Transformation strategies for stable expression of complex hetero‐multimeric proteins like secretory immunoglobulin A in plants

Abstract

Summary Plant expression systems have proven to be exceptional in producing high‐value complex polymeric proteins such as secretory IgAs ( SI gAs). However, polymeric protein production requires the expression of multiple genes, which can be transformed as single or multiple T‐ DNA units to generate stable transgenic plant lines. Here, we evaluated four strategies to stably transform multiple genes and to obtain high expression of all components. Using the in‐seed expression of a simplified secretory IgA ( sSIgA ) as a reference molecule, we conclude that it is better to spread the genes over two T‐ DNA s than to contain them in a single T‐ DNA , because of the presence of homologous recombination events and gene silencing. These T‐ DNA s can be cotransformed to obtain transgenic plants in one transformation step. However, if time permits, more transformants with high production levels of the polymeric protein can be obtained either by sequential transformation or by in‐parallel transformation followed by crossing of transformants independently selected for excellent expression of the genes in each T‐DNA.

High Level Production of Monoclonal Antibodies Using an Optimized Plant Expression System

“…In many cases, co-expression of two or more proteins is required to produce biologically active pharmaceutical proteins, such as monoclonal antibodies, IgA, or multi-component virus like particles (Thuenemann et al, 2013;Palaci et al, 2019). To further improve the utility of plant-based systems, we have FIGURE 7 | Characterization of c2A10G6.…”

Prevention of Shiga toxin 1-caused colon injury by plant-derived recombinant IgA

“…As to several mucosal pathogens, it has been reported that pathogen-specific IgA can prevent infection 31 , 41 – 45 . As to Stx, previous studies indicated that mucosal vaccination could prevent disease and induce production of toxin-specific IgA antibodies, however, the effect of passively administered Stx-specific IgA was not clarified 33 , 39 , 46 .…”

Simplified monomeric VHH-Fc antibodies provide new opportunities for passive immunization

“…To obtain simplified SIgA antibodies (sSIgAs), the J chain and secretory component have to be co-expressed with the VHH-IgA molecule [7][8][9]. Several methods could be used for introducing the three genes in the same cell, such as simultaneous introduction -as a tandem construct or triple co-transformation, or rather successive introduction -either by successive supertransformation or genetic crossing.…”

“…Several methods could be used for introducing the three genes in the same cell, such as simultaneous introduction -as a tandem construct or triple co-transformation, or rather successive introduction -either by successive supertransformation or genetic crossing. In dicot seeds, the highest level of sSIgA expression has been obtained by first identifying transformants expressing high amounts of both the J chain and the secretory component, and super-transforming or crossing them with plants expressing the bivalent monomeric VHH-IgA molecule [8]. Indeed, expression of several genes in the same plant cell, even if the same promoter is used, does not necessarily result in the same amounts of transcripts or co-translated constituent protein chains of the sSIgA.…”