scholarly journals The Saccharomyces cerevisiae MADS-box transcription factor Rlm1 is a target for the Mpk1 mitogen-activated protein kinase pathway.

1997 ◽  
Vol 17 (4) ◽  
pp. 1848-1859 ◽  
Author(s):  
E Dodou ◽  
R Treisman
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Transcription Factor ◽  
Dna Binding ◽  
Transcriptional Activation ◽  
Mapk Pathway ◽  
Consensus Sequence ◽  
Binding Domain ◽  
Mitogen Activated Protein Kinase ◽  
Dna Binding Domain ◽  
Mads Box

Mutation of Saccharomyces cerevisiae RLM1, which encodes a MADS-box transcription factor, confers resistance to the toxic effects of constitutive activity of the Mpk1 mitogen-activated kinase (MAPK) pathway. The Rlm1 DNA-binding domain, which is similar to that of the metazoan MEF2 transcription factors, is also closely related to that of a second S. cerevisiae protein, Smp1 (second MEF2-like protein), encoded by the YBR182C open reading frame (N. Demolis et al., Yeast 10:1511-1525, 1994; H. Feldmann et al., EMBO J. 13:5795-5809, 1994). We show that Rlm1 and Smp1 have MEF2-related DNA-binding specificities: Rlm1 binds with the same specificity as MEF2, CTA(T/A)4TAG, while SMP1 binds a more extended consensus sequence, ACTACTA(T/A)4TAG. The two DNA-binding domains can heterodimerize with each other and with MEF2A. Deletion of RLM1 enhances resistance to cell wall disruptants, increases saturation density, reduces flocculation, and inactivates reporter genes controlled by the Rlm1 consensus binding site. Deletion of SMP1 neither causes these phenotypes nor enhances the Rlm1 deletion phenotype. However, overexpression of the DNA-binding domain of either protein causes an osmoremedial phenotype. Synthetic and naturally occurring MEF2 consensus sequences exhibit strong RLM1- and MPK1-dependent upstream activation sequence activity. Transcriptional activation by Rlm1 requires its C-terminal sequences, and Gal4 fusion proteins containing Rlm1 C-terminal sequences also act as MPK1-dependent transcriptional activators. These results establish the Rlm1 C-terminal sequences as a target for the Mpk1 MAPK pathway.

scholarly journals Adaptive evolution of transcription factor binding affinities

2017 ◽  
Author(s):  
Jungeui Hong ◽  
Nathan Brandt ◽  
Ally Yang ◽  
Tim Hughes ◽  
David Gresham
Keyword(s):  
Gene Expression ◽  
Transcription Factor ◽  
Dna Binding ◽  
Adaptive Evolution ◽  
Consensus Sequence ◽  
Binding Domain ◽  
Dna Binding Domain ◽  
Missense Mutations ◽  
Binding Affinities ◽  
Synonymous Mutations

Understanding the molecular basis of gene expression evolution is a central problem in evolutionary biology. However, connecting changes in gene expression to increased fitness, and identifying the functional basis of those changes, remains challenging. To study adaptive evolution of gene expression in real time, we performed long term experimental evolution (LTEE) of Saccharomyces cerevisiae (budding yeast) in ammonium-limited chemostats. Following several hundred generations of continuous selection we found significant divergence of nitrogen-responsive gene expression in lineages with increased fitness. In multiple independent lineages we found repeated selection for non-synonymous mutations in the zinc finger DNA binding domain of the activating transcription factor (TF), GAT1, that operates within incoherent feedforward loops to control expression of the nitrogen catabolite repression (NCR) regulon. Missense mutations in the DNA binding domain of GAT1 reduce its binding affinity for the GATAA consensus sequence in a promoter-specific manner, resulting in increased expression of ammonium permease genes via both direct and indirect effects, thereby conferring increased fitness. We find that altered transcriptional output of the NCR regulon results in antagonistic pleiotropy in alternate environments and that the DNA binding domain of GAT1 is subject to purifying selection in natural populations. Our study shows that adaptive evolution of gene expression can entail tuning expression output by quantitative changes in TF binding affinities while maintaining the overall topology of a gene regulatory network.

