The Schizosaccharomyces pombe Corepressor Tup11 Interacts with the Iron-responsive Transcription Factor Fep1

Abstract

The Schizosaccharomyces pombe fep1 ؉ gene encodes a GATA transcription factor that represses the expression of iron transport genes in response to elevated iron concentrations. This transcriptional response is altered only in strains harboring a combined deletion of both tup11 ؉ and tup12 ؉ genes. This suggests that Tup11 is capable of negatively regulating iron transport gene expression in the absence of Tup12 and vice versa. The tup11 ؉ -and tup12 ؉ -encoded proteins resemble the Saccharomyces cerevisiae Tup1 corepressor. Using yeast two-hybrid analysis we show that Tup11 and Fep1 physically interact with each other. The C-terminal region from amino acids 242 to 564 of Fep1 is required for interaction with Tup11. Within this region, a minimal domain encompassing amino acids 405-541 was sufficient for Tup11-Fep1 association. Deletion mapping analysis revealed that the WD40-repeat sequence motifs of Tup11 are necessary for its interaction with Fep1. Analysis of Tup11 mutants with single amino acid substitutions in the WD40 repeats suggested that the Fep1 transcription factor interacts with a putative flat upper surface on the predicted ␤-propeller structure of this motif. Further analysis by in vivo coimmunoprecipitation showed that Tup11 and Fep1 are physically associated. In vitro pull-down experiments further verified a direct interaction between the Fep1 C terminus and the Tup11 C-terminal WD40 repeat domain. Taken together, these results describe the first example of a physical interaction between a corepressor and an iron-sensing factor controlling the expression of iron uptake genes.

Cti6 Is an Rpd3-Sin3 Histone Deacetylase-associated Protein Required for Growth under Iron-limiting Conditions in Saccharomyces cerevisiae

“…In addition, studies in S. cerevisiae and Candida albicans have shown that cells defective in Tup1 de-repress the iron-siderophore uptake system (53,54). Furthermore, Tup1 homologues in S. pombe (Tup11 and Tup12) are required for transcriptional repression of components of the iron transport machinery through a mechanism involving direct interaction with the GATA transcription factor Fep1, which is involved in repression of iron transport in response to elevated iron concentrations (25,55). Our genomic microarray and RNA blot analysis for wild type and cti6 mutant cells under conditions of iron limitation shows that several Tup1 target genes including the RNR genes, involved in DNA damage and replication, and the PAU and DAN families, expressed under anaerobic conditions, are up-regulated in the cti6 mutant, suggesting that Cti6 mediates transcriptional repression through the Ssn6-Tup1 corepressor complex.…”

“…This is possible because the Cti6 protein interacts simultaneously with the Ssn6-Tup1 and SAGA complexes, and mediates SAGA and TBP recruitment, histone acetylation, and transcriptional activation (17). Interestingly, recent studies in S. cerevisiae and Schizosaccharomyces pombe have also shown that the Tup1 complex (Tup11/Tup12 in S. pombe) modulates the expression of iron-regulated genes by associating with the Aft1 (Fep1 in S. pombe) transcription factor (25,26). …”

Functional Comparison of the Tup11 and Tup12 Transcriptional Corepressors in Fission Yeast

“…The fission yeast is unusual compared to other fungi because it contains two genes encoding Tup1-like corepressors. Evidence that the S. pombe proteins function similarly to the Tup1-Ssn6 complex in S. cerevisiae has been reported recently (12,14,16,23,24,27,45). Tup11 has been shown to repress target genes containing LexA binding sites when fused to the LexA DNAbinding domain, and binding studies have shown that Tup11 is able to interact with the S. cerevisiae transcription factor Mat␣2 (24).…”

“…Examples of known target genes for Tup11 and Tup12 are the intracellular cation transporter cta3 ϩ , which is activated under high-salt conditions (12). Tup11 has also been reported to interact directly with Fep1, a transcription factor that represses the expression of the iron transport genes fio1 ϩ and fip1 ϩ in response to high iron concentrations (27,45). Gene duplication is thought to be an important mechanism involved in the evolution of biological diversity.…”

Simon Labbé’s work on iron and copper homeostasis

“…These studies also demonstrated, for the first time, a central role for Fep1 in coordinating regulation of genes that encode components of the reductive and non-reductive iron transport systems [4] . Using a genetic approach to find GATA-factor-associated proteins, his research group identified the co-repressors Tup11 and Tup12 as physi-ological partners of Fep1 [5] . This is significant because these results describe the first example of a physical interaction between a co-repressor and an iron-sensing factor that control the expression of iron uptake genes [6,7] .…”