Superinfection exclusion by p28 of turnip crinkle virus is separable from its replication function

Authors: GUO, QIN - The Ohio State University; ZHANG, SHAOYAN - The Ohio State University; YAO, XIAOLONG - The Ohio State University; Tatineni, Satyanarayana - Ts; MEULIA, TEA - The Ohio State University; QU, FENG - The Ohio State University

Submitted to: Molecular Plant-Microbe Interactions
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 11/8/2019
Publication Date: 2/1/2020
Citation: Guo, Q., Zhang, S., Yao, X., Tatineni, S., Meulia, T., Qu, F. 2020. Superinfection exclusion by p28 of turnip crinkle virus is separable from its replication function. Molecular Plant-Microbe Interactions. 33(2):364-375. https://doi.org/10.1094/MPMI-09-19-0258-R.
DOI: https://doi.org/10.1094/MPMI-09-19-0258-R

Interpretive Summary: Superinfection exclusion (SIE) insulates virus-infected cells from subsequent invasion by the same or closely related viruses. SIE occurs in both animal and plant virus-infected cells. Therefore, a thorough understanding of how SIE is achieved at the molecular level is expected to inspire novel strategies for combating virus infections in humans, animals, and plants. Our group has been using Turnip crinkle virus (TCV) to explain the molecular interactions critical for SIE triggering. The current study builds on our previous observation that TCV SIE is driven by one single TCV-encoded protein (p28), and further identifies key regions and amino acids that are needed for SIE. This study identifies key amino acid changes that decouple the replication and SIE functions of p28 and provides valuable resources for in-depth evaluations of SIE.

Technical Abstract: We recently reported that the p28 auxiliary replication protein encoded by turnip crinkle virus (TCV) is also responsible for eliciting superinfection exclusion (SIE) against superinfecting TCV. However, it remains unresolved whether the replication function of p28 could be separated from its ability to elicit SIE. Here, we report the identification of two single amino acid mutations that decouple these two functions. Using an Agrobacterium infiltration-based delivery system, we transiently expressed a series of p28 deletion and point mutants, and tested their ability to elicit SIE against a cointroduced TCV replicon. We found that substituting alanine (A) for valine (V) and phenylalanine (F) at p28 positions 181 and 182, respectively, modestly compromised SIE in transiently expressed p28 derivatives. Upon incorporation into TCV replicons, V181A and F182A decoupled TCV replication and SIE diametrically. Although V181A impaired SIE without detectably compromising replication, F182A abolished TCV replication but had no effect on SIE once the replication of the defective replicon was restored through complementation. Both mutations diminished accumulation of p28 protein, suggesting that p28 must reach a concentration threshold in order to elicit a strong SIE. Importantly, the severe reduction of F182A protein levels correlated with a dramatic loss in the number of intracellular p28 foci formed by p28–p28 interactions. Together, these findings not only decouple the replication and SIE functions of p28 but also unveil a concentration dependence for p28 coalescence and SIE elicitation. These data further highlight the role of p28 multimerization in driving the exclusion of secondary TCV infections.