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目的通过观察博尔纳病毒(borna disease virus,BDV)磷蛋白(P24)及其编码核酸不同时期的表达和细胞定位,探索BDV潜伏感染的机制。方法构建包含BDV GFP-P24质粒的OL细胞模型,检测并比较不同时期P24核酸和蛋白的表达;对构建的OL细胞模型和各种阴性对照进行逆转录原位PCR的检测,观察P24蛋白编码核酸的细胞定位;使用倒置荧光显微镜观察OL细胞中P24蛋白的细胞定位。结果成功构建了含BDV GFP-P24质粒的OL细胞,发现转染后20代以内细胞P24核酸和蛋白的表达差异不显著(P>0.05);BDV的P24重组蛋白基因始终定位于细胞核,其蛋白早期在细胞核出现,反复传代约15次后可同时在细胞核和细胞质内出现。结论BDV的重组蛋白P24在病毒感染过程中起关键作用,其编码基因始终存在于宿主细胞核内,但表达的蛋白可以在一定时期通过核膜进入细胞质。
Objective To explore the mechanism of latent infection with BDV by observing the expression and cellular localization of P24 and its encoding nucleic acid in borna disease virus (BDV) at different stages. Methods The OL cell model containing GFP-P24 BDV was constructed and the expression of P24 nucleic acid and protein was detected and compared. The constructed OL cell model and various negative controls were detected by reverse transcription in situ PCR. The expression of P24 protein-encoding nucleic acid Cell localization of P24 protein in OL cells was observed using an inverted fluorescent microscope. Results The cells containing BDV GFP-P24 plasmid were successfully constructed. The results showed that there was no significant difference in the expression of P24 nucleic acid and protein between cells transfected with GFP-P24 plasmid (P> 0.05). P24 recombinant protein of BDV was always located in the nucleus and its protein Early appear in the nucleus, repeated passages about 15 times after the nucleus and cytoplasm can occur simultaneously. Conclusion The recombinant protein P24 of BDV plays a key role in viral infection. The coding gene of BDV is always present in the nucleus of host cells, but the expressed protein can enter the cytoplasm through the nuclear membrane at a certain period.