scholarly journals Two domains of ISGF3 gamma that mediate protein-DNA and protein-protein interactions during transcription factor assembly contribute to DNA-binding specificity.

1993 ◽  
Vol 13 (1) ◽  
pp. 196-206 ◽  
Author(s):  
S A Veals ◽  
T Santa Maria ◽  
D E Levy
Keyword(s):  
Amino Acids ◽  
Transcription Factor ◽  
Dna Binding ◽  
Protein Interaction ◽  
Binding Specificity ◽  
Binding Domain ◽  
Dna Binding Domain ◽  
Response Element ◽  
Ifn Alpha ◽  
Dna Binding Specificity

Alpha interferon (IFN-alpha) induces the transcription of a large set of genes through activation of multimeric transcription factor ISGF3. This factor can be dissociated into two protein components, termed ISGF3 gamma and ISGF3 alpha. ISGF3 gamma is a 48-kDa protein related at the amino terminus to members of the IFN-regulatory factor (IRF) and Myb families of DNA-binding proteins; ISGF3 alpha consists of three polypeptides of 84, 91, and 113 kDa that self-assemble to form an activated component in response to IFN-alpha. DNA-binding studies indicated that ISGF3 gamma binds DNA alone, recognizing the IFN-stimulated response element, while the ISGF3 alpha polypeptides alone display no specific interactions with DNA. A complex between ISGF3 gamma and activated ISGF3 alpha binds the IFN-stimulated response element with much greater affinity than does the 48-kDa ISGF3 gamma protein alone. The DNA-binding domain of ISGF3 gamma and regions responsible for protein-protein interaction with ISGF3 alpha were identified by using deleted forms of ISGF3 gamma expressed in vitro. The amino-terminal region of ISGF3 gamma homologous to the IRF and Myb proteins was sufficient for interaction with DNA and displayed the binding specificity of the intact protein; phosphorylation of this region was necessary for activity. A second region of 160 amino acids separated from the DNA-binding domain by over 100 amino acids contained a domain capable of associating with ISGF3 alpha and was sufficient to confer specific ISGF3 alpha interaction to a heterologous protein. Interaction of the ISGF3 alpha component with the protein interaction domain of ISGF3 gamma altered the DNA-binding specificity of the resulting complex, suggesting that one or more of the ISGF3 alpha polypeptides make base-specific contacts with DNA. This interaction defines a mechanism through which IRF-like proteins complexed with regulatory components can display novel DNA-binding specificities.

Two domains of ISGF3 gamma that mediate protein-DNA and protein-protein interactions during transcription factor assembly contribute to DNA-binding specificity

1993 ◽  
Vol 13 (1) ◽  
pp. 196-206
Author(s):  
S A Veals ◽  
T Santa Maria ◽  
D E Levy
Keyword(s):  
Amino Acids ◽  
Transcription Factor ◽  
Dna Binding ◽  
Protein Interaction ◽  
Binding Specificity ◽  
Binding Domain ◽  
Dna Binding Domain ◽  
Response Element ◽  
Ifn Alpha ◽  
Dna Binding Specificity

Alpha interferon (IFN-alpha) induces the transcription of a large set of genes through activation of multimeric transcription factor ISGF3. This factor can be dissociated into two protein components, termed ISGF3 gamma and ISGF3 alpha. ISGF3 gamma is a 48-kDa protein related at the amino terminus to members of the IFN-regulatory factor (IRF) and Myb families of DNA-binding proteins; ISGF3 alpha consists of three polypeptides of 84, 91, and 113 kDa that self-assemble to form an activated component in response to IFN-alpha. DNA-binding studies indicated that ISGF3 gamma binds DNA alone, recognizing the IFN-stimulated response element, while the ISGF3 alpha polypeptides alone display no specific interactions with DNA. A complex between ISGF3 gamma and activated ISGF3 alpha binds the IFN-stimulated response element with much greater affinity than does the 48-kDa ISGF3 gamma protein alone. The DNA-binding domain of ISGF3 gamma and regions responsible for protein-protein interaction with ISGF3 alpha were identified by using deleted forms of ISGF3 gamma expressed in vitro. The amino-terminal region of ISGF3 gamma homologous to the IRF and Myb proteins was sufficient for interaction with DNA and displayed the binding specificity of the intact protein; phosphorylation of this region was necessary for activity. A second region of 160 amino acids separated from the DNA-binding domain by over 100 amino acids contained a domain capable of associating with ISGF3 alpha and was sufficient to confer specific ISGF3 alpha interaction to a heterologous protein. Interaction of the ISGF3 alpha component with the protein interaction domain of ISGF3 gamma altered the DNA-binding specificity of the resulting complex, suggesting that one or more of the ISGF3 alpha polypeptides make base-specific contacts with DNA. This interaction defines a mechanism through which IRF-like proteins complexed with regulatory components can display novel DNA-binding specificities.

scholarly journals The C6 zinc finger and adjacent amino acids determine DNA-binding specificity and affinity in the yeast activator proteins LAC9 and PPR1.

