Repressors and upstream repressing sequences of the stress-regulated ENA1 gene in Saccharomyces cerevisiae: bZIP protein Sko1p confers HOG-dependent osmotic regulation.
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.
Proft
M
M
Instituto de Biología Molecular y Celular de Plantas, Universidad Politécnica de Valencia-CSIC, 46022 Valencia, Spain. mproft@ibmcp.upv.es
Serrano
R
R
D000251:Adenosine Triphosphatases / Q000235:genetics*; D001483:Base Sequence; D050976:Basic-Leucine Zipper Transcription Factors; D001665:Binding Sites; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism; D027682:Cation Transport Proteins; D004271:DNA, Fungal; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D015966:Gene Expression Regulation, Fungal; D016350:Leucine Zippers; D020928:Mitogen-Activated Protein Kinases; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009687:Nuclear Proteins; D010641:Phenotype; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins; D000254:Sodium-Potassium-Exchanging ATPase; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D014882:Water-Electrolyte Balance
D050976:Basic-Leucine Zipper Transcription Factors; C065455:CYC8 protein, S cerevisiae; D027682:Cation Transport Proteins; D004271:DNA, Fungal; D004268:DNA-Binding Proteins; C086375:ENA1 protein, S cerevisiae; D005656:Fungal Proteins; C052130:GAL4 protein, S cerevisiae; C085976:MIG1 protein, S cerevisiae; D009687:Nuclear Proteins; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; C077832:SKO1 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; C066502:TUP1 protein, S cerevisiae; D014157:Transcription Factors; D017871:Calcium-Calmodulin-Dependent Protein Kinases; C088589:HOG1 protein, S cerevisiae; D020928:Mitogen-Activated Protein Kinases; D000251:Adenosine Triphosphatases; D000254:Sodium-Potassium-Exchanging ATPase