scholarly journals Functional domains of interferon regulatory factor I (IRF-1)

1998 ◽  
Vol 335 (1) ◽  
pp. 147-157 ◽  
Author(s):  
Fred SCHAPER ◽  
Sabine KIRCHHOFF ◽  
Guido POSERN ◽  
Mario KÖSTER ◽  
André OUMARD ◽  
...  
Keyword(s):  
Dna Binding ◽  
Transcriptional Activation ◽  
Mammalian Cells ◽  
Fluorescent Protein ◽  
Nuclear Translocation ◽  
Consensus Sequence ◽  
Binding Domain ◽  
Functional Domains ◽  
Dna Binding Domain ◽  
Transcriptional Activation Domain

Interferon (IFN) regulatory factors (IRFs) are a family of transcription factors among which are IRF-1, IRF-2, and IFN consensus sequence binding protein (ICSBP). These factors share sequence homology in the N-terminal DNA-binding domain. IRF-1 and IRF-2 are further related and have additional homologous sequences within their C-termini. Whereas IRF-2 and ICSBP are identified as transcriptional repressors, IRF-1 is an activator. In the present work, the identification of functional domains in murine IRF-1 with regard to DNA-binding, nuclear translocation, heterodimerization with ICSBP and transcriptional activation are demonstrated. The minimal DNA-binding domain requires the N-terminal 124 amino acids plus an arbitrary C-terminal extension. By using mutants of IRF-1 fusion proteins with green fluorescent protein and monitoring their distribution in living cells, a nuclear location signal (NLS) was identified and found to be sufficient for nuclear translocation. Heterodimerization was confirmed by a two-hybrid system adapted to mammalian cells. The heterodimerization domain in IRF-1 was defined by studies in vitroand was shown to be homologous with a sequence in IRF-2, suggesting that IRF-2 also heterodimerizes with ICSBP through this sequence. An acidic domain in IRF-1 was found to be required and to be sufficient for transactivation. Epitope mapping of IRF-1 showed that regions within the NLS, the heterodimerization domain and the transcriptional activation domain are exposed for possible contacts with interacting proteins.

scholarly journals The Schizosaccharomyces pombe mei4+ Gene Encodes a Meiosis-Specific Transcription Factor Containing a forkhead DNA-Binding Domain

1998 ◽  
Vol 18 (4) ◽  
pp. 2118-2129 ◽  
Author(s):  
S. Horie ◽  
Y. Watanabe ◽  
K. Tanaka ◽  
S. Nishiwaki ◽  
H. Fujioka ◽  
...  
Keyword(s):  
Amino Acids ◽  
Transcription Factor ◽  
Dna Binding ◽  
Schizosaccharomyces Pombe ◽  
Consensus Sequence ◽  
Binding Domain ◽  
Dna Binding Domain ◽  
Specific Transcription Factor ◽  
Amino Terminal ◽  
Diploid Cells

ABSTRACT The mei4 + gene of the fission yeastSchizosaccharomyces pombe was cloned by functional complementation. The mei4 disruptant failed to complete meiosis-I but could proliferate normally. mei4 +was transcribed only in meiosis-proficient diploid cells after premeiotic DNA replication. The mei4 + open reading frame encodes a 57-kDa serine-rich protein comprised of 517 amino acids with a forkhead/HNF3 DNA-binding domain in the amino-terminal region. Transcription of spo6 +, a gene required for sporulation, was dependent on themei4 + function. Two copies of the GTAAAYA consensus sequence, proposed as the binding site for human forkhead proteins, were found in the promoter region ofspo6 +. A gel mobility shift assay demonstrated the sequence-dependent binding of the GST-Mei4 forkhead domain fusion protein to DNA fragments with one of the consensus elements. Deletion of this consensus element from the spo6 promoter abolished the transcription of spo6 + and resulted in a sporulation deficiency. One-hybrid assay of Mei4 which was fused to the Gal4 DNA-binding domain localized the transcriptional activation domain in the C-terminal 140 amino acids of Mei4. These results indicate that Mei4 functions as a meiosis-specific transcription factor of S. pombe.

