Viruses can cause severe infectious symptoms not only in humans and animals but also in plants serving as a threat to the production of agricultural crops. Therefore, prevention of plant virus infections is a major objective in crop protection. There are various traditional strategies for controlling the viral diseases such as crop rotation, pathogen free plant production through tissue culture and integrated vector management, however, direct control strategy for viral infection in plants are limited.

Advancement in technology has resulted in a novel approach, which uses peptide aptamers for controlling plant viruses. Selection and screening of peptide aptamers against plant viral proteins is usually carried out using yeast two-hybrid system. Peptide aptamers are effective in conferring high level of specificity to the viral target such as capsid proteins (CP), nucleoproteins (N) and movement proteins (MP). This in turn inhibits the process of replication or viral assembly by expressing a dominant interfering peptide (Figure 1).

Figure 1. Flow chart displaying broad-spectrum mechanism for controlling the viral infection in plants by using peptide aptamers (Adapted from Yadav et al., 2019).

This aptamer strategy also offers additional advantages because of the difficulties associated with the expression of antibodies in the plant cytoplasm. In order to devise a new knowledge based strategy to engineer virus-resistant plants, following researchers investigated the use of peptide aptamers to produce broad-spectrum viral resistance in plants.

Novel strategy for generating resistance to plant viruses using peptide aptamers

Rudolph et al., 2003 for the first time, reported the feasibility of using peptide aptamers as an in vivo tool to control viral infection namely, tomato spotted wilt virus tospovirus (TSWV) in higher plants.  Peptide aptamers specific for the target TSWV nucleoprotein (TSWV N) were selected using yeast two-hybrid system in which a TSWV N mutant with no homopolymerization ability was used as the bait. The selected overlapping peptides covering C-terminal of TSWV N protein were isolated and named as T220/248. The isolated peptide aptamer was later fused with beta-glucuronidase (Gus) as a carrier protein and transgenic plants expressing the fusion protein (GusT) was then generated. These transgenic lines showed strong interaction and resistance to TSWV N protein and other tospovirus species, whereas, all inoculated control lines developed symptoms and eventually died (Figure 2).

Figure 2. (Top image) – Interaction between GusT peptide fusion and controls with TSWV N protein was investigated using yeast two hybrid system and β- galactosidase activity assay. (Bottom left) – TSWV inoculated plants evaluated 23 days after inoculation (A) Wild type (B) Gus expressing unmodified glucuronidase (C) GusT expressing the β- glucuronidase peptide fusion. (Bottom right) – Average plant height of transgenic and wild type plant lines after inoculation with different Tospovirus species (23 dpi).

Applications of peptide aptamers for resistance against geminivirus in plants

Geminiviruses (GV), Tomato yellow leaf curl virus (TYLCV) and Tomato mottle virus (ToMoV) constitutes a large family of ssDNA viruses that cause diseases in food and plants creating serious losses in important crops worldwide. To overcome these challenges, Reyes et al., 2013 created a novel resistance strategy using peptide aptamers to target the conserved motifs in the N terminus of GV Rep protein, which has been an essential player in GV infection. With this attractive approach, peptide aptamers could inhibit functions of viral proteins responsible for infections and act as antiviral agents. Two sets of peptide aptamers A22 and A64 were able to bind the Rep/AL1 proteins of nine different viruses in a yeast two-hybrid assay (Figure 3).

Figure 3.  (Top image) – Yeast two-hybrid growth assay. Selected peptide aptamers tested for interaction with TGMV Rep in yeast cells. (Bottom image) – ToMoV infected A22 and A64 expressing tomato lines have milder symptoms and lower viral DNA loads.

The authors further demonstrated that peptide aptamers could be extended to produce virus resistance in tomato. Transgenic tomato lines expressing peptide aptamers A22 and A64 and inoculated with TSWV and TYLCV exhibited delayed viral DNA accumulation and contained lower levels of viral DNA, displaying reduced symptoms as compared to the wild type tomato. Together these results established the efficacy of using Rep-binding peptide aptamers to develop crops that are resistant to diverse geminiviruses. This peptide aptamer approach could also be expanded to other geminivirus proteins and viral processes to enhance the resistance of the phenotype.

Overall, peptide aptamers have served as a promising approach to battle new infection species or new variations related to plant viruses. Moreover, because of their useful properties, peptide aptamers have significant potential in plant biotechnology and food safety, ranging from broad viral resistance in plants to biosensors for detecting contamination in food and feed.

At Aptamer Group Ltd, projects involving viral protein targets along with small molecules are carried out with customer defined end applications in mind. For example, we isolate aptamers that are able to bind to the target in buffered cell lysate or culture harvest. After harvest, we then use customer defined elution conditions to ensure that the aptamers release the target in a buffer that is compatible with the target and downstream applications. If you would like more information on how aptamers can be used in your research, please contact us using the form below.

References:

Reyes MI, Nash TE, Dallas MM, Ascencio-Ibáñez JT, Hanley-Bowdoin L. Peptide aptamers that bind to geminivirus replication proteins confer a resistance phenotype to tomato yellow leaf curl virus and tomato mottle virus infection in tomato. J Virol. (2013);87 (17):9691-9706. doi:10.1128/JVI.01095-13

Rudolph C, Schreier PH, Uhrig JF. Peptide-mediated broad-spectrum plant resistance to tospoviruses. Proc Natl Acad Sci U S A. (2003);100 (8):4429-4434.   doi:10.1073/pnas.0730832100

Yadav PK, Kumar S, Yadav S, Kumar S.Role of Aptamers in Plant Defense Mechanism Against Viral Diseases (2019). In: Yadav G., Kumar V., Aggarwal N. (eds) Aptamers. Springer, Singapore

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