Cross-strand binding of TFAM to a single mtDNA molecule forms the mitochondrial nucleoid
Cross-strand binding of TFAM to a single mtDNA molecule forms the mitochondrial nucleoid
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DOI：
10.1073/pnas.1512131112
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发表时间：
2015-09-08
影响因子：
11.1
通讯作者：
Larsson, Nils-Goeran
中科院分区：
文献类型：
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作者：
Kukat, Christian;Davies, Karen M.;Larsson, Nils-Goeran
哺乳动物线粒体DNA(MtDNA)由线粒体转录因子A(TFAM)包装成线粒体类核，在控制mtDNA的传递和表达中起着关键作用。类核的超微结构定义不清，因此我们结合使用生化、超分辨显微镜和电子显微镜来显示线粒体类核具有不规则的椭圆形，通常含有单一拷贝的mtDNA。旋转阴影电子显微镜显示，体外类核的形成是一个由TFAM聚集和跨链结合启动的多步骤过程。培养细胞的超分辨显微镜显示，mtDNA拷贝数的增加增加了类核细胞的数量，而不改变它们的大小。电子冷冻断层扫描显示了分离的哺乳动物线粒体中高分辨率的类核物质，并证实了细胞系超分辨率显微镜观察到的大小。我们得出的结论是，线粒体类核的基本组织单位是由TFAM压缩的mtDNA的单个拷贝，并提出了一种包装机制。
Mammalian mitochondrial DNA (mtDNA) is packaged by mitochondrial transcription factor A (TFAM) into mitochondrial nucleoids that are of key importance in controlling the transmission and expression of mtDNA. Nucleoid ultrastructure is poorly defined, and therefore we used a combination of biochemistry, super-resolution microscopy, and electron microscopy to show that mitochondrial nucleoids have an irregular ellipsoidal shape and typically contain a single copy of mtDNA. Rotary shadowing electron microscopy revealed that nucleoid formation in vitro is a multistep process initiated by TFAM aggregation and cross-strand binding. Superresolution microscopy of cultivated cells showed that increased mtDNA copy number increases nucleoid numbers without altering their sizes. Electron cryo-tomography visualized nucleoids at high resolution in isolated mammalian mitochondria and confirmed the sizes observed by superresolution microscopy of cell lines. We conclude that the fundamental organizational unit of the mitochondrial nucleoid is a single copy of mtDNA compacted by TFAM, and we suggest a packaging mechanism.