The Experts below are selected from a list of 294 Experts worldwide ranked by ideXlab platform
Benno Mullerhill - One of the best experts on this subject based on the ideXlab platform.
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genetic studies of the lac Repressor xv 4000 single amino acid substitutions and analysis of the resulting phenotypes on the basis of the protein structure
Journal of Molecular Biology, 1996Co-Authors: Jorg Suckow, Jeffrey H. Miller, Peter Markiewicz, Lynn G Kleina, Brigitte Kisterswoike, Benno MullerhillAbstract:Abstract Each amino acid from position 2 to 329 of Lac Repressor was replaced by 12 or 13 of the 20 natural occurring amino acids. The resulting phenotypes are discussed on the basis of (1) the recently published structure of the Lac Repressor core complexed with the inducer IPTG and (2) a model of the dimeric Lac Repressor built by homology modelling from the X-ray structure of the purine Repressor-coRepressor-operator complex. This phenotype analysis, based on 4000 well-defined mutants, yields a functional description of each amino acid position of Lac Repressor. In most cases, mutant effects can be directly correlated with the structure and function of the protein. This connection between the amino acid position and the structure and function of the protein is in most cases direct and not complicated: amino acids which are directly involved in sugar binding are affected in Lac Repressor mutants of the I S type; small amino acids which can only be replaced by other small acids are located in the core of the protein; positions at which nearly all amino acids are tolerated are in most cases located on the surface of the protein. Amino acids which are highly conserved throughout the LacI family of Repressors, and not directly involved in specific functions of the protein like DNA recognition or sugar binding, form a network of contacts with other amino acids. Such amino acids are either located inside one subunit, mostly at the interface between secondary structure elements, or are involved in the dimerisation interface.
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the side chain of the amino acid residue in position 110 of the lac Repressor influences its allosteric equilibrium
Journal of Molecular Biology, 1996Co-Authors: Herbert Mullerhartmann, Benno MullerhillAbstract:Abstract Binding of the Lac Repressor to its operator DNA controls the expression of the genes of the lac operon of Escherichia coli . Lac Repressor's affinity for the lac operator is diminished by an inducer that affects the structure of the Repressor tetramer. Here we report the cloning and sequencing of the mutant Lac Repressor i t gene, whose product, the LacR t Repressor, shows a higher affinity for the inducer isopropyl-β- D -thiogalactopyranoside (IPTG) and a lower affinity for the lac operator than the wild-type Repressor. We show that the altered phenotype is due to a single amino acid residue replacement; the alanine residue at position 110 in the wild-type is replaced by threonine in i t . Other amino acid residues in position 110 have been shown to result in an i S phenotype. For the i S -substitution of alanine 110 with lysine we demonstrate an increase in the affinity for operator DNA and a decrease in the affinity for IPTG. Thus, A110→K shows the opposite effect to A110→T on the Repressor protein. We explain the phenotype of the LacR mutants by displacements of the conformational equilibrium for the dimeric Repressor unit between RR (high operator affinity, low inducer affinity) and R*R* (low operator affinity, high inducer affinity) towards R*R* in the i t and towards RR in the i S mutant in position 110 with respect to the wild-type. The putative structures of the wild-type and mutant Lac Repressors confirm this conclusion.
Oscar R Colamonici - One of the best experts on this subject based on the ideXlab platform.
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deletion of the p107 p130 binding domain of mip130 lin 9 bypasses the requirement for cdk4 activity for the dissociation of mip130 lin 9 from p107 p130 e2f4 complex
Experimental Cell Research, 2009Co-Authors: Raudel Sandoval, Mark Pilkinton, Oscar R ColamoniciAbstract:Abstract Mip130/LIN-9 is part of a large complex that includes homologs of the Drosophila dREAM (drosophila RB-like, E2F, and Myb) and C. elegans DRM complexes. This complex also includes proteins such as Mip40/LIN-37, Mip120/LIN-54, and LIN-52. In mammalian cells, Mip130/LIN-9 specifically associates with the p107/p130-E2F4 Repressor complex in G0/G1 and with B-Myb in S-phase. However, little is known about how the transition occurs and whether Mip130/LIN-9 contributes to the Repressor effect of p107/p130. In this report, we demonstrate that Mip130/LIN-9, Mip40/LIN-37, Mip120/LIN-54, and Sin3b form a core complex, the Mip Core Complex or LIN Complex (MCC/LINC), which is detectable in all phases of the cell cycle. This complex specifically recruits transcriptional Repressors such as p107, p130, E2F4 and HDAC1 in G0/G1, and B-Myb in S-phase. Importantly, we provide strong evidence that the transition between Repressors and activators of transcription is mediated by CDK4, through the phosphorylation of the pocket proteins, p107 and p130. The requirement for CDK4 activity is bypassed by the deletion of the first 84 amino acids (Mip130/LIN-9Δ84), since this mutant is unable to interact with p107/p130 in G0/G1, while maintaining its association with B-Myb. Importantly, the Mip130/LIN-9Δ84 allele rescues the low expression of G1/S genes observed in CDK4−/− MEFs demonstrating that Mip130/LIN-9 contributes to the repression of these E2F-regulated genes in G0/G1.
