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Journal of Bacteriology, November 1998, p. 5961-5967, Vol. 180, No. 22
0021-9193/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.
Transcriptional Activation of the Bacillus
subtilis ackA Gene Requires Sequences Upstream of the
Promoter
Andrew J.
Turinsky,1
Frank J.
Grundy,2
Jeong-Ho
Kim,3
Glenn H.
Chambliss,3 and
Tina
M.
Henkin2,*
Department of Microbiology, The Ohio State
University, Columbus, Ohio 432102;
Department of Biochemistry and Molecular Biology, Albany
Medical College, Albany, New York 122081;
and
Department of Bacteriology, University of Wisconsin,
Madison, Wisconsin 537063
Received 7 April 1998/Accepted 8 September 1998
Transcriptional activation of the Bacillus subtilis
ackA gene, encoding acetate kinase, was previously shown to
require catabolite control protein A (CcpA) and sequences upstream of
the ackA promoter. CcpA, which is responsible for
catabolite repression of a number of secondary carbon source
utilization genes in B. subtilis and other gram-positive
bacteria, recognizes a cis-acting consensus sequence,
designated cre (catabolite response element), generally located within or downstream of the promoter of the repressed gene. Two
sites resembling this sequence are centered at positions
116.5 and
56.5 of the ackA promoter and have been termed
cre1 and cre2, respectively. Synthesis of
acetate kinase, which is involved in the conversion of acetyl coenzyme
A to acetate, is induced when cells are grown in the presence of an
easily metabolized carbon source such as glucose. In this study,
cre2, the site closer to the promoter, and the region
upstream of cre2 were shown to be indispensable for
CcpA-dependent transcriptional activation of ackA, whereas
cre1 was not required. In addition, insertion of 5 bp
between cre2 and the promoter disrupted activation, while 10 bp was tolerated, suggesting face-of-the-helix dependence of the
position of cre2 and/or upstream sequences. DNase
footprinting experiments demonstrated binding of CcpA in vitro to
cre2 but not cre1, consistent with the genetic
data. Activation of ackA transcription was blocked in a
ptsH1/crh double mutant, suggesting involvement of this
pathway in CcpA-mediated transcriptional activation.
*
Corresponding author. Mailing address: Department of
Microbiology, The Ohio State University, 484 W. 12th Ave., Columbus, OH
43210. Phone: (614) 688-3831. Fax: (614) 292-8120. E-mail: henkin.3{at}osu.edu.
Journal of Bacteriology, November 1998, p. 5961-5967, Vol. 180, No. 22
0021-9193/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.
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