The TobamovirusTurnip Vein Clearing Virus30-Kilodalton Movement Protein Localizes to Novel Nuclear Filaments To Enhance Virus Infection

Abstract

Plant viruses overcome the barrier of the plant cell wall by encoding cell-to-cell movement proteins (MPs), which direct newly replicated viral genomes to, and across, the wall. The paradigm for how a single MP regulates and coordinates these activities is the Tobacco mosaic virus (TMV) 30-kDa protein (MP TMV ). Detailed studies demonstrate that TMV multiplies exclusively in the cytoplasm and have documented associations of MP TMV with endoplasmic reticulum (ER) membrane, microtubules, and plasmodesmata throughout the course of infection. As TMV poorly infects Arabidopsis thaliana, Turnip vein clearing virus (TVCV) is the tobamovirus of choice for studies in this model plant. A key problem, which has contributed to confusion in the field, is the unproven assumption that the TVCV and TMV life cycles are identical. We engineered an infectious TVCV replicon that expressed a functional fluorescence-tagged MP TVCV and report here the unexpected discovery that MP TVCV , beyond localizing to ER membrane and plasmodesmata, targeted to the nucleus in a nuclear localization signal (NLS)-dependent manner, where it localized to novel F-actin-containing filaments that associated with chromatin. The MP TVCV NLS appeared to be conserved in the subgroup 3 tobamoviruses, and our mutational analyses showed that nuclear localization of MP TVCV was necessary for efficient TVCV cell-to-cell movement and systemic infection in Nicotiana benthamiana and Arabidopsis thaliana. Our studies identify a novel nuclear stage in TVCV infection and suggest that nuclear MP encoded by TVCV and other subgroup 3 tobamoviruses interacts with F-actin and chromatin to modulate host defenses or cellular physiology to favor virus movement and infection.

Importin-α-Mediated Nucleolar Localization of Potato Mop-Top Virus TRIPLE GENE BLOCK1 (TGB1) Protein Facilitates Virus Systemic Movement, Whereas TGB1 Self-Interaction Is Required for Cell-to-Cell Movement in Nicotiana benthamiana

“…These proteins contain NLS motifs and protein-protein and proteinnucleic acid (RNA and/or DNA) interaction domains that allow them to bind to host promoters (Lukhovitskaya et al, 2013a) or to recruit the viral RNA, other viral proteins, and/or host factors to facilitate viral cell-tocell and/or systemic movement through the phloem (see references below). Well-characterized plant virus proteins of this category include the ORF3 protein of Groundnut rosette virus (GRV; unassigned family, genus Umbravirus), the CP of Potato leaf roll virus (family Luteoviridae), the genome-linked viral protein of Potato virus A (family Potyviridae), the 2b suppressor of RNA silencing of Cucumber mosaic virus (family Bromoviridae), the P7a movement protein of Beet black scorch virus (family Tombusviridae), the CP of Alfalfa mosaic virus (family Bromoviridae), the P30 movement protein encoded by Turnip vein clearing virus (family Virgaviridae), and the P14 suppressor of RNA silencing and P25 of Beet necrotic yellow vein virus (family Virgaviridae; Haupt et al, 2005b;Kim et al, 2007;Rajamäki and Valkonen, 2009;González et al, 2010;Herranz et al, 2012;Wang et al, 2012;Chiba et al, 2013;Levy et al, 2013). Other viral proteins found in the nucleolus include the P3 protein of Tobacco etch virus (family Potyviridae) and the CP of Ourmia melon virus (unassigned family, genus Ourmiavirus), but the function of these nucleolar localizations is unknown (Taliansky et al, 2010;Crivelli et al, 2011;Rossi et al, 2014).…”

Tobacco mosaic virus-directed reprogramming of auxin/indole acetic acid protein transcriptional responses enhances virus phloem loading

“…Furthermore, the P30 protein from the Tobamovirus turnip vein clearing virus (TVCV) encodes a nuclear localization signal required for both cellto-cell and systemic virus movement (19). Association of the TVCV P30 protein with nuclear F-actin filaments was hypothesized to alter gene expression and promote virus infection (19). These studies provide evidence that virus interactions with host-associated transcriptional regulators can modulate systemic virus movement.…”

“…Similarly, overexpression of KELP resulted in the partial relocation of P30 to the nucleus and prevented ToMV cell-to-cell and systemic movement. Furthermore, the P30 protein from the Tobamovirus turnip vein clearing virus (TVCV) encodes a nuclear localization signal required for both cellto-cell and systemic virus movement (19). Association of the TVCV P30 protein with nuclear F-actin filaments was hypothesized to alter gene expression and promote virus infection (19).…”

Turnip vein clearing virus movement protein nuclear activity: Do Tobamovirus movement proteins play a role in immune response suppression?

“…To answer this question, we engineered a TVCV replicon that express a functional fusion of MP TVCV with GFP or RFP, and used these infectious clones to examine the distribution of MP TVCV during TVCV infection in Arabidopsis Col-0 and N. benthamiana, both susceptible hosts. 5 Surprisingly, we found that TVCV exhibits three distinct differences from TMV: (1) …”