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Tec1 mediates the pheromone response of the white phenotype of Candida albicans: insights into the evolution of new signal transduction pathways.

Sahni N, Yi S, Daniels KJ, Huang G, Srikantha T, Soll DR - PLoS Biol. (2010)

Bottom Line: The three portions of this new regulatory pathway appear to have been derived from three different ancestral programs still functional in C. albicans.The upstream portion, including signals, receptors, the trimeric G protein complex, and the MAP kinase cascade, was derived intact from the upstream portion of the opaque pheromone response pathway of the mating process; Tec1, the transcription factor targeted by the MAP kinase pathway, was derived from a filamentation pathway; and the white-specific downstream target genes were derived from an ancestral biofilm process.The evolution of this pheromone response pathway provides a possible paradigm for how such signal transduction pathways evolve.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, The University of Iowa, Iowa City, Iowa, United States of America.

ABSTRACT
The way in which signal transduction pathways evolve remains a mystery, primarily because we have few examples of ones that have newly evolved. There are numerous examples of how signal transduction pathways in the same organism selectively share components, most notably between the signal transduction pathways in Saccharomyces cerevisiae for the mating process, the filamentation process, cell wall integrity, ascospore formation, and osmoregulation. These examples, however, have not provided insights into how such pathways evolve. Here, through construction of an overexpression library for 107 transcription factors, and through mutational analyses, we have identified the transcription factor Tec1 as the last component of the newly evolved signal transduction pathway that regulates the pheromone response of the white cell phenotype in Candida albicans. The elucidation of this last component, Tec1, establishes a comprehensive description of the pheromone response pathway in the white cell phenotype of C. albicans, providing a unique perspective on how new signal transduction pathways may evolve. The three portions of this new regulatory pathway appear to have been derived from three different ancestral programs still functional in C. albicans. The upstream portion, including signals, receptors, the trimeric G protein complex, and the MAP kinase cascade, was derived intact from the upstream portion of the opaque pheromone response pathway of the mating process; Tec1, the transcription factor targeted by the MAP kinase pathway, was derived from a filamentation pathway; and the white-specific downstream target genes were derived from an ancestral biofilm process. The evolution of this pheromone response pathway provides a possible paradigm for how such signal transduction pathways evolve.

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Tec1 directly interacts with WPRE-containing promoter regions and localizes in the nucleus.(A) ChIP analysis of gene promoters that bind to Tec1. The gene categories for the promoters screened for are presented. The primers used for the promoter regions are listed in Table S6. An anti-myc antibody was used to immunoprecipitate chromatin. “Input” represents chromatin preparation before immunoprecipitation, and “−Ab” and “+Ab” represent the absence and presence, respectively, of anti-myc antibody. (B) Opaque and white cells of strain tec1/tec1-TEC1, in which TEC1 is tagged with GFP, stained for DNA using DAPI, and imaged for DAPI staining and GFP fluorescence.
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pbio-1000363-g004: Tec1 directly interacts with WPRE-containing promoter regions and localizes in the nucleus.(A) ChIP analysis of gene promoters that bind to Tec1. The gene categories for the promoters screened for are presented. The primers used for the promoter regions are listed in Table S6. An anti-myc antibody was used to immunoprecipitate chromatin. “Input” represents chromatin preparation before immunoprecipitation, and “−Ab” and “+Ab” represent the absence and presence, respectively, of anti-myc antibody. (B) Opaque and white cells of strain tec1/tec1-TEC1, in which TEC1 is tagged with GFP, stained for DNA using DAPI, and imaged for DAPI staining and GFP fluorescence.

Mentions: Since the transcription factor Tec1 mediates α-pheromone–induced expression of genes in white cells, and WPRE is the presumed cis-acting regulatory sequence [23] that mediates up-regulation, we tested whether there was a direct interaction between Tec1 and the promoters of these select genes using a chromatin immunoprecipitation (ChIP) assay [43]–[45]. TEC1 was tagged with myc at the 3′ end in the heterozygous deletion mutant to generate the strain tec1/TEC1-myc. White cells of this strain were treated with α-pheromone to induce the putative interaction between Tec1-myc and the promoters of regulated genes. An antibody against myc was then used to immunoprecipitate chromatin fragments bound to Tec1-myc. Immunoprecipitated DNA was then amplified by the polymerase chain reaction with primers designed to span approximately 400 base pairs of the promoter region harboring the WPRE in the case of white-specific genes and the OPRE in the case of opaque-specific genes (Table S6). The white-specific gene promoters tested for were CSH1, PBR1, RBT5, and WH11, and the opaque-specific gene promoters tested for were KAR4 and MFA1 [10],[17],[23],[42]. We also tested for the promoters of STE2 and RBT1, which contain both a WPRE and OPRE [23]. Finally, we tested for the promoter of ACT1, which contains neither a WPRE nor an OPRE [23]. The promoters coimmunoprecipitated by the anti-myc antibody were those of genes selectively up-regulated by α-pheromone in white but not opaque cells (CSH1, PBR1, RBT5, and WH11), and those up-regulated by α-pheromone in both white and opaque cells (STE2, RBT1) (Figure 4A). The promoters of genes up-regulated through Cph1 in opaque cells only (KAR4, MFA1) and the promoter of the gene ACT1 were not coimmunoprecipitated by the anti-myc antibody (Figure 4A). These results demonstrate that Tec1 binds selectively to WPRE-containing promoters of the genes up-regulated by α-pheromone in white cells, but not to OPRE-containing promoters of genes up-regulated by α-pheromone only in opaque cells.


