Ferraro Paola, Nicolosi Luca, Bernardi Paolo, Reichard Peter, Bianchi Vera
Departments of Biology and Biomedical Sciences, University of Padua, I-35131 Padua, Italy.
Proc Natl Acad Sci U S A. 2006 Dec 5;103(49):18586-91. doi: 10.1073/pnas.0609020103. Epub 2006 Nov 21.
Dividing cultured cells contain much larger pools of the four dNTPs than resting cells. In both cases the sizes of the individual pools are only moderately different. The same applies to mitochondrial (mt) pools of cultured cells. Song et al. [Song S, Pursell ZF, Copeland WC, Longley MJ, Kunkel TA, Mathews CK (2005) Proc Natl Acad Sci USA 102:4990-4995] reported that mt pools of rat tissues instead are highly asymmetric, with the dGTP pool in some cases being several-hundred-fold larger than the dTTP pool, and suggested that the asymmetry contributes to increased mutagenesis during mt DNA replication. We have now investigated this discrepancy and determined the size of each dNTP pool in mouse liver mitochondria. We found large variations in pool sizes that closely followed variations in the ATP pool and depended on the length of time spent in the preparation of mitochondria. The proportion between dNTPs was in all cases without major asymmetries and similar to those found earlier in cultured resting cells. We also investigated the import and export of thymidine phosphates in mouse liver mitochondria and provide evidence for a rapid, highly selective, and saturable import of dTMP, not depending on a functional respiratory chain. At nM external dTMP the nucleotide is concentrated 100-fold inside the mt matrix. Export of thymidine phosphates was much slower and possibly occurred at the level of dTDP.
与静息细胞相比，处于分裂状态的培养细胞含有大得多的四种脱氧核苷三磷酸（dNTP）池。在这两种情况下，各个池的大小仅有适度差异。培养细胞的线粒体（mt）池情况也是如此。宋等人[宋S，珀塞尔ZF，科普兰WC，朗利MJ，昆克尔TA，马修斯CK（2005年）《美国国家科学院院刊》102:4990 - 4995]报告称，大鼠组织的线粒体池则高度不对称，在某些情况下，dGTP池比dTTP池大数百倍，并表明这种不对称性导致线粒体DNA复制过程中诱变增加。我们现在研究了这种差异，并确定了小鼠肝线粒体中每个dNTP池的大小。我们发现池大小存在很大差异，这些差异紧密跟随ATP池的变化，并取决于制备线粒体所花费的时间长度。在所有情况下，dNTP之间的比例没有明显不对称，与早期在培养的静息细胞中发现的比例相似。我们还研究了小鼠肝线粒体中胸苷磷酸的进出，并为dTMP的快速、高度选择性和可饱和性导入提供了证据，这种导入不依赖于功能性呼吸链。在细胞外dTMP浓度为纳摩尔时，该核苷酸在线粒体基质内浓缩100倍。胸苷磷酸的输出要慢得多，可能发生在dTDP水平。
