The Zygosaccharomyces bailii transcription factor Haa1 is required for acetic acid and copper stress responses suggesting subfunctionalization of the ancestral bifunctional protein Haa1/Cup2.

BACKGROUND: The food spoilage yeast species Zygosaccharomyces bailii exhibits an extraordinary capacity to tolerate weak acids, in particular acetic acid. In Saccharomyces cerevisiae, the transcription factor Haa1 (ScHaa1) is considered the main player in genomic expression reprogramming in response to acetic acid stress, but the role of its homologue in Z. bailii (ZbHaa1) is unknown. RESULTS: In this study it is demonstrated that ZbHaa1 is a ScHaa1 functional homologue by rescuing the acetic acid susceptibility phenotype of S. cerevisiae haa1Δ. The disruption of ZbHAA1 in Z. bailii IST302 and the expression of an extra ZbHAA1 copy confirmed ZbHAA1 as a determinant of acetic acid tolerance. ZbHaa1 was found to be required for acetic acid stress-induced transcriptional activation of Z. bailii genes homologous to ScHaa1-target genes. An evolutionary analysis of the Haa1 homologues identified in 28 Saccharomycetaceae species genome sequences, including Z bailii, was carried out using phylogenetic and gene neighbourhood approaches. Consistent with previous studies, this analysis revealed a group containing pre-whole genome duplication species Haa1/Cup2 single orthologues, including ZbHaa1, and two groups containing either Haa1 or Cup2 orthologues from post-whole genome duplication species. S. cerevisiae Cup2 (alias Ace1) is a transcription factor involved in response and tolerance to copper stress. Taken together, these observations led us to hypothesize and demonstrate that ZbHaa1 is also involved in copper-induced transcriptional regulation and copper tolerance. CONCLUSIONS: The transcription factor ZbHaa1 is required for adaptive response and tolerance to both acetic acid and copper stresses. The subfunctionalization of the single ancestral Haa1/Cup2 orthologue that originated Haa1 and Cup2 paralogues after whole genome duplication is proposed.

Phylogenetic and syntenic analyses of the 12-spanner drug:H(+) antiporter family 1 (DHA1) in pathogenic Candida species: evolution of MDR1 and FLU1 genes.

Candida albicans and other pathogenic Candida species can develop resistance to clinical fungicides through active drug export mediated by multidrug efflux pumps, in particular by members of the drug:H(+) antiporter family 1 (DHA1). The DHA1 proteins encoded in the genomes of 31 hemiascomycetous strains from 25 species were identified and homology relationships between these proteins and the functionally characterised DHA1 in the model yeast Saccharomyces cerevisiae were established. Gene neighbourhood analysis allowed the reconstruction of sixteen DHA1 lineages conserved during the CTG complex species evolution. The evolutionary history of C. albicans MDR1 and FLU1 genes and Candida dubliniensis, Candida tropicalis and Candida parapsilosis MDR1 genes was detailed. Candida genomes show an abundant number of MDR1 and FLU1 homologues but the chromosome environment where MDR1 homologues reside was poorly conserved during evolution. Gene duplication and loss are major mechanisms underlying the evolution of the DHA1 genes in Candida species.

The drug:H⁺ antiporters of family 2 (DHA2), siderophore transporters (ARN) and glutathione:H⁺ antiporters (GEX) have a common evolutionary origin in hemiascomycete yeasts.

