Exosomes mediate horizontal transmission of viral pathogens from insect vectors to plant phloem
- Vector-borne Virus Research Center, Fujian Province Key Laboratory of Plant Virology, Institute of Plant Virology, Fujian Agriculture and Forestry University, China
Figures
Figure 1 with 2 supplements
Transmission electron micrographs showing that RDV traverses the apical plasmalemma into the salivary cavities via the exosomal release pathway.
At 14 days padp, virus-infected salivary glands of N. cincticeps were examined by transmission electron microscopy. (A) Salivary cavities in type III cells. (B–C) Virus-containing MVBs at the …
Figure 1—figure supplement 1
RDV dissemination into the salivary cavities in the PSG.
(A) Transmission light micrographs of salivary glands of the leafhopper vector N. cincticeps. Leafhopper salivary glands consist of a pair of PSGs, each containing six types of cells (I to IV), and …
Figure 1—figure supplement 2
Immunogold labeling of RDV antigens on viral particles within the MVBs or exosomes in the leafhopper salivary glands or cultured cells.
Virus-infected salivary glands (A–C) or cultured cells (D) were immunolabeled with virus-specific IgG as primary antibodies, followed by treatment with 15 nm gold particle-conjugated goat antibodies …
Figure 2
RDV dissemination into the salivary cavities via Rab27a-involved exosomal release pathway.
(A–B) Immunogold labeling of Rab27a (A) or CD63 (B) on virus-containing MVBs or exosomes in the salivary glands of N. cincticeps. Virus-infected (panels I and II) or uninfected (panels III) salivary …
Figure 3
Knockdown of Rab27a expression reduces viral dissemination from salivary glands via the exosomal release pathway.
(A) RT-qPCR assay showing the transcript levels of RDV P8 and Rab27a expression in the salivary glands of dsRab27a- or dsGFP-treated insects. Results were normalized against the actin expression …
Figure 4 with 1 supplement
The secretion of RDV from cultured insect vector cells via the exosomal release pathway.
(A) At 48 hpi, virus-infected cultured leafhopper cells were examined by transmission electron microscopy. RDV virions were sequestered in the endosomal compartments (I), which then developed into …
Figure 4—figure supplement 1
Effects of treatment with dsRab27a, dsGFP, GW4869 (20 μM), or DMSO on the accumulation of cell-associated and extracellular viruses in cultured leafhopper cells infected with RDV (MOI of 10) at 48 hpi.
The relative transcript levels of RDV P8 were examined in the cell-associated and extracellular viruses after different treatments. Results were normalized against the actin expression level; the …
Figure 5 with 3 supplements
RDV P2 interacts with endosomal Rab5 of leafhopper.
(A) Schematic representation of the structures of RDV particle and P2 protein. P2 is composed of an L-shaped flexible structure with a 15 nm domain (aa 1–688, P2N) toward the exterior, and a 10 nm …
Figure 5—figure supplement 1
Bioinformatic analyses of Rab5 of N. cincticeps.
(A) Schematic representation of N. cincticeps Rab5. (B) Phylogenetic relationships of N. cincticeps Rab5 (in red) with its counterparts. The available sequences were aligned using ClustalW, and a …
Figure 5—figure supplement 2
Bioinformatic analyses of Rab27a of N. cincticeps.
(A) Schematic representation of N. cincticeps Rab27a. (B) Phylogenetic relationships of N. cincticeps Rab27a (in red) with its counterparts. The available sequences were aligned using ClustalW, and …
Figure 5—figure supplement 3
Basic Local Alignment Search Tool of Protein Databases (BLASTP) showing the similarity of Rab27a (A) or Rab5 (B) from N. cincticeps with homologues from Drosophila.
Figure 6 with 2 supplements
RDV P2N is associated with the endosomal compartments in Sf9 cells.
(A) Immunofluorescence assay showing the colocalization of RDV P2N with Rab5 of N. cincticeps in Sf9 cells. RDV P2N-His and Rab5-Myc were expressed alone or co-expressed. Sf9 cells were fixed at 48 …
Figure 6—figure supplement 1
The effects of chloroquine or GW4869 on the expression of P2N-His in Sf9 cells.
(A) Immunofluorescence microscopy showing the expression of P2N-His in Sf9 cells treated with chloroquine or GW4869 at different concentrations. Each panel is representative of three independent …
Figure 6—figure supplement 2
The effects of chloroquine or GW4869 on the expression of P2N-GFP in Sf9 cells.
(A–B) Immunofluorescence microscopy showing the expression of P2N-GFP in Sf9 cells. Bars, 10 μm (A) and 50 μm (B). (C) The fluorescence intensity of P2N-GFP in cells treated with chloroquine or …
Figure 7 with 1 supplement
Exosome-mediated RDV delivery to the phloem of rice plants via leafhopper feeding.
(A–C) Immunofluorescence microscopy showing the distribution of RDV antigens, Rab27a, CD63 or Rab5 in the phloem of rice plants after 2 day feeding of viruliferous or nonviruliferous leafhoppers. …
Figure 7—figure supplement 1
Schematic illustration of the feeding cage used for collecting components released from leafhopper salivary glands.
Fifteen viruliferous leafhoppers were confined for 2 days, respectively, in pipe-like cages (2.5 cm in diameter by 4 cm in height), in which one end was covered by mesh and the other end was fixed to …
Figure 8
Proposed model of RDV hijacking the exosomal release pathway to be released from salivary glands of leafhopper vectors into rice phloem.
In the proposed model, the specific interaction of RDV P2N and Rab5 causes the internalization and delivery of viruses to endosomes. Then, these virus-containing endosomes are engulfed in the MVBs …
