Plant A20/AN1 protein serves as the important hub to mediate antiviral immunity

Abstract

Salicylic acid (SA) is a key phytohormone that mediates a broad spectrum of resistance against a diverse range of viruses; however, the downstream pathway of SA governed antiviral immune response remains largely to be explored. Here, we identified an orchid protein containing A20 and AN1 zinc finger domains, designated Pha13. Pha13 is up-regulated upon virus infection, and the transgenic monocot orchid and dicot Arabidopsis overexpressing orchid Pha13 conferred greater resistance to different viruses. In addition, our data showed that Arabidopsis homolog of Pha13, AtSAP5, is also involved in virus resistance. Pha13 and AtSAP5 are early induced by exogenous SA treatment, and participate in the expression of SA-mediated immune responsive genes, including the master regulator gene of plant immunity, NPR1, as well as NPR1-independent virus defense genes. SA also induced the proteasome degradation of Pha13. Functional domain analysis revealed that AN1 domain of Pha13 is involved in expression of orchid NPR1 through its AN1 domain, whereas dual A20/AN1 domains orchestrated the overall virus resistance. Subcellular localization analysis suggested that Pha13 can be found localized in the nucleus. Self-ubiquitination assay revealed that Pha13 confer E3 ligase activity, and the main E3 ligase activity was mapped to the A20 domain. Identification of Pha13 interacting proteins and substrate by yeast two-hybrid screening revealed mainly ubiquitin proteins. Further detailed biochemical analysis revealed that A20 domain of Pha13 binds to various polyubiquitin chains, suggesting that Pha13 may interact with multiple ubiquitinated proteins. Our findings revealed that Pha13 serves as an important regulatory hub in plant antiviral immunity, and uncover a delicate mode of immune regulation through the coordination of A20 and/or AN1 domains, as well as through the modulation of E3 ligase and ubiquitin chain binding activity of Pha13.

Tomato SlSAP3, a member of the stress‐associated protein family, is a positive regulator of immunity against Pseudomonas syringae pv. tomato DC3000

“…Such different expression patterns imply that the SA signalling pathway is required for the function of SlSAP3 in immunity against Pst DC3000. This is consistent with a recent observation that orchid SAP protein Pha13 positively regulates the expression of SA‐mediated immune responsive genes (Chang et al ., ), but is different from a previous observation that AtSAP9 positively acts in JA signalling and negatively acts in SA pathway in response to a non‐host pathogen challenge (Kang et al ., ).…”

“…In plants, the Arabidopsis AtSAP5 and AtSAP9, rice OsiSAP7 and orchid Pha13, all of which contain both A20 and AN1 domains, were found to exhibit ubiquitin E3 ligase activity (Kang et al ., , ; Sharma et al ., ). Furthermore, the A20 domains in AtSAP5 and Pha13 were responsible for both E3 ligase and ubiquitin binding ability (Chang et al ., ; Kang et al ., ). It was also reported that the AN1 domain in AtSAP5 had strong ubiquitin E3 ligase activity (Choi et al ., ).…”

“…phaseolicola , indicating that AtSAP9 plays key roles in basal resistance (Kang et al ., ). Recently, an orchid SAP protein Pha13 and its Arabidopsis homologue AtSAP5 were reported to serve as an important regulatory hub in plant antiviral immunity (Chang et al ., ). It was shown that SAPs regulate various stress responses by modulating phytohormone signalling cascades, which are mediated by JA, SA, ET and ABA.…”

“…It was shown that SAPs regulate various stress responses by modulating phytohormone signalling cascades, which are mediated by JA, SA, ET and ABA. Interestingly, Arabidopsis AtSAP5 and AtSAP9 and rice OsSAP7 and OsSAP11 prefer to negatively regulate phytohormone signalling pathways (Kang et al ., , ; Liu et al ., ; Sharma et al ., ), while Pha13 positively regulates the expression of two SA responsive genes PhaRdR1 and PhaGRX (Chang et al ., ).…”

“…Besides, some SAPs were found to interact with UPS components, such as ubiquitin receptors RAD23s, which are capable of targeting ubiquitylated substrates to UPS (Farmer et al ., ; Fu et al ., ; Saeki, ). For instance, Arabidopsis AtSAP5, Prunus PpSAP1 and orchid Pha13 were found to interact with polyubiquitinated proteins (Chang et al ., ; Choi et al ., ; Lloret et al ., ). AtSAP9 was found to interact with RAD23b and RAD23d, which act as shuttling factors of ubiquitin conjugates (Farmer et al ., ; Guzder et al ., ; Kang et al ., ).…”