Reduced cold tolerance of viral-infected leafhoppers attenuates viral persistent epidemics.

Most arthropod-borne viruses produce intermittent epidemics in infected plants. However, the underlying mechanisms of these epidemics are unclear. Here, we demonstrated that rice stripe mosaic virus (RSMV), a viral pathogen, significantly increases the mortality of its overwintering vector, the leafhopper species Recilia dorsalis. Cold-stress assays indicated that RSMV reduces the cold tolerance of leafhoppers, a process associated with the downregulation of leafhopper cuticular protein genes. An RSMV-derived small RNA (vsiR-t00355379) was found to facilitate the downregulation of a leafhopper endocuticle gene that is mainly expressed in the abdomen (named RdABD-5) and is conserved across dipteran species. The downregulation of RdABD-5 expression in R. dorsalis resulted in fewer and thinner endocuticle lamellae, leading to decreased cold tolerance. This effect was correlated with a reduced incidence rate of RSMV in early-planted rice plants. These findings contribute to our understanding of the mechanism by which viral pathogens reduce cold tolerance in arthropod vectors and suggest an approach to managing the fluctuating prevalence of arboviruses. IMPORTANCE: Increasing arthropod vector dispersal rates have increased the susceptibility of crop to epidemic viral diseases. However, the incidence of some viral diseases fluctuates annually. In this study, we demonstrated that a rice virus reduces the cold tolerance of its leafhopper vector, Recilia dorsalis. This effect is linked to the virus-derived small RNA-mediated downregulation of a gene encoding a leafhopper abdominal endocuticle protein. Consequently, the altered structural composition of the abdominal endocuticle reduces the overwinter survival of leafhoppers, resulting in a lower incidence of RSMV infection in early-planted rice plants. Our findings illustrate the important roles of RNA interference in virus-vector insect-environment interactions and help explain the annual fluctuations of viral disease epidemics in rice fields.

Maintenance of persistent transmission of a plant arbovirus in its insect vector mediated by the Toll-Dorsal immune pathway.

Throughout evolution, arboviruses have developed various strategies to counteract the host's innate immune defenses to maintain persistent transmission. Recent studies have shown that, in addition to bacteria and fungi, the innate Toll-Dorsal immune system also plays an essential role in preventing viral infections in invertebrates. However, whether the classical Toll immune pathway is involved in maintaining the homeostatic process to ensure the persistent and propagative transmission of arboviruses in insect vectors remain unclear. In this study, we revealed that the transcription factor Dorsal is actively involved in the antiviral defense of an insect vector (Laodelphax striatellus) by regulating the target gene, zinc finger protein 708 (LsZN708), which mediates downstream immune-related effectors against infection with the plant virus (Rice stripe virus, RSV). In contrast, an antidefense strategy involving the use of the nonstructural-protein (NS4) to antagonize host antiviral defense through competitive binding to Dorsal from the MSK2 kinase was employed by RSV; this competitive binding inhibited Dorsal phosphorylation and reduced the antiviral response of the host insect. Our study revealed the molecular mechanism through which Toll-Dorsal-ZN708 mediates the maintenance of an arbovirus homeostasis in insect vectors. Specifically, ZN708 is a newly documented zinc finger protein targeted by Dorsal that mediates the downstream antiviral response. This study will contribute to our understanding of the successful transmission and spread of arboviruses in plant or invertebrate hosts.

Two different viral proteins suppress NUCLEAR FACTOR-YC-mediated antiviral immunity during infection in rice.

