RNAi suppression of rice endogenous storage proteins enhances the production of rice-based Botulinum neutrotoxin type A vaccine.
RNAi suppression of rice endogenous storage proteins enhances the production of rice-based Botulinum neutrotoxin type A vaccine.
复制标题
RNAi 抑制水稻内源性储存蛋白可增强基于水稻的 A 型肉毒杆菌中性毒素疫苗的生产。
DOI：
10.1016/j.vaccine.2012.04.064
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发表时间：
2012
期刊：
影响因子：
5.5
通讯作者：
Kiyono H.
中科院分区：
文献类型：
--
作者：
Yuki Y;Mejima M;Kurokawa S;Hiroiwa T. Kong IG;Kuroda M;Takahashi Y;Nochi T;Tokuhara D;Kohda T;Kozaki S;Kiyono H.
基于大米的粘蛋白疫苗（MucoRice）为接种大规模人群以对抗粘膜感染提供了一种非常实用且具有成本效益的策略。然而，高分子量疫苗抗原的低表达和产量的限制仍有待克服。在这里，我们引入RNAi技术，通过共引入编码内源水稻贮藏蛋白的基因的反义序列来推进MucoRice系统，以最大限度地减少贮藏蛋白的产生，并为疫苗抗原在水稻种子中的积累提供更多的空间。当我们使用RNAi抑制T-DNA载体中的主要水稻内源贮藏蛋白、13 kDa醇溶蛋白和谷蛋白A的组合时，我们可以以平均100μg/种子高度表达包含A型肉毒杆菌神经毒素（BoHc）重链的45 kDa C-末端一半的疫苗（MucoRice-BoHc）。MucoRice-Hc是水溶性的，在水稻种子的细胞质中表达，而不在蛋白体I或II中表达。因此，我们对RNAi系统的改造提高了具有高分子量的疫苗抗原的产量。当检查纯化的MucoRice-BoHc的粘膜免疫原性时，疫苗在小鼠中诱导针对A型肉毒杆菌神经毒素攻击的保护性免疫。这些发现证明了先进的MucoRice系统作为用于产生高分子量抗原的高免疫原性粘膜疫苗的创新疫苗生产系统的效率和实用性。
Mucosal vaccines based on rice (MucoRice) offer a highly practical and cost-effective strategy for vaccinating large populations against mucosal infections. However, the limitation of low expression and yield of vaccine antigens with high molecular weight remains to be overcome. Here, we introduced RNAi technology to advance the MucoRice system by co-introducing antisense sequences specific for genes encoding endogenous rice storage proteins to minimize storage protein production and allow more space for the accumulation of vaccine antigen in rice seed. When we used RNAi suppression of a combination of major rice endogenous storage proteins, 13kDa prolamin and glutelin A in a T-DNA vector, we could highly express a vaccine comprising the 45kDa C-terminal half of the heavy chain of botulinum type A neurotoxin (BoHc), at an average of 100μg per seed (MucoRice-BoHc). The MucoRice-Hc was water soluble, and was expressed in the cytoplasm but not in protein body I or II of rice seeds. Thus, our adaptation of the RNAi system improved the yield of a vaccine antigen with a high molecular weight. When the mucosal immunogenicity of the purified MucoRice-BoHc was examined, the vaccine induced protective immunity against a challenge with botulinum type A neurotoxin in mice. These findings demonstrate the efficiency and utility of the advanced MucoRice system as an innovative vaccine production system for generating highly immunogenic mucosal vaccines of high-molecular-weight antigens.