Plant-produced mouse zona pellucida 3 peptide, its immunogenicity and contraceptive efficacy

Abstract

Fertility control is a promising method for controlling the population of highly fertile species such as wild mice. Immunization with the mouse-specific contraceptive zona pellucida 3 peptide (mZP3) reduces fertility in some mouse strains, including wild mice. Here, we employed a plant expression system for the production of glycosylated mZP3 and evaluated the immunogenicity of plant-produced mZP3-based antigens in a female BALB/c mouse model. In the mZP3-1 antigen, mZP3 fused with a T-cell epitope of tetanus toxoid, a histidine tag and a SEKDEL sequence. A fusion antigen (GFP-mZP3-1) and a polypeptide antigen containing three repeats of mZP3 (mZP3-3) were also examined. Glycosylation of mZP3 should be achieved by targeting proteins to the endoplasmic reticulum. Agrobacterium-mediated transient expression of antigens resulted in successful production of mZP3 in Nicotiana benthamiana leaves. Compared with mZP3-1, GFP-mZP3-1 and mZP3-3 increased the production of the mZP3 peptide by more than 20 and 25 times, respectively. The glycosylation of the proteins was indicated by their size and their binding to a carbohydrate-binding protein. Both plant-produced GFP-mZP3-1 and mZP3-3 antigens were immunogenic in mice; however, mZP3-3 generated significantly higher levels of serum antibodies against mZP3. Although, as previously reported, the formation of antibodies did not lead to a significant reduction in fertility in BALB/c mice, strong and specific binding of antibodies to wild mouse zona pellucida was observed. Therefore, we assume that the plants might be an efficient system for the production of mZP3-3 as a contraceptive vaccine to reduce fertility in wild mice.

Immunogenicity and contraceptive efficacy of plant-produced putative mouse-specific contraceptive peptides

“…In accordance with our previous report that three repeats of mZP3 peptide significantly increased the antigen yield (Ghasemian et al, 2023b), the production of mZP2 and mIzumo1 in the form of three-peptide antigens resulted in sufficient amounts of the peptides in plants in this study. Our results confirmed the successful expression of the C-terminal His-tagged mZP2-3 construct.…”

“…Moreover, plant-produced peptides can be bioencapsulated in plant cells and protected from the gastro-intestinal environment that can limit the efficiency of free orally-delivered peptides (Kwon and Daniell, 2016;Pantazica et al, 2021). In addition, expression of contraceptive peptides in plants in fusion with highly stable proteins such as virus like particles can significantly improve their stability and immunogenicity to induce systemic and mucosal immunity in orally vaccinated mice (Ghasemian et al, 2023a).…”

“…Peptide vaccines are less effective compared with protein vaccines (Ferro and Mordini, 2004). However, incorporation of adjuvants and multiple epitopes into antigens can improve the immunogenicity and efficacy of peptide-based antigens (Sadler et al, 2002;Ferro and Mordini, 2004;Hardy et al, 2004;Ghasemian et al, 2023a). Production of peptide-based mouse-specific contraceptive antigens facilitates the oral administration of vaccines to widely dispersed animals.…”