Plant viruses have multiple strategies to counter and evade the host's antiviral immune response. However, limited research has been conducted on the antiviral defense mechanisms commonly targeted by distinct types of plant viruses. In this study, we discovered that NUCLEAR FACTOR-YC (NF-YC) and NUCLEAR FACTOR-YA (NF-YA), 2 essential components of the NF-Y complex, were commonly targeted by viral proteins encoded by 2 different rice (Oryza sativa L.) viruses, rice stripe virus (RSV, Tenuivirus) and southern rice black streaked dwarf virus (SRBSDV, Fijivirus). In vitro and in vivo experiments showed that OsNF-YCs associate with OsNF-YAs and inhibit their transcriptional activation activity, resulting in the suppression of OsNF-YA-mediated plant susceptibility to rice viruses. Different viral proteins RSV P2 and SRBSDV SP8 directly disrupted the association of OsNF-YCs with OsNF-YAs, thereby suppressing the antiviral defense mediated by OsNF-YCs. These findings suggest an approach for conferring broad-spectrum disease resistance in rice and reveal a common mechanism employed by viral proteins to evade the host's antiviral defense by hindering the antiviral capabilities of OsNF-YCs.

Alternative Splicing Landscape of Small Brown Planthopper and Different Response of JNK2 Isoforms to Rice Stripe Virus Infection.

Alternative splicing (AS) is a frequent posttranscriptional regulatory event occurring in response to various endogenous and exogenous stimuli in most eukaryotic organisms. However, little is known about the effects of insect-transmitted viruses on AS events in insect vectors. The present study used third-generation sequencing technology and RNA sequencing (RNA-Seq) to evaluate the AS response in the small brown planthopper Laodelphax striatellus to rice stripe virus (RSV). The full-length transcriptome of L. striatellus was obtained using single-molecule real-time sequencing technology (SMRT). Posttranscriptional regulatory events, including AS, alternative polyadenylation, and fusion transcripts, were analyzed. A total of 28,175 nonredundant transcript isoforms included 24,950 transcripts assigned to 8,500 annotated genes of L. striatellus, and 5,000 of these genes (58.8%) had AS events. RNA-Seq of the gut samples of insects infected by RSV for 8 d identified 3,458 differentially expressed transcripts (DETs); 2,185 of these DETs were transcribed from 1,568 genes that had AS events, indicating that 31.4% of alternatively spliced genes responded to RSV infection of the gut. One of the c-Jun N-terminal kinase (JNK) genes, JNK2, experienced exon skipping, resulting in three transcript isoforms. These three isoforms differentially responded to RSV infection during development and in various organs. Injection of double-stranded RNAs targeting all or two isoforms indicated that three or at least two JNK2 isoforms facilitated RSV accumulation in planthoppers. These results implied that AS events could participate in the regulation of complex relationships between viruses and insect vectors. IMPORTANCE Alternative splicing (AS) is a regulatory mechanism that occurs after gene transcription. AS events can enrich protein diversity to promote the reactions of the organisms to various endogenous and exogenous stimulations. It is not known how insect vectors exploit AS events to cope with transmitted viruses. The present study used third-generation sequencing technology to obtain the profile of AS events in the small brown planthopper Laodelphax striatellus, which is an efficient vector for rice stripe virus (RSV). The results indicated that 31.4% of alternatively spliced genes responded to RSV infection in the gut of planthoppers. One of the c-Jun N-terminal kinase (JNK) genes, JNK2, produced three transcript isoforms by AS. These three isoforms showed different responses to RSV infection, and at least two isoforms facilitated viral accumulation in planthoppers. These results implied that AS events could participate in the regulation of complex relationships between viruses and insect vectors.

Membrane association of importin α facilitates viral entry into salivary gland cells of vector insects.

