Entry created on 1 July 2019 (Revision 1.0) Annotator: Rita Pancsa; Orsolya Kovács
Basic protein information
Accession O60885
Common name BRD4
Gene BRD4
Organism Homo sapiens
Uniprot name Bromodomain-containing protein 4
Basic LLPS information
Organelle enhanceosome; nuclear body; nuclear bodies that occur at super enhancers in mESCs
Type of experimental evidence
Protein region(s) mediating LLPS
674
-
1351
P/Q-rich IDR, acidic and basic regions
Based on the experimental results of the following publication: 29930091
Molecular features viewer
PDB structures
View on PDBe
Extended LLPS information
Functional description
Enhancers are gene regulatory elements bound by transcription factors (TFs) and other components of the transcription apparatus that function to regulate expression of cell type-specific genes. Super enhancers (SEs) – clusters of enhancers that are occupied by exceptionally high densities of transcriptional machinery – regulate genes with especially important roles in cell identity. Two key components of SEs, BRD4 and MED1, form nuclear condensates at sites of SE-driven transcription. The IDRs of BRD4 and MED1 are sufficient to form phase-separated droplets in vitro. BRD4 is compartmentalized and therefore concentrated in MED1-IDR droplets, which compartmentalize and concentrate other transcriptional components in a transcriptionally competent nuclear extract as well. This offers insights into mechanisms involved in the control of key cell-identity genes since a study of RNA Pol II clusters which may be phase-separated condensates, suggests a correlation between condensate lifetime and transcriptional output (PMID:29930091). A mouse model expressing the BRD4 protein without the LCD domain helped establish an essential role of the BRD4 C-terminal LCD in vivo (PMID:31065677).
Literature supporting the LLPS: 29930091, 31065677
Functional class of membraneless organelle: activation/nucleation/signal amplification/bioreactor
Binding partners (at biological protein concentrations)
N/A
Type of RNA(s) required/used for the LLPS at biological protein concentrations
RNA not required.
Molecular interaction types contributing to LLPS
simple coacervation of hydrophobic residues (PMID:29930091) electrostatic (cation-anion) interaction (PMID:29930091)
Determinants of phase separation and droplet properties
1) protein concentration of BRD4 2) salt concentration
Membrane cluster No
Partner-dependent No
RNA-dependent No
PTM required No
Domain-motif interactions No
Discrete oligomerization No
Regulation and disease
Post-translational modifications affecting LLPS
Position Residue PTM Effect Reference Modifying enzyme Notes
Isoforms known to affect LLPS
Isoform Effect Reference
All known isoforms containing sequence changes in the LLPS region(s)
Position type Isoform names from UniProt
Disease mutations affecting LLPS
Mutation dbSNP Disease OMIM Effect Reference Notes
Experimental information
Experimental techniques applied to prove/investigate LLPS
In vitro immunofluorescent tagging of the proteins BRD4 and MED1 with anti-bodies in fixed murine embryonic stem cells (mESCs) revealed nuclear puncta for both factors (protein localization). The mEGFP-fused proteins showed similar localization in vivo by epifluorescence microscopy (particle size and count). ChIP-seq (chromatinimmunoprecipitation followed by sequencing) data for BRD4 and MED1 show that superenhancers are especially enriched in these coactivators (protein co-localization). BRD4 and MED1 puncta was found consistently overlapped with the DNA-FISH foci or RNA-FISH foci for the genomic region containing the Nanog gene. Based o FRAP measurements and 1,6-hexanediol treatment BRD4 and MED1 nuclear puncta exhibited liquid properties. The mEGFP-fused MED1 and BRD4 IDRs samples showed a change in optical properties (turbidity) with the addition of a crowding agent. The number and size of droplets (particle size and count) formed by the mEGFP-fused MED1 and BRD4 IDRs in vitro were depending on changes in protein concentration, salt concentration and crowding agent. The MED1 IDR droplets could incorporate and concentrate BRD4-IDR in vitro under conditions where the BRD4 IDR could not form droplets on its own (protein co-localization) as assessed by epifluorescence microscopy. PMID:29930091.
Experimental observations supporting the liquid material state of the condensate
dynamic movement/reorganization of molecules within the droplet (PMID:29930091) sensitivity to 1,6-hexanediol (PMID:29930091) morphological traits (PMID:29930091)