Quantitative imaging of endosome acidification and single retrovirus fusion with distinct pools of early endosomes
Quantitative imaging of endosome acidification and single retrovirus fusion with distinct pools of early endosomes
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DOI：
10.1073/pnas.1211714109
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发表时间：
2012-10-23
影响因子：
11.1
通讯作者：
Melikyan, Gregory B.
中科院分区：
文献类型：
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作者：
Padilla-Parra, Sergi;Matos, Pedro M.;Melikyan, Gregory B.
不同的包膜病毒通过内吞作用进入宿主细胞，并在遇到酸性pH时与内体膜融合。目前，携带病毒的内体中的pH动力学以及酸化与病毒融合之间的关系的特征很差。在这里，我们研究了鸟类逆转录病毒的进入，需要两个连续的刺激结合到同源受体和低pH值进行融合。使用并入病毒膜中的遗传编码传感器来测量携带病毒的内体中的pH。酸诱导的病毒融合被可视化为荧光病毒含量标记物释放到胞质溶胶中。在早期酸性内体运输病毒的pH值范围从5.6至6.5，但随着时间的推移相对稳定的一个给定的囊泡。对病毒运动性和管腔pH的分析表明，表达受体（TVA 950）的跨膜亚型的细胞优先将病毒分选到缓慢运输的酸性较低的内体中。相反，由表达GPI锚定亚型（TVA 800）的细胞内化的病毒均匀分布在固定和移动的区室之间。我们发现，酸化和融合之间的滞后时间显着缩短，融合孔更大的动态内体比在更固定的隔间。尽管表达替代受体亚型的细胞的移动的区室内的平均pH相同，但TVA 950支持比TVA 800受体更快的融合。总的来说，我们的结果表明，低pH触发器下游的融合步骤受到携带病毒的细胞内隔室的特性的调节。
Diverse enveloped viruses enter host cells through endocytosis and fuse with endosomal membranes upon encountering acidic pH. Currently, the pH dynamics in virus-carrying endosomes and the relationship between acidification and viral fusion are poorly characterized. Here, we examined the entry of avian retrovirus that requires two sequential stimuli-binding to a cognate receptor and low pH-to undergo fusion. A genetically encoded sensor incorporated into the viral membrane was used to measure the pH in virus-carrying endosomes. Acid-induced virus fusion was visualized as the release of a fluorescent viral content marker into the cytosol. The pH values in early acidic endosomes transporting the virus ranged from 5.6 to 6.5 but were relatively stable over time for a given vesicle. Analysis of viral motility and luminal pH showed that cells expressing the transmembrane isoform of the receptor (TVA950) preferentially sorted the virus into slowly trafficking, less acidic endosomes. In contrast, viruses internalized by cells expressing the GPI-anchored isoform (TVA800) were uniformly distributed between stationary and mobile compartments. We found that the lag times between acidification and fusion were significantly shorter and fusion pores were larger in dynamic endosomes than inmore stationary compartments. Despite the same average pH within mobile compartments of cells expressing alternative receptor isoforms, TVA950 supported faster fusion than TVA800 receptor. Collectively, our results suggest that fusion steps downstream of the low-pH trigger are modulated by properties of intracellular compartments harboring the virus.