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Single-chain antibodies against a plant viral RNA-dependent RNA polymerase confer virus resistance

Abstract

Crop loss due to viral diseases is still a major problem for agriculture today. We present a strategy to achieve virus resistance based on the expression of single-chain Fv fragments (scFvs) against a conserved domain in a plant viral RNA-dependent RNA polymerase (RdRp), a key enzyme in virus replication. The selected scFvs inhibited complementary RNA synthesis of different plant virus RdRps in vitro and virus replication in planta. Moreover, the scFvs also bound to the RdRp of the distantly related hepatitis C virus. T1 and T2 progeny of transgenic lines of Nicotiana benthamiana expressing different scFvs either in the cytosol or in the endoplasmic reticulum showed varying degrees of resistance against four plant viruses from different genera, three of which belong to the Tombusviridae family. Virus resistance based on antibodies to RdRps adds another tool to the repertoire for combating plant viruses.

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Figure 1: scFvs inhibit the activities of viral RdRps based on in vitro assays.
Figure 2: Selected scFvs target motif E of RdRp.
Figure 3: Binding of three scFvs to HCV RdRp in vitro.
Figure 4: ScFv-mediated inhibition of the replication of TBSV-BS3 in N. benthamiana.
Figure 5: T1 and T2 scFv-transgenic N. benthamiana plants show resistance to TBSV and CNV and partial resistance to TCV and RCNMV.

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Acknowledgements

We thank Judit Pogany for providing CNV RdRp and K.S. Rajendran for providing TCV RdRp. We also thank Gudrun Mönke for anti-FUS3-scFv and Ralf Bartenschlager for the HCV NS5B preparation. We thank Jan-Peter Nap (Hanze Hogeschool, Groningen), Michael Wassenegger and Pascal Cobanov for critically reviewing this manuscript and for helpful discussion. We are grateful to Sasithorn Chotiwutmontri and Claudia Eisenhauer for technical assistance. This work was supported by a grant from the Stiftung Rheinland-Pfalz für Innovation awarded to G.K. and in part by a National Science Foundation award to P.D.N.

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Correspondence to Gabi Krczal.

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Boonrod, K., Galetzka, D., Nagy, P. et al. Single-chain antibodies against a plant viral RNA-dependent RNA polymerase confer virus resistance. Nat Biotechnol 22, 856–862 (2004). https://doi.org/10.1038/nbt983

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