Synergistic activation of transcription is mediated by the N-terminal domain of Drosophila fushi tarazu homeoprotein and can occur without DNA binding by the protein

Synergistic activation of transcription by Drosophila segmentation genes in tissue culture cells provides a model with which to study combinatorial regulation. We examined the synergistic activation of an engrailed-derived promoter by the pair-rule proteins paired (PRD) and fushi tarazu (FTZ). Synergistic activation by PRD requires regions of the homeodomain or adjacent sequences, and that by FTZ requires the first 171 residues. Surprisingly, deletion of the FTZ homeodomain does not reduce the capacity of the protein for synergistic activation, although this mutation abolishes any detectable DNA-binding activity. This finding suggests that FTZ can function through protein-protein interactions with PRD or other components of the homeoprotein transcription complex, adding a new layer of mechanisms that could underlie the functional specificities and combinatorial regulation of homeoproteins.

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DNA Damage Induced Hyperphosphorylation of Replication Protein A. 2. Characterization of DNA Binding Activity, Protein Interactions, and Activity in DNA Replication and Repair†

Biochemistry ◽

10.1021/bi048057b ◽

2005 ◽

Vol 44 (23) ◽

pp. 8438-8448 ◽

Cited By ~ 34

Author(s):

Steve M. Patrick ◽

Greg G. Oakley ◽

Kathleen Dixon ◽

John J. Turchi

Keyword(s):

Dna Damage ◽

Dna Replication ◽

Dna Binding ◽

Protein Interactions ◽

Protein A ◽

Replication Protein A ◽

Binding Activity ◽

Dna Binding Activity ◽

Dna Replication And Repair

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Transcriptional and DNA Binding Activity of the Foxp1/2/4 Family Is Modulated by Heterotypic and Homotypic Protein Interactions

Molecular and Cellular Biology ◽

10.1128/mcb.24.2.809-822.2004 ◽

2004 ◽

Vol 24 (2) ◽

pp. 809-822 ◽

Cited By ~ 191

Author(s):

Shanru Li ◽

Joel Weidenfeld ◽

Edward E. Morrisey

Keyword(s):

Dna Binding ◽

Protein Interactions ◽

Leucine Zipper ◽

Binding Activity ◽

Transcriptional Repressors ◽

Transcription Analysis ◽

Dna Binding Activity ◽

The Family ◽

Two Hybrid ◽

Repression Domain

ABSTRACT Foxp1, Foxp2, and Foxp4 are large multidomain transcriptional regulators belonging to the family of winged-helix DNA binding proteins known as the Fox family. Foxp1 and Foxp2 have been shown to act as transcriptional repressors, while regulatory activity of the recently identified Foxp4 has not been determined. Given the importance of this Fox gene subfamily in neural and lung development, we sought to elucidate the mechanisms by which Foxp1, Foxp2, and Foxp4 repress gene transcription. We show that like Foxp1 and Foxp2, Foxp4 represses transcription. Analysis of the N-terminal repression domain in Foxp1, Foxp2, and Foxp4 shows that this region contains two separate and distinct repression subdomains that are highly homologous termed subdomain 1 and subdomain 2. However, subdomain 2 is not functional in Foxp4. Screening for proteins that interact with subdomains 1 and 2 of Foxp2 using yeast two-hybrid analysis revealed that subdomain 2 binds to C-terminal binding protein 1, which can synergistically repress transcription with Foxp1 and Foxp2, but not Foxp4. Subdomain 1 contains a highly conserved leucine zipper similar to that found in N-myc and confers homo- and heterodimerization to the Foxp1/2/4 family members. These interactions are dependent on the conserved leucine zipper motif. Finally, we show that the integrity of this subdomain is essential for DNA binding, making Foxp1, Foxp2, and Foxp4 the first Fox proteins that require dimerization for DNA binding. These data reveal a complex regulatory mechanism underlying Foxp1, Foxp2, and Foxp4 activity, demonstrating that Foxp1, Foxp2, and Foxp4 are the first Fox proteins reported whose activity is regulated by homo- and heterodimerization.