1990 ◽  
Vol 10 (10) ◽  
pp. 5128-5137 ◽  
Author(s):  
M M Witte ◽  
R C Dickson
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Dna Binding ◽  
Zinc Finger ◽  
Binding Specificity ◽  
Binding Domain ◽  
Dna Binding Domain ◽  
Transcription Activator ◽  
Activator Proteins ◽  
Dna Binding Specificity

LAC9 is a DNA-binding protein that regulates transcription of the lactose-galactose regulon in Kluyveromyces lactis. The DNA-binding domain is composed of a zinc finger and nearby amino acids (M. M. Witte and R. C. Dickson, Mol. Cell. Biol. 8:3726-3733, 1988). The single zinc finger appears to be structurally related to the zinc finger of many other fungal transcription activator proteins that contain positively charged residues and six conserved cysteines with the general form Cys-Xaa2-Cys-Xaa6-Cys-Xaa6-9-Cys-Xaa2-Cys-Xaa 6-Cys, where Xaan indicates a stretch of the indicated number of any amino acids (R. M. Evans and S. M. Hollenberg, Cell 52:1-3, 1988). The function(s) of the zinc finger and other amino acids in DNA-binding remains unclear. To determine which portion of the LAC9 DNA-binding domain mediates sequence recognition, we replaced the C6 zinc finger, amino acids adjacent to the carboxyl side of the zinc finger, or both with the analogous region from the Saccharomyces cerevisiae PPR1 or LEU3 protein. A chimeric LAC9 protein, LAC9(PPR1 34-61), carrying only the PPR1 zinc finger, retained the DNA-binding specificity of LAC9. However, LAC9(PPR1 34-75), carrying the PPR1 zinc finger and 14 amino acids on the carboxyl side of the zinc finger, gained the DNA-binding specificity of PPR1, indicating that these 14 amino acids are necessary for specific DNA binding. Our data show that C6 fingers can substitute for each other and allow DNA binding, but binding affinity is reduced. Thus, in a qualitative sense C6 fingers perform a similar function(s). However, the high-affinity binding required by natural C6 finger proteins demands a unique C6 finger with a specific amino acid sequence. This requirement may reflect conformational constraints, including interactions between the C6 finger and the carboxyl-adjacent amino acids; alternatively or in addition, it may indicate that unique, nonconserved amino acid residues in zinc fingers make sequence-specifying or stabilizing contacts with DNA.

The C6 zinc finger and adjacent amino acids determine DNA-binding specificity and affinity in the yeast activator proteins LAC9 and PPR1

1990 ◽  
Vol 10 (10) ◽  
pp. 5128-5137
Author(s):  
M M Witte ◽  
R C Dickson
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Dna Binding ◽  
Zinc Finger ◽  
Binding Specificity ◽  
Binding Domain ◽  
Dna Binding Domain ◽  
Transcription Activator ◽  
Activator Proteins ◽  
Dna Binding Specificity

LAC9 is a DNA-binding protein that regulates transcription of the lactose-galactose regulon in Kluyveromyces lactis. The DNA-binding domain is composed of a zinc finger and nearby amino acids (M. M. Witte and R. C. Dickson, Mol. Cell. Biol. 8:3726-3733, 1988). The single zinc finger appears to be structurally related to the zinc finger of many other fungal transcription activator proteins that contain positively charged residues and six conserved cysteines with the general form Cys-Xaa2-Cys-Xaa6-Cys-Xaa6-9-Cys-Xaa2-Cys-Xaa 6-Cys, where Xaan indicates a stretch of the indicated number of any amino acids (R. M. Evans and S. M. Hollenberg, Cell 52:1-3, 1988). The function(s) of the zinc finger and other amino acids in DNA-binding remains unclear. To determine which portion of the LAC9 DNA-binding domain mediates sequence recognition, we replaced the C6 zinc finger, amino acids adjacent to the carboxyl side of the zinc finger, or both with the analogous region from the Saccharomyces cerevisiae PPR1 or LEU3 protein. A chimeric LAC9 protein, LAC9(PPR1 34-61), carrying only the PPR1 zinc finger, retained the DNA-binding specificity of LAC9. However, LAC9(PPR1 34-75), carrying the PPR1 zinc finger and 14 amino acids on the carboxyl side of the zinc finger, gained the DNA-binding specificity of PPR1, indicating that these 14 amino acids are necessary for specific DNA binding. Our data show that C6 fingers can substitute for each other and allow DNA binding, but binding affinity is reduced. Thus, in a qualitative sense C6 fingers perform a similar function(s). However, the high-affinity binding required by natural C6 finger proteins demands a unique C6 finger with a specific amino acid sequence. This requirement may reflect conformational constraints, including interactions between the C6 finger and the carboxyl-adjacent amino acids; alternatively or in addition, it may indicate that unique, nonconserved amino acid residues in zinc fingers make sequence-specifying or stabilizing contacts with DNA.

scholarly journals Preferences in a trait decision determined by transcription factor variants

