Mitochondrial RNA Polymerase Is Needed for Activation of the Origin of Light-Strand DNA Replication
Mitochondrial RNA Polymerase Is Needed for Activation of the Origin of Light-Strand DNA Replication
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DOI：
10.1016/j.molcel.2009.12.021
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发表时间：
2010-01-15
期刊：
影响因子：
16
通讯作者：
Falkenberg, Maria
中科院分区：
文献类型：
--
作者：
Fuste, Javier Miralles;Wanrooij, Sjoerd;Falkenberg, Maria
线粒体DNA通过独特的酶机制复制，这与用于复制核基因组的复制装置不同。我们在这里研究的机制起源特异性启动滞后链DNA合成在人类线粒体。我们证明，线粒体RNA聚合酶（POLRMT）是启动DNA合成所需的引物从轻链DNA复制起点（OriL）。仅使用纯化的POLRMT和DNA复制因子，我们就可以在体外忠实地重建OriL依赖的起始。前导链DNA合成起始于DNA复制的重链起点并通过OriL。单链OriL裸露，呈茎环结构。在该阶段，POLRMT从单链环区域中的聚-dT延伸起始引物合成。约25 nt后，POLRMT被DNA聚合酶γ取代，DNA合成开始。我们的研究结果表明，POLRMT可以作为一个起源特异性引物在哺乳动物线粒体。
Mitochondrial DNA is replicated by a unique enzymatic machinery, which is distinct from the replication apparatus used for copying the nuclear genome. We examine here the mechanisms of origin-specific initiation of lagging-strand DNA synthesis in human mitochondria. We demonstrate that the mitochondrial RNA polymerase (POLRMT) is the primase required for initiation of DNA synthesis from the light-strand origin of DNA replication (OriL). Using only purified POLRMT and DNA replication factors, we can faithfully reconstitute OriL-dependent initiation in vitro. Leading-strand DNA synthesis is initiated from the heavy-strand origin of DNA replication and passes OriL. The single-stranded OriL is exposed and adopts a stem-loop structure. At this stage, POLRMT initiates primer synthesis from a poly-dT stretch in the single-stranded loop region, After about 25 nt, POLRMT is replaced by DNA polymerase gamma, and DNA synthesis commences. Our findings demonstrate that POLRMT can function as an origin-specific primase in mammalian mitochondria.