Posiri Pratsaneeyaporn, Kondo Hidehiro, Hirono Ikuo, Panyim Sakol, Ongvarrasopone Chalermporn
Institute of Molecular Biosciences, Mahidol University (Salaya Campus), Nakhon Pathom 73170 Thailand.
Laboratory of Genome Science, Tokyo University of Marine Science and Technology, Tokyo, Japan.
Aquaculture. 2015 Jan 1;435:480-487. doi: 10.1016/j.aquaculture.2014.10.018. Epub 2014 Oct 22.
Viral disease caused by the Yellow head virus (YHV) had great impact on economic loss in the aquaculture industry. Prevention or curing YHV disease is still not possible due to the lack of understanding of the basic mechanisms of YHV infection. In this report, the endocytosis inhibitors (chlorpromazine (CPZ), amiloride and methyl-β-cyclodextrin (MβCD)) were used to identify the cellular entry pathway of YHV. Pretreating shrimp with CPZ but not amiloride or MβCD followed by YHV challenge resulted in a significant reduction of YHV levels, suggesting that YHV entered the shrimp cells via clathrin-mediated endocytosis. Next, the major component of the clathrin-coated vesicle, clathrin heavy chain (CHC) was cloned and characterized. The complete coding sequence of PmCHC is 5055 bp encoding a putative protein of 1684 amino acids. Specific silencing of PmCHC mRNA by dsRNA-PmCHC showed an inhibition of YHV replication for 48 h post YHV injection as well as exhibiting a delay in shrimp mortality. These results indicated that PmCHC was an essential component for YHV infection of shrimp cells.
由黄头病毒（YHV）引起的病毒性疾病对水产养殖业造成了巨大的经济损失。由于对YHV感染的基本机制缺乏了解，预防或治愈YHV疾病仍然不可能。在本报告中，使用内吞作用抑制剂（氯丙嗪（CPZ）、氨氯吡咪和甲基-β-环糊精（MβCD））来确定YHV的细胞进入途径。用CPZ预处理虾，但不用氨氯吡咪或MβCD，然后进行YHV攻击，导致YHV水平显著降低，这表明YHV通过网格蛋白介导的内吞作用进入虾细胞。接下来，克隆并鉴定了网格蛋白包被囊泡的主要成分——网格蛋白重链（CHC）。PmCHC的完整编码序列为5055 bp，编码一个推定的1684个氨基酸的蛋白质。通过dsRNA-PmCHC对PmCHC mRNA进行特异性沉默，结果显示在YHV注射后48小时抑制了YHV复制，并且虾的死亡率出现延迟。这些结果表明，PmCHC是虾细胞YHV感染的必需成分。
