DNA conformation-dependent activities of human mitochondrial RNA polymerase.
Mitochondrial RNA polymerase (POLRMT) is a core protein for mitochondrial DNA (mtDNA) transcription. In addition, POLRMT is assumed to be involved in replication, although its exact role is not yet clearly elucidated. We have found novel properties of human POLRMT using a reconstituted transcription system. Various lengths of RNA molecules were synthesized from templates even without a defined promoter sequence, when we used supercoiled circular double-stranded DNA as a template. This promoter-independent activity was as strong as the promoter-dependent one. Promoter-independent DNA conformation-dependent transcription required TFB2M. On supercoiled templates, the promoter-independent activity was strongly suppressed by a putatively physiological amount of TFAM, while promoter-dependent transcription was inhibited to a lesser extent. These different inhibition patterns by TFAM may be important for prevention of random RNA synthesis in vivo. Promoter-independent activity was also observed on relaxed circular single-stranded DNA, where its activity no longer required TFB2M. RNA synthesis on single-stranded DNA was weakly suppressed by a putatively physiological amount of TFAM but restored by the addition of mitochondrial single-stranded DNA binding protein. We suggest that these properties of POLRMT could explain the characteristic features of mammalian mtDNA transcription and replication.
Fukuoh
Atsushi
A
Department of Clinical Chemistry and Laboratory Medicine, Kyushu University Graduate School of Medical Sciences, Fukuoka 812-8582, Japan.
Ohgaki
Kippei
K
Hatae
Hinako
H
Kuraoka
Isao
I
Aoki
Yoshimasa
Y
Uchiumi
Takeshi
T
Jacobs
Howard T
HT
Kang
Dongchon
D
D004261:DNA Replication; D004272:DNA, Mitochondrial / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D004278:DNA, Superhelical; D004268:DNA-Binding Proteins; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism*; D006801:Humans; D008780:Methyltransferases; D008928:Mitochondria / Q000201:enzymology*; D024101:Mitochondrial Proteins; D009690:Nucleic Acid Conformation; D011401:Promoter Regions, Genetic; D014157:Transcription Factors; D014158:Transcription, Genetic
D004272:DNA, Mitochondrial; D004278:DNA, Superhelical; D004268:DNA-Binding Proteins; D024101:Mitochondrial Proteins; C499631:TFAM protein, human; D014157:Transcription Factors; D008780:Methyltransferases; C462443:TFB2M protein, human; D012321:DNA-Directed RNA Polymerases