During plant virus transmission by piercing-sucking insects, most viruses are inoculated into the plant phloem via the insect’s secreted saliva (Arcà and Ribeiro, 2018; Conway et al., 2016; Wu et al., 2022). Thus, insect saliva acts as an interface for the virus–insect–host tripartite interaction and can directly promote viral transmission to, and infection of, the host plants (Sun et al., 2020; Wu et al., 2022). However, despite the importance of insect salivary proteins in this tripartite interaction, there is still much to learn about how these proteins enable successful viral infection.
Previous studies have revealed that there are two ways in which insect saliva facilitates viral infection. One is an indirect approach whereby the saliva modulates the host microenvironment at the feeding site, and saliva effectors work together to allow the arthropod to go unnoticed while it feeds on the host plant (Acevedo et al., 2019; Arcà and Ribeiro, 2018; Sun et al., 2020). For example, our work indicated that a Laodelphax striatellus mucin protein, LssaMP, enables the formation of the salivary sheath and facilitates the transmission of rice stripe virus (RSV) into the rice phloem (Huo et al., 2022). The study on leafhoppers revealed that the expression of a saliva calcium-binding protein is inhibited by rice gall dwarf virus (RGDV), thus causing an increase of cytosolic Ca2+ levels in rice and triggering callose deposition and H2O2 production. This increases the frequency of insect probing, thereby enhancing viral horizontal transmission into the rice phloem (Wu et al., 2022). The other mechanism by insect saliva to facilitate virus infection is direct regulation, whereby saliva proteins promote virus transmission through specific molecular interactions (Wen et al., 2019). Direct saliva protein–pathogen interactions have been reported in animal pathogens. For example, during transmission of Borrelia burgdorferi by Ixodes scapularis, the saliva protein Salp15 of I. scapularis binds to the bacterial outer surface protein C, which prevents the bacterium from being recognized by the animal immune system. In this way, the saliva protein enables the pathogen to infect the animal host (Ramamoorthi et al., 2005; Schuijt et al., 2008). Although most plant viruses are heavily dependent on insect vectors for plant-to-plant transmission (Gray, 2008), the direct function of insect saliva proteins in mediating virus transmission remains largely uninvestigated.
During sap-feeding, arthropods produce two distinct types of saliva at different stages of the feeding process: gel saliva and watery saliva (Bonaventure, 2012; Lou et al., 2019). The former forms a salivary sheath to provide a smooth path for the stylet penetration (Lou et al., 2019). The latter is mainly secreted into the phloem sieve elements to prevent them from plugging up and suppress plant defense responses. Some salivary components may act as herbivore-associated molecular patterns that can trigger pattern-triggered immunity, and certain salivary effectors may be recognized by plant resistance proteins to induce effector-triggered immunity, etc. (Huang et al., 2017; Ji et al., 2017; Yi et al., 2021).
For phloem-feeding insects, callose deposited on phloem sieve plate and plasmodesmata of sieve elements functions as a defense mechanism by reducing insect feeding and preventing viral movement (Hao et al., 2008; Hipper et al., 2013; Will and Vilcinskas, 2015; Zavaliev et al., 2011; Yue et al., 2022). Callose is a β-(1,3)-D-glucan polysaccharide that is synthesized by callose synthases and degraded by β-(1,3)-glucanases. Plants defend themselves by depositing callose at the sieve plates and plasmodesmata in response to virus infection, whereas viruses counter this defense by activating β-(1,3)-glucanases to degrade callose (Bucher et al., 2001; Hao et al., 2008; Wu et al., 2022; Zavaliev et al., 2011).
RSV is the causative agent of rice stripe disease, a serious disease of rice crops that has occurred repeatedly in China, Japan, and Korea (Xu et al., 2021). RSV is completely dependent on insect vectors for transmission among its host plants, and L. striatellus is the main vector (Xu et al., 2021; Zhao et al., 2017). L. striatellus transmits RSV in a persistent-propagative manner. The virus initially infects the midgut, then disperses from the hemolymph into the salivary glands and is inoculated into the plant host during L. striatellus feeding (Huo et al., 2022). L. striatellus belongs to the order Hemiptera, whose members mainly feed from sieve tubes through their mouthparts (stylets) that penetrate plant tissues and reach sieve tubes to ingest the phloem sap (Tjallingii, 2006; van Bel and Will, 2016). RSV is mainly secreted into the rice phloem via the watery saliva (Huo et al., 2022; Wang and Blanc, 2021).
In this study, we identified a molecular interaction among RSV, an L. striatellus saliva protein, and a plant β-1,3-glucanase. The insect saliva protein directly binds to the RSV nucleocapsid protein (NP) and then binds to a rice thaumatin-like protein to activate its β-1,3-glucanase activity. The activation of β-1,3-glucanase helps RSV infection by inhibiting callose deposition in response to viral infection.