CAJ | 학술논문

为研究猪轮状病毒非结构蛋白 NSP4的细胞定位及其蛋白功能，将构建的重组质粒 pNSP4-EGFP，pviroporin-EGFP 和 pEGFP-N1空载体质粒转染到 SIEC02细胞株，通过与内质网定位蛋白的共聚焦分析、实时荧光定量 PCR 检测和 Western blot 分析，发现 NSP4主要分布于内质网，少数定位于细胞核；NSP4蛋白可显著上调 Bax 基因水平（P ＜0．01）和 caspase-3蛋白表达水平。说明猪轮状病毒 NSP4蛋白具有类似 viroporin 的功能，主要定位于内质网上；NSP4依赖或部分依赖 caspase 的激活和 Bax 的活化促进细胞凋亡。
위연구저륜상병독비결구단백 NSP4적세포정위급기단백공능，장구건적중조질립 pNSP4-EGFP，pviroporin-EGFP 화 pEGFP-N1공재체질립전염도 SIEC02세포주，통과여내질망정위단백적공취초분석、실시형광정량 PCR 검측화 Western blot 분석，발현 NSP4주요분포우내질망，소수정위우세포핵；NSP4단백가현저상조 Bax 기인수평（P ＜0．01）화 caspase-3단백표체수평。설명저륜상병독 NSP4단백구유유사 viroporin 적공능，주요정위우내질망상；NSP4의뢰혹부분의뢰 caspase 적격활화 Bax 적활화촉진세포조망。
The study was to explore the localization of porcine rotavirus nonstructural protein 4(NSP4 )in the immortalized swine intestinal epithelial cells (SIEC02)established by our laboratory,as well as its pro-tein function.The eukaryotic recombinant plasmids pNSP4-EGFP,pViroporin-EGFP,and pEGFP-N1 vec-tor were transfected transiently into SIEC02 cells.By means of confocal microscopic analysis with endoplas-mic reticulum (ER)positioning vector,real-time fluorescent quantitative PCR and Western blot analysis, the results showed that NSP4 were mainly localized in the ER and few were localized in the cell nucleus;the relative mRNA levels of Bax and the protein expression of caspase-3 were upregulated significantly in the group of NSP4 overexpression (P <0.01).This illustrates that porcine rotavirus NSP4 has similar function with viroporin,mainly localized in the ER;NSP4-induced apoptosis at least partly depended on the caspase and Bax activation in the immortalized swine intestinal epithelial cells.