Differential Requirement of the Ribosomal Protein S6 and Ribosomal Protein S6 Kinase for Plant-Virus Accumulation and Interaction of S6 Kinase with Potyviral VPg

Abstract

Ribosomal protein S6 (RPS6) is an indispensable plant protein regulated, in part, by ribosomal protein S6 kinase (S6K) which, in turn, is a key regulator of plant responses to stresses and developmental cues. Increased expression of RPS6 was detected in Nicotiana benthamiana during infection by diverse plant viruses. Silencing of the RPS6 and S6K genes in N. benthamiana affected accumulation of Cucumber mosaic virus, Turnip mosaic virus (TuMV), and Potato virus A (PVA) in contrast to Turnip crinkle virus and Tobacco mosaic virus. In addition, the viral genome-linked protein (VPg) of TuMV and PVA interacted with S6K in plant cells, as detected by bimolecular fluorescence complementation assay. The VPg-S6K interaction was detected in cytoplasm, nucleus, and nucleolus, whereas the green fluorescent protein-tagged S6K alone showed cytoplasmic localization only. These results demonstrate that the requirement for RPS6 and S6K differs for diverse plant viruses with different translation initiation strategies and suggest that potyviral VPg-S6K interaction may affect S6K functions in both the cytoplasm and the nucleus.

Ribosome profiles and riboproteomes of healthy and Potato virus A‐ and Agrobacterium‐infected Nicotiana benthamiana plants

“…Virus infections can affect the phosphorylation status of r‐proteins and induce unusual phosphorylation events (Diaz et al ., ). Our riboproteome data contained phosphopeptides for RPS6 in healthy and Agrobacterium ‐infected ribosome samples only, but it has been shown recently that the levels of phosphorylated RPS6 are comparable in PVA‐ and TuMV‐infected and healthy N. benthamiana plants (Rajamäki et al ., ). We found hits for phosphopeptides of RPP1 and RPP2 in the riboproteomes of Agrobacterium ‐ and PVA‐infected plants, but not in healthy plants.…”

“…Of the r‐protein transcripts, PVA infection increases the accumulation of the transcript encoding acidic r‐protein P0 in potato ( Solanum tuberosum ssp. andigena ) (Vuorinen et al ., ), whereas infected N. benthamiana leaves show increased levels of rps6 mRNAs (Rajamäki et al ., ). Potyvirus‐induced effects on r‐proteins at a protein level have been less well studied.…”

“…Potyvirus‐induced effects on r‐proteins at a protein level have been less well studied. Soybean mosaic virus infection represses the accumulation of both RPS12 mRNA and protein (Yang et al ., ), and PVA and TuMV infections induce the accumulation of RPS6 (Rajamäki et al ., ). Importantly, potyvirus infections are sensitive to the silencing of several rps and rpl genes (Rajamäki et al ., ; Yang et al ., ).…”

“…Soybean mosaic virus infection represses the accumulation of both RPS12 mRNA and protein (Yang et al ., ), and PVA and TuMV infections induce the accumulation of RPS6 (Rajamäki et al ., ). Importantly, potyvirus infections are sensitive to the silencing of several rps and rpl genes (Rajamäki et al ., ; Yang et al ., ). Although silencing of P0 affects PVA infection substantially, this defect may relate mostly to the non‐ribosomal functions of P0 during infection (Hafren et al ., ).…”

“…However, r‐proteins have been identified by LC‐MS/MS from HCPro‐induced RNA granules, HCPro‐containing high‐molecular‐weight complexes and the viral replication complexes isolated from PVA‐infected N. benthamiana plants (Hafren et al ., ; Ivanov et al ., ; Lohmus et al ., ). In addition, it has been shown that PVA VPg co‐localizes with RPS6 in the nucleus and nucleolus (Rajamäki et al ., ), and HCPro co‐localizes with P0 in PVA‐induced RNA granules (Hafren et al ., ). Our riboproteome analysis focused on translating ribosomes that lacked translation initiation factors, which are released from preinitiation complexes on 80S formation (Unbehaun et al ., ).…”