含缬酪肽蛋白促进IBV的胞内运输

Valosin-containing Protein Promotes RNA Virus Infection

含缬酪肽蛋白(VCP)是三磷酸腺苷酶超家族中的一员,主要参与蛋白降解、膜融合、DNA修复与复制、细胞周期调控、NF-κB的激活等细胞生理活动。已有报道显示:siRNA干扰VCP的表达,导致传染性支气管炎病毒(IBV)滞留在早期内体,显著降低IBV的感染。为进一步探讨VCP在IBV感染过程中发挥的作用,本研究通过干扰或者外源表达VCP,检测病毒RNA的复制水平和病毒蛋白表达水平。结果表明:干扰VCP的表达,可有效地抑制IBV的增殖;外源表达GFP-VCP,则促进IBV的增殖,但对IBV的内吞没有显著影响。IBV发生内吞后,通过内体系统进行胞内运输,到达晚期内体或溶酶体,发生膜融合和病毒基因组的释放。通过免疫荧光实验观察外源表达的GFP-VCP与内体标志物在IBV感染过程中的共定位情况,发现GFP-VCP与早期内体标志物Rab5有明显共定位,而跟晚期内体标志物Rab7或溶酶体标志物LAMP1没有共定位,证明VCP可能参与早期内体的成熟,帮助病毒的胞内运输。

Valosin-containing protein(VCP)is a member of the adenosine triphosphate superfamily,and associated with various cellular processes in cells.It plays important roles in protein degradation,membrane fusion,DNA repair and replication,regulation of cell cycle,and activation of NF-κB pathway.Previous studies have shown that knock down of VCP results in accumulation of avian infectious bronchitis virus(IBV)in early endosome and suppression of IBV proliferation.To further investigate whether VCP plays important roles in the infection of IBV,we manipulated VCP expression by knocking down or overexpression following by IBV infection.Results showed that interfering the expression of VCP signifi cantly reduced viral RNA level and protein expression.In consistence,overexpression of GFP-VCP greatly promoted the levels of viral RNA and viral protein.Interestingly,the internalized viral RNA level was not affected by VCP overexpression.We further checked colocalization of GFP-VCP and intracellular transport system by immunofl uorescence.It was found that during IBV infection,GFP-VCP colocalized well with early endosome marker Rab5,but not with late endosome marker Rab7 or lysosome marker LAMP1.These results suggested that VCP might be involved in early endosome maturation and thereby helped virus intracellular traffi cking.