The principal goal of this grant proposal is to obtain information about the control of transcription of mitochondrial DNA. The purpose is to gain better insight to the mechanisms that control mitochondrial biogenesis in both the lower and higher eukaryotes. The work is particularly relevant to cardiac tissue in which metabolism is almost completely dependent upon the aerobic synthesis of ATP generated by mitochondria.
Our specific aims are to characterize the transcriptional apparatus of the yeast mitochondria, which serve as an important model of mammalian mitochondria. In particular we aim to purify the mitochondrial RNA polymerase, to characterize its interaction with mitochondrial promoter and other mitochondrial DNA sequences. We plan to isolate the genes of the polymerase subunits, with special emphasis on a 70 kd peptide transcription factor. In vivo mutagenesis of these subunit genes will be undertaken. The generation of mutants in these genes will enable use to undertake structure function studies and to isolate other factors involved in mitochondrial transcription. The biogenesis and import into the mitochondria of the RNA polymerase will be studied intensively both in vivo and in vitro using a new procedure recently developed in our laboratory for the measurement of transcripts. Attenuation of mitochondrial transcription has recently been discovered in our laboratory and we will develop the tools to study the site and possible mechanism of transcriptional attenuation.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL004442-30
Application #
3334177
Study Section
Molecular Biology Study Section (MBY)
Project Start
1977-06-01
Project End
1992-05-31
Budget Start
1988-06-01
Budget End
1989-05-31
Support Year
30
Fiscal Year
1988
Total Cost
Indirect Cost
Name
University of Chicago
Department
Type
Schools of Medicine
DUNS #
225410919
City
Chicago
State
IL
Country
United States
Zip Code
60637
Sanyal, A; Getz, G S (1995) Import of transcription factor MTF1 into the yeast mitochondria takes place through an unusual pathway. J Biol Chem 270:11970-6
Harington, A; Herbert, C J; Tung, B et al. (1993) Identification of a new nuclear gene (CEM1) encoding a protein homologous to a beta-keto-acyl synthase which is essential for mitochondrial respiration in Saccharomyces cerevisiae. Mol Microbiol 9:545-55
Tung, B S; Unger, E R; Levin, B et al. (1991) Use of an unsaturated fatty acid auxotroph of Saccharomyces cerevisiae to modify the lipid composition and function of mitochondrial membranes. J Lipid Res 32:1025-38
Tonsgard, J H; Tung, B; Kornafel, K S et al. (1990) Environmentally induced differential amplification of mitochondrial populations. Biochem J 270:511-8
Biswas, T K; Getz, G S (1988) Promoter-promoter interactions influencing transcription of the yeast mitochondrial gene, Oli 1, coding for ATPase subunit 9. Cis and trans effects. J Biol Chem 263:4844-51
Ticho, B S; Getz, G S (1988) The characterization of yeast mitochondrial RNA polymerase. A monomer of 150,000 daltons with a transcription factor of 70,000 daltons. J Biol Chem 263:10096-103
Backer, J S; Getz, G S (1987) Identification of a new promoter within the tRNA gene cluster of the mitochondrial DNA of Saccharomyces cerevisiae. Nucleic Acids Res 15:9309-24
Mueller, D M; Biswas, T K; Backer, J et al. (1987) Temperature sensitive pet mutants in yeast Saccharomyces cerevisiae that lose mitochondrial RNA. Curr Genet 11:359-67
Biswas, T K; Ticho, B; Getz, G S (1987) In vitro characterization of the yeast mitochondrial promoter using single-base substitution mutants. J Biol Chem 262:13690-6
Mueller, D M; Getz, G S (1987) Direct determination of the specific activity of RNA uniformly labeled with 32P. Anal Biochem 162:521-8

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