| 2001 |
Human EAF1 was identified as a direct binding partner of ELL via yeast two-hybrid screen and confirmed by co-immunoprecipitation of endogenous proteins from multiple cell lines. EAF1 also interacts with ELL2. EAF1 and ELL colocalize in a nuclear speckled (Cajal body) pattern, and expression of MLL-ELL fusion protein delocalizes EAF1 from this nuclear speckled distribution to a diffuse nucleoplasmic pattern. |
Yeast two-hybrid screen, co-immunoprecipitation of endogenous proteins, confocal microscopy, transfection of MLL-ELL fusion gene |
Blood |
High |
11418481
|
| 2003 |
ELL and EAF1 are components of Cajal bodies (CBs) and colocalize with p80 coilin, though without direct physical interaction with coilin. Localization of ELL and EAF1 in CBs is dependent on active RNA Pol II transcription, as treatment with actinomycin D, DRB, or alpha-amanitin disperses them from CBs. In MLL-ELL leukemia cells, CBs are disrupted and EAF1 and p80 coilin are delocalized from CBs, with diminished nuclear expression of EAF1. |
Confocal microscopy, co-immunoprecipitation, transcription inhibitor treatment (actinomycin D, DRB, alpha-amanitin), nuclear/cytoplasmic fractionation |
Molecular biology of the cell |
High |
12686606
|
| 2002 |
EAF1 binds to the carboxy-terminus of ELL, whereas EAF2 does not; both EAF1 and EAF2 bind an amino-terminal interaction domain of ELL that is disrupted in MLL-ELL fusion protein formation, so MLL-ELL retains an interaction domain for EAF1 but not EAF2. EAF1 contains a transcriptional activation domain in a serine/aspartate/glutamate-rich region homologous to MLL translocation partners. |
Co-immunoprecipitation, domain mapping with ELL truncation/fusion constructs, functional transcriptional activation assays |
Blood |
Medium |
12446457
|
| 2008 |
Yeast Eaf1 serves as the central platform/scaffold for assembly of the NuA4 histone acetyltransferase complex. Deletion of EAF1 causes collapse of the entire NuA4 complex in vivo, while depletion of other subunits does not. Eaf1 is found exclusively associated with NuA4 (not other complexes) in vivo. Expression of a chimeric Eaf1-Swr1 protein recreates a single human TIP60-like complex in yeast, demonstrating structural and functional equivalence. NuA4 and SWR1 show strong genetic interactions and NuA4 affects H2AZ incorporation/acetylation in vivo. |
Genetic deletion/depletion, co-immunoprecipitation/complex purification, chimeric protein expression, genetic interaction analysis, in vivo histone acetylation assays, chromatin immunoprecipitation |
Molecular and cellular biology |
High |
18212047 18212056
|
| 2015 |
Loss of yeast Eaf1 affects H2A.Z (Htz1) incorporation predominantly at promoters normally highly enriched in this histone variant. NuA4 (via Eaf1 scaffold) directly interacts with the Bas1 transcription factor activation domain, and NuA4-dependent acetylation presets ADE gene promoter nucleosomes (enriched in Htz1 and acetylated) for rapid derepression of purine biosynthesis genes. |
Chromatin immunoprecipitation (ChIP), expression arrays, genome-wide Htz1 mapping, co-immunoprecipitation of NuA4 with Bas1 activation domain |
Eukaryotic cell |
Medium |
25841019
|
| 2009 |
Zebrafish eaf1 (and eaf2) act upstream of noncanonical Wnt signaling to mediate convergence and extension movements. Morpholino knockdown of eaf1 reduces expression of wnt11 and wnt5, and cell tracing experiments demonstrate cell migration defects. Rescue with wnt11/wnt5 mRNA (and more effectively with rhoA mRNA, a convergence point of both) restores normal morphogenesis. Ectopic expression of wnt11/wnt5 does not affect eaf1/eaf2 expression, placing eaf1 upstream. |
Morpholino knockdown, mRNA rescue injection, in situ hybridization, cell tracing with kaeda mRNA |
The Journal of biological chemistry |
Medium |
19380582
|
| 2010 |
EAF1 and EAF2 are transcriptionally upregulated by Wnt4 signaling, and in turn, EAF1 and EAF2 directly bind to the Wnt4 promoter to suppress its expression, forming an auto-regulatory negative feedback loop. This was demonstrated by chromatin immunoprecipitation in both zebrafish embryos and mammalian cells. |
Chromatin immunoprecipitation (ChIP), mRNA/morpholino gain- and loss-of-function in zebrafish, reporter assays in mammalian cells |
PloS one |
Medium |
20161747
|
| 2013 |