2018 ◽  
Vol 115 (34) ◽  
pp. E7997-E8006 ◽  
Author(s):  
Michael W. Dorrity ◽  
Josh T. Cuperus ◽  
Jolie A. Carlisle ◽  
Stanley Fields ◽  
Christine Queitsch
Keyword(s):  
Transcription Factor ◽  
Dna Binding ◽  
Fungal Pathogens ◽  
Binding Domain ◽  
Single Amino Acid ◽  
Environmental Cues ◽  
Dna Binding Domain ◽  
Independent Manner ◽  
Dna Binding Specificity ◽  
The Many

Few mechanisms are known that explain how transcription factors can adjust phenotypic outputs to accommodate differing environments. In Saccharomyces cerevisiae, the decision to mate or invade relies on environmental cues that converge on a shared transcription factor, Ste12. Specificity toward invasion occurs via Ste12 binding cooperatively with the cofactor Tec1. Here, we determine the range of phenotypic outputs (mating vs. invasion) of thousands of DNA-binding domain variants in Ste12 to understand how preference for invasion may arise. We find that single amino acid changes in the DNA-binding domain can shift the preference of yeast toward either mating or invasion. These mutations define two distinct regions of this domain, suggesting alternative modes of DNA binding for each trait. We characterize the DNA-binding specificity of wild-type Ste12 to identify a strong preference for spacing and orientation of both homodimeric and heterodimeric sites. Ste12 mutants that promote hyperinvasion in a Tec1-independent manner fail to bind cooperative sites with Tec1 and bind to unusual dimeric Ste12 sites composed of one near-perfect and one highly degenerate site. We propose a model in which Ste12 alone may have evolved to activate invasion genes, which could explain how preference for invasion arose in the many fungal pathogens that lack Tec1.

Functional characterisation of a WRKY transcription factor of wheat and its expression analysis during leaf rust pathogenesis

2014 ◽  
Vol 41 (12) ◽  
pp. 1295 ◽  
Author(s):  
Dhananjay Kumar ◽  
Anjali Kapoor ◽  
Dharmendra Singh ◽  
Lopamudra Satapathy ◽  
Ashwini Kumar Singh ◽  
...  
Keyword(s):  
Amino Acids ◽  
Transcription Factor ◽  
Dna Binding ◽  
Leaf Rust ◽  
Stress Responses ◽  
Rust Fungus ◽  
Wrky Transcription Factor ◽  
Binding Domain ◽  
Leaf Rust Resistance Gene ◽  
Dna Binding Domain

WRKY proteins are a large family of plant-specific transcription factors associated with regulation of biotic and abiotic stress responses, but how they respond to cereal rust pathogens has never been explored at the molecular level. Full-length cDNA of TaWRKY1B was obtained from a wheat cultivar HD2329 derivative containing leaf rust resistance gene Lr28 based on domain characteristics. The unique feature of this WRKY transcription factor gene was the close proximity of the DNA-binding domain and consensus DNA element W-Box within the open reading frame. Infection with a virulent race of leaf rust fungus resulted in 146-fold induction of the gene in resistant plants, but only 12-fold in the susceptible plants as compared with mock-inoculated controls. Docking models of 74 amino acids DNA-binding domain and 26 bp W-Box element showed that the WRKY domain, located on the β1 strand, only interacts with the W-Box at positions corresponding to W125, R126, K127 and Y128 amino acids. A truncated recombinant protein of 9.0 kD, encompassing the DNA-binding domain also showed binding specificity to the 32 bp W-Box element in electrophoretic mobility shift assays. The protein–DNA ensemble was also characterised using high-resolution atomic force microscopic imaging. The results contribute to an understanding of the molecular structure and function of a previously uncharacterised WRKY transcription factor in wheat that can be manipulated to improve biotic stress tolerance.

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 Dynamic evolution of the major transcription factor DNA binding domain, and protein-protein interaction families during the evolution of the avian lineage

2017 ◽  
Author(s):  
Allie M. Graham ◽  
Jason S. Presnell
Keyword(s):  
Transcription Factor ◽  
Transcription Factors ◽  
Dna Binding ◽  
Protein Interaction ◽  
Phenotypic Diversity ◽  
Domain Architecture ◽  
Binding Domain ◽  
Dna Binding Domain ◽  
Interaction Domain ◽  
Protein Protein Interaction

ABSTRACTTranscription factors are characterized by their domain architecture, including DNA binding and protein-protein interaction domain combinations, which regulate their binding specificity, as well as their ability to effect a change on gene expression of their downstream targets. Transcription factors are central to organismal development, thus they potentially are instrumental in producing phenotypic diversity. Transcription factor abundance was estimated via 49 major DNA binding domain families, as well as 34 protein-protein interaction domain families, in 48 bird genomes, which were then compared with 6 available reptile genomes, in an effort to assess the degree to which these domains are potentially connected to increased phenotypic diversity in the avian lineage. We hypothesized that there would be increased abundance in multiple transcription factor domain families, as well as domains associated with protein-protein interactions, that would correlate with the increased phenotypic diversity found in birds; instead, this data shows a general loss/contraction of major domain families, with the largest losses in domain families associated with multiple developmental (feather, body-plan, immune) and metabolic processes. Ultimately, the results of this analyses represent a general characterization of domain family composition in birds, thus the specific domain composition of TF families should be probed further, especially those with the largest reductions seen in this study.

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