Use of virus vectors for the expression in plants of active full‐length and single chain anti‐coronavirus antibodies

Abstract

To extend the potential of antibodies and their derivatives to provide passive protection against enteric infections when supplied orally in crude plant extracts, we have expressed both a small immune protein (SIP) and a full-length antibody in plants using two different plant virus vectors based on potato virus X (PVX) and cowpea mosaic virus (CPMV). The alphaSIP molecule consisted of a single chain antibody (scFv) specific for the porcine coronavirus, transmissible gastroenteritis virus (TGEV) linked to the alpha-CH3 domain from human IgA. To express the full-length IgA, the individual light and heavy chains from the TGEV-specific mAb 6A.C3 were inserted into separate PVX constructs and plants were co-infected with both constructs. Western blot analysis revealed the efficient expression of both the SIP and IgA molecules. Analysis of crude plant extracts revealed that both the plant-expressed alphaSIP and IgA molecules could bind to and neutralize TGEV in tissue culture, indicating that active molecules were produced. Oral administration of crude extracts from antibody-expressing plant tissue to 2-day-old piglets showed that both the alphaSIP and full-length IgA molecules can provide in vivo protection against TGEV.

Expression of multiple proteins using full‐length and deleted versions of cowpea mosaic virus RNA‐2

“…Verch et al . (1998) and Alamillo et al . (2006) used vectors based on tobacco mosaic virus (TMV) and potato virus X (PVX), respectively, and, in each case, infected plants with separate constructs expressing the heavy and light chains.…”

“…This is a significant drawback when using the natural host of CPMV, cowpea ( Vigna unguiculata ), which possesses a thick cuticle, which must be extensively damaged to allow constructs to be agroinfiltrated into leaves. However, cowpea has the advantage as an expression system that it is an edible plant and has already been used for the expression of HBcAg and SIPs (Alamillo et al ., 2006; Mechtcheriakova et al ., 2006; Monger et al ., 2006). We have recently conducted preliminary experiments on the expression of a human anti‐human immunodeficiency virus (anti‐HIV) IgG in cowpea, which have shown that deleted RNA‐2 molecules can be used to produce assembled antibodies in this host (F. Sainsbury and G. P. Lomonossoff, unpubl.…”

“…The resulting RNA‐2 molecules are capable of spreading both within and between plants. This strategy has been used to express a number of recombinant proteins, such as the hepatitis B core antigen (HBcAg) and small immune proteins (SIPs), in cowpea plants (Alamillo et al ., 2006; Mechtcheriakova et al ., 2006; Monger et al ., 2006). The plant‐expressed HBcAg has been shown to self‐assemble into core particles, and the SIP molecules, consisting of a single‐chain antibody (scFv) fused to a domain from the constant region of a heavy chain, are able to dimerize in planta and neutralize infectivity in vivo .…”

Production systems for recombinant antibodies

“…(149). Rapid transient antibody productions in plants has also been performed using modified plant viral vectors (150,151) with production levels of up to 0.7 % of total protein. Monocotyledonous plants are mostly transfected by particle gun (152).…”

Virus‐Specific Read‐Through Codon Preference Affects Infectivity of Chimeric Cucumber Green Mottle Mosaic Viruses Displaying a Dengue Virus Epitope

“…The development of plant virus vectors as in planta expression systems for foreign genes provides an attractive alternative biotechnological approach for peptide expression [1][2][3][4][5]. This method has been exploited in vaccine production, where small foreign peptides are expressed as a fusion with the viral coat proteins.…”