P2X1 Receptor Antagonists Inhibit HIV-1 Fusion by Blocking Virus-Coreceptor Interactions
P2X1 Receptor Antagonists Inhibit HIV-1 Fusion by Blocking Virus-Coreceptor Interactions
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DOI：
10.1128/jvi.01178-15
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发表时间：
2015-09-01
影响因子：
5.4
通讯作者：
Melikyan, Gregory B.
中科院分区：
文献类型：
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作者：
Giroud, Charline;Marin, Mariana;Melikyan, Gregory B.
HIV-1 Env糖蛋白介导的融合在Env与CD 4和辅助受体CXCR 4或CCR 5顺序结合后开始。虽然这些相互作用被认为是必要的，足以促进HIV-1融合，其他宿主因素可以调节这一过程。先前的研究报道了选择性P2 X1受体拮抗剂（包括NF 279）对HIV-1融合的有效抑制，并表明这些受体在HIV-1进入中发挥作用。在这里，我们研究了NF 279的抗病毒活性的机制，发现这种化合物不会通过阻止P2 X1通道的激活来抑制HIV-1融合，而是有效地阻断病毒与CXCR 4或CCR 5的结合。NF 279对HIV-1融合的脱靶效应的概念得到融合实验中使用的细胞中P2 X1受体的可检测表达的缺乏以及培养基中ATP的添加或ATP的酶促消耗不调节病毒融合的事实的支持。重要的是，当在Env-CD 4-辅助受体接合下游的中间阶段加入时，NF 279不能抑制HIV-1与细胞系和原代巨噬细胞的融合。相反，在NF 279的存在下，HIV-1融合在CD 4结合的下游但在辅助受体接合之前被阻止。NF 279还拮抗CCR 5、CXCR 4和另一种趋化因子受体的信号传导功能，如通过抑制由特异性配体和重组gp 120引起的钙应答所证明的。总的来说，我们的研究结果表明，NF 279是一种双重的HIV-1辅助受体抑制剂，干扰CCR 5和CXCR 4的功能参与的Env。重要的是抑制P2 X受体活性抑制HIV-1融合和复制，这表明P2 X信号参与HIV-1的进入。然而，在这项研究中进行的机制实验表明，P2 X1受体在靶细胞中不表达，也不参与病毒融合。相反，我们发现特异性P2 X1受体拮抗剂NF 279对HIV-1融合的抑制作用是由于阻断了病毒与CXCR 4和CCR 5共受体的相互作用。NF 279消除由相应趋化因子诱导的细胞钙信号传导的能力表明，该化合物作为双辅受体拮抗剂。因此，P2 X1受体拮抗剂可以代表一类新的双辅助受体抑制剂，其结构和作用机制与已知的HIV-1辅助受体拮抗剂不同。
HIV-1 Env glycoprotein-mediated fusion is initiated upon sequential binding of Env to CD4 and the coreceptor CXCR4 or CCR5. Whereas these interactions are thought to be necessary and sufficient to promote HIV-1 fusion, other host factors can modulate this process. Previous studies reported potent inhibition of HIV-1 fusion by selective P2X1 receptor antagonists, including NF279, and suggested that these receptors play a role in HIV-1 entry. Here we investigated the mechanism of antiviral activity of NF279 and found that this compound does not inhibit HIV-1 fusion by preventing the activation of P2X1 channels but effectively blocks the binding of the virus to CXCR4 or CCR5. The notion of an off-target effect of NF279 on HIV-1 fusion is supported by the lack of detectable expression of P2X1 receptors in cells used in fusion experiments and by the fact that the addition of ATP or the enzymatic depletion of ATP in culture medium does not modulate viral fusion. Importantly, NF279 fails to inhibit HIV-1 fusion with cell lines and primary macrophages when added at an intermediate stage downstream of Env-CD4-coreceptor engagement. Conversely, in the presence of NF279, HIV-1 fusion is arrested downstream of CD4 binding but prior to coreceptor engagement. NF279 also antagonizes the signaling function of CCR5, CXCR4, and another chemokine receptor, as evidenced by the suppression of calcium responses elicited by specific ligands and by recombinant gp120. Collectively, our results demonstrate that NF279 is a dual HIV-1 coreceptor inhibitor that interferes with the functional engagement of CCR5 and CXCR4 by Env.IMPORTANCEInhibition of P2X receptor activity suppresses HIV-1 fusion and replication, suggesting that P2X signaling is involved in HIV-1 entry. However, mechanistic experiments conducted in this study imply that P2X1 receptor is not expressed in target cells or involved in viral fusion. Instead, we found that inhibition of HIV-1 fusion by a specific P2X1 receptor antagonist, NF279, is due to the blocking of virus interactions with both the CXCR4 and CCR5 coreceptors. The ability of NF279 to abrogate cellular calcium signaling induced by the respective chemokines showed that this compound acts as a dual-coreceptor antagonist. P2X1 receptor antagonists could thus represent a new class of dual-coreceptor inhibitors with a structure and a mechanism of action that are distinct from those of known HIV-1 coreceptor antagonists.