scholarly journals Crystal Structure of the DNA-Binding Domain of Human Herpesvirus 6A Immediate Early Protein 2

2017 ◽  
Vol 91 (21) ◽  
Author(s):  
Mitsuhiro Nishimura ◽  
Junjie Wang ◽  
Aika Wakata ◽  
Kento Sakamoto ◽  
Yasuko Mori
Keyword(s):  
Gene Expression ◽  
Dna Binding ◽  
Binding Site ◽  
Human Herpesvirus ◽  
Binding Domain ◽  
Immediate Early ◽  
Dna Binding Domain ◽  
Factor Binding ◽  
Early Protein ◽  
Dna Binding Site

ABSTRACT Immediate early proteins of human herpesvirus 6A (HHV-6A) are expressed at the outset of lytic infection and thereby regulate viral gene expression. Immediate early protein 2 (IE2) of HHV-6A is a transactivator that drives a variety of promoters. The C-terminal region of HHV-6A IE2 is shared among IE2 homologs in betaherpesviruses and is involved in dimerization, DNA binding, and transcription factor binding. In this study, the structure of the IE2 C-terminal domain (IE2-CTD) was determined by X-ray crystallography at a resolution of 2.5 Å. IE2-CTD forms a homodimer stabilized by a β-barrel core with two interchanging long loops. Unexpectedly, the core structure resembles those of the gammaherpesvirus factors EBNA1 of Epstein-Barr virus and LANA of Kaposi sarcoma-associated herpesvirus, but the interchanging loops are longer in IE2-CTD and form helix-turn-helix (HTH)-like motifs at their tips. The HTH and surrounding α-helices form a structural feature specific to the IE2 group. The apparent DNA-binding site (based on structural similarity with EBNA1 and LANA) resides on the opposite side of the HTH-like motifs, surrounded by positive electrostatic potential. Mapping analysis of conserved residues on the three-dimensional structure delineated a potential factor-binding site adjacent to the expected DNA-binding site. The predicted bi- or tripartite functional sites indicate a role for IE2-CTD as an adapter connecting the promoter and transcriptional factors that drive gene expression. IMPORTANCE Human herpesvirus 6A (HHV-6A) and HHV-6B belong to betaherpesvirus subfamily. Both viruses establish lifelong latency after primary infection, and their reactivation poses a significant risk to immunocompromised patients. Immediate early protein 2 (IE2) of HHV-6A and HHV-6B is a transactivator that triggers viral replication and contains a DNA-binding domain shared with other betaherpesviruses such as human herpesvirus 7 and human cytomegalovirus. In this study, an atomic structure of the DNA-binding domain of HHV-6A IE2 was determined and analyzed, enabling a structure-based understanding of the functions of IE2, specifically DNA recognition and interaction with transcription factors. Unexpectedly, the dimeric core resembles the DNA-binding domain of transcription regulators from gammaherpesviruses, showing structural conservation as a DNA-binding domain but with its own unique structural features. These findings facilitate further characterization of this key viral transactivator.

Crystallization and Preliminary X-ray Analysis of the DNA Binding Domain of the Hin Recombinase with Its DNA Binding Site

1993 ◽  
Vol 232 (3) ◽  
pp. 982-986
Author(s):  
Jin-An Feng ◽  
Melvin Simon ◽  
David P. Mack ◽  
Peter B. Dervan ◽  
Reid C. Johnson ◽  
...  
Keyword(s):  
Dna Binding ◽  
Binding Site ◽  
Binding Domain ◽  
Dna Binding Domain ◽  
X Ray ◽  
Dna Binding Site ◽  
Hin Recombinase

scholarly journals Hesx1 homeodomain protein represses transcription as a monomer and antagonises transactivation of specific sites as a homodimer

