Coordinate expression of antibody subunit genes yields high levels of functional antibodies in roots of transgenic tobacco

Plant Mol Biol. 1994 Dec;26(6):1701-10. doi: 10.1007/BF00019485.

Abstract

To explore the feasibility of employing antibodies to obtain disease resistance against plant root pathogens, we have studied the expression of genes encoding antibodies in roots of transgenic plants. A model monoclonal antibody was used that binds to a fungal cutinase. Heavy and light chain cDNAs were amplified by PCR, fused to a signal sequence for secretion and cloned behind CaMV 35S and TR2' promoters in a single T-DNA. The chimeric genes were cloned both in tandem and in a divergent orientation. The roots of tobacco plants transformed with these constructs produced antibodies that were able to bind antigen in an ELISA. Immunoblotting showed assembly to a full-size antibody. In addition, a F(ab')2-like fragment was observed, which is probably formed by proteolytic processing. Both antibody species were properly targeted to the apoplast, but the full-size antibody was partially retained by the wall of suspension cells. The construct with divergent promoters showed a better performance than the construct with promoters in tandem. It directed the accumulation of functional antibodies to a maximum of 1.1% of total soluble protein, with half of the plants having levels higher than 0.35%. The high efficiency of this construct probably results from coordinated and balanced expression of light and heavy chain genes, as evidenced by RNA blot hybridization.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Antibodies, Fungal / biosynthesis*
  • Antibodies, Fungal / genetics
  • Antibodies, Monoclonal / biosynthesis
  • Antibodies, Monoclonal / genetics
  • Antigen-Antibody Reactions
  • Base Sequence
  • Carboxylic Ester Hydrolases / immunology*
  • Cloning, Molecular
  • Enzyme-Linked Immunosorbent Assay
  • Genetic Engineering
  • Immunoglobulin Heavy Chains / biosynthesis
  • Immunoglobulin Heavy Chains / genetics
  • Immunoglobulin Light Chains / biosynthesis
  • Immunoglobulin Light Chains / genetics
  • Molecular Sequence Data
  • Mycoses / prevention & control
  • Plant Diseases
  • Plant Roots / immunology*
  • Plant Roots / metabolism
  • Plants, Genetically Modified
  • Plants, Toxic*
  • Promoter Regions, Genetic / genetics
  • Protein Processing, Post-Translational
  • Tobacco / genetics
  • Tobacco / immunology*
  • Tobacco / metabolism
  • Transformation, Genetic

Substances

  • Antibodies, Fungal
  • Antibodies, Monoclonal
  • Immunoglobulin Heavy Chains
  • Immunoglobulin Light Chains
  • Carboxylic Ester Hydrolases
  • cutinase

Associated data

  • GENBANK/L35037
  • GENBANK/L35138