Tec1 mediates the pheromone response of the white phenotype of Candida albicans: insights into the evolution of new signal transduction pathways.

Sahni N, Yi S, Daniels KJ, Huang G, Srikantha T, Soll DR - PLoS Biol. (2010)

Tec1 directly interacts with WPRE-containing promoter regions and localizes in the nucleus.(A) ChIP analysis of gene promoters that bind to Tec1. The gene categories for the promoters screened for are presented. The primers used for the promoter regions are listed in Table S6. An anti-myc antibody was used to immunoprecipitate chromatin. “Input” represents chromatin preparation before immunoprecipitation, and “−Ab” and “+Ab” represent the absence and presence, respectively, of anti-myc antibody. (B) Opaque and white cells of strain tec1/tec1-TEC1, in which TEC1 is tagged with GFP, stained for DNA using DAPI, and imaged for DAPI staining and GFP fluorescence.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2864266&req=5

pbio-1000363-g004: Tec1 directly interacts with WPRE-containing promoter regions and localizes in the nucleus.(A) ChIP analysis of gene promoters that bind to Tec1. The gene categories for the promoters screened for are presented. The primers used for the promoter regions are listed in Table S6. An anti-myc antibody was used to immunoprecipitate chromatin. “Input” represents chromatin preparation before immunoprecipitation, and “−Ab” and “+Ab” represent the absence and presence, respectively, of anti-myc antibody. (B) Opaque and white cells of strain tec1/tec1-TEC1, in which TEC1 is tagged with GFP, stained for DNA using DAPI, and imaged for DAPI staining and GFP fluorescence.
Mentions: Since the transcription factor Tec1 mediates α-pheromone–induced expression of genes in white cells, and WPRE is the presumed cis-acting regulatory sequence [23] that mediates up-regulation, we tested whether there was a direct interaction between Tec1 and the promoters of these select genes using a chromatin immunoprecipitation (ChIP) assay [43]–[45]. TEC1 was tagged with myc at the 3′ end in the heterozygous deletion mutant to generate the strain tec1/TEC1-myc. White cells of this strain were treated with α-pheromone to induce the putative interaction between Tec1-myc and the promoters of regulated genes. An antibody against myc was then used to immunoprecipitate chromatin fragments bound to Tec1-myc. Immunoprecipitated DNA was then amplified by the polymerase chain reaction with primers designed to span approximately 400 base pairs of the promoter region harboring the WPRE in the case of white-specific genes and the OPRE in the case of opaque-specific genes (Table S6). The white-specific gene promoters tested for were CSH1, PBR1, RBT5, and WH11, and the opaque-specific gene promoters tested for were KAR4 and MFA1 [10],[17],[23],[42]. We also tested for the promoters of STE2 and RBT1, which contain both a WPRE and OPRE [23]. Finally, we tested for the promoter of ACT1, which contains neither a WPRE nor an OPRE [23]. The promoters coimmunoprecipitated by the anti-myc antibody were those of genes selectively up-regulated by α-pheromone in white but not opaque cells (CSH1, PBR1, RBT5, and WH11), and those up-regulated by α-pheromone in both white and opaque cells (STE2, RBT1) (Figure 4A). The promoters of genes up-regulated through Cph1 in opaque cells only (KAR4, MFA1) and the promoter of the gene ACT1 were not coimmunoprecipitated by the anti-myc antibody (Figure 4A). These results demonstrate that Tec1 binds selectively to WPRE-containing promoters of the genes up-regulated by α-pheromone in white cells, but not to OPRE-containing promoters of genes up-regulated by α-pheromone only in opaque cells.

Bottom Line: The three portions of this new regulatory pathway appear to have been derived from three different ancestral programs still functional in C. albicans.The upstream portion, including signals, receptors, the trimeric G protein complex, and the MAP kinase cascade, was derived intact from the upstream portion of the opaque pheromone response pathway of the mating process; Tec1, the transcription factor targeted by the MAP kinase pathway, was derived from a filamentation pathway; and the white-specific downstream target genes were derived from an ancestral biofilm process.The evolution of this pheromone response pathway provides a possible paradigm for how such signal transduction pathways evolve.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, The University of Iowa, Iowa City, Iowa, United States of America.

ABSTRACT
The way in which signal transduction pathways evolve remains a mystery, primarily because we have few examples of ones that have newly evolved. There are numerous examples of how signal transduction pathways in the same organism selectively share components, most notably between the signal transduction pathways in Saccharomyces cerevisiae for the mating process, the filamentation process, cell wall integrity, ascospore formation, and osmoregulation. These examples, however, have not provided insights into how such pathways evolve. Here, through construction of an overexpression library for 107 transcription factors, and through mutational analyses, we have identified the transcription factor Tec1 as the last component of the newly evolved signal transduction pathway that regulates the pheromone response of the white cell phenotype in Candida albicans. The elucidation of this last component, Tec1, establishes a comprehensive description of the pheromone response pathway in the white cell phenotype of C. albicans, providing a unique perspective on how new signal transduction pathways may evolve. The three portions of this new regulatory pathway appear to have been derived from three different ancestral programs still functional in C. albicans. The upstream portion, including signals, receptors, the trimeric G protein complex, and the MAP kinase cascade, was derived intact from the upstream portion of the opaque pheromone response pathway of the mating process; Tec1, the transcription factor targeted by the MAP kinase pathway, was derived from a filamentation pathway; and the white-specific downstream target genes were derived from an ancestral biofilm process. The evolution of this pheromone response pathway provides a possible paradigm for how such signal transduction pathways evolve.

Show MeSH
Related in: MedlinePlus