EAF1 and EAF2 negatively regulate canonical Wnt/β-catenin signaling. By immunoprecipitation, Eaf1 and Eaf2 bind to the Armadillo repeat region and C-terminus of β-catenin, as well as to c-Jun, Tcf, and Axin (components of the β-catenin transcription complex). The N-terminus of Eaf1 binds β-catenin and exhibits dominant-negative activity; the C-terminus harbors a suppression domain or recruits a repressor. Both termini must be intact for full suppressive activity. |
Immunoprecipitation, zebrafish morpholino/mRNA gain- and loss-of-function, β-catenin reporter assays in cultured cells, domain deletion analysis |
Development (Cambridge, England) |
Medium |
23364330
|
| 2014 |
Zebrafish eaf1 suppresses foxo3b expression; microarray analysis identified foxo3b as a target gene suppressed by eaf1. Dominant-negative foxo3b partially rescues primitive hematopoietic defects in eaf1 morphants. Foxo3b inhibits transcriptional activity of gata1 and spi1 through protein-protein interaction, establishing the pathway: eaf1 → (suppresses) foxo3b → (inhibits) gata1/spi1. |
Morpholino knockdown, microarray analysis, mRNA rescue, dominant-negative construct, protein-protein interaction assay |
Developmental biology |
Medium |
24445282
|
| 2017 |
The NMR solution structure of MED26 N-terminal domain (NTD) revealed a 4-helix bundle. EAF1 (residues 239-268) binds the same groove on MED26-NTD (formed by H3 and H4 helices) as TAF7. Both interactions occur with dissociation constants in the 10-μM range and involve a folding-upon-binding mechanism generating an EAF1 helix (N247-S260). Ala mutations of charged residues in the C-terminal disordered part of EAF1 peptide reduced binding affinity ~10-fold. |
NMR structure determination, NMR chemical shift perturbation mapping, NOE contacts, mutagenesis (Ala substitutions), binding affinity measurements |
Journal of molecular biology |
High |
28893534
|
| 2017 |
Eaf1 and Eaf2 inhibit TGF-β signaling in zebrafish and mammalian cells. Loss-of-function expands mesoderm and endoderm, while gain-of-function reduces them. Using TGF-β reporters and eaf1/2-engrailed fusion proteins (repressor domain fusions), Eaf1/2 suppress TGF-β transduction synergistically via both P53-dependent and P53-independent pathways. Eaf1/2 co-localize and interact with TGF-β transcriptional factors as repressors in the transcriptional complex. |
Morpholino/mRNA gain- and loss-of-function in zebrafish, TGF-β reporter assays, engrailed repressor domain fusions, co-localization/interaction experiments, P53 mutant analysis |
Biochimica et biophysica acta. Gene regulatory mechanisms |
Medium |
28887217
|
| 2020 |
EAF1 and EAF2 interact with Super Elongation Complex (SEC) components in an ELL1/2-dependent manner and negatively regulate HIV-1 transcription. EAF1/2 compete with the scaffolding subunit AFF1 for binding to ELL, thereby reducing SEC formation and its occupancy on HIV-1 proviral DNA. Depletion of EAF1/2 increases SEC formation and Tat-dependent HIV-1 transactivation. |
Co-immunoprecipitation, ChIP on HIV-1 proviral DNA, knockdown of EAF1/2, HIV-1 transcription reporter assays |
Biochimica et biophysica acta. Gene regulatory mechanisms |
Medium |
32087315
|
| 2022 |
EAF1 regulates ELL protein stability by competing with HDAC3 for binding at the N-terminus of ELL; reduced HDAC3 binding causes increased ELL acetylation, leading to reduced ubiquitylation-mediated ELL degradation. EAF1 and DBC1 form a negative feedback loop: increased DBC1 reduces EAF1 level via TRIM28-mediated ubiquitylation, while increased EAF1 reduces DBC1 level through transcriptional repression. During genotoxic stress, the EAF1-dependent pathway is preferentially used for ELL level maintenance over the DBC1-dependent pathway. |
Co-immunoprecipitation, knockdown experiments, ubiquitylation assays, acetylation assays, transcriptional reporter assays, genotoxic stress treatments |
Molecular and cellular biology |
Medium |
36036574
|
| 2022 |
During genotoxic stress, ATM phosphorylates ELL, which enhances ELL's interaction with EAF1. This enhanced EAF1 association promotes ELL self-association, which reduces ELL interaction with other SEC components (including AFF1), leading to global transcriptional inhibition. EAF1 alone forms a complex with ELL distinct from SEC and LEC, and contrary to in vitro studies, EAF1 inhibits ELL-dependent RNA Pol II transcription in vivo. |
Co-immunoprecipitation, ATM kinase inhibition, genotoxic stress treatments, ELL phosphorylation assays, transcription run-on/nascent RNA assays, complex purification |
Nucleic acids research |
Medium |
36305813
|