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J Biol Chem, Vol. 274, Issue 6, 3897-3903, February 5, 1999
From the Plant mitochondrial genes are often transcribed
into complex sets of RNAs, resulting from multiple initiation sites and
processing steps. To elucidate the role of initiation in generating the
more than 10 cox2 transcripts found in maize mitochondria,
we surveyed sequences upstream of cox2 for active
promoters. Because the cox2 coding region is immediately
downstream of a 0.7-kb recombination repeat, cox2 is under
the control of two different sets of potential expression signals.
Using an in vitro transcription assay, we localized four
promoters upstream of the coding region in the so-called master
chromosome, and two promoters upstream of the coding region in the
recombinant subgenome. Ribonuclease protection analysis of labeled
primary transcripts confirmed that all but one of these promoters is
active in vivo. Primer extension was used to identify the
promoter sequences and initiation sites, which agree with the consensus
established earlier for maize mitochondria. This study identified two
unusual promoters, the core sequences of which were composed entirely
of adenines and thymines, and one of which was a complex promoter
consisting of seven overlapping units. Deletion mutagenesis of the
complex promoter suggested that each of its units was recognized
independently by RNA polymerase. While each active promoter fit the
maize core consensus sequence YRTAT, not all such sequences surveyed
supported initiation. We conclude that in vitro
transcription is a powerful tool for locating mitochondrial promoters
and that, in the case of cox2, promoter multiplicity
contributes strongly to transcript complexity.
The Maize Mitochondrial cox2 Gene Has Five Promoters
in Two Genomic Regions, Including a Complex Promoter Consisting of
Seven Overlapping Units
§,, and
Boyce Thompson Institute for Plant Research
and the § Section of Genetics and Development, Cornell
University, Ithaca, New York 14853-1801
Copyright © 1999 by The American Society for Biochemistry and Molecular Biology, Inc.
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