The importin α family belongs to the conserved nuclear transport pathway in eukaryotes. However, the biological functions of importin α in the plasma membrane are still elusive. Here, we report that importin α, as a plasma membrane-associated protein, is exploited by the rice stripe virus (RSV) to enter vector insect cells, especially salivary gland cells. When the expression of three importin α genes was simultaneously knocked down, few virions entered the salivary glands of the small brown planthopper, Laodelphax striatellus Through hemocoel inoculation of virions, only importin α2 was found to efficiently regulate viral entry into insect salivary-gland cells. Importin α2 bound the nucleocapsid protein of RSV with a relatively high affinity through its importin ß-binding (IBB) domain, with a dissociation constant KD of 9.1 µM. Furthermore, importin α2 and its IBB domain showed a distinct distribution in the plasma membrane through binding to heparin in heparan sulfate proteoglycan. When the expression of importin α2 was knocked down in viruliferous planthoppers or in nonviruliferous planthoppers before they acquired virions, the viral transmission efficiency of the vector insects in terms of the viral amount and disease incidence in rice was dramatically decreased. These findings not only reveal the specific function of the importin α family in the plasma membrane utilized by viruses, but also provide a promising target gene in vector insects for manipulation to efficiently control outbreaks of rice stripe disease.

Laodelphax striatellus Atg8 facilitates Rice stripe virus infection in an autophagy-independent manner.

Rice stripe virus (RSV) is the causative agent of rice stripe disease and is completely dependent on insect vectors for its plant-to-plant transmission. Laodelphax striatellus is the major insect vector for RSV. In this study, we explored the interactions between RSV infection and L. striatellus autophagy, a potential intrinsic antiviral mechanism in insects. We found that L. striatellus autophagic activity did not affect RSV infection; however, the autophagy-related-8 (Atg8) gene significantly enhanced virus infection. During RSV initial infection within the L. striatellus midgut, silencing of Atg8 expression significantly decreased the phosphorylation of c-Jun N-terminal kinase (p-JNK); however, when RSV infection is absent, silencing of Atg8 did not alter p-JNK levels. These results indicated that Atg8 might activate the JNK machinery by allowing more virus infection into cells. We further revealed that Atg8-deficiency significantly decreased RSV accumulation on the surface of the insect midgut epithelial cells, suggesting a receptor trafficking function of the γ-aminobutyric acid receptor-associated protein family. Using the RSV ovary entry as a model, in which vitellogenin receptor (VgR) mediates RSV cell entry, we clarified that Atg8-deficiency decreased the abundance of VgR localizing on the cytomembrane and disturbed the attachment of RSV in the germarium zones. Collectively, these results revealed an autophagy-independent function of L. striatellus Atg8 that enhances RSV initial infection by increasing virus attachment on the infection sites.

The α-tubulin of Laodelphax striatellus mediates the passage of rice stripe virus (RSV) and enhances horizontal transmission.

Rice stripe virus (RSV, genus Tenuivirus, family Phenuiviridae) is the causal agent of rice stripe disease transmitted by the small brown planthopper (SBPH, Laodelphax striatellus) in a persistent propagative manner. The midgut and salivary glands of SBPH are the first and last barriers to the viral circulation and transmission processes, respectively; however, the precise mechanisms used by RSV to cross these organs and transmit to rice plants have not been fully elucidated. We obtained the full-length cDNA sequence of L. striatellus α-tubulin 2 (LsTUB) and found that RSV infection increased the level of LsTUB in vivo. Furthermore, LsTUB was shown to co-localize with RSV nonstructural protein 3 (NS3) in vivo and bound NS3 at positions 74-76 and 80-82 in vitro. Transient gene silencing of LsTUB expression caused a significant reduction in detectable RSV loads and viral NS3 expression levels, but had no effect on NS3 silencing suppressor activity and viral replication in insect cells. However, suppression of LsTUB attenuated viral spread in the bodies of SBPHs and decreased RSV transmission rates to rice plants. Electrical penetration graphs (EPG) showed that LsTUB knockdown by RNAi did not impact SBPH feeding; therefore, the reduction in RSV transmission rates was likely caused by a decrease in viral loads inside the planthopper. These findings suggest that LsTUB mediates the passage of RSV through midgut and salivary glands and leads to successful horizontal transmission.

Ubiquitin-Conjugating Enzyme E2 E Inhibits the Accumulation of Rice Stripe Virus in Laodelphax striatellus (Fallén).

