Establishment ofpredidive method for shift of coreceptor usage in HIV 1-infected individnals

HIV-1感染的疾病进展在很大程度上与HIV-1从CCR5到CXCR4的辅助受体使用转换有关。尽管大多数使用CXCR4的病毒（称为R5X4病毒）保留了CCR5的使用，但在同时表达CCR5和CXCR4的细胞中，单独使用CXCR4抑制剂可以抑制R5X4病毒的复制，这表明这些病毒优先使用CXCR4。为了确定Env的哪些区域与CXCR4的优先使用相关，我们从R5X4病毒（89.6株）中分离出CXCR4拮抗剂T140存在的逃逸突变体。一个分离的t140 -逃逸突变体在gp120的V3区域（Arginine to Serine, R308S）有一个氨基酸替换。带有突变体Env的荧光素酶报告蛋白HIV-1假型表明，在同时表达CCR5和CXCR4的细胞感染中，这种替代完全赋予了对CXCR4拮抗剂的抗性，但增加了对CCR5拮抗剂TAK-779的敏感性，表明CCR5的优先使用。使用表达单一辅助受体的细胞进行分析表明，突变体Env主要有效地利用了CCR5而不是CXCR4，并保留了R5X4表型。这些结果表明，R5X4病毒对CXCR4的优先利用是由gp120 V3区的单个氨基酸决定的。

The disease progression of HIV-1 infection is largely associated with a switch of coreceptor usage by HIV-1 from CCR5 to CXCR4. Although most of CXCR4-using viruses retain CCR5 usage called R5X4 viruses, the replication of R5X4 viruses were inhibited by CXCR4 inhibitors alone in the cells expressing both CCR5 and CXCR4, indicating the preferential usage of CXCR4 by these viruses. In order to determine which regions of the Env are associated with the preferential usage of CXCR4, we isolated an escape mutant in the presence of a CXCR4 antagonist T140 from an R5X4 virus (89.6 strain). An isolated T140-escape mutant harbored a single amino acid substitution in the V3 region of gp120(Arginine to Serine, R308S). Luciferase-reporter HIV-1 pseudotyped with the mutant Env showed that the substitution totally conferred resistance to CXCR4 antagonists but increased sensitivity to a CCR5 antagonist TAK-779 in the infection of the cells expressing both CCR5 and CXCR4, indicating the preferential usage of CCR5. Analyses using the cells expressing a single coreceptor showed that the mutant Env predominantly and efficiently utilized CCR5 than CXCR4 with retaining R5X4 phenotype. These results indicated that the preferential usage of CXCR4 by the R5X4 virus was determined by the single amino acid in the V3 region of gp120.