Understanding the mechanisms by which viruses evade host cell immune defenses is important for developing improved antiviral therapies. In an unusual twist, human cytomegalovirus co-opts the antiviral radical SAM enzyme viperin (virus-inhibitory protein, endoplasmic reticulum-associated, interferon-inducible) to enhance viral infectivity. This process involves translocation of viperin to the mitochondrion, where it binds the β-subunit (HADHB) of the mitochondrial trifunctional enzyme complex that catalyzes thiolysis of β-ketoacyl-CoA esters as part of fatty acid β-oxidation. Here we investigated how the interaction between these two enzymes alters their activities and affects cellular ATP levels. Experiments with purified enzymes indicated that viperin inhibits the thiolase activity of HADHB, but, unexpectedly, HADHB activates viperin, leading to synthesis of the antiviral nucleotide 3'-deoxy-3',4'-didehydro-CTP. Measurements of enzyme activities in lysates prepared from transfected HEK293T cells expressing these enzymes mirrored the findings obtained with purified enzymes. Thus, localizing viperin to mitochondria decreased thiolase activity, and coexpression of HADHB significantly increased viperin activity. Furthermore, targeting viperin to mitochondria also increased the rate at which HADHB is retrotranslocated out of mitochondria and degraded, providing an additional mechanism by which viperin reduces HADHB activity. Targeting viperin to mitochondria decreased cellular ATP levels by more than 50%, consistent with the enzyme disrupting fatty acid catabolism. These results provide biochemical insight into the mechanism by which human cytomegalovirus subverts viperin; they also provide a biochemical rationale for viperin's recently discovered role in regulating thermogenesis in adipose tissues.

中文翻译：

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将 viperin 靶向线粒体会抑制三功能酶复合物的硫解酶活性。

了解病毒逃避宿主细胞免疫防御的机制对于开发改进的抗病毒疗法很重要。在一个不寻常的转折中，人类巨细胞病毒与抗病毒自由基 SAM 酶 viperin（病毒抑制蛋白、内质网相关、干扰素诱导型）结合以增强病毒感染性。该过程涉及蝰蛇易位到线粒体，在那里它与线粒体三功能酶复合物的 β-亚基 (HADHB) 结合，催化 β-酮酰基辅酶 A 酯的硫解，作为脂肪酸 β-氧化的一部分。在这里，我们研究了这两种酶之间的相互作用如何改变它们的活性并影响细胞 ATP 水平。用纯化酶进行的实验表明，viperin 抑制了 HADHB 的硫解酶活性，但出人意料的是，HADHB 激活了 viperin，导致抗病毒核苷酸 3'-deoxy-3',4'-didehydro-CTP 的合成。从表达这些酶的转染 HEK293T 细胞制备的裂解物中酶活性的测量反映了用纯化酶获得的结果。因此，将 viperin 定位到线粒体会降低硫解酶活性，而 HADHB 的共表达显着增加了 viperin 活性。此外，将 viperin 靶向线粒体也增加了 HADHB 逆转录出线粒体并降解的速率，提供了一种额外的机制，通过这种机制，viperin 降低了 HADHB 的活性。将 viperin 靶向线粒体可使细胞 ATP 水平降低 50% 以上，这与破坏脂肪酸分解代谢的酶一致。这些结果为人类巨细胞病毒破坏蝰蛇的机制提供了生化洞察；它们还为 viperin 最近发现的调节脂肪组织产热作用提供了生化原理。