The ubiquitin-proteasome system (UPS) is an essential protagonist in host-pathogen interactions. Among the three classes of enzymes in the UPS, ubiquitin-conjugating enzyme E2 plays a dual role in viral pathogenesis; however, the role of insect E2s in interactions with plant viruses is unclear. Twenty E2-encoding genes in Laodelphax striatellus, the small brown planthopper, were identified and classified into 17 groups by transcriptomic and phylogenetic analysis. Full-length cDNAs of four LstrE2s (LstrE2 A/E/G2/H) were obtained by rapid-amplification of cDNA ends (RACE-PCR) analysis. Expression of the four LstrE2s showed tissue- and development-specific patterns. RT-qPCR analyses revealed that Rice stripe viruse (RSV) infection increased the level of LstrE2 A/E/G2/H. Further study indicated that repression of LstrE2 E via RNAi caused significant increases in the expression of RSV coat protein mRNA and protein levels. These findings suggest that LstrE2 E inhibits RSV accumulation in the planthopper body. Understanding the function of LstrE2 E in RSV accumulation may ultimately result in the development of novel antiviral strategies.

Planthopper Transmission of Ramu Stunt Virus, a Tenuivirus Causing the Sugarcane Disease Ramu Stunt, and its Distribution in Papua New Guinea.

Ramu stunt is a serious disease of sugarcane, currently only reported from Papua New Guinea. It is found in both commercial sugarcane grown on the Ramu Agri Industries Limited (RAIL) estate and in chewing canes (Saccharum officinarum L.) grown in village gardens. The vector of Ramu stunt disease is the island sugarcane planthopper, Eumetopina flavipes Muir. Here we report on the successful transmission of Ramu stunt using E. flavipes and verify that the disease is caused by Ramu stunt virus, a virus with homology to the genus Tenuivirus. Diagnostic reverse transcription PCR screening, with partial genome sequencing and viral protein characterization, was used for confirmation. Disease surveys were undertaken on the RAIL estate, along roadsides, and in village gardens in parts of Papua New Guinea. When the disease was identified, partial genome sequencing of the virus was performed to assess the extent of genome variability among isolates. The disease was less common than predicted from early surveys based on symptoms alone, and genotypic variation was associated with geographic location.

Immune function of an angiotensin-converting enzyme against Rice stripe virus infection in a vector insect.

Angiotensin-converting enzyme (ACE) plays diverse roles in the animal kingdom. However, whether ACE plays an immune function against viral infection in vector insects is unclear. In this study, an ACE gene (LsACE) from the small brown planthopper was found to respond to Rice stripe virus (RSV) infection. The enzymatic activities of LsACE were characterized at different pH and temperature. Twenty planthopper proteins were found to interact with LsACE. RSV infection significantly upregulated LsACE expression in the testicle and fat body. When the expression of LsACE in viruliferous planthoppers was inhibited, the RNA level of the RSV SP gene was upregulated 2-fold in planthoppers, and all RSV genes showed higher RNA levels in the rice plants consumed by these planthoppers, leading to a higher viral infection rate and disease rating index. These results indicate that LsACE plays a role in the immune response against RSV transmission by planthoppers.

Rice stripe virus hitchhikes the vector insect vitellogenin ligand-receptor pathway for ovary entry.

It is known that plant arboviruses infect insect vector cells by endocytosis; however, the cellular receptors that mediate endocytosis have not been well defined. In our recently published work and this study, by clarifying the vertical transmission mechanism of Rice stripe virus (RSV) in Laodelphax striatellus, we provide a novel paradigm for how arboviruses enter insect germ-line cells. Instead of direct interaction with a viral receptor, the virus binds to a secreted ligand protein, hitchhiking the ligand-receptor pathway to achieve cell entry. Vitellogenin (Vg) is an indispensable protein for embryo development that is synthesized extra-ovarially and taken up by germ-line cells through Vg receptor (VgR)-mediated endocytosis. After revealing that RSV invades L. striatellus ovary by a specific molecular interaction with the insect Vg in haemolymph, this study addressed VgR's function in mediating the RSV invasion of the germarium nurse cells, further confirming the ligand's receptor-mediated viral cell-invasion mechanism. Understanding the viral ovary-entry pathways in vectors will help to find suitable measures to block the trans-generation transmission of the viruses. This article is part of the theme issue 'Biotic signalling sheds light on smart pest management'.