2002 ◽  
Vol 28 (3) ◽  
pp. 193-205 ◽  
Author(s):  
J Quirk ◽  
P Brown
Keyword(s):  
Gene Expression ◽  
Dna Binding ◽  
Binding Site ◽  
Anterior Pituitary ◽  
Homeodomain Protein ◽  
Dependent Manner ◽  
Gene Promoters ◽  
Differentiation Markers ◽  
Proximal Half ◽  
Dna Binding Site

The homeobox repressor Hesx1, expressed throughout Rathke's pouch and required for normal pituitary development, has been implicated in anterior pituitary pathogenesis in man. Prolonged expression of Hesx1 delays the appearance of anterior pituitary terminal differentiation markers in mice, particularly the gonadotroph hormones. We tested if Hesx1 could modulate gonadotrophin gene expression directly, and found that Hesx1 repressed both common alpha subunit (alpha GSU) and luteinising hormone beta-subunit (LH beta) gene promoters. Repression mapped to the Pitx1 homeodomain protein transactivation site in the proximal alpha GSU promoter, but did not map to the equivalent site on LH beta. Hesx1 repression of the alpha GSU Pitx1 site was overridden by co-transfection of Pitx1. In contrast, Hesx1 antagonised Pitx1 transactivation of LH beta in a dose-dependent manner. This was due to monomeric binding of Hesx1 on alpha GSU and homodimerisation on LH beta. The homodimerisation site comprises the Pitx1 DNA binding site and a proximal binding site, and mutation of either inhibited homodimer formation. Conversion of the LH beta Pitx1 DNA binding site to an alpha GSU-type did not promote homodimer formation, arguing that Hesx1 has pronounced site selectivity. Furthermore, mutation of the proximal half of the homodimerisation site blocked Hesx1 antagonisation of Pitx1 transactivation. We conclude that Hesx1 monomers repress gene expression, and homodimers block specific transactivation sites.

Steroid receptor-mediated inhibition of rat prolactin gene expression does not require the receptor DNA-binding domain

Cell ◽  
1988 ◽  
Vol 52 (5) ◽  
pp. 685-695 ◽  
Author(s):  
Stuart Adler ◽  
Marian L. Waterman ◽  
Xi He ◽  
Michael G. Rosenfeld
Keyword(s):  
Gene Expression ◽  
Dna Binding ◽  
Steroid Receptor ◽  
Binding Domain ◽  
Dna Binding Domain ◽  
Prolactin Gene

scholarly journals Cell Cycle Localization, Dimerization, and Binding Domain Architecture of the Telomere Protein cPot1

2004 ◽  
Vol 24 (5) ◽  
pp. 2091-2102 ◽  
Author(s):  
Chao Wei ◽  
Carolyn M. Price
Keyword(s):  
Cell Cycle ◽  
Dna Binding ◽  
Binding Site ◽  
Domain Architecture ◽  
Binding Domain ◽  
Binding Motif ◽  
Dna Binding Domain ◽  
Telomere Protein ◽  
Ob Fold

ABSTRACT Pot1 is a single-stranded-DNA-binding protein that recognizes telomeric G-strand DNA. It is essential for telomere capping in Saccharomyces pombe and regulates telomere length in humans. Human Pot1 also interacts with proteins that bind the duplex region of the telomeric tract. Thus, like Cdc13 from S. cerevisiae, Pot 1 may have multiple roles at the telomere. We show here that endogenous chicken Pot1 (cPot1) is present at telomeres during periods of the cell cycle when t loops are thought to be present. Since cPot1 can bind internal loops and directly adjacent DNA-binding sites, it is likely to fully coat and protect both G-strand overhangs and the displaced G strand of a t loop. The minimum binding site of cPot1 is double that of the S. pombe DNA-binding domain. Although cPot can self associate, dimerization is not required for DNA binding and hence does not explain the binding-site duplication. Instead, the DNA-binding domain appears to be extended to contain a second binding motif in addition to the conserved oligonucleotide-oligosaccharide (OB) fold present in other G-strand-binding proteins. This second motif could be another OB fold. Although dimerization is inefficient in vitro, it may be regulated in vivo and could promote association with other telomere proteins and/or telomere compaction.

