Bascom Palmer Eye Institute, Miller School of Medicine, University of Miami, Miami, FL 33136, USA.
Proc Natl Acad Sci U S A. 2012 May 15;109(20):E1238-47. doi: 10.1073/pnas.1119577109. Epub 2012 Apr 20.
To introduce DNA into mitochondria efficiently, we fused adenoassociated virus capsid VP2 with a mitochondrial targeting sequence to carry the mitochondrial gene encoding the human NADH ubiquinone oxidoreductase subunit 4 (ND4). Expression of WT ND4 in cells with the G11778A mutation in ND4 led to restoration of defective ATP synthesis. Furthermore, with injection into the rodent eye, human ND4 DNA levels in mitochondria reached 80% of its mouse homolog. The construct expressed in most inner retinal neurons, and it also suppressed visual loss and optic atrophy induced by a mutant ND4 homolog. The adenoassociated virus cassette accommodates genes of up to ∼5 kb in length, thus providing a platform for introduction of almost any mitochondrial gene and perhaps even allowing insertion of DNA encompassing large deletions of mtDNA, some associated with aging, into the organelle of adults.
为了高效地将 DNA 导入线粒体，我们将腺相关病毒衣壳 VP2 与一个线粒体靶向序列融合，以携带编码人类 NADH 泛醌氧化还原酶亚单位 4（ND4）的线粒体基因。在具有 ND4 中 G11778A 突变的细胞中表达 WT ND4 导致缺陷的 ATP 合成得到恢复。此外，通过注射到啮齿动物眼睛中，线粒体中人 ND4 DNA 水平达到其小鼠同源物的 80%。该构建体在大多数内视网膜神经元中表达，并且还抑制了由突变 ND4 同源物诱导的视觉丧失和视神经萎缩。腺相关病毒盒可容纳长达约 5kb 的基因，因此为引入几乎任何线粒体基因提供了一个平台，甚至可能允许将包含与衰老相关的 mtDNA 大片段缺失的 DNA 插入到成年细胞器中。
