scholarly journals Multivalent and Bidirectional Binding of Transcriptional Transactivation Domains to the MED25 Coactivator

Biomolecules ◽  
2020 ◽  
Vol 10 (9) ◽  
pp. 1205
Author(s):  
Heather M. Jeffery ◽  
Robert O. J. Weinzierl
Keyword(s):  
Transcriptional Activation ◽  
Site Directed Mutagenesis ◽  
Viral Gene ◽  
Interaction Patterns ◽  
Hydrophobic Residues ◽  
Activation Domains ◽  
Intrinsically Disordered ◽  
Transactivation Domains ◽  
Transcriptional Activation Domains ◽  
Dynamics Simulations

The human mediator subunit MED25 acts as a coactivator that binds the transcriptional activation domains (TADs) present in various cellular and viral gene-specific transcription factors. Previous studies, including on NMR measurements and site-directed mutagenesis, have only yielded low-resolution models that are difficult to refine further by experimental means. Here, we apply computational molecular dynamics simulations to study the interactions of two different TADs from the human transcription factor ETV5 (ERM) and herpes virus VP16-H1 with MED25. Like other well-studied coactivator-TAD complexes, the interactions of these intrinsically disordered domains with the coactivator surface are temporary and highly dynamic (‘fuzzy’). Due to the fact that the MED25 TAD-binding region is organized as an elongated cleft, we specifically asked whether these TADs are capable of binding in either orientation and how this could be achieved structurally and energetically. The binding of both the ETV5 and VP16-TADs in either orientation appears to be possible but occurs in a conformationally distinct manner and utilizes different sets of hydrophobic residues present in the TADs to drive the interactions. We propose that MED25 and at least a subset of human TADs specifically evolved a redundant set of molecular interaction patterns to allow binding to particular coactivators without major prior spatial constraints.

scholarly journals The regulatory domain of human heat shock factor 1 is sufficient to sense heat stress.

1996 ◽  
Vol 16 (3) ◽  
pp. 839-846 ◽  
Author(s):  
E M Newton ◽  
U Knauf ◽  
M Green ◽  
R E Kingston
Keyword(s):  
Amino Acids ◽  
Heat Shock ◽  
Transcriptional Activation ◽  
Heat Shock Factor ◽  
Acid Activation ◽  
Regulatory Domain ◽  
Activation Domain ◽  
Control Temperature ◽  
Activation Domains ◽  
Transcriptional Activation Domains

Heat shock factor (HSF) activates transcription in response to cellular stress. Human HSF1 has a central regulatory domain which can repress the activity of its activation domains at the control temperature and render them heat shock inducible. To determine whether the regulatory domain works in tandem with specific features of the HSF1 transcriptional activation domains, we first used deletion and point mutagenesis to define these activation domains. One of the activation domains can be reduced to just 20 amino acids. A GAL4 fusion protein containing the HSF 1 regulatory domain and this 20-amino-acid activation domain is repressed at the control temperature but potently activates transcription in response to heat shock. No specific amino acids in this activation domain are required for response to the regulatory domain; in particular, none of the potentially phosphorylated serine and threonine residues are required for heat induction, implying that heat-induced phosphorylation of the transcriptional activation domains is not required for induction. The regulatory domain is able to confer heat responsiveness to an otherwise completely heterologous chimeric activator that contains a portion of the VP16 activation domain, suggesting that the regulatory domain can sense heat in the absence of other portions of HSF1.

RNA polymerase II cofactor PC2 facilitates activation of transcription by GAL4-AH in vitro

1994 ◽  
Vol 14 (6) ◽  
pp. 3927-3937
Author(s):  
M Kretzschmar ◽  
G Stelzer ◽  
R G Roeder ◽  
M Meisterernst
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Transcriptional Activation ◽  
Activation Domains ◽  
Positive Effects ◽  
Activation Of Transcription ◽  
Transcriptional Activation Domains ◽  
Necessary And Sufficient ◽  
Specific Pathway

We have isolated from a crude Hela cell cofactor fraction (USA) a novel positive cofactor that cooperates with the general transcription machinery to effect efficient stimulation of transcription by GAL4-AH, a derivative of the Saccharomyces cerevisiae regulatory factor GAL4. PC2 was shown to be a 500-kDa protein complex and to be functionally and biochemically distinct from native TFIID and previously identified cofactors. In the presence of native TFIID and other general factors, PC2 was necessary and sufficient for activation by GAL4-AH. Cofactor function was specific for transcriptional activation domains of GAL4-AH. The repressor histone H1 further potentiated but was not required for activation of transcription by GAL4-AH. On the basis of the observation that PC2 exerts entirely positive effects on transcription, we propose a model in which PC2 increases the activity of the preinitiation complex in the presence of an activator, thereby establishing a specific pathway during activation of RNA polymerase II.

scholarly journals Identification and Characterization of ART-27, a Novel Coactivator for the Androgen Receptor N Terminus

2002 ◽  
Vol 13 (2) ◽  
pp. 670-682 ◽  
Author(s):  
Steven M. Markus ◽  
Samir S. Taneja ◽  
Susan K. Logan ◽  
Wenhui Li ◽  
Susan Ha ◽  
...  
Keyword(s):  
Androgen Receptor ◽  
Transcriptional Activation ◽  
Mammalian Cells ◽  
Hybrid Approach ◽  
Independent Manner ◽  
Activation Domains ◽  
Cell Library ◽  
Nuclear Extracts ◽  
N Terminus ◽  
Transcriptional Activation Domains

