Diaz-Griffero Felipe, Hoschander Steven Ari, Brojatsch Jürgen
Department of Microbiology and Immunology, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461, USA.
J Virol. 2002 Dec;76(24):12866-76. doi: 10.1128/jvi.76.24.12866-12876.2002.
The avian leukosis virus (ALV) entry mechanism is controversial, with evidence for and against a low-pH requirement for viral fusion. To further address this question, we tested the entry of human immunodeficiency virus type 1 (HIV-1) pseudotyped with the envelope protein of subgroup B ALV (ALV-B) in the presence of three different lysosomotropic agents. These lysosomotropic agents were able to block the entry of wild-type and pseudotyped ALV-B in two different cell lines, strongly suggesting that ALV-B requires a low-pH step for entry. ALV-B and pH-dependent Semliki Forest virus (SFV) entered cells with slower uptake kinetics than HIV-1, which is pH independent. These slow uptake rates support the theory that ALV-B utilizes endocytic pathways to enter cells. Using immunofluorescence and electron microscopy analysis, we visualized the colocalization of virus particles with the endosomal marker transferrin and demonstrated virus particles in clathrin-coated vesicles and endosome-like structures. Surprisingly, a low-pH treatment did not overcome the inhibition of ALV-B entry by lysosomotropic agents. This indicates that, in contrast to SFV, ALV-B is unable to fuse at the cellular surface, even at a low pH. Taken together, our findings suggest that endocytosis and a subsequent low-pH step are critical for successful ALV-B infection.
禽白血病病毒（ALV）的进入机制存在争议，有支持和反对病毒融合需要低pH值的证据。为了进一步解决这个问题，我们在三种不同的溶酶体促渗剂存在的情况下，测试了用B亚群ALV（ALV-B）包膜蛋白假型化的1型人类免疫缺陷病毒（HIV-1）的进入情况。这些溶酶体促渗剂能够在两种不同的细胞系中阻断野生型和假型化ALV-B的进入，强烈表明ALV-B进入需要一个低pH步骤。与不依赖pH值的HIV-1相比，ALV-B和依赖pH值的Semliki森林病毒（SFV）以较慢的摄取动力学进入细胞。这些缓慢的摄取速率支持了ALV-B利用内吞途径进入细胞的理论。通过免疫荧光和电子显微镜分析，我们观察到病毒颗粒与内体标记转铁蛋白的共定位，并在网格蛋白包被的囊泡和类内体结构中发现了病毒颗粒。令人惊讶的是，低pH处理并没有克服溶酶体促渗剂对ALV-B进入的抑制作用。这表明，与SFV不同，即使在低pH值下，ALV-B也无法在细胞表面融合。综上所述，我们的研究结果表明，内吞作用和随后的低pH步骤对于ALV-B的成功感染至关重要。