scholarly journals The DNA-binding domain of CaNdt80p is required to activate CDR1 involved in drug resistance in Candida albicans

2006 ◽  
Vol 55 (10) ◽  
pp. 1403-1411 ◽  
Author(s):  
Jang-Shiun Wang ◽  
Yun-Liang Yang ◽  
Chin-Jung Wu ◽  
Karen J. Ouyang ◽  
Kuo-Yun Tseng ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Drug Resistance ◽  
Transcription Factor ◽  
Candida Albicans ◽  
Dna Binding ◽  
Efflux Pump ◽  
Binding Domain ◽  
Null Mutation ◽  
Dna Binding Domain ◽  
Positive Regulator

CaNdt80p, the Candida albicans homologue of the Saccharomyces cerevisiae transcription factor ScNdt80p, has been identified as a positive regulator of CDR1, which encodes an efflux pump involved in drug resistance in C. albicans. To investigate the involvement of the putative DNA-binding domain of CaNdt80p in drug resistance, chimeras of CaNdt80p and ScNdt80p were constructed. Interestingly, the DNA-binding domain of ScNdt80p could functionally complement that of CaNdt80p to activate CDR1p–lacZ in S. cerevisiae. Consistently, CaNdt80p containing a mutation in the DNA-binding domain failed to activate CDR1p–lacZ in S. cerevisiae. Furthermore, a copy of CaNDT80 with the same mutation also failed to complement the drug-sensitive phenotype caused by a null mutation in C. albicans. Thus, the DNA-binding domain of CaNdt80p is critical for its function in drug resistance in C. albicans.

scholarly journals Cloning and characterization of GETS-1, a goldfish Ets family member that functions as a transcriptional repressor in muscle

1998 ◽  
Vol 335 (2) ◽  
pp. 267-275 ◽  
Author(s):  
Daniel GOLDMAN ◽  
Mohan K. SAPRU ◽  
Scott STEWART ◽  
Joshua PLOTKIN ◽  
Towia A. LIBERMANN ◽  
...  
Keyword(s):  
Dna Binding ◽  
Map Kinase ◽  
Family Member ◽  
Transcriptional Activation ◽  
Phosphorylation Site ◽  
Binding Domain ◽  
Mitogen Activated Protein Kinase ◽  
Dna Binding Domain ◽  
Subunit Gene ◽  
Activation Domain

An Ets transcription factor family member, GETS-1, was cloned from a goldfish retina cDNA library. GETS-1 contains a conserved Ets DNA-binding domain at its N-terminus and is most similar to ternary complex factor (TCF) serum-response-factor protein-1a (SAP-1a). GETS-1 is expressed in many tissues, but is enriched in retina and brain. As with the TCFs SAP-1a and ets-related protein (ERP), overexpression of the GETS-1 promoter suppresses nicotinic acetylcholine receptor ε-subunit gene expression in cultured muscle cells. A consensus Ets binding site sequence in the promoter of the ε-subunit gene is required for GETS-1-mediated repression. GETS-1 repressor activity is abrogated by overexpression of an activated Ras/mitogen-activated protein kinase (MAP kinase) or by mutation of Ser-405, a MAP kinase phosphorylation site in GETS-1. Fusion proteins created between GETS-1 and the Gal4 DNA-binding domain show that, like other TCFs, GETS-1 contains a C-terminal activation domain that is activated by a Ras/MAP kinase signalling cascade. Interestingly, mutation of Ser-405 located within this activation domain abrogated transcriptional activation of the fusion protein.

