Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2023 Jan 15:324:199019.
doi: 10.1016/j.virusres.2022.199019. Epub 2022 Dec 8.

Voltage-dependent anion channel 2 (VDAC2) facilitates the accumulation of rice stripe virus in the vector Laodelphax striatellus

Lu Zhang  1 , Linying Li  1 , Lijun Huang  1 , Xinyi Li  1 , Chengzhu Xu  1 , Wenxing Hu  1 , Yixuan Sun  1 , Fang Liu  2 , Yao Li  3
Affiliations

Voltage-dependent anion channel 2 (VDAC2) facilitates the accumulation of rice stripe virus in the vector Laodelphax striatellus

Lu Zhang et al. Virus Res. .

Abstract

Rice stripe virus (RSV) causes enormous losses in rice production and is transmitted by the small brown planthopper, Laodelphax striatellus, in a persistent-propagative manner. RSV accumulation within the gut lumen of the vector is indispensable for the successful transmission to rice and insects. In this study, we obtained a 1464 bp full-length cDNA of a voltage-dependent anion channel 2 from L. striatellus (LsVDAC2), which encodes a 283 amino acid protein. RSV infection increased the expression of LsVDAC2 in the midguts and ovaries of L. striatellus by 260% and 228%, respectively. Silencing of LsVDAC2 resulted in a 88% reduction of RSV loads at 24 h after RNAi, indicating that LsVDAC2 facilitates RSV accumulation in the vector. Yeast two-hybrid and GST pulldown assays demonstrated that LsVDAC2 interacted with RSV RNA-dependent RNA polymerase, RdRp. Furthermore, experiments in vivo and in vitro showed that LsVDAC2 induced the apoptotic response in RSV-infected insects and tissues. Silencing of LsVDAC2 via RNAi significantly reduced the expression of genes for apoptosis-related caspases 1a and 1c by 62% and 78%, respectively, in RSV-infected vectors. Whether LsVDAC2-induced RSV accumulation is related to RSV RdRp and LsVDAC2-induced cell apoptosis deserves further investigation.

Keywords: Laodelphax striatellus; RNA-dependent RNA polymerase; Rice stripe virus; apoptosis; viral accumulation; voltage-dependent anion channel 2.

