Cat8 and Sip4 mediate regulated transcriptional activation of the yeast malate dehydrogenase gene MDH2 by three carbon source-responsive promoter elements
S. Roth et Hj. Schuller, Cat8 and Sip4 mediate regulated transcriptional activation of the yeast malate dehydrogenase gene MDH2 by three carbon source-responsive promoter elements, YEAST, 18(2), 2001, pp. 151-162
Malate dehydrogenase isoenzymes are localized in different cellular compart
ments and fulfil important functions in intermediary metabolism. In the yea
st Saccharomyces cerevisiae, three malate dehydrogenase genes, MDH1, MDH2 a
nd MDH3, encoding mitochondrial, cytosolic and peroxisomal variants, have b
een identified. We demonstrate the importance of transcriptional activators
Hap4, Cats and Pip2 for the carbon source-dependent regulation of MDH1, MD
H2 and MDH3, respectively. The control region of the MDH2 gene required for
gluconeogenic growth with C-2 substrates contains three sequence elements
similar to the previously identified carbon source-responsive element (CSRE
), In a synthetic test system, each of these sequences turned out to be a w
eak UAS element showing a strong synergism when present in multiple copies.
Cumulative mutagenesis of the natural MDH2 promoter confirmed the contribu
tion of all three elements to transcriptional derepression under non-fermen
tative growth conditions. The DNA-binding domains of zinc fluster proteins
Cats and Sip4 synthesized in Escherichia coil could interact in vitro with
CSRE motifs of MDH2, This result was confirmed by binding assays using prot
ein extracts from yeast. Deregulated variants of Cats and Sip4 modified by
heterologous transcriptional activation domains were able to alleviate gluc
ose repression of MDH2 substantially. Although Sip4 turned out as the less
effective activator, our findings demonstrate the general significance of b
oth proteins for expression of gluconeogenic structural genes. Copyright (C
) 2000 John Wiley & Sons, Ltd.