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Dissection of Functional Modules of AT-HOOK MOTIF NUCLEAR LOCALIZED PROTEIN 4 in the Development of the Root Xylem

Frontiers in Plant Science ◽

10.3389/fpls.2021.632078 ◽

2021 ◽

Vol 12 ◽

Author(s):

Minji Seo ◽

Ji-Young Lee

Keyword(s):

Protein Interactions ◽

Binding Activity ◽

Fine Tuning ◽

Root Apical Meristem ◽

Xylem Differentiation ◽

Protein Protein Interactions ◽

Xylem Development ◽

Dna Binding Activity ◽

C Terminus ◽

Plant Hormone Signaling

Xylem development in the Arabidopsis root apical meristem requires a complex cross talk between plant hormone signaling and transcriptional factors (TFs). The key processes involve fine-tuning between neighboring cells, mediated via the intercellular movement of signaling molecules. As an example, we previously reported that AT-HOOK MOTIF NUCLEAR LOCALIZED PROTEIN (AHL) 4 (AHL4), a member of the 29 AT-hook family TFs in Arabidopsis, moves into xylem precursors from their neighbors to determine xylem differentiation. As part of the effort to understand the molecular functions of AHL4, we performed domain swapping analyses using AHL1 as a counterpart, finding that AHL4 has three functionally distinctive protein modules. The plant and prokaryotes conserved (PPC) domain of AHL4 acts as a mediator of protein–protein interactions with AHL members. The N-terminus of AHL4 is required for the regulation of xylem development likely via its unique DNA-binding activity. The C-terminus of AHL4 confers intercellular mobility. Our characterization of modules in the AHL4 protein will augment our understanding of the complexity of regulation and the evolution of intercellular mobility in AHL4 and its relatives.

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Efficient expression and purification of human replication fork-stabilizing factor, Claspin, from mammalian cells: DNA-binding activity and novel protein interactions

Genes to Cells ◽

10.1111/j.1365-2443.2011.01535.x ◽

2011 ◽

Vol 16 (8) ◽

pp. 842-856 ◽

Cited By ~ 27

Author(s):

Syuzi Uno ◽

Hisao Masai

Keyword(s):

Dna Binding ◽

Protein Interactions ◽

Mammalian Cells ◽

Replication Fork ◽

Binding Activity ◽

Expression And Purification ◽

Dna Binding Activity ◽

Efficient Expression ◽

Novel Protein

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Modulation of Hsf1 activity by novobiocin and geldanamycin

Biochemistry and Cell Biology ◽

10.1139/o09-049 ◽

2009 ◽

Vol 87 (6) ◽

pp. 845-851 ◽

Cited By ~ 53

Author(s):

Renaud Conde ◽

Zachery R. Belak ◽

Manoj Nair ◽

Ruth F. O’Carroll ◽

Nick Ovsenek

Keyword(s):

Heat Shock ◽

Dna Binding ◽

Molecular Mechanisms ◽

Binding Pocket ◽

Binding Activity ◽

Chaperone Activity ◽

Atp Binding ◽

Dna Binding Activity ◽

Culture Cells ◽

Dose Dependent Decrease

Since Hsp90 is a known modulator of HSF1 activity, we examined the effects of two pharmacological inhibitors of Hsp90, novobiocin and geldanamycin, on HSF1 DNA-binding activity in the Xenopus oocyte model system. Novobiocin exhibits antiproliferative activity in culture cells and interacts with a C-terminal ATP-binding pocket on Hsp90, inhibiting Hsp90 autophosphorylation. Treatment of oocytes with novobiocin followed by heat shock results in a dose-dependent decrease in HSF1 DNA-binding and transcriptional activity. Immunoprecipitation experiments demonstrate novobiocin does not alter HSF1 activity through dissociation of Hsp90 from either monomeric or trimerized HSF1, suggesting that the effect of novobiocin on HSF1 is mediated through alterations in Hsp90 autophosphorylation. Geldanamycin binds the N-terminal ATPase site of Hsp90 and inhibits chaperone activity. Geldanamycin treatment of oocytes resulted in a dose-dependant increase in stability of active HSF1 trimers during submaximal heat shock and a delay in disassembly of trimers during recovery. The results suggest that Hsp90 chaperone activity is required for disassembly of HSF1 trimers. The data obtained with novobiocin suggests the C-terminal ATP-binding activity of Hsp90 is required for the initial steps of HSF1 trimerization, whereas the effects of geldanamycin suggest N-terminal ATPase and chaperone activities are required for disassembly of activated trimers. These data provide important insight into the molecular mechanisms by which pharmacological inhibitors of Hsp90 affect the heat shock response.