The hypersensitive induced reaction 3 (HIR3) gene contributes to plant basal resistance via an EDS1 and salicylic acid-dependent pathway.

The hypersensitive-induced reaction (HIR) gene family is associated with the hypersensitive response (HR) that is a part of the plant defense system against bacterial and fungal pathogens. The involvement of HIR genes in response to viral pathogens has not yet been studied. We now report that the HIR3 genes of Nicotiana benthamiana and Oryza sativa (rice) were upregulated following rice stripe virus (RSV) infection. Silencing of HIR3s in N. benthamiana resulted in an increased accumulation of RSV RNAs, whereas overexpression of HIR3s in N. benthamiana or rice reduced the expression of RSV RNAs and decreased symptom severity, while also conferring resistance to Turnip mosaic virus, Potato virus X, and the bacterial pathogens Pseudomonas syringae and Xanthomonas oryzae. Silencing of HIR3 genes in N. benthamiana reduced the content of salicylic acid (SA) and was accompanied by the downregulated expression of genes in the SA pathway. Transient expression of the two HIR3 gene homologs from N. benthamiana or the rice HIR3 gene in N. benthamiana leaves caused cell death and an accumulation of SA, but did not do so in EDS1-silenced plants or in plants expressing NahG. The results indicate that HIR3 contributes to plant basal resistance via an EDS1- and SA-dependent pathway.

Olfactory co-receptor Orco stimulated by Rice stripe virus is essential for host seeking behavior in small brown planthopper.

BACKGROUND: Laodelphax striatellus, the small brown planthopper (SBPH), is an economically important pest, besides sucking damage, which transmits rice viruses to cause severe damages to rice. In the process of virus transmission, the host orientation behavior of insect is mainly driven by olfaction. In this context, the molecular basis of olfaction in SBPH is of particular interest. RESULTS: Here, we identified the gene that encodes olfactory receptor co-receptor (Orco) and analyzed its expression profiles in Rice stripe virus (RSV)-infected and RSV-free SBPH. It was found that LstrOrco shared high identity with other Orcos from different order insects. LstrOrco was mainly expressed in the head of SBPH, and its expression was significantly stimulated by RSV-infection. The behavioral bioassay revealed that viruliferous SBPH might have a stronger olfactory and seeking ability for rice than RSV-free insect. After silencing of LstrOrco expression, the olfaction and seeking behavior of nymphs for rice seedlings was significantly inhibited, mainly in the increase of the 'no response' percent and the prolongation of the response time. CONCLUSION: These results suggested that Orco played an important role in olfactory signaling and seeking behavior of SBPH, which provided a basic for future development of olfactory-based agriculture pest management strategies. © 2018 Society of Chemical Industry.

Immune responses induced by different genotypes of the disease-specific protein of Rice stripe virus in the vector insect.

Persistent plant viruses circulate between host plants and vector insects, possibly leading to the genetic divergence in viral populations. We analyzed the single nucleotide polymorphisms (SNPs) of Rice stripe virus (RSV) when it incubated in the small brown planthopper and rice. Two SNPs, which lead to nonsynonymous substitutions in the disease-specific protein (SP) of RSV, produced three genotypes, i.e., GG, AA and GA. The GG type mainly existed in the early infection period of RSV in the planthoppers and was gradually substituted by the other two genotypes during viral transmission. The two SNPs did not affect the interactions of SP with rice PsbP or with RSV coat protein. The GG genotype of SP induced stronger immune responses than those of the other two genotypes in the pattern recognition molecule and immune-responsive effector pathways. These findings demonstrated the population variations of RSV during the circulation between the vector insect and host plant.

