title: Mammalian transcription factor A is a core component of the mitochondrial transcription machinery.
abstract: 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 tC(O)-tC(nitro), 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.
authors.affiliation: Department of Medical Biochemistry and Cell Biology, University of Gothenburg, Gothenburg, Sweden.
authors.forename: Yonghong
authors.initials: Y
authors.lastname: Shi
authors.forename: Anke
authors.initials: A
authors.lastname: Dierckx
authors.forename: Paulina H
authors.initials: PH
authors.lastname: Wanrooij
authors.forename: Sjoerd
authors.initials: S
authors.lastname: Wanrooij
authors.forename: Nils-Göran
authors.initials: NG
authors.lastname: Larsson
authors.forename: L Marcus
authors.initials: LM
authors.lastname: Wilhelmsson
authors.forename: Maria
authors.initials: M
authors.lastname: Falkenberg
authors.forename: Claes M
authors.initials: CM
authors.lastname: Gustafsson
mesh_terms: D000818:Animals; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D004264:DNA Topoisomerases, Type I / Q000378:metabolism; D004272:DNA, Mitochondrial / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D004278:DNA, Superhelical / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D015151:Immunoblotting; D008928:Mitochondria / Q000235:genetics* / Q000378:metabolism; D024101:Mitochondrial Proteins / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D009154:Mutation; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding / Q000187:drug effects; D061987:Sf9 Cells; D012965:Sodium Chloride / Q000494:pharmacology; D013053:Spectrophotometry; D018411:Spodoptera; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism; D014158:Transcription, Genetic
chemical_list: D004272:DNA, Mitochondrial; D004278:DNA, Superhelical; D004268:DNA-Binding Proteins; D024101:Mitochondrial Proteins; C499631:TFAM protein, human; D014157:Transcription Factors; D012965:Sodium Chloride; D004264:DNA Topoisomerases, Type I