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Rice-derived SARS-CoV-2 glycoprotein S1 subunit vaccine elicits humoral and cellular immune responses
Saturday, 2025/04/12 | 08:45:00

Li SongYaya WenYu ZhouHui ZhangYuqi TianJing WangYaodan CuiRuimeng TanDan XiongChuang MengYan ZhouQianfeng LiZhiming PanQiaoquan LiuXinan Jiao

Plant Biotechnology Journal; First published: 04 April 2025; https://doi.org/10.1111/pbi.70077

Summary

Since 2019, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the virus causing COVID-19, has been spreading and mutating globally despite the expedited approval of many commercial vaccines. Therefore, developing safe, effective and affordable vaccines remains essential to meet the global demand, particularly in developing countries. Transgenic plants have emerged as a promising platform to express recombinant proteins for pharmaceutical and vaccine applications. Two binary vectors, pCAMBIA1300Gt1-S1 and pCAMBIA1300Actin-S1, containing distinct promoters, were constructed and transformed into rice via Agrobacterium. Overall, 56 independent transgenic rice lines were regenerated. Expression analysis revealed that the rice-derived S1 (rS1) protein could be expressed in pGt1::S1 transgenic rice seeds. rS1 protein expression levels reached up to 282 μg/g dry weight, with S1 gene insertion having no effect on grain size and weight. The rS1 protein exhibited a high affinity for human angiotensin-converting enzyme 2 (ACE2) in vitro. Moreover, the immunogenicity of purified rS1 protein co-administered with various adjuvants demonstrated that mice vaccinated with Alum-adjuvant rS1 generated enhanced humoral immune responses with high serum IgG, IgG1 and neutralizing antibody levels. Salmonella Typhimurium flagellin (FliC)-adjuvanted rS1 elicited stronger S1-specific IgG2a levels, promoted splenocyte proliferation and induced mixed Th1/Th2/Th17 cytokine responses. This was evidenced by increased proportions of antigen-specific interferon (IFN)-γ, interleukin-4 (IL-4) and IL-17A-positive CD4+ T lymphocytes, suggesting its potential to induce both humoral and cellular immune responses. These findings suggest that rS1 protein offers a promising approach for affordable COVID-19 subunit vaccine production, and this strategy can be universally applied to other viral vaccines.

 

See https://onlinelibrary.wiley.com/doi/full/10.1111/pbi.70077

 

Figure 1: Genetic construction and detection of rS1 protein. (a) Schematic diagram of the recombinant S1 antigen design. Gt1pro: Gt1 Promoter, a rice seed storage protein glutelin gene promoter; Kozak sequence: Sequence enhancing translation initiation; Gt1sp: Gt1 Signal peptide; Codon-optimized S1 gene: Sequence adapted for expression in rice; KDEL: Endoplasmic reticulum retention signal; NOSter: Nopaline synthase gene terminator; Actpro: Actin promoter, a constitutive promoter. (b) Expression of the rS1 protein in T2 transgenic rice seeds. Total protein from T2 generation transgenic rice seeds was extracted for rS1 protein identification using Western blot analysis. Hsp protein was used as the internal control for gene expression normalization. M: Protein marker; WT: Non-transgenic control. (c) N-glycosylation modification of the rS1 protein. M: Protein marker; 1: T2-2 Transgenic rice seed extract; 2: T2-2 Transgenic rice seed extract treated with PNGase F. (d) Mass spectrometry analysis of the rS1 protein. Matched peptides in the S1 protein are highlighted in Bold Red. (e) Representative diagram of homozygous seeds from T2 transgenic rice.

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