Turnip mosaic virus selectively subverts a PR-5 thaumatin-like, plasmodesmal protein to promote viral infection

Introduction

Plant viruses infect almost all crops, causing serious diseases world-wide and threatening global food security. Plant viruses usually have small genomes that encode a few proteins and, thus, largely depend on host cellular machinery and factors to complete their infection cycle (Nagy & Pogany, 2012; Wang, 2015; Hyodo & Okuno, 2020; He et al., 2023). Unlike animal viruses that enter adjacent cells through receptor-mediated endocytosis or membrane fusion, plant viruses move to neighboring cells through plasmodesmata (PD) to establish systemic infection. PD are plasma membrane-lined pores, a specialized intercellular organelle that enables cytoplasmic and endomembrane continuity between adjacent cells (Cheval & Faulkner, 2018; Petit et al., 2020; Li et al., 2021; Burch-Smith, 2024; Tee & Faulkner, 2024). As intercellular channels, PD are dynamic structures, potentially allowing passage of small metabolites, assumably signal molecules and even macromolecules between neighboring cells, and the primary limiting factor of this capacity is the size of the plasmodesmal aperture (Nicolas et al., 2017; Tee & Faulkner, 2024). Due to this limitation, viral intercellular movement via PD requires the coordinated action of virus-encoded proteins and host factors, especially PD-localized ones (Wang, 2021). Molecular identification and functional characterization of PD-specific host proteins may assist in the development of novel strategies to control plant viral diseases for the sustainable crop production (Liu et al., 2021).

Most known viruses have a positive-sense single-stranded (+ss) RNA genome. Potyviruses (viruses in the genus Potyvirus in the family Potyviridae) constitute the largest group of plant-infecting +ssRNA viruses including several agriculturally and economically important ones such as potato virus Y, plum pox virus, soybean mosaic virus and turnip mosaic virus (TuMV) (Revers & García, 2015; Yang et al., 2021). The potyviral +ssRNA genome is an RNA of c. 10 kb that contains a single large open reading frame (ORF). During viral genome replication, transcriptional slippage at the coding region for the third protein (P3) leads to the generation of a small percentage of viral subpopulation whose genome encodes a short ORF due to frame shift (Cui & Wang, 2019; Yang et al., 2021). After translation, the long and short polyproteins are proteolytically processed into 11 mature proteins and various intermediate precursor proteins (Revers & García, 2015; Yang et al., 2021). Among them, the second 6-kDa protein, 6K2 is an integral membrane protein that targets and remodels the endoplasmic reticulum (ER) to initiate the formation of ER-derived membranous vesicles for viral replication (Schaad et al., 1997; Wei & Wang, 2008; Cotton et al., 2009).

To better understand the involvement of PD in viral infection, our lab conducted a quantitative, comparative proteomic study between the PD-enriched fractions from leaf tissues of Nicotiana benthamiana L. plants infected by TuMV and healthy leaves of mock-inoculated plants (Park et al., 2017). Three osmotin (OSM)-like proteins (OLPs) were identified to be significantly differentially accumulated in TuMV-infected leaves, compared to their amounts in the healthy control. Together with thaumatins, zeamatin and their like proteins (TLPs and ZLPs), OSMs and OLPs belong to the PR-5 family (the fifth class of pathogenesis-related proteins) (also known as TLP family) (Kumar et al., 2015; Hakim et al., 2018). The PR-5 family proteins are produced in plants in response to various biotic and abiotic stresses and play important roles in the plant immune system to confer tolerance to them (Sinha et al., 2014). Osmotin has been recognized as a plant defense tool (reviewed by Hakim et al., 2018). In the present study, we used the Arabidopsis orthologs (AtOSM34 and AtOLP) of the three OLPs identified from N. benthamiana to investigate their possible roles in TuMV infection. We show that both AtOSM34 and AtOLP are PD-located proteins and are differentially regulated in response to TuMV infection, but only AtOSM34 functions as a proviral factor. AtOSM34 is recruited to the viral replication complex (VRC) likely via its interaction with the TuMV 6K2 protein to promote viral replication. We further present evidence that AtOSM34 also promotes viral cell-to-cell movement. Moreover, we demonstrate that AtOMS34 suppresses antiviral resistance mediated by reactive oxygen species (ROS) burst.