Biological Resource Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), 181 Ipsin-gil, Jeongeup, Jeonbuk, 56212, Republic of Korea.
Department of Bioscience and Biotechnology, Chungnam National University, Daejeon, 34134, Republic of Korea.
Planta. 2018 Apr;247(4):973-985. doi: 10.1007/s00425-017-2836-z. Epub 2018 Jan 8.
We demonstrated successful overexpression of porcine reproductive and respiratory syndrome virus (PRRSV)-derived GP4D and GP5D antigenic proteins in Arabidopsis. Pigs immunized with transgenic plants expressing GP4D and GP5D proteins generated both humoral and cellular immune responses to PRRSV. Porcine reproductive and respiratory syndrome virus (PRRSV) causes PRRS, the most economically significant disease affecting the swine industry worldwide. However, current commercial PRRSV vaccines (killed virus or modified live vaccines) show poor efficacy and safety due to concerns such as reversion of virus to wild type and lack of cross protection. To overcome these problems, plants are considered a promising alternative to conventional platforms and as a vehicle for large-scale production of recombinant proteins. Here, we demonstrate successful production of recombinant protein vaccine by expressing codon-optimized and transmembrane-deleted recombinant glycoproteins (GP4D and GP5D) from PRRSV in planta. We generated transgenic Arabidopsis plants expressing GP4D and GP5D proteins as candidate antigens. To examine immunogenicity, pigs were fed transgenic Arabidopsis leaves expressing the GP4D and GP5D antigens (three times at 2-week intervals) and then challenged with PRRSV at 6-week post-initial treatment. Immunized pigs showed significantly lower lung lesion scores and reduced viremia and viral loads in the lung than pigs fed Arabidopsis leaves expressing mYFP (control). Immunized pigs also had higher titers of PRRSV-specific antibodies and significantly higher levels of pro-inflammatory cytokines (TNF-α and IL-12). Furthermore, the numbers of IFN-γ-producing cells were higher, and those of regulatory T cells were lower, in GP4D and GP5D immunized pigs than in control pigs. Thus, plant-derived GP4D and GP5D proteins provide an alternative platform for producing an effective subunit vaccine against PRRSV.
我们成功地在拟南芥中表达了猪繁殖与呼吸综合征病毒(PRRSV)衍生的 GP4D 和 GP5D 抗原蛋白。用表达 GP4D 和 GP5D 蛋白的转基因植物免疫的猪产生了针对 PRRSV 的体液和细胞免疫反应。猪繁殖与呼吸综合征病毒(PRRSV)引起 PRRS,这是全球养猪业最重要的经济疾病。然而,由于病毒返祖为野生型和缺乏交叉保护等问题,目前的商业 PRRSV 疫苗(灭活病毒或减毒活疫苗)显示出较差的效果和安全性。为了克服这些问题,植物被认为是传统平台的一种有前途的替代方案,也是大规模生产重组蛋白的载体。在这里,我们通过在植物体内表达经过密码子优化和跨膜缺失的重组糖蛋白(GP4D 和 GP5D)来展示重组蛋白疫苗的成功生产。我们生成了表达 GP4D 和 GP5D 蛋白的转基因拟南芥植物作为候选抗原。为了研究免疫原性,将转基因拟南芥叶片中的 GP4D 和 GP5D 抗原(每两周喂食三次)喂食给猪,然后在初次治疗后 6 周用 PRRSV 进行攻毒。与喂食表达 mYFP(对照)的拟南芥叶片的猪相比,免疫猪的肺部病变评分显著降低,肺部病毒血症和病毒载量降低。免疫猪还具有更高滴度的 PRRSV 特异性抗体和更高水平的促炎细胞因子(TNF-α 和 IL-12)。此外,GP4D 和 GP5D 免疫猪中 IFN-γ 产生细胞的数量更高,调节性 T 细胞的数量更低。因此,植物来源的 GP4D 和 GP5D 蛋白为生产针对 PRRSV 的有效亚单位疫苗提供了替代平台。