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Molecular and Cellular Biology, January 1999, p. 537-546, Vol. 19, No. 1
0270-7306/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.
Repressors and Upstream Repressing Sequences of the
Stress-Regulated ENA1 Gene in Saccharomyces
cerevisiae: bZIP Protein Sko1p Confers HOG-Dependent
Osmotic Regulation
Markus
Proft* and
Ramón
Serrano
Instituto de Biología Molecular y
Celular de Plantas, Universidad Politécnica de Valencia-CSIC,
46022 Valencia, Spain
Received 14 May 1998/Returned for modification 22 June
1998/Accepted 9 October 1998
The yeast ENA1/PMR2A gene encodes a cation extrusion
ATPase in Saccharomyces cerevisiae which is essential for
survival under salt stress conditions. One important mechanism of
ENA1 transcriptional regulation is based on repression
under normal growth conditions, which is relieved by either osmotic
induction or glucose starvation. Analysis of the ENA1
promoter revealed a Mig1p-binding motif (
533 to
544) which was
characterized as an upstream repressing sequence (URSMIG-ENA1) regulated by carbon source. Its
function was abolished in a mig1 mig2 double-deletion
strain as well as in either ssn6 or tup1 single
mutants. A second URS at
502 to
513 is responsible for
transcriptional repression regulated by osmotic stress and is similar
to mammalian cyclic AMP response elements (CREs) that are recognized by
CREB proteins. This URSCRE-ENA1 element
requires for its repression function the yeast CREB homolog Sko1p
(Acr1p) as well as the integrity of the Ssn6p-Tup1p corepressor complex. When targeted to the GAL1 promoter by fusing with
the Gal4p DNA-binding domain, Sko1p acts as an Ssn6/Tup1p-dependent repressor regulated by osmotic stress. A glutathione
S-transferase-Sko1 fusion protein binds specifically to
the URSCRE-ENA1 element. Furthermore, a
hog1 mitogen-activated protein kinase deletion strain could
not counteract repression on URSCRE-ENA1 during
osmotic shock. The loss of SKO1 completely restored
ENA1 expression in a hog1 mutant and partially
suppressed the osmotic stress sensitivity, qualifying Sko1p as a
downstream effector of the HOG pathway. Our results indicate that
different signalling pathways (HOG osmotic pathway and glucose
repression pathway) use distinct promoter elements of ENA1
(URSCRE-ENA1 and URSMIG-ENA1) via specific transcriptional
repressors (Sko1p and Mig1/2p) and via the general Ssn6p-Tup1p complex.
The physiological importance of the relief from repression during salt
stress was also demonstrated by the increased tolerance of sko1 or ssn6 mutants to Na+ or
Li+ stress.
*
Corresponding author. Mailing address: Instituto de
Biología Molecular y Celular de Plantas, Universidad
Politécnica de Valencia-CSIC, Camino de Vera s/n, 46022 Valencia,
Spain. Phone: 34-96-3877860. Fax: 34-96-3877859. E-mail:
mproft{at}ibmcp.upv.es.
Molecular and Cellular Biology, January 1999, p. 537-546, Vol. 19, No. 1
0270-7306/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.
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