BACKGROUND: The Saccharomyces cerevisiae 14-spanner Drug:H+ Antiporter family 2 (DHA2) are transporters of the Major Facilitator Superfamily (MFS) involved in multidrug resistance (MDR). Although poorly characterized, DHA2 family members were found to participate in the export of structurally and functionally unrelated compounds or in the uptake of amino acids into the vacuole or the cell. In S. cerevisiae, the four ARN/SIT family members encode siderophore transporters and the two GEX family members encode glutathione extrusion pumps. The evolutionary history of DHA2, ARN and GEX genes, encoding 14-spanner MFS transporters, is reconstructed in this study. RESULTS: The translated ORFs of 31 strains from 25 hemiascomycetous species, including 10 pathogenic Candida species, were compared using a local sequence similarity algorithm. The constraining and traversing of a network representing the pairwise similarity data gathered 355 full size proteins and retrieved ARN and GEX family members together with DHA2 transporters, suggesting the existence of a close phylogenetic relationship among these 14-spanner major facilitators. Gene neighbourhood analysis was combined with tree construction methodologies to reconstruct their evolutionary history and 7 DHA2 gene lineages, 5 ARN gene lineages, and 1 GEX gene lineage, were identified. The S. cerevisiae DHA2 proteins Sge1, Azr1, Vba3 and Vba5 co-clustered in a large phylogenetic branch, the ATR1 and YMR279C genes were proposed to be paralogs formed during the Whole Genome Duplication (WGD) whereas the closely related ORF YOR378W resides in its own lineage. Homologs of S. cerevisiae DHA2 vacuolar proteins Vba1, Vba2 and Vba4 occur widespread in the Hemiascomycetes. Arn1/Arn2 homologs were only found in species belonging to the Saccharomyces complex and are more abundant in the pre-WGD species. Arn4 homologs were only found in sub-telomeric regions of species belonging to the Sacharomyces sensu strictu group (SSSG). Arn3 type siderophore transporters are abundant in the Hemiascomycetes and form an ancient gene lineage extending to the filamentous fungi. CONCLUSIONS: The evolutionary history of DHA2, ARN and GEX genes was reconstructed and a common evolutionary root shared by the encoded proteins is hypothesized. A new protein family, denominated DAG, is proposed to span these three phylogenetic subfamilies of 14-spanner MFS transporters.

Evolution of the 12-spanner drug:H+ antiporter DHA1 family in hemiascomycetous yeasts.

Frequently, although not exclusively, multidrug resistance (MDR) results from the action of drug-efflux pumps, which are thought to be able to catalyze the active expulsion of several unrelated cytotoxic compounds out of the cell or their intracellular partitioning. The transporters of the major facilitator superfamily (MFS) presumably involved in MDR belong to the 12-spanner drug:H(+) antiporter DHA1 or to the 14- spanner drug:H(+) antiporter DHA2 families. The expression of most Saccharomyces cerevisiae DHA1 family members was found to confer broad chemoprotection. The evolution of the hemiascomycetous DHA1 proteins, belonging to the Génolevures GL3C007 family, was studied using a combined phylogenetic and gene neighborhood approach. The phylogenetic analysis of 189 DHA1 proteins belonging to the genome of 13 hemiascomycetous species identified 20 clusters. Eleven clusters contained no S. cerevisiae members. The phylogenetic clusters were analyzed by the IONS method developed for Identification of Orthologues by Neighborhood and Similarity. This allowed reconstructing the evolutionary history of most DHA1 members within 10 main gene lineages, spanning the whole hemiascomycetes clade, encompassing an evolutionary history of about 350 million years. In addition, five other more species specific lineages, spanning only two hemiascomycetous species, were identified. It is concluded that 57 out of the 143 members of the DHA1 hemiascomycetous members originated from gene duplication events. In half of these duplicates, the two members belong to different phylogenetic clusters, indicating that at least one of them has sufficiently differentiated to provide potential novel functions to this complex family from which most physiological substrates remain unknown.

Yeast adaptation to mancozeb involves the up-regulation of FLR1 under the coordinate control of Yap1, Rpn4, Pdr3, and Yrr1.

FLR1 gene, encoding a multidrug resistance (MDR) transporter of the major facilitator superfamily (MFS) was found to confer resistance to the fungicide mancozeb in Saccharomyces cerevisiae. This agrochemical has been linked to the development of Parkinson disease and cancer. Yeast response to mancozeb was proved to involve the strong activation of FLR1 transcription (20-fold) during the fungicide-induced growth latency. This activation of FLR1 transcription is fully dependent on Yap1p and is reduced (by 50%) in the absence of Rpn4p, Yrr1p or Pdr3p. A model for the coordinate action over FLR1 transcription activation, in response to mancozeb, of these transcription factors that mediate oxidative stress response (Yap1p), proteasome gene expression (Rpn4p), and pleiotropic drug resistance (Pdr3p and Yrr1p), is proposed.