BRD4: a general regulator of transcription elongation

Abstract

Transcription elongation by RNA polymerase II (Pol II) has emerged as a regulatory hub in gene expression. A key control point occurs during early transcription elongation when Pol II pauses in the promoter-proximal region at the majority of genes in mammalian cells and at a large set of genes in Drosophila. An increasing number of trans-acting factors have been linked to promoter-proximal pausing. Some factors help to establish the pause, whereas others are required for the release of Pol II into productive elongation. A dysfunction of this elongation control point leads to aberrant gene expression and can contribute to disease development. The BET bromodomain protein BRD4 has been implicated in elongation control. However, only recently direct BRD4-specific functions in Pol II transcription elongation have been uncovered. This mainly became possible with technological advances that allow selective and rapid ablation of BRD4 in cells along with the availability of approaches that capture the immediate consequences on nascent transcription. This review sheds light on the experimental breakthroughs that led to the emerging view of BRD4 as a general regulator of transcription elongation.

Keywords: BET proteins; BRD4; PROTAC; RNA polymerase II; promoter-proximal pausing; transcription elongation.

Figure 1.

Classic model of the role of BRD4 in Pol II transcription regulation. BRD4 recruits P-TEFb to the promoter-proximal gene region through its binding to acetylated chromatin. Chromatin-bound P-TEFb phosphorylates different components of the Pol II transcription machinery indicated by gray arrows, leading to pause release. The DNA and nascent RNA are indicated as dark blue or red lines, respectively. The CTD is indicated as a gray tail of Pol II. Phosphorylation is depicted as a yellow bubble. TSS: transcription start site; Ac: Acetylation; (adapted from [79]).

Figure 2.

Experimental strategies applied to perturb BRD4 in mammalian cells. BRD4 protein-specific depletion methods include the auxin inducible degron (AID) (137) and the degradation tag (dTAG) (138) systems.

Figure 3.

Methods used to study the immediate impact of BRD4 perturbation on transcription. ChIP-Rx: chromatin immunoprecipitation with reference exogenous genome (106, 111, 139); nascONT-seq: nascent Oxford nanopore sequencing (106) and similar approaches (140, 141); NET-seq: native elongating transcript sequencing (25, 142, 143); SI-NET-seq: spike-in controlled NET-seq (106); p(a): polyadenylation; SLAM-seq: thiol(SH)-linked alkylation for the metabolic sequencing of RNA (110, 144); T: thymine; C: cytosine; 4sU: 4-thiouridine.

Figure 4.

Emerging direct functions of BRD4 in Pol II transcription elongation control. BRD4 helps to assemble a functional Pol II elongation complex and recruits 3’-end RNA processing factors (CPSF, CstF) during a general 5’-elongation control point to allow productive elongation and proper RNA processing at the 3’-end of genes. The scheme also includes the SPT5 (DSIF) phosphorylation cycle [131–133]. Although elongation factors can contact the Pol II CTD and nascent RNA they are shown at a different location for clarity. Although NELF and PAF cannot bind to Pol II at the same time in vitro [9], the model depicts both factors to illustrate that they interact with BRD4 and are present at the promoter-proximal region of genes in cells [114,134–144]. Despite accumulating evidence that BRD4 binds to acetylated chromatin (Figure 1) it is not shown for clarity. The color code for nascent RNA, DNA and phosphorylations is as in Figure 1. The Pol II CTD and P-TEFb phosphorylation targets are indicated as in Figure 1. The torpedo termination factor XRN2 is shown as a brown pac-man. TSS: transcription start site; S2: Serine 2 residue of the CTD; S5P: phosphorylated serine 5 residue of CTD, S2P: phosphorylated serine 2 residue of the CTD; P: phosphorylation; pA: polyadenylation site; PAS: polyadenylation signal.