rRNA Transcription: Mechanisms, Regulation, and Cellular Impact

Cells rely on ribosomes to synthesize proteins, and the production of ribosomal RNA (rRNA) is a key step in this process. rRNA transcription is tightly regulated to meet cellular demands for protein synthesis, ensuring proper growth and adaptation to environmental conditions. Disruptions can lead to developmental abnormalities and diseases such as cancer.

Understanding how rRNA transcription is controlled provides insight into fundamental aspects of gene expression and cellular function.

Genetic Organization of Ribosomal DNA

Ribosomal DNA (rDNA) is arranged in highly repetitive clusters within the genome, encoding the rRNA molecules that form the core of ribosomes. In eukaryotic cells, these rDNA sequences are found in tandem arrays at specific chromosomal loci, known as nucleolar organizer regions (NORs). In humans, rDNA is located on the short arms of acrocentric chromosomes (13, 14, 15, 21, and 22), where hundreds of copies of the rRNA genes are arranged in head-to-tail orientation. This organization ensures high transcriptional output to meet the demand for ribosome production.

Each rDNA repeat unit consists of a transcriptional unit and intergenic spacer regions that regulate expression. The transcriptional unit includes the genes for the 18S, 5.8S, and 28S rRNAs, which are transcribed as a single precursor (45S pre-rRNA) before undergoing processing. Upstream of the transcriptional unit lies the promoter region, which contains binding sites for transcription factors and RNA polymerase I, the enzyme responsible for rRNA synthesis. The intergenic spacers contain regulatory elements such as enhancers and terminator sequences that influence transcription efficiency and chromatin organization.

The chromatin state of rDNA determines transcriptional activity. Active rDNA repeats are typically unmethylated and associated with histone acetylation, allowing efficient transcription. In contrast, silenced rDNA copies are often hypermethylated and enriched in repressive histone marks, forming condensed chromatin that prevents transcription. This regulation enables cells to adjust rRNA synthesis in response to metabolic needs and environmental cues.

Transcription Mechanism and Major Steps

The synthesis of rRNA involves transcription factors, chromatin remodelers, and RNA polymerase I. This process consists of three main stages: initiation, elongation, and termination.

Initiation

Initiation begins with the recruitment of RNA polymerase I to the rDNA promoter. This process is mediated by the upstream binding factor (UBF) and the selectivity factor 1 (SL1) complex, which includes TATA-binding protein (TBP) and several TBP-associated factors (TAFs). UBF binds