The vitellogenin receptor has an essential role in vertical transmission of rice stripe virus during oogenesis in the small brown plant hopper.

BACKGROUND: The small brown plant hopper (SBPH), Laodelphax striatellus Fallén, is one of the most destructive pests on rice. This pest transmits rice stripe virus (RSV) both horizontally and vertically, leading to major yield and economic losses in rice production. However, the way that RSV particles enter oocytes of SBPH remains largely unknown. Thus, identification of key factors involved in the interaction between SBPH and RSV in the ovary is crucial. RESULTS: Transcriptome of non-viruliferous (NV) or high viruliferous (HV) SBPH ovaries at 24 and 48 h of emergence was sequenced. Differentially expressed genes analysis showed that vitellogenin receptor was significantly highly expressed in the ovary of the HV SBPH strains compared to NV strains. Quantitative real-time polymer chain reaction showed that the vitellogenin receptor in L. striatellus (LsVgR) was highly expressed in the ovaries of female adults and maintained a high level of expression at the early stage of ovary development. By using RNA interference, the expression of LsVgR in the ovaries of the HV strain was significantly decreased by 98.1%. RSV titer was reduced by 60.9% as quantified by viral RNA3 intergenic region and the transcripts of nucleocapsid protein gene (CP) reduced by 46.3%. The numbers of offspring hatched were significantly reduced in dsRNA-treated groups. The transcripts of CP were not affected by silencing LsVgR, whereas the abundance of RNA-dependent RNA polymerase increased by 15-fold in the member of surviving progenies. CONCLUSION: Our results suggest that vitellogenin receptor participates in regulating RSV replication during oogenesis. © 2018 Society of Chemical Industry.

New insights on the transmission mechanism of tenuiviruses by their vector insects.

Tenuiviruses, which cause serious diseases in rice, wheat, maize and other gramineae crops, recently have been assigned to the family Phenuiviridae in the order Bunyavirales. Transmission of tenuiviruses to host plants depends on the specific vector planthoppers. The interaction between the virus and insect offers critical points for developing an efficient management strategy. This review focuses on recent advancements in our understanding of the interactions between the virus and insect components. Vector components such as various proteins play major roles in virus replication, stability and transovarial transmission. The virus can either directly interact with these proteins or regulate expression of genes that encode them to alter the metabolism or defense mechanisms of the insect vectors. However, the vector components that are involved in virus infection and movement in midgut and salivary glands are not as well explored and are targets for further study.

NS3 Protein from Rice stripe virus affects the expression of endogenous genes in Nicotiana benthamiana.

BACKGROUND: Rice stripe virus (RSV) belongs to the genus Tenuivirus. It is transmitted by small brown planthoppers in a persistent and circulative-propagative manner and causes rice stripe disease (RSD). The NS3 protein of RSV, encoded by the viral strand of RNA3, is a viral suppressor of RNA silencing (VSR). NS3 plays a significant role in viral infection, and NS3-transgenic plants manifest resistance to the virus. METHODS: The stability and availability of NS3 produced by transgenic Nicotiana benthamiana was investigated by northern blot analysis. The accumulation of virus was detected by western blot analysis. Transcriptome sequencing was used to identify differentially expressed genes (DEGs) in NS3-transgenic N. benthamiana. RESULTS: When the host plants were inoculated with RSV, symptoms and viral accumulation in NS3-transgenic N. benthamiana were reduced compared with the wild type. Transcriptome analysis identified 2533 differentially expressed genes (DEGs) in the NS3-transgenic N. benthamiana, including 597 upregulated genes and 1936 downregulated genes. These DEGs were classified into three Gene Ontology (GO) categories and were associated with 43 GO terms. KEGG pathway analysis revealed that these DEGs were involved in pathways associated with ribosomes (ko03010), photosynthesis (ko00195), photosynthesis-antenna proteins (ko00196), and carbon metabolism (ko01200). More than 70 DEGs were in these four pathways. Twelve DEGs were selected for RT-qPCR verification and subsequent analysis. The results showed that NS3 induced host resistance by affecting host gene expression. CONCLUSION: NS3, which plays dual roles in the process of infection, may act as a VSR during RSV infection, and enable viral resistance in transgenic host plants. NS3 from RSV affects the expression of genes associated with ribosomes, photosynthesis, and carbon metabolism in N. benthamiana. This study enhances our understanding of the interactions between VSRs and host plants.

