title: Two residues in the basic region of the yeast transcription factor Yap8 are crucial for its DNA-binding specificity.
abstract: In Saccharomyces cerevisiae, the transcription factor Yap8 is a key determinant in arsenic stress response. Contrary to Yap1, another basic region-leucine zipper (bZIP) yeast regulator, Yap8 has a very restricted DNA-binding specificity and only orchestrates the expression of ACR2 and ACR3 genes. In the DNA-binding basic region, Yap8 has three distinct amino acids residues, Leu26, Ser29 and Asn31, at sites of highly conserved positions in the other Yap family of transcriptional regulators and Pap1 of Schizosaccharomyces pombe. To evaluate whether these residues are relevant to Yap8 specificity, we first built a homology model of the complex Yap8bZIP-DNA based on Pap1-DNA crystal structure. Several Yap8 mutants were then generated in order to confirm the contribution of the residues predicted to interact with DNA. Using bioinformatics analysis together with in vivo and in vitro approaches, we have identified several conserved residues critical for Yap8-DNA binding. Moreover, our data suggest that Leu26 is required for Yap8 binding to DNA and that this residue together with Asn31, hinder Yap1 response element recognition by Yap8, thus narrowing its DNA-binding specificity. Furthermore our results point to a role of these two amino acids in the stability of the Yap8-DNA complex.
authors.affiliation: Genomics and Stress Laboratory, Instituto de Tecnologia Química e Biológica António Xavier, Universidade Nova de Lisboa, Oeiras, Portugal.
authors.forename: Catarina
authors.initials: C
authors.lastname: Amaral
authors.affiliation: Genomics and Stress Laboratory, Instituto de Tecnologia Química e Biológica António Xavier, Universidade Nova de Lisboa, Oeiras, Portugal.
authors.forename: Catarina
authors.initials: C
authors.lastname: Pimentel
authors.affiliation: Control of Gene Expression Laboratory, Instituto de Tecnologia Química e Biológica António Xavier, Universidade Nova de Lisboa, Oeiras, Portugal.
authors.forename: Rute G
authors.initials: RG
authors.lastname: Matos
authors.affiliation: Control of Gene Expression Laboratory, Instituto de Tecnologia Química e Biológica António Xavier, Universidade Nova de Lisboa, Oeiras, Portugal.
authors.forename: Cecília M
authors.initials: CM
authors.lastname: Arraiano
authors.affiliation: Biomolecular NMR Laboratory, Instituto de Tecnologia Química e Biológica António Xavier, Universidade Nova de Lisboa, Oeiras, Portugal.
authors.forename: Manolis
authors.initials: M
authors.lastname: Matzapetakis
authors.affiliation: Genomics and Stress Laboratory, Instituto de Tecnologia Química e Biológica António Xavier, Universidade Nova de Lisboa, Oeiras, Portugal.
authors.forename: Claudina
authors.initials: C
authors.lastname: Rodrigues-Pousada
mesh_terms: D000595:Amino Acid Sequence; D050976:Basic-Leucine Zipper Transcription Factors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D017124:Conserved Sequence; D004247:DNA / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D000073718:Pancreatitis-Associated Proteins; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs / Q000235:genetics; D020218:Response Elements; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D012568:Schizosaccharomyces / Q000235:genetics / Q000378:metabolism; D017385:Sequence Homology; D014157:Transcription Factors / Q000235:genetics
chemical_list: C494591:ARR1 protein, S cerevisiae; D050976:Basic-Leucine Zipper Transcription Factors; D004268:DNA-Binding Proteins; D000073718:Pancreatitis-Associated Proteins; C000612141:REG3A protein, human; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; C086677:YAP1 protein, S cerevisiae; D004247:DNA