The influenza A virus PB1-F2 protein targets the inner mitochondrial membrane via a predicted basic amphipathic helix that disrupts mitochondrial function
The influenza A virus PB1-F2 protein targets the inner mitochondrial membrane via a predicted basic amphipathic helix that disrupts mitochondrial function
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DOI：
10.1128/jvi.77.13.7214-7224.2003
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发表时间：
2003-07-01
影响因子：
5.4
通讯作者：
Yewdell, JW
中科院分区：
文献类型：
--
作者：
Gibbs, JS;Malide, D;Yewdell, JW
第11个甲型流感病毒基因产物是一种由87个氨基酸组成的蛋白质，暂时命名为PB 1-F2（因为。其由与PB 1开放阅读帧重叠的开放阅读帧编码）。在甲型流感病毒感染的细胞中，PB 1-F2的显著部分定位于线粒体内膜。PB 1-F2似乎以细胞类型依赖性方式增强病毒诱导的细胞死亡。对于目前的通信，我们已经确定和表征了一个区域附近的COOH端的PB 1-F2是必要的和足够的内部线粒体膜的定位，确定了瞬时表达的嵌合蛋白组成的元素PB 1-F2基因融合增强绿色荧光蛋白（EGFP）在HeLa细胞。通过该序列将EGFP靶向线粒体导致线粒体内膜电位的丧失，从而导致细胞死亡。线粒体靶向序列（MTS）被预测形成带正电荷的两亲性et-螺旋，并且因此类似于人T细胞白血病病毒1型的p13（II）蛋白的MTS。我们正式证明了PB 1-F2线粒体定位的两个序列的功能互补性。PB 1-F2中假定的两亲性螺旋的突变分析表明，用Ala替换5个碱性氨基酸废除了线粒体靶向，而两个高度保守的Leu突变为Ala则没有。这些发现表明PB 1-F2具有与其他病毒蛋白类似的MTS，并且当与EGFP融合时，该MTS能够独立地损害线粒体功能和细胞活力。
The 11th influenza A virus gene product is an 87-amino-acid protein provisionally named PB1-F2 (because. it is encoded by an open reading frame overlapping the PB1 open reading frame). A significant fraction of PB1-F2 localizes to the inner mitochondrial membrane in influenza A virus-infected cells. PB1-F2 appears to enhance virus-induced cell death in a cell type-dependent manner. For the present communication we have identified and characterized a region near the COOH terminus of PB1-F2 that is necessary and sufficient for its inner mitochondrial membrane localization, as determined by transient expression of chimeric proteins consisting of elements of PB1-F2 genetically fused to enhanced green fluorescent protein (EGFP) in HeLa cells. Targeting of EGFP to mitochondria by this sequence resulted in the loss of the inner mitochondrial membrane potential, leading to cell death. The mitochondrial targeting sequence (MTS) is predicted to form a positively charged amphipathic et-helix and, as such, is similar to the MTS of the p13(II) protein of human T-cell leukemia virus type 1. We formally demonstrate the functional interchangeability of the two sequences for mitochondrial localization of PB1-F2. Mutation analysis of the putative amphipathic helix in the PB1-F2 reveals that replacement of five basic amino acids with Ala abolishes mitochondrial targeting, whereas mutation of two highly conserved Leu to Ala does not. These findings demonstrate that PB1-F2 possesses an MTS similar to other viral proteins and that this MTS, when fused to EGFP, is capable of independently compromising mitochondrial function and cellular viability.