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J. Biol. Chem., Vol. 277, Issue 19, 16952-16959, May 10, 2002
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From the Understanding mitochondrial transcription is a
requisite first step toward understanding the regulation of
mitochondrial gene expression in kinetoplastids. Here we report the
identification and functional characterization of a mitochondrial RNA
polymerase (mtRNAP) from Trypanosoma brucei, the first
trans-acting factor involved in kinetoplast mitochondrial
transcription to be identified. Using sequences conserved among the
catalytic domains of the single-subunit mtRNAPs, we were able to obtain
a full-length sequence for a candidate mtRNAP from T. brucei. Sequence comparison indicates that it shares homology in
its catalytic domain with other single-subunit mtRNAPs, including
functionally conserved residues that are identical in all
single-subunit RNAPs. We used RNA interference to functionally knock
out the gene product to determine whether the candidate gene represents
an mtRNAP. As predicted for a mitochondrial specific RNA polymerase,
reduction of the gene product resulted in a specific decrease of
mitochondrial versus nuclear transcripts. Additionally, similar to the mtRNAP of other organisms, the mtRNAP characterized here
is involved in replication of the mitochondrial genome. Thus, based on
sequence comparison and functional studies, we have cloned an mtRNAP
from trypanosomes.
A Trypanosome Mitochondrial RNA Polymerase Is Required for
Transcription and Replication*
,¶
Department of Biochemistry and Molecular
Genetics, Schools of Medicine and Dentistry, University of Alabama,
Birmingham, Alabama 35294 and the § Department of Biological
Chemistry, Johns Hopkins School of Medicine,
Baltimore, Maryland 21205
*
This work was supported by National Institutes
of Health Grant AI21401 (to S. L. H.), Medical Scientist
Training Program Grant 5T32GM08361 (to J. G.), and National
Institutes of Health Grant GM27608 (to P. T. E.).The costs of publication of this
article were defrayed in part by the
payment of page charges. The article must therefore be hereby marked
"advertisement" in
accordance with 18 U.S.C. Section
1734 solely to indicate this fact.
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