- RNA Polymerase II (Pol II) is a critical enzyme in eukaryotic cells, primarily responsible for transcribing protein-coding genes into messenger RNA (mRNA). It also synthesizes several non-coding RNAs, including small nuclear RNAs (snRNAs) and microRNAs (miRNAs), which play essential roles in RNA processing and gene regulation.
- As a multi-subunit complex composed of 12 core subunits, RNA Polymerase II is at the heart of the transcription machinery and is tightly regulated at multiple levels to ensure precise control of gene expression.
- The structure of RNA Polymerase II resembles a crab claw, with a deep central cleft that accommodates DNA and the nascent RNA transcript. The largest subunit, RPB1, contains a unique and essential feature known as the C-terminal domain (CTD), which consists of multiple repeats of a heptapeptide sequence (YSPTSPS). The CTD is dynamically phosphorylated during different stages of the transcription cycle, acting as a regulatory hub that coordinates transcription with RNA processing events such as capping, splicing, and polyadenylation.
- Initiation of transcription by RNA Polymerase II requires the assembly of a large pre-initiation complex (PIC) at the promoter region of genes. This assembly involves multiple general transcription factors (GTFs), including TFIID (which contains the TATA-binding protein or TBP), TFIIA, TFIIB, TFIIF, TFIIE, and TFIIH. TFIID first recognizes and binds the promoter’s core elements, followed by sequential recruitment of the other GTFs and Pol II. TFIIH possesses helicase activity to unwind DNA and kinase activity to phosphorylate the CTD, marking the transition from initiation to elongation.
- During transcription elongation, Pol II moves along the DNA template strand, synthesizing RNA in the 5′ to 3′ direction. Elongation is supported and regulated by elongation factors that help Pol II navigate chromatin structures and pause sites. The CTD becomes progressively phosphorylated, particularly at Ser2 and Ser5 residues, to recruit RNA-processing enzymes that add a 5’ cap, remove introns, and prepare the 3’ end for polyadenylation. These modifications are essential for RNA stability, nuclear export, and translation.
- Termination of transcription involves cleavage of the nascent RNA and dissociation of Pol II from the DNA. This process is tightly linked to 3′ end processing and is mediated by cleavage and polyadenylation specificity factors (CPSFs), which recognize polyadenylation signals. Once the RNA is cleaved and polyadenylated, Pol II disengages and may be recycled for new rounds of transcription.
- RNA Polymerase II function is also intricately connected to chromatin remodeling and epigenetic regulation. It must contend with nucleosomes, the basic units of chromatin, during transcription. Chromatin remodelers and histone-modifying enzymes assist in opening the chromatin to allow Pol II access to DNA. Furthermore, enhancers, silencers, and mediator complexes interact with Pol II to modulate transcription in a gene-specific and context-dependent manner.
