A型流感病毒NS1与突触后密度蛋白-95的相互作用

目的 A型流感病毒NS1蛋白是一种多功能的致病因子,能够与被感染细胞中的多种蛋白相互结合,影响并干扰宿主细胞内的信号转导、蛋白质合成及抗病毒反应。突触后密度蛋白(Postsynaptic density protein95,PSD-95)主要存在于神经元及SH-SY-5Y等神经来源的细胞株中。假设NS1能够与PSD-95结合,则更有利于了解A型流感病毒对神经元及相关细胞的作用机制。方法通过酵母双杂交,GST-pull down及免疫荧光技术分别从体外和体内两方面检测NS1与PSD-95的相互作用。结果酵母双杂交表明,仅转染PGAD-NS51/PGBK-PSD-95的QDO有菌落生长,且α-半乳糖苷酶活性显著高于阳性对照;而转染PGAD-NS32/PGBK-PSD-95的QDO无菌落生长;GST-pull down表明仅NS51与PSD-95孵育后,能够被Western-blot检测到;免疫荧光表明NS51与PSD-95可能存在共定位,而NS32与PSD-95则不存在共定位。结论 H5N1(A/chicken/Guangdong/1/2005)的NS1能够与PSD-95结合;反之,H3N2(A/Shantou/602/06)的NS1则不能。

Different bindings between influenza A virus NS1 and postsynaptic density protein-95

Objective The NS1 protein of influenza A virus is a multifunction virulence factor. It is able to bind with many proteins which affect cellular signal transduction, protein synthesis and counteracts from antiviral responses in infected cells. Postsynaptic density proteins （ PSD-95 ） mainly expressed in neurons and SH-SY-5Y ceils. It is benefit for us to recognize the function mechanism of influenza A virus. Methods We conducted a yeast two-hybrid analysis, GST-pull down assayand immunofluorescence to detecting interaction in between NSI and PSD-95. Results Yeast two-hybrid analysis shows that yeast is transformed with PGAD-NS$1 in combination with PGBK-PSD-95 is able to grown in QDO media, and c~-galactosidase activity is higher than that of positive control, but PGAD-NS32 in combination with PGBK-PSD-95 is una- ble to grown. GST-pull down shows that only NSS1 is identified by western blotting. Immunofluorescence also shows that co-location may be existed between NS51 and PSD-95. Conclusion NS1 of H5N1 （ A/chicken/Guangdong/1/2005 ） is a- ble to bind to PSD-95, but the NS1 of H3N2 （A/Shantou/602/06） is unable to do so.