the function(s) of p15, its subcellular localization, role as a symptom determinant and capacity to act as a viral suppressor of RNA silencing (VSR) were analysed. Results showed that GFP-tagged p15 was distributed in the cytoplasm, nucleus and nucleolus. Expression of p15 from PVX caused additional systemic foliar malformation and led to increased accumulation of PVX, showing that p15 is a virulence factor for reconstructed PVX-p15. Moreover, using a transient agro-infiltration patch assay and a Turnip crinkle virus (TCV) movement complementation assay, it was demonstrated that p15 possesses weak RNA silencing suppressor activity. Removal of an amino acid motif resembling a nuclear localization signal (NLS) prevented p15 from accumulating in the nucleus but did not abolish its silencing suppression activity. This study provides the first insights into the multiple functions of the GarVX p15 protein.

p24G1 Encoded by Grapevine Leafroll-Associated Virus 1 Suppresses RNA Silencing and Elicits Hypersensitive Response-Like Necrosis in Nicotiana Species

“…The subcellular localization assay revealed that p24 G1 mainly accumulates in the nucleus, in agreement with the distribution pattern of some other viral RSSs [ 16 , 17 , 18 , 19 ]. Consistent with previous reports for CMV 2b [ 17 ], GVX p15 [ 18 ] and CVB p12 [ 19 ], our data also indicate that the nuclear distribution pattern of p24 G1 is not essential for its RSS activity, because ∆1–21 showed impaired nuclear localization ( Figure 4 ) but could block RNA silencing ( Figure 6 ).…”

“…The subcellular localization assay revealed that p24 G1 mainly accumulates in the nucleus, in agreement with the distribution pattern of some other viral RSSs [ 16 , 17 , 18 , 19 ]. Consistent with previous reports for CMV 2b [ 17 ], GVX p15 [ 18 ] and CVB p12 [ 19 ], our data also indicate that the nuclear distribution pattern of p24 G1 is not essential for its RSS activity, because ∆1–21 showed impaired nuclear localization ( Figure 4 ) but could block RNA silencing ( Figure 6 ). Moreover, similar to the canonical importin α/β nuclear import pathway adopted by CMV 2b [ 43 ] and p6 of cauliflower mosaic virus (CaMV) [ 16 ], p24 G1 also interacted with NbIMPα1 ( Figure 4 b), suggesting that the nuclear import of p24 G1 is mediated by importin α1, although other importin α1-independent nuclear transport pathways cannot be excluded.…”

“…For example, many viral RSSs act as dimers to block RNA silencing [ 9 , 10 , 11 , 12 , 13 ], while the monomers of tomato golden mosaic virus AL2 and beet curly top virus L2 can also suppress RNA silencing [ 14 ]. Similarly, in some instances, the nuclear distribution of nuclear-localized viral RSSs is essential for their suppressive function [ 15 , 16 ], whereas the RSS activity of CMV 2b [ 17 ], garlic virus X (GVX) p15 [ 18 ] and chrysanthemum virus B (CVB) p12 [ 19 ] is not correlated with their nuclear distribution pattern.…”

The nanovirus U2 protein suppresses RNA silencing via three conserved cysteine residues

“…The CRPs of several plant viruses have been shown to be pathogenicity determinants (Zhang et al, 2018 ). To examine whether the U2 protein is a pathogenicity factor, we inserted the U2 and mutants (U2C43A, U2C79A, U2C82A and U2‐3C) into an Agrobacterium ‐delivered infectious clone of PVX.…”

“…Cysteine‐rich proteins (CRPs) that have multiple cysteine residues in their amino acid sequences but no other consistent amino acid sequence similarity occurs in viruses from different genera, including Allexivirus , Benyvirus , Carlavirus , Furovirus , Hordeivirus , Pecluvirus , Pomovirus and Tobravirus (Andika et al, 2012 ; Donald & Jackson, 1994 ; Fujita et al, 2018 ; Koonin et al, 1991 ; Liu et al, 2002 ; Lukhovitskaya et al, 2005 ; Te et al, 2005 ; Zhang et al, 2018 ). Many CRPs have been shown to play a role in VSR and induction of viral symptoms (Andika et al, 2012 ; Donald & Jackson, 1994 ; Fujita et al, 2018 ; Ghazala et al, 2008 ; Liu et al, 2002 ; Lukhovitskaya et al, 2005 ; Te et al, 2005 ; Yelina et al, 2005 ; Zhang et al, 2018 ). Furthermore, cysteine residues consistently occur in the putative zinc‐finger domains of the CRPs of hordeiviruses, carlaviruses and potyviruses (Bragg & Jackson, 2004 ; Lukhovitskaya et al, 2009 ; Pasin et al, 2022 ; Senshu et al, 2011 ; Xu et al, 2021 ).…”