Functional characterization of KlHAP1: a model to foresee different mechanisms of transcriptional regulation by Hap1p in yeasts.
In this work we have cloned and characterized the Kluyveromyces lactis HAP1 gene and we have found that, contrary to data previously described for the homologous gene of Saccharomyces cerevisiae, i.) the function of this gene does not affect growth in media with carbon sources used by fermentative or respiratory pathways ii) in aerobiosis, KlHap1p is not a transcriptional activator of the expression of genes related to respiration, cholesterol biosynthesis or oxidative stress defence analyzed in this study. The comparison of homology between specific regions of ScHap1p and KlHap1p reveals that the dimerization domain is poorly conserved and we have verified that this domain, cloned in the two plasmids of the two hybrid system, does not reconstitute S. cerevisiae Gal4p activity. Since the COOH-terminal transcriptional activation domain of KlHap1p is active when fused to the Gal4p-DNA binding domain, we hypothesize that differences in the capacity to form dimers could contribute to allow different functions of the protein in K. lactis and S. cerevisiae. Transcriptional expression of KlHAP1 is dependent on oxygen availability, increasing its expression in hypoxia. Deletion of KlHAP1 increases the resistance to oxidative stress or cadmium and the induction of KlYAP1 and KlTSA1 by the addition of 0.5 mM H(2)O(2) is repressed by KlHap1p. These data are discussed in reference to the evolution of respiro-fermentative metabolism in yeasts.
Lamas-Maceiras
Mónica
M
Dpto. Biología Celular y Molecular. Universidad de A Coruña, F. Ciencias, Campus de A Zapateira s/n 15075, A Coruña, Spain.
Núñez
Laura
L
Rodríguez-Belmonte
Esther
E
González-Siso
María Isabel
MI
Cerdán
María Esperanza
ME
D000595:Amino Acid Sequence; D001483:Base Sequence; D002104:Cadmium / Q000494:pharmacology; D017931:DNA Primers; D015966:Gene Expression Regulation, Fungal; D005800:Genes, Fungal; D006861:Hydrogen Peroxide / Q000494:pharmacology; D008957:Models, Genetic; D008969:Molecular Sequence Data; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D017386:Sequence Homology, Amino Acid; D014158:Transcription, Genetic
D017931:DNA Primers; D002104:Cadmium; D006861:Hydrogen Peroxide