The androgen receptor (AR) is a ligand-regulated transcription factor that stimulates cell growth and differentiation in androgen-responsive tissues. The AR N terminus contains two activation functions (AF-1a and AF-1b) that are necessary for maximal transcriptional enhancement by the receptor; however, the mechanisms and components regulating AR transcriptional activation are not fully understood. We sought to identify novel factors that interact with the AR N terminus from an androgen-stimulated human prostate cancer cell library using a yeast two-hybrid approach designed to identify proteins that interact with transcriptional activation domains. A 157-amino acid protein termed ART-27 was cloned and shown to interact predominantly with the AR153–336, containing AF-1a and a part of AF-1b, localize to the nucleus and increase the transcriptional activity of AR when overexpressed in cultured mammalian cells. ART-27 also enhanced the transcriptional activation by AR153–336 fused to the LexA DNA-binding domain but not other AR N-terminal subdomains, suggesting that ART-27 exerts its effect via an interaction with a defined region of the AR N terminus. ART-27 interacts with AR in nuclear extracts from LNCaP cells in a ligand-independent manner. Interestingly, velocity gradient sedimentation of HeLa nuclear extracts suggests that native ART-27 is part of a multiprotein complex. ART-27 is expressed in a variety of human tissues, including sites of androgen action such as prostate and skeletal muscle, and is conserved throughout evolution. Thus, ART-27 is a novel cofactor that interacts with the AR N terminus and plays a role in facilitating receptor-induced transcriptional activation.

scholarly journals Three functional classes of transcriptional activation domain.

1996 ◽  
Vol 16 (5) ◽  
pp. 2044-2055 ◽  
Author(s):  
J Blau ◽  
H Xiao ◽  
S McCracken ◽  
P O'Hare ◽  
J Greenblatt ◽  
...  
Keyword(s):  
Rna Polymerase Ii ◽  
Transcriptional Activation ◽  
Transcription Initiation ◽  
Type I ◽  
Rnase Protection ◽  
Activation Domains ◽  
Transcriptional Activation Domain ◽  
Transactivation Domains ◽  
Functional Classes

We have studied the abilities of different transactivation domains to stimulate the initiation and elongation (postinitiation) steps of RNA polymerase II transcription in vivo. Nuclear run-on and RNase protection analyses revealed three classes of activation domains: Sp1 and CTF stimulated initiation (type I); human immunodeficiency virus type 1 Tat fused to a DNA binding domain stimulated predominantly elongation (type IIA); and VP16, p53, and E2F1 stimulated both initiation and elongation (type IIB). A quadruple point mutation of VP16 converted it from a type IIB to a type I activator. Type I and type IIA activators synergized with one another but not with type IIB activators. This observation implies that synergy can result from the concerted action of factors stimulating two different steps in transcription: initiation and elongation. The functional differences between activators may be explained by the different contacts they make with general transcription factors. In support of this idea, we found a correlation between the abilities of activators, including Tat, to stimulate elongation and their abilities to bind TFIIH.

scholarly journals A heat shock-responsive domain of human HSF1 that regulates transcription activation domain function.

1995 ◽  
Vol 15 (6) ◽  
pp. 3354-3362 ◽  
Author(s):  
M Green ◽  
T J Schuetz ◽  
E K Sullivan ◽  
R E Kingston
Keyword(s):  
Heat Shock ◽  
Dna Binding ◽  
Transcriptional Activation ◽  
Binding Domain ◽  
Chimeric Proteins ◽  
Dna Binding Domain ◽  
Regulatory Domain ◽  
Activation Domain ◽  
Activation Domains ◽  
Transcriptional Activation Domains

Human heat shock factor 1 (HSF1) stimulates transcription from heat shock protein genes following stress. We have used chimeric proteins containing the GAL4 DNA binding domain to identify the transcriptional activation domains of HSF1 and a separate domain that is capable of regulating activation domain function. This regulatory domain conferred heat shock inducibility to chimeric proteins containing the activation domains. The regulatory domain is located between the transcriptional activation domains and the DNA binding domain of HSF1 and is conserved between mammalian and chicken HSF1 but is not found in HSF2 or HSF3. The regulatory domain was found to be functionally homologous between chicken and human HSF1. This domain does not affect DNA binding by the chimeric proteins and does not contain any of the sequences previously postulated to regulate DNA binding of HSF1. Thus, we suggest that activation of HSF1 by stress in humans is controlled by two regulatory mechanisms that separately confer heat shock-induced DNA binding and transcriptional stimulation.

Dimeric ligands define a role for transcriptional activation domains in reinitiation

Nature ◽  
1996 ◽  
Vol 382 (6594) ◽  
pp. 822-826 ◽  
Author(s):  
Steffan N. Ho ◽  
Stephen R. Biggar ◽  
David M. Spencer ◽  
Stuart L. Schreiber ◽  
Gerald R. Crabtree
Keyword(s):  
Transcriptional Activation ◽  
Activation Domains ◽  
Transcriptional Activation Domains
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