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 Dissection of a carboxy-terminal region of the yeast regulatory protein RAP1 with effects on both transcriptional activation and silencing

1992 ◽  
Vol 12 (3) ◽  
pp. 1209-1217
Author(s):  
C F Hardy ◽  
D Balderes ◽  
D Shore
Keyword(s):  
Dna Binding ◽  
Binding Site ◽  
Transcriptional Activation ◽  
Binding Sites ◽  
Binding Protein ◽  
Binding Domain ◽  
Dominant Negative Effect ◽  
Dna Binding Domain ◽  
Wild Type ◽  
Carboxy Terminal

RAP1 is an essential sequence-specific DNA-binding protein in Saccharomyces cerevisiae whose binding sites are found in a large number of promoters, where they function as upstream activation sites, and at the silencer elements of the HMR and HML mating-type loci, where they are important for repression. We have examined the involvement of specific regions of the RAP1 protein in both repression and activation of transcription by studying the properties of a series of hybrid proteins containing RAP1 sequences fused to the DNA-binding domain of the yeast protein GAL4 (amino acids 1 to 147). GAL4 DNA-binding domain/RAP1 hybrids containing only the carboxy-terminal third of the RAP1 protein (which lacks the RAP1 DNA-binding domain) function as transcriptional activators of a reporter gene containing upstream GAL4 binding sites. Expression of some hybrids from the strong ADH1 promoter on multicopy plasmids has a dominant negative effect on silencers, leading to either partial or complete derepression of normally silenced genes. The GAL4/RAP1 hybrids have different effects on wild-type and several mutated but functional silencers. Silencers lacking either an autonomously replicating sequence consensus element or the RAP1 binding site are strongly derepressed, whereas the wild-type silencer or a silencer containing a deletion of the binding site for another silencer-binding protein, ABF1, are only weakly affected by hybrid expression. By examining a series of GAL4 DNA-binding domain/RAP1 hybrids, we have mapped the transcriptional activation and derepression functions to specific parts of the RAP1 carboxy terminus.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals AguR Is Required for Induction of the Streptococcus mutans Agmatine Deiminase System by Low pH and Agmatine

2009 ◽  
Vol 75 (9) ◽  
pp. 2629-2637 ◽  
Author(s):  
Yaling Liu ◽  
Lin Zeng ◽  
Robert A. Burne
Keyword(s):  
Gene Expression ◽  
Dna Binding ◽  
Streptococcus Mutans ◽  
Low Ph ◽  
Binding Domain ◽  
Negative Regulator ◽  
Dna Binding Domain ◽  
Mutant Proteins ◽  
Acidic Conditions ◽  
Agmatine Deiminase

ABSTRACT Acidic conditions and the presence of exogenous agmatine are required to achieve maximal expression of the agmatine deiminase system (AgDS) of Streptococcus mutans. Here we demonstrate that the transcriptional activator of the AgDS, AguR, is required for the responses to agmatine and to low pH. Linker scanning mutagenesis was used to create a panel of mutated aguR genes that were utilized to complement an aguR deletion mutant of S. mutans. The level of production of the mutant proteins was shown to be comparable to that of the wild-type AguR protein. Mutations in the predicted DNA binding domain of AguR eliminated activation of the agu operon. Insertions into the region connecting the DNA binding domain to the predicted extracellular and transmembrane domains were well tolerated. In contrast, a variety of mutants were isolated that had a diminished capacity to respond to low pH but retained the ability to activate AgDS gene expression in response to agmatine, and vice versa. Also, a number of mutants were unable to respond to either agmatine or low pH. AguD, which is a predicted agmatine-putrescine antiporter, was found to be a negative regulator of AgDS gene expression in the absence of exogenous agmatine but was not required for low-pH induction of the AgDS genes. This study reveals that the control of AgDS gene expression by both agmatine and low pH is coordinated through the AguR protein and begins to identify domains of the protein involved in sensing and signaling.

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