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A Capped Tudor Domain within a Core Subunit of the Sin3L/Rpd3L Histone Deacetylase Complex Binds Nucleic Acids

10.1101/2021.08.09.455673 ◽

2021 ◽

Author(s):

Ryan Dale Marcum ◽

Joseph Hsieh ◽

Maksim Giljen ◽

Yongbo Zhang ◽

Ishwar Radhakrishnan

Keyword(s):

Nucleic Acids ◽

Histone Deacetylase ◽

Protein Interactions ◽

Binding Activity ◽

Protein Protein Interactions ◽

Dna Binding Activity ◽

Tudor Domain ◽

Unknown Structure ◽

Sites Of Action ◽

And Function

Chromatin-modifying complexes containing histone deacetylase (HDAC) activities play critical roles in the regulation of gene transcription in eukaryotes. These complexes are thought to lack intrinsic DNA-binding activity, but according to a well-established paradigm, they are recruited via protein-protein interactions by gene-specific transcription factors and post-translational histone modifications to their sites of action on the genome. The mammalian Sin3L/Rpd3L complex, comprising more than a dozen different polypeptides, is an ancient HDAC complex found in diverse eukaryotes. The subunits of this complex harbor conserved domains and motifs of unknown structure and function. Here we show that Sds3, a constitutively associated subunit critical for the proper functioning of the complex, harbors a type of Tudor domain that we designate the capped Tudor domain (CTD). Unlike canonical Tudor domains that bind modified histones, the Sds3 CTD binds to nucleic acids that can form higher-order structures such as G-quadruplexes, and shares similarities with the knotted Tudor domain of the Esa1 histone acetyltransferase (HAT) that was previously shown to bind single-stranded RNA. Our findings expand the range of macromolecules capable of recruiting the Sin3L/Rpd3L complex and draws attention to potentially new roles for this HDAC complex in transcription biology.

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Specific residues in the Pbx homeodomain differentially modulate the DNA-binding activity of Hox and Engrailed proteins

Development ◽

10.1242/dev.124.5.1089 ◽

1997 ◽

Vol 124 (5) ◽

pp. 1089-1098 ◽

Cited By ~ 1

Author(s):

L.T. Peltenburg ◽

C. Murre

Keyword(s):

Dna Binding ◽

Cell Fate ◽

Protein Interactions ◽

Control Cell ◽

Binding Activity ◽

Homeodomain Protein ◽

Gene Products ◽

Homeodomain Proteins ◽

Dna Binding Activity ◽

Multicellular Organisms

Two classes of homeodomain proteins, Hox and Engrailed, have been shown to act in concert with the atypical homeodomain proteins Pbx and extradenticle. We now show that specific residues located within the Pbx homeodomain are essential for cooperative DNA binding with Hox and Engrailed gene products. Within the N-terminal region of the Pbx homeodomain, we have identified a residue that is required for cooperative DNA binding with three Hox gene products but not for cooperativity with Engrailed-2 (En-2). Furthermore, there are similarities between heterodimeric interactions involving the yeast mating type proteins MATa1 and MATalpha2 and those that allow the formation of Pbx/Hox and Pbx/En-2 heterodimers. Specifically, residues located in the a1 homeodomain that were previously shown to form a hydrophobic pocket allowing the alpha2 C-terminal tail to bind, are also required for Pbx/Hox and Pbx/En-2 cooperativity. Furthermore, we show that three residues located in the turn between helix 1 and helix 2, characteristic of many atypical homeodomain proteins, are required for cooperative DNA binding involving both Hox and En-2. Replacement of the three residues located in the turn between helix 1 and helix 2 of the Pbx homeodomain with those of the atypical homeodomain proteins controlling cell fate in the basidiomycete Ustilago maydis, bE5 and bE6, allows cooperative DNA binding with three Hox members but abolishes interactions with En-2. The data suggest that the molecular mechanism of homeodomain protein interactions that control cell fate in Saccharomyces cerevisiae and in the basidiomycetes may well be conserved in part in multicellular organisms.

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