Insect tissue-specific vitellogenin facilitates transmission of plant virus.

Insect vitellogenin (Vg) has been considered to be synthesized in the fat body. Here, we found that abundant Vg protein is synthesized in Laodelphax striatellus hemocytes as well. We also determined that only the hemocyte-produced Vg binds to Rice stripe virus (RSV) in vivo. Examination of the subunit composition of L. striatellus Vg (LsVg) revealed that LsVg was processed differently after its expression in different tissues. The LsVg subunit able to bind to RSV exist stably only in hemocytes, while fat body-produced LsVg lacks the RSV-interacting subunit. Nymph and male L. striatellus individuals also synthesize Vg but only in hemocytes, and the proteins co-localize with RSV. We observed that knockdown of LsVg transcripts by RNA interference decreased the RSV titer in the hemolymph, and thus interfered with systemic virus infection. Our results reveal the sex-independent expression and tissue-specific processing of LsVg and also unprecedentedly connect the function of this protein in mediating virus transmission to its particular molecular forms existing in tissues previously known as non-Vg producing.

Altered accumulation of osa-miR171b contributes to rice stripe virus infection by regulating disease symptoms.

Viral infection affects the pattern of plant miRNA expression. It has been presumed that reduction of miR171 and several other miRNAs influences viral symptoms in plants. We here experimentally demonstrate the association of osa-miR171b with rice stripe virus (RSV) symptoms in rice. Inhibition of osa-miR171b caused stunting with reduced chlorophyll content in leaves similar to viral symptoms. Overexpression of osa-miR171b by an artificial miRNA extended vegetative growth and enhanced chlorophyll accumulation in leaves. Tillers were thicker, and panicles were longer with more spikelets in plants overexpressing osa-miR171b than in controls, but there were no differences in tiller numbers. Targets of osa-miR171b, OsSCL6-IIa, OsSCL6-IIb, and OsSCL6-IIc, were respectively up- and down-regulated in plants where osa-miR171b was inhibited or overexpressed. In plants overexpressing osa-miR171b, five positive regulators for heading development, Ehd1, Ehd2, Ehd3, Ehd4, and Hd3a were up-regulated, while the negative regulator Ghd7 was down-regulated. Plants overexpressing osa-miR171b were less susceptible to RSV and virus symptoms were attenuated. Taken together, the results reveal that a reduction of osa-miR171b in RSV-infected rice contributes to RSV symptoms, and provide more insight into the roles of osa-miR171b in rice.

The c-Jun N-terminal kinase pathway of a vector insect is activated by virus capsid protein and promotes viral replication.

No evidence has shown whether insect-borne viruses manipulate the c-Jun N-terminal kinase (JNK) signaling pathway of vector insects. Using a system comprising the plant virus Rice stripe virus (RSV) and its vector insect, the small brown planthopper, we have studied the response of the vector insect's JNK pathway to plant virus infection. We found that RSV increased the level of Tumor Necrosis Factor-α and decreased the level of G protein Pathway Suppressor 2 (GPS2) in the insect vector. The virus capsid protein competitively bound GPS2 to release it from inhibiting the JNK activation machinery. We confirmed that JNK activation promoted RSV replication in the vector, whereas JNK inhibition caused a significant reduction in virus production and thus delayed the disease incidence of plants. These findings suggest that inhibition of insect vector JNK may be a useful strategy for controling the transmission of plant viruses.