Expression of a Single-Chain Variable-Fragment Antibody Against a Fusarium virguliforme Toxin Peptide Enhances Tolerance to Sudden Death Syndrome in Transgenic Soybean Plants

Abstract

Plants do not produce antibodies. However, plants can correctly assemble functional antibody molecules encoded by mammalian antibody genes. Many plant diseases are caused by pathogen toxins. One such disease is the soybean sudden death syndrome (SDS). SDS is a serious disease caused by the fungal pathogen Fusarium virguliforme. The pathogen, however, has never been isolated from diseased foliar tissues. Thus, one or more toxins produced by the pathogen have been considered to cause foliar SDS. One of these possible toxins, FvTox1, was recently identified. We investigated whether expression of anti-FvTox1 single-chain variable-fragment (scFv) antibody in transgenic soybean can confer resistance to foliar SDS. We have created two scFv antibody genes, Anti-FvTox1-1 and Anti-FvTox1-2, encoding anti-FvTox1 scFv antibodies from RNAs of a hybridoma cell line that expresses mouse monoclonal anti-FvTox1 7E8 antibody. Both anti-FvTox1 scFv antibodies interacted with an antigenic site of FvTox1 that binds to mouse monoclonal anti-FvTox1 7E8 antibody. Binding of FvTox1 by the anti-FvTox1 scFv antibodies, expressed in either Escherichia coli or transgenic soybean roots, was initially verified on nitrocellulose membranes. Expression of anti-FvTox1-1 in stable transgenic soybean plants resulted in enhanced foliar SDS resistance compared with that in nontransgenic control plants. Our results suggest that i) FvTox1 is an important pathogenicity factor for foliar SDS development and ii) expression of scFv antibodies against pathogen toxins could be a suitable biotechnology approach for protecting crop plants from toxin-induced diseases.

Analyses of the Xylem Sap Proteomes Identified Candidate Fusarium virguliforme Proteinacious Toxins

“…Brar and M.K. Bhattacharyya, unpublished) and shown to be involved in SDS development in foliar tissues [16]. Most likely FvTox1 is produced at much lower concentration which was insufficient for detection in our LC-ESI-MS/MS study.…”

“…We have shown that FvTox1 is a major virulence factor that is involved in foliar SDS in soybean [13], [16], [17]. The foliar SDS was not completely absent in fvtox1 infected soybean plants [17] suggesting that additional toxins may be involved in foliar SDS development; and it is conceivable that such toxins may be present in xylem sap of infected soybean plants.…”

“…This toxin can rapidly cause foliar SDS-like symptoms in leaf discs of soybean lines, highly susceptible to F. virguliforme [13]. Expression of a single chain variable fragment (scFv) antibody against FvTox1 enhanced foliar SDS resistance in transgenic soybean plants supporting the role of FvTox1 in foliar SDS development [16]. Investigation of fvtox1 mutants suggested that FvTox1 is a major virulence factor involved in foliar SDS.…”

Advancements in Transgenic Soy: From Field to Bedside

“…In fact, antibody production in soybean was first demonstrated in 1998 [30]. A similar antibody approach was recently taken by Brar and Bhattacharyya to control Fusarium virguliforme which is responsible for soybean sudden death syndrome (SDS) [31]. Using the pathogenic toxin Tox1 as a target, soybeans were transformed with an antibody gene encoding scFv anti-FvTox1 to create transgenic lines with enhanced foliar SDS resistance compared to control plants.…”

Investigation of the Fusarium virguliforme Transcriptomes Induced during Infection of Soybean Roots Suggests that Enzymes with Hydrolytic Activities Could Play a Major Role in Root Necrosis

“…virguliforme culture media [11]. Expression of a single-chain variable fragment (scFv) antibody against this toxin enhanced foliar SDS resistance in transgenic soybean plants [16]. Knockout mutants created by homologous recombination established that FvTox1 is a major virulence factor for foliar SDS development in soybean [17].…”