Department of Pediatrics, Division of Infectious Diseases, Emory Children's Center, Atlanta, GA 30322, USA.
Proc Natl Acad Sci U S A. 2012 Oct 23;109(43):17627-32. doi: 10.1073/pnas.1211714109. Epub 2012 Oct 9.
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 in more 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.
多种包膜病毒通过内吞作用进入宿主细胞，并在遇到酸性 pH 值时与内体膜融合。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还没有得到很好的描述。在这里，我们研究了需要两个连续刺激——与同源受体结合和低 pH 值——才能发生融合的禽反转录病毒的进入。一种遗传编码的传感器被整合到病毒膜中，用于测量携带病毒的内体中的 pH 值。酸性诱导的病毒融合可以作为荧光病毒内容标记物释放到细胞质中而被可视化。运输病毒的早期酸性内体中的 pH 值范围为 5.6 至 6.5，但对于给定的囊泡来说，相对稳定。对病毒运动和腔室 pH 值的分析表明，表达跨膜受体同工型 (TVA950) 的细胞优先将病毒分拣到转运速度较慢、酸性较弱的内体中。相比之下，由表达 GPI 锚定同工型 (TVA800) 的细胞内化的病毒均匀分布于固定和移动隔间。我们发现，在酸化和融合之间的延迟时间在动态内体中比在更固定的隔间中更短，融合孔更大。尽管在表达替代受体同工型的细胞的移动隔间中具有相同的平均 pH 值，但 TVA950 支持的融合速度比 TVA800 受体更快。总的来说，我们的结果表明，低 pH 值触发下游的融合步骤受到携带病毒的细胞内区室特性的调节。