scholarly journals Two Regulators of Ste12p Inhibit Pheromone-Responsive Transcription by Separate Mechanisms

2000 ◽  
Vol 20 (12) ◽  
pp. 4199-4209 ◽  
Author(s):  
K. Amy Olson ◽  
Chris Nelson ◽  
Georgia Tai ◽  
Wesley Hung ◽  
Carl Yong ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Transcription Factor ◽  
Dna Binding ◽  
Transcriptional Activation ◽  
Dna Binding Domain ◽  
Wild Type ◽  
C Terminus ◽  
Multiple Regions ◽  
Vp16 Fusion

ABSTRACT The yeast Saccharomyces cerevisiae transcription factor Ste12p is responsible for activating genes in response to MAP kinase cascades controlling mating and filamentous growth. Ste12p is negatively regulated by two inhibitor proteins, Dig1p (also called Rst1p) and Dig2p (also called Rst2p). The expression of a C-terminal Ste12p fragment (residues 216 to 688) [Ste12p(216–688)] from aGAL promoter causes FUS1 induction in a strain expressing wild-type STE12, suggesting that this region can cause the activation of endogenous Ste12p. Residues 262 to 594 are sufficient to cause STE12-dependent FUS1induction when overexpressed, and this region of Ste12p was found to bind Dig1p but not Dig2p in yeast extracts. In contrast, recombinant glutathione S-transferase–Dig2p binds to the Ste12p DNA-binding domain (DBD). Expression of DIG2, but notDIG1, from a GAL promoter inhibits transcriptional activation by an Ste12p DBD-VP16 fusion. Furthermore, disruption of dig1, but not dig2, causes elevated transcriptional activation by a LexA–Ste12p(216–688) fusion. Ste12p has multiple regions within the C terminus (flanking residue 474) that can promote multimerization in vitro, and we demonstrate that these interactions can contribute to the activation of endogenous Ste12p by overproduced C-terminal fragments. These results demonstrate that Dig1p and Dig2p do not function by redundant mechanisms but rather inhibit pheromone-responsive transcription through interactions with separate regions of Ste12p.

scholarly journals Positive and Negative Regulation of the Cardiovascular Transcription Factor KLF5 by p300 and the Oncogenic Regulator SET through Interaction and Acetylation on the DNA-Binding Domain

2003 ◽  
Vol 23 (23) ◽  
pp. 8528-8541 ◽  
Author(s):  
Saku Miyamoto ◽  
Toru Suzuki ◽  
Shinsuke Muto ◽  
Kenichi Aizawa ◽  
Akatsuki Kimura ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Dna Binding ◽  
Transcriptional Activation ◽  
Affinity Purification ◽  
Gene Activation ◽  
Negative Regulation ◽  
Binding Domain ◽  
Negative Regulator ◽  
The Other ◽  
Dna Binding Domain

ABSTRACT Here we show a novel pathway of transcriptional regulation of a DNA-binding transcription factor by coupled interaction and modification (e.g., acetylation) through the DNA-binding domain (DBD). The oncogenic regulator SET was isolated by affinity purification of factors interacting with the DBD of the cardiovascular transcription factor KLF5. SET negatively regulated KLF5 DNA binding, transactivation, and cell-proliferative activities. Down-regulation of the negative regulator SET was seen in response to KLF5-mediated gene activation. The coactivator/acetylase p300, on the other hand, interacted with and acetylated KLF5 DBD, and activated its transcription. Interestingly, SET inhibited KLF5 acetylation, and a nonacetylated mutant of KLF5 showed reduced transcriptional activation and cell growth complementary to the actions of SET. These findings suggest a new pathway for regulation of a DNA-binding transcription factor on the DBD through interaction and coupled acetylation by two opposing regulatory factors of a coactivator/acetylase and a negative cofactor harboring activity to inhibit acetylation.

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