Masaru Ohmetakagi - One of the best experts on this subject based on the ideXlab platform.
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arabidopsis wuschel is a bifunctional transcription factor that acts as a Repressor in stem cell regulation and as an activator in floral patterning
The Plant Cell, 2009Co-Authors: Miho Ikeda, Nobutaka Mitsuda, Masaru OhmetakagiAbstract:Most transcription factors act either as activators or Repressors, and no such factors with dual function have been unequivocally identified and characterized in plants. We demonstrate here that the Arabidopsis thaliana protein WUSCHEL (WUS), which regulates the maintenance of stem cell populations in shoot meristems, is a bifunctional transcription factor that acts mainly as a Repressor but becomes an activator when involved in the regulation of the AGAMOUS (AG) gene. We show that the WUS box, which is conserved among WOX genes, is the domain that is essential for all the activities of WUS, namely, for regulation of stem cell identity and size of floral meristem. All the known activities of WUS were eliminated by mutation of the WUS box, including the ability of WUS to induce the expression of AG. The mutation of the WUS box was complemented by fusion of an exogenous repression domain, with resultant induction of somatic embryogenesis in roots and expansion of floral meristems as observed upon ectopic expression of WUS. By contrast, fusion of an exogenous activation domain did not result in expanded floral meristems but induced flowers similar to those induced by the ectopic expression of AG. Our results demonstrate that WUS acts mainly as a Repressor and that its function changes from that of a Repressor to that of an activator in the case of regulation of the expression of AG.
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dominant repression of target genes by chimeric Repressors that include the ear motif a repression domain in arabidopsis
Plant Journal, 2003Co-Authors: Keiichiro Hiratsu, Kyoko Matsui, Tomotsugu Koyama, Masaru OhmetakagiAbstract:The redundancy of genes for plant transcription factors often interferes with efforts to identify the biologic functions of such factors. We show here that four different transcription factors fused to the EAR motif, a repression domain of only 12 amino acids, act as dominant Repressors in transgenic Arabidopsis and suppress the expression of specific target genes, even in the presence of the redundant transcription factors, with resultant dominant loss-of-function phenotypes. Chimeric EIN3, CUC1, PAP1, and AtMYB23 Repressors that included the EAR motif dominantly suppressed the expression of their target genes and caused insensitivity to ethylene, cup-shaped cotyledons, reduction in the accumulation of anthocyanin, and absence of trichomes, respectively. This chimeric Repressor silencing technology (CRES-T), exploiting the EAR-motif repression domain, is simple and effective and can overcome genetic redundancy. Thus, it should be useful not only for the rapid analysis of the functions of redundant plant transcription factors but also for the manipulation of plant traits via the suppression of gene expression that is regulated by specific transcription factors.
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repression domains of class ii erf transcriptional Repressors share an essential motif for active repression
The Plant Cell, 2001Co-Authors: Masaru Ohta, Kyoko Matsui, Keiichiro Hiratsu, Hideaki Shinshi, Masaru OhmetakagiAbstract:We reported previously that three ERF transcription factors, tobacco ERF3 (NtERF3) and Arabidopsis AtERF3 and AtERF4, which are categorized as class II ERFs, are active Repressors of transcription. To clarify the roles of these Repressors in transcriptional regulation in plants, we attempted to identify the functional domains of the ERF Repressor that mediates the repression of transcription. Analysis of the results of a series of deletions revealed that the C-terminal 35 amino acids of NtERF3 are sufficient to confer the capacity for repression of transcription on a heterologous DNA binding domain. This repression domain suppressed the intermolecular activities of other transcriptional activators. In addition, fusion of this repression domain to the VP16 activation domain completely inhibited the transactivation function of VP16. Comparison of amino acid sequences of class II ERF Repressors revealed the conservation of the sequence motif L / F DLN L / F (x)P. This motif was essential for repression because mutations within the motif eliminated the capacity for repression. We designated this motif the ERF-associated amphiphilic repression (EAR) motif, and we identified this motif in a number of zinc-finger proteins from wheat, Arabidopsis, and petunia plants. These zinc finger proteins functioned as Repressors, and their repression domains were identified as regions that contained an EAR motif.
