Release of the yeast mitochondrial RNA polymerase specificity factor from transcription complexes.
The yeast mitochondrial RNA polymerase is composed of two nuclear encoded subunits, a catalytic core (Rpo41p), which resembles the enzymes from bacteriophage T7 and T3, and a specificity factor required for promoter recognition (Mtf1p), which is similar to members of the eubacterial sigma factor family. Using mitochondrial RNA polymerase reconstituted from highly purified subunits, we have determined that Rpo41p and Mtf1p interact to form a holoenzyme in solution prior to DNA binding and promoter recognition. We analyzed the composition of the polymerase during and after the initiation of transcription and found that, like the eubacterial sigma factors, Mtf1p is released after initiation and is available to catalyze transcription on a second template. By analyzing gel mobility shift complexes of the RNA polymerase and DNA at different stages of the transcription reaction, we found that both subunits were associated with DNA prior to initiation and after the formation of two phosphodiester bonds. After the formation of a 13-nucleotide transcript, Mtf1p is no longer associated with Rpo41p on the DNA. These data establish that Mtf1p is functionally as well as structurally similar to eubacterial sigma factors.
Mangus
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DA
Department of Biology, Indiana University, Bloomington 47405.
Jang
S H
SH
Jaehning
J A
JA
D001483:Base Sequence; D004247:DNA / Q000378:metabolism; D012321:DNA-Directed RNA Polymerases / Q000032:analysis / Q000502:physiology*; D005656:Fungal Proteins / Q000302:isolation & purification / Q000502:physiology*; D008928:Mitochondria / Q000201:enzymology*; D024101:Mitochondrial Proteins; D008969:Molecular Sequence Data; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D029701:Saccharomyces cerevisiae Proteins; D012808:Sigma Factor / Q000502:physiology; D014157:Transcription Factors / Q000502:physiology*; D014158:Transcription, Genetic
D005656:Fungal Proteins; C071720:MTF1 protein, S cerevisiae; D024101:Mitochondrial Proteins; D029701:Saccharomyces cerevisiae Proteins; D012808:Sigma Factor; D014157:Transcription Factors; D004247:DNA; D012321:DNA-Directed RNA Polymerases