Department of Microbiology and Immunology, Indiana University School of Medicine, Indianapolis, Indiana, USA.
J Virol. 2010 Mar;84(5):2563-72. doi: 10.1128/JVI.00342-09. Epub 2009 Dec 16.
We previously demonstrated that the naturally occurring splice variant stromal cell-derived factor 1gamma/CXCL12gamma is the most potent CXCL12 isoform in blocking X4 HIV-1, with weak chemotactic activity. A conserved BBXB domain (B for basic and X for any residue) located in the N terminus ((24)KHLK(27)) is found in all six isoforms of CXCL12. To determine whether the potent antiviral activity of CXCL12gamma is due to the presence of the extra C-terminal BBXB domains, we mutated each domain individually as well as in combination. Although binding of CXCL12gamma to heparan sulfate proteoglycan (HSPG) was 10-fold higher than that observed with CXCL12alpha, the results did not demonstrate a direct correlation between HSPG binding and the potent antiviral activity. CXCL12gamma mutants lacking the conserved BBXB domain (designated gammaB1) showed increased binding to HSPG but reduced anti-HIV activity. In contrast, the mutants lacking the C-terminal second and/or third BBXB domain but retaining the conserved domain (designated B2, B3, and B23) showed decreased binding to HSPG but increased anti-HIV activity. The B2, B3, and B23 mutants were associated with enhanced CXCR4 binding, receptor internalization, and restored chemotaxis. Internalization of CXCR4 was more potent with CXCL12gamma than with CXCL12alpha and was significantly reduced when the conserved BBXB domain was mutated. We concluded that the observed potent anti-HIV-1 activity of CXCL12gamma is due to increased affinity for CXCR4 and to efficient receptor internalization.
我们之前已经证明，天然存在的基质细胞衍生因子 1gamma/CXCL12gamma 是阻断 X4 HIV-1 的最有效 CXCL12 同工型，其趋化活性较弱。位于 N 端的保守 BBXB 结构域（B 代表碱性，X 代表任何残基）存在于 CXCL12 的所有六个同工型中。为了确定 CXCL12gamma 的强大抗病毒活性是否归因于额外的 C 末端 BBXB 结构域的存在，我们分别突变了每个结构域以及组合。尽管 CXCL12gamma 与硫酸乙酰肝素蛋白聚糖（HSPG）的结合比观察到的 CXCL12alpha 高 10 倍，但结果并未表明 HSPG 结合与强大的抗病毒活性之间存在直接相关性。缺乏保守 BBXB 结构域的 CXCL12gamma 突变体（指定为 gammaB1）显示出与 HSPG 的结合增加，但抗 HIV 活性降低。相比之下，缺乏 C 末端第二个和/或第三个 BBXB 结构域但保留保守结构域的突变体（指定为 B2、B3 和 B23）显示出与 HSPG 的结合减少，但抗 HIV 活性增加。B2、B3 和 B23 突变体与增强的 CXCR4 结合、受体内化和恢复趋化性相关。与 CXCL12alpha 相比，CXCL12gamma 引起的 CXCR4 内化更有效，当保守的 BBXB 结构域发生突变时，内化作用明显减少。我们得出的结论是，CXCL12gamma 观察到的强大抗 HIV-1 活性是由于与 CXCR4 的亲和力增加和有效的受体内化所致。