Raudel Sandoval - One of the best experts on this subject based on the ideXlab platform.
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deletion of the p107 p130 binding domain of mip130 lin 9 bypasses the requirement for cdk4 activity for the dissociation of mip130 lin 9 from p107 p130 e2f4 complex
Experimental Cell Research, 2009Co-Authors: Raudel Sandoval, Mark Pilkinton, Oscar R ColamoniciAbstract:Abstract Mip130/LIN-9 is part of a large complex that includes homologs of the Drosophila dREAM (drosophila RB-like, E2F, and Myb) and C. elegans DRM complexes. This complex also includes proteins such as Mip40/LIN-37, Mip120/LIN-54, and LIN-52. In mammalian cells, Mip130/LIN-9 specifically associates with the p107/p130-E2F4 Repressor complex in G0/G1 and with B-Myb in S-phase. However, little is known about how the transition occurs and whether Mip130/LIN-9 contributes to the Repressor effect of p107/p130. In this report, we demonstrate that Mip130/LIN-9, Mip40/LIN-37, Mip120/LIN-54, and Sin3b form a core complex, the Mip Core Complex or LIN Complex (MCC/LINC), which is detectable in all phases of the cell cycle. This complex specifically recruits transcriptional Repressors such as p107, p130, E2F4 and HDAC1 in G0/G1, and B-Myb in S-phase. Importantly, we provide strong evidence that the transition between Repressors and activators of transcription is mediated by CDK4, through the phosphorylation of the pocket proteins, p107 and p130. The requirement for CDK4 activity is bypassed by the deletion of the first 84 amino acids (Mip130/LIN-9Δ84), since this mutant is unable to interact with p107/p130 in G0/G1, while maintaining its association with B-Myb. Importantly, the Mip130/LIN-9Δ84 allele rescues the low expression of G1/S genes observed in CDK4−/− MEFs demonstrating that Mip130/LIN-9 contributes to the repression of these E2F-regulated genes in G0/G1.
David P Giedroc - One of the best experts on this subject based on the ideXlab platform.
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tuning site specific dynamics to drive allosteric activation in a pneumococcal zinc uptake regulator
eLife, 2018Co-Authors: Daiana A Capdevila, Fidel Huerta, Katherine A Edmonds, David P GiedrocAbstract:MarR (multiple antibiotic resistance Repressor) family proteins are bacterial Repressors that regulate transcription in response to a wide range of chemical signals. Although specific features of MarR family function have been described, the role of atomic motions in MarRs remains unexplored thus limiting insights into the evolution of allostery in this ubiquitous family of Repressors. Here, we provide the first experimental evidence that internal dynamics play a crucial functional role in MarR proteins. Streptococcus pneumoniae AdcR (adhesin-competence Repressor) regulates ZnII homeostasis and ZnII functions as an allosteric activator of DNA binding. ZnII coordination triggers a transition from somewhat independent domains to a more compact structure. We identify residues that impact allosteric activation on the basis of ZnII-induced perturbations of atomic motions over a wide range of timescales. These findings appear to reconcile the distinct allosteric mechanisms proposed for other MarRs and highlight the importance of conformational dynamics in biological regulation.
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tuning site specific dynamics to drive allosteric activation in a pneumococcal zinc uptake regulator
bioRxiv, 2018Co-Authors: Daiana A Capdevila, Fidel Huerta, Katherine A Edmonds, David P GiedrocAbstract:MarR (multiple antibiotic resistance Repressor) family proteins are bacterial Repressors that regulate transcription in response to a wide range of chemical signals. Although specific features of MarR family function have been described, the role of atomic motions in MarRs remains unexplored thus limiting insights into the evolution of allostery in this ubiquitous family of Repressors. Here, we provide the first experimental evidence that internal dynamics play a crucial functional role in MarR proteins. Streptococcus pneumoniae AdcR (adhesin-competence Repressor) regulates Zn II homeostasis and Zn II functions as an allosteric activator of DNA binding. Zn II coordination triggers a transition from independent domains to a more compact structure. We identify residues that impact allosteric activation on the basis of Zn II -induced perturbations of atomic motions over a wide range of timescales. These findings reconcile the distinct allosteric mechanisms proposed for other MarRs and highlight the importance of conformational dynamics in biological regulation.
