Role of RNA Polymerase II Promoter-Proximal Pausing in Viral Transcription

Abstract

The promoter-proximal pause induced by the binding of the DRB sensitivity-inducing factor (DSIF) and the negative elongation factor (NELF) to RNAP II is a key step in the regulation of metazoan gene expression. It helps maintain a permissive chromatin landscape and ensures a quick transcriptional response from stimulus-responsive pathways such as the innate immune response. It is also involved in the biology of several RNA viruses such as the human immunodeficiency virus (HIV), the influenza A virus (IAV) and the hepatitis delta virus (HDV). HIV uses the pause as one of its mechanisms to enter and maintain latency, leading to the creation of viral reservoirs resistant to antiretrovirals. IAV, on the other hand, uses the pause to acquire the capped primers necessary to initiate viral transcription through cap-snatching. Finally, the HDV RNA genome is transcribed directly by RNAP II and requires the small hepatitis delta antigen to displace NELF from the polymerase and overcome the transcriptional block caused by RNAP II promoter-proximal pausing. In this review, we will discuss the RNAP II promoter-proximal pause and the roles it plays in the life cycle of RNA viruses such as HIV, IAV and HDV.

Keywords: DSIF; HDV; HIV; IAV; NELF; RNA polymerase II; promoter-proximal pause.

Figure 1

Transcription by RNAP II is paused after initiation as DSIF and NELF are recruited to the polymerase. This pause creates a hub for the capping enzyme, allowing the efficient capping of the nascent transcript. Interaction between P-TEFb and either BRD4 or SEC allows its activation and dissociation from the 7SK snRNP, where it is sequestered in its inactive state. The BRD4- or SEC-P-TEFb complexes are then recruited to the stalled polymerase where the kinase subunit of P-TEFb, CDK9, phosphorylates Spt5, NELF-E and the RNAP II CTD at Ser2. This leads to the dissociation of NELF from the polymerase, the transformation of DSIF into a positive elongation factor and entry into productive elongation.

Figure 2

(Active state) Transcription on the provirus is paused shortly after RNAP II transcribes through the TAR element. The interaction between P-TEFb and Tat leads to the activation and dissociation of P-TEFb from the 7SK snRNP, where it is sequestered in its inactive form. Tat-P-TEFb complexes are then recruited to the stalled polymerase through the interaction between Tat and TAR. The kinase subunit of P-TEFb, CDK9, and then phosphorylates Spt5, NELF-E and the RNAP II CTD at Ser2, leading to entry into productive elongation. (Latent state) Low levels of Tat in the infected cell due to insufficient amounts of cellular TFs, promoter mutations and low concentrations of nucleotides impedes basal HIV transcription. Therefore, P-TEFb remains sequestered by 7SK snRNP, and the polymerase remains stalled in a promoter-proximal manner, leading to latency.

Figure 3

The IAV RdRp interacts with the paused RNAP II through its PA and PB2 subunit. The PA subunit binds Ser5-P on the CTD, opening the cap-binding domain of PB2. Once the cap structure is bound by PB2, the endonuclease domain of PA cleaves the nascent pre-mRNA, which is then inserted in the catalytic center of PB1 and extended using the vRNA as a template.

Figure 4

RNAP II initiates transcription on the HDV RNA genome. Transcription is paused ~40 nucleotides after initiation, following recruitment of DSIF and NELF to the polymerase. Once the nascent transcript is capped, HDAg-S is recruited to the stalled complex where it competes with NELF-A for a common binding site on RNAP II. HDAg-S displaces NELF from RNAP II, stimulating processivity of the polymerase and therefore entry into productive elongation.