Replication factors transiently associate with mtDNA at the mitochondrial inner membrane to facilitate replication.
Replication factors transiently associate with mtDNA at the mitochondrial inner membrane to facilitate replication.
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DOI：
10.1093/nar/gkt988
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发表时间：
2014-01
影响因子：
14.9
通讯作者：
Spelbrink JN
中科院分区：
文献类型：
--
作者：
Rajala N;Gerhold JM;Martinsson P;Klymov A;Spelbrink JN
线粒体DNA（mtDNA）是由离散的蛋白质-DNA复合物组成的，即类核，通常被认为是与细胞内膜相关的。在这里，我们解决了复制因子与类核的关联，并表明内源性mtDNA解旋酶Twinkle和单链DNA结合蛋白，mtSSB，共定位仅与一个子集的类核。使用核苷酸类似物原位鉴定复制的mtDNA，闪烁/mtSSB阳性的标记阳性类核的分数在最短的标记脉冲下最高。此外，mtSSB的募集被证明是Twinkle依赖的。因此，这些蛋白质以有序的方式与mtDNA短暂结合以促进复制。为了了解mtDNA复制复合物的性质，我们研究了类核蛋白膜缔合，并表明即使在没有mtDNA的情况下，内源性Twinkle也是牢固的膜缔合，而在膜结合和可溶性组分中均发现了mtSSB和其他类核相关蛋白。同样，大量的mtDNA被发现是可溶的或松散的膜结合。我们发现，通过操纵Twinkle水平，mtDNA膜缔合部分依赖于Twinkle。因此，我们的研究结果表明，Twinkle招募或与线粒体DNA在内膜组装，形成一个复制平台，并首次明确证明类核在组成和并发活动方面都是动态的。
Mitochondrial DNA (mtDNA) is organized in discrete protein–DNA complexes, nucleoids, that are usually considered to be mitochondrial-inner-membrane associated. Here we addressed the association of replication factors with nucleoids and show that endogenous mtDNA helicase Twinkle and single-stranded DNA-binding protein, mtSSB, co-localize only with a subset of nucleoids. Using nucleotide analogs to identify replicating mtDNA in situ, the fraction of label-positive nucleoids that is Twinkle/mtSSB positive, is highest with the shortest labeling-pulse. In addition, the recruitment of mtSSB is shown to be Twinkle dependent. These proteins thus transiently associate with mtDNA in an ordered manner to facilitate replication. To understand the nature of mtDNA replication complexes, we examined nucleoid protein membrane association and show that endogenous Twinkle is firmly membrane associated even in the absence of mtDNA, whereas mtSSB and other nucleoid-associated proteins are found in both membrane-bound and soluble fractions. Likewise, a substantial amount of mtDNA is found as soluble or loosely membrane bound. We show that, by manipulation of Twinkle levels, mtDNA membrane association is partially dependent on Twinkle. Our results thus show that Twinkle recruits or is assembled with mtDNA at the inner membrane to form a replication platform and amount to the first clear demonstration that nucleoids are dynamic both in composition and concurrent activity.
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DOI：
10.1083/jcb.93.1.97
发表时间：
1982-04
期刊：
The Journal of cell biology
影响因子：
--
作者：
Fujiki Y;Hubbard AL;Fowler S;Lazarow PB
通讯作者：
Lazarow PB
具有DNA特性的室内纤维。 I.固定和电子染色反应。
影响因子：
7.8
作者：
NASS, M M;NASS, S
通讯作者：
NASS, S
影响因子：
4
作者：
Arnoult, Damien;Soares, Fraser;Girardin, Stephen E.
通讯作者：
Girardin, Stephen E.
影响因子：
14.9
作者：
Davis, AF;Ropp, PA;Copeland, WC
通讯作者：
Copeland, WC
影响因子：
5.3
作者：
Gilquin, Benoit;Taillebourg, Emmanuel;Baudier, Jacques
通讯作者：
Baudier, Jacques