期刊名称：Proceedings of the National Academy of Sciences
印刷版ISSN：0027-8424
电子版ISSN：1091-6490
出版年度：2012
卷号：109
期号：41
页码：16510-16515
DOI：10.1073/pnas.1119738109
语种：English
出版社：The National Academy of Sciences of the United States of America
摘要：Transcription factor A (TFAM) functions as a DNA packaging factor in mammalian mitochondria. TFAM also binds sequence-specifically to sites immediately upstream of mitochondrial promoters, but there are conflicting data regarding its role as a core component of the mitochondrial transcription machinery. We here demonstrate that TFAM is required for transcription in mitochondrial extracts as well as in a reconstituted in vitro transcription system. The absolute requirement of TFAM can be relaxed by conditions that allow DNA breathing, i.e., low salt concentrations or negatively supercoiled DNA templates. The situation is thus very similar to that described in nuclear RNA polymerase II-dependent transcription, in which the free energy of supercoiling can circumvent the need for a subset of basal transcription factors at specific promoters. In agreement with these observations, we demonstrate that TFAM has the capacity to induce negative supercoils in DNA, and, using the recently developed nucleobase analog FRET-pair tCO-tCnitro, we find that TFAM distorts significantly the DNA structure. Our findings differ from recent observations reporting that TFAM is not a core component of the mitochondrial transcription machinery. Instead, our findings support a model in which TFAM is absolutely required to recruit the transcription machinery during initiation of transcription.