PubMed Disclaimer

Conflict of interest statement

Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Figures

Fig. 1
Fig. 1
Motif architecture and phylogenetic analysis of VDAC2 in L. striatellus. (A) The full-length cDNA, deduced amino acid sequence and motifs of LsVDAC2. A four-element eukaryotic porin signature is demarcated by blue ovals and open rectangles in the diagram and amino acid sequence, respectively. A glycine-leucine-lysine (GLK) motif is represented by a red rectangle and underscored residues in the diagram and amino acid sequence, respectively. A VKAKV-like motif is shown by an orange rectangle in the diagram and in the sequence by shaded amino acids. (B) The maximum-likelihood phylogenetic tree was constructed based on the sequences of LsVDAC2 and VDACs from other insects (N. lugens, D. melanogaster, A. aegypti, A. glabripennis and S. oryzae) and mammals (VDAC1, VDAC2 and VDAC3 from M. ochrogaster, L. africana, and E. caballus, respectively). Numbers above branches represent bootstrap values.
Fig. 2
Fig. 2
LsVDAC2 transcription and protein production in non-infected and RSV-infected L. striatellus. (A) RT-qPCR analysis of LsVDAC2 transcription in whole bodies of non-infected and RSV-infected adults. (B) Western blot analysis of LsVDAC2 protein production in whole bodies of non-infected and RSV-infected adults. GAPDH was used as a control. Image shows the bands from three independent samples, and the mean values and significance of relative intensities were marked above three repeated bands. (C) LsVDAC2 mRNA expression in ovaries, midguts, salivary glands and hemolymph of non-infected and RSV-infected adults. (D) Western blot analysis of LsVDAC2 protein production in ovaries, midguts, salivary glands and hemolymph of non-infected and RSV-infected adults. Image shows the relative intensities of protein bands from two independent samples. Means ± SE, t-test analysis: * P < 0.05, ** P < 0.01.
Fig. 3
Fig. 3
dsVDAC treatment inhibits RSV loads in L. striatellus.In this experiment, RSV-infected planthoppers were microinjected with dsGFP or dsVDAC. Whole bodies of RSV-infected planthoppers were analyzed for LsVDAC2 expression (A) and viral loads (RSV NP expression) (B) at 6, 12, 18, 24 and 48 h after RNAi treatment by RT-qPCR. (C) Protein levels of LsVDAC2 and RSV NP at 6-48 h post dsVDAC treatment were analyzed by immunoblotting, and GAPDH was used as control. The band intensity from three independent samples shows above each band. (D) Immunofluorescence microscopy of midguts (a–b”’), ovaries (c-d’”) and salivary glands (e–f”’) of RSV-infected adults. Images were obtained 24 h after treatment with dsVDAC or dsGFP (control) and were immunolabeled with anti-RSV NP (Alexa Fluor 488, green) or anti-LsVDAC2 (Alexa Fluor 555, red). Values represent means ± SE, and the student's t-test was used to analyze significance. * P < 0.05, ** P < 0.01. Bars, 50 μm.
Fig. 4
Fig. 4
Interactions between LsVDAC2 and RSV proteins in yeast two-hybrid and GST pull-down assays.(A) Analysis of interactions between LsVDAC2 and seven RSV proteins using Y2H assays. Yeast cells co-transfected with LsVDAC2 and one of the seven RSV proteins (diluted 10−1 to 10−4) were cultured on QDO (SD/-Ade/-His/-leu/-Trp-20 mM3-AT) medium and evaluated for β-galactosidase activity (blue color). AD-T + BD-53 was regarded as a positive control, and AD-T + BD-Lam was the negative control. The interaction of LsVDAC2 with RSV RdRp protein was evaluated by dividing the latter into five functional regions, namely: RdRP1 (1-486 aa); RdRp2 (480-988 aa); RdRp3 (982-1811 aa); RdRp4 (1805-2388 aa); and RdRp5 (2381-2920 aa). Abbreviations: AD, activation domain, cloned in pGADT7; BD, bait domain, cloned in pGBKT7. (B) GST pull-down assays were conducted to confirm the interaction of LsVDAC2 and RSV RdRp1. RdRp1 fused with GST was incubated with Non-infected adult extracts. Similarly, insect extracts were incubated with GST as a negative control. Blots were probed with the anti-LsVDAC2 or anti-GST antibodies.
Fig. 5
Fig. 5
The effect of LsVDAC2 on RSV-induced apoptosis in L. striatellus. (A) Apoptotic cells were labeled by TUNEL (green) in non-infected, RSV-infected, and dsGFP/dsVDAC treated RSV-infected adults. Nucleoli were then immunolabeled with DAPI/ antifade solution (blue) and observed by confocal microscopy. Bars, 500 μm. (B) Apoptotic cells were detected by TUNEL signals (green) in RSV-infected Sf9 cells transfected with LsVDAC2, and nucleoli were stained with propidium iodide (PI, red). RSV-infected cells transfected with eGFP served as a control. Bars, 50 μm. (C) Relative mRNA expression of three apoptosis-related genes (Caspase1a, Capase1c, and Caspase8) by RT-qPCR. Values represent means ± SE. The student's t-test was used to analyze significance: (*), P<0.05, (**), P<0.01.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

References

    1. Bolter B., Soll J. Ion channels in the outer membranes of chloroplasts and mitochondria: open doors or regulated gates? EMBO. J. 2001;20(5):935–940. - PMC - PubMed
    1. Chang J.S., Chi S.C. Grouper voltage-dependent anion selective channel protein 2 is required for nervous necrosis virus infection. Fish Shellfish Immunol. 2015;46(2):315–322. - PubMed
    1. Chanturiya A.N., Basanez G., Schubert U., Henklein P., Yewdell J.W., Zimmerberg J. PB1-F2, an influenza A virus-encoded proapoptotic mitochondrial protein, creates variably sized pores in planar lipid membranes. J. Virol. 2004;78(12):6304–6312. - PMC - PubMed
    1. Chen Q., Wei T. Cell biology during infection of plant viruses in insect vectors and plant hosts. Mol. Plant. Microbe. Interact. 2020;33(1):18–25. - PubMed
    1. Chen Q., Zheng L., Mao Q., Liu J., Wang H., Jia D., Chen H., Wu W., Wei T. Fibrillar structures induced by a plant reovirus target mitochondria to activate typical apoptotic response and promote viral infection in insect vectors. PLoS. Pathog. 2019;15(1) - PMC - PubMed

Publication types

Substances

-