Cholesterol Binds the Amphipathic Helix of IFITM3 and Regulates Antiviral Activity.
Cholesterol Binds the Amphipathic Helix of IFITM3 and Regulates Antiviral Activity.
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胆固醇结合IFITM3的两亲性阻碍并调节抗病毒活性。
DOI：
10.1016/j.jmb.2022.167759
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发表时间：
2022-10-15
影响因子：
5.6
通讯作者：
Compton, Alex A.
中科院分区：
文献类型：
--
作者：
Rahman, Kazi;Datta, Siddhartha A. K.;Beaven, Andrew H.;Jolley, Abigail A.;Sodt, Alexander J.;Compton, Alex A.
干扰素诱导的跨膜（IFITM）蛋白通过抑制病毒-细胞膜融合广泛地抑制多种致病性病毒的进入，包括甲型流感病毒（IAV）、寨卡病毒、HIV-1和SARS冠状病毒。IFITM 3以前被证明可以破坏胆固醇的运输，但IFITM 3和胆固醇之间的功能关系仍不清楚。我们以前表明，抑制IAV进入IFITM 3与它的能力，以促进细胞膜的刚性，这些活动是功能上连接的一个共同的要求，在IFITM 3的膜内结构域（IMD）中发现的两亲性螺旋（AH）。此外，已显示IFITM 3的AH以胆固醇依赖性方式在体外改变脂质膜。因此，我们旨在更详细地阐明IFITM 3与胆固醇之间的关系。使用基于荧光的体外结合测定，我们发现，从IFITM 3的AH衍生的肽直接与胆固醇类似物NBD-胆固醇相互作用，而IFITM 3 IMD的其他区域没有，并且天然胆固醇与这种相互作用竞争。此外，重组全长IFITM 3蛋白也表现出NBD-胆固醇结合活性。重要的是，先前表征的强烈抑制抗病毒功能的IFITM 3 AH内的突变（F63 Q和F67 Q）破坏了溶液中的AH结构，抑制了体外胆固醇结合，并限制了计算机模拟的双层插入。我们的数据表明，与胆固醇的直接相互作用可能有助于IFITM 3抑制膜融合孔的形成。这些发现可能有助于设计用于广谱抗病毒治疗的治疗肽。
The interferon-induced transmembrane (IFITM) proteins broadly inhibit the entry of diverse pathogenic viruses, including Influenza A virus (IAV), Zika virus, HIV-1, and SARS coronaviruses by inhibiting virus-cell membrane fusion. IFITM3 was previously shown to disrupt cholesterol trafficking, but the functional relationship between IFITM3 and cholesterol remains unclear. We previously showed that inhibition of IAV entry by IFITM3 is associated with its ability to promote cellular membrane rigidity, and these activities are functionally linked by a shared requirement for the amphipathic helix (AH) found in the intramembrane domain (IMD) of IFITM3. Furthermore, it has been shown that the AH of IFITM3 alters lipid membranes in vitro in a cholesterol-dependent manner. Therefore, we aimed to elucidate the relationship between IFITM3 and cholesterol in more detail. Using a fluorescence-based in vitro binding assay, we found that a peptide derived from the AH of IFITM3 directly interacted with the cholesterol analog, NBD-cholesterol, while other regions of the IFITM3 IMD did not, and native cholesterol competed with this interaction. In addition, recombinant full-length IFITM3 protein also exhibited NBD-cholesterol binding activity. Importantly, previously characterized mutations within the AH of IFITM3 that strongly inhibit antiviral function (F63Q and F67Q) disrupted AH structure in solution, inhibited cholesterol binding in vitro, and restricted bilayer insertion in silico. Our data suggest that direct interactions with cholesterol may contribute to the inhibition of membrane fusion pore formation by IFITM3. These findings may facilitate the design of therapeutic peptides for use in broad-spectrum antiviral therapy.
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