近日，國(guó)際學(xué)術(shù)期刊《PANS》在線發(fā)表了病毒學(xué)國(guó)家重點(diǎn)實(shí)驗(yàn)室崔宗強(qiáng)教授團(tuán)隊(duì)的最新研究成果，論文題為Real-time dissection of dynamic uncoating of individual influenzaviruses（實(shí)時(shí)動(dòng)態(tài)解析單個(gè)流感病毒脫殼過程）。該團(tuán)隊(duì)利用量子點(diǎn)特異性標(biāo)記病毒的基因組和單顆粒示蹤技術(shù)，實(shí)時(shí)動(dòng)態(tài)解析了單個(gè)流感病毒的脫殼過程，揭示了病毒八節(jié)段RNA以單體形式釋放到細(xì)胞質(zhì)并進(jìn)入細(xì)胞核及其在細(xì)胞核內(nèi)的動(dòng)態(tài)行為及機(jī)制。

結(jié)果速覽

該團(tuán)隊(duì)首先建立了量子點(diǎn)位點(diǎn)特異性標(biāo)記甲型流感病毒基因組技術(shù)。在病毒感染過程中，病毒核糖核蛋白復(fù)合物（vRNP）頂端的聚合酶蛋白PA被可控生物素化，與鏈霉親和素修飾的量子點(diǎn)特異性結(jié)合，并組裝進(jìn)入成熟的子代病毒粒子，從而獲得基因組被量子點(diǎn)標(biāo)記的流感病毒。基于量子點(diǎn)的高亮度和耐光漂白等特點(diǎn)，利用單顆粒示蹤技術(shù)，結(jié)合病毒包膜和細(xì)胞組分熒光標(biāo)記，對(duì)單顆流感病毒感染宿主細(xì)胞時(shí)的脫殼過程進(jìn)行實(shí)時(shí)動(dòng)態(tài)可視化分析。實(shí)時(shí)動(dòng)態(tài)揭示了流感病毒在感染后30-90分鐘，病毒由早期內(nèi)體進(jìn)入晚期內(nèi)體發(fā)生膜融合、基因組與病毒包膜分離、單個(gè)節(jié)段基因從晚期內(nèi)體中分別釋放、解離釋放的單個(gè)節(jié)段基因入核、及其在細(xì)胞核內(nèi)運(yùn)輸?shù)膭?dòng)態(tài)過程。發(fā)現(xiàn)流感病毒八個(gè)vRNP在脫殼過程中以單獨(dú)而非組團(tuán)的方式從晚期內(nèi)體中釋放進(jìn)入細(xì)胞質(zhì)，這些單獨(dú)的vRNP以特征性三階段主動(dòng)運(yùn)輸機(jī)制進(jìn)入細(xì)胞核，入核后的vRNP以兩種不同的擴(kuò)散模式運(yùn)輸?shù)狡鋸?fù)制/轉(zhuǎn)錄位點(diǎn)等重要過程與機(jī)制。該研究為世界上首次記錄到單個(gè)流感病毒脫殼的實(shí)時(shí)動(dòng)態(tài)過程，對(duì)深入理解流感病毒在宿主細(xì)胞中的生命周期具有重要意義，并將為開發(fā)新的抗病毒途徑提供思路。

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ABSTRACT: Uncoating is an obligatory step in the virus life cycle that serves as an antiviral target. Unfortunately, it is challenging to study viral uncoating due to methodology limitations for detecting this transient and dynamic event. The uncoating of influenza A virus (IAV), which contains an unusual genome of eight segmented RNAs, is particularly poorly understood. Here, by encapsulating quantum dot (QD)-conjugated viral ribonucleoprotein complexes (vRNPs) within infectious IAV virions and applying single-particle imaging, we tracked the uncoating process of individual IAV virions. Approximately 30% of IAV particles were found to undergo uncoating through fusion with late endosomes in the “around-nucleus” region at 30 to 90 minutes postinfection. Inhibition of viral M2 proton channels and cellular endosome acidification prevented IAV uncoating. IAV vRNPs are released separately into the cytosol after virus uncoating. Then, individual vRNPs undergo a three-stage movement to the cell nucleus and display two diffusion patterns when inside the nucleus. These findings reveal IAV uncoating and vRNP trafficking mechanisms, filling a critical gap in knowledge about influenza viral infection.

中國(guó)科學(xué)院武漢病毒研究所崔宗強(qiáng)研究員為該論文通訊作者，秦沖博士為第一作者。該研究得到了中國(guó)科學(xué)院戰(zhàn)略性先導(dǎo)科技專項(xiàng)、國(guó)家重點(diǎn)研發(fā)計(jì)劃、國(guó)家自然科學(xué)基金、中國(guó)科學(xué)院青年創(chuàng)新促進(jìn)會(huì)等基金項(xiàng)目的資助。

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參考：

1.原文鏈接：

https://doi.org/10.1073/pnas.1812632116

2.病毒學(xué)國(guó)家重點(diǎn)實(shí)驗(yàn)室官網(wǎng)：

https://klv.whu.edu.cn/index.php/View/321.html
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文章內(nèi)容來(lái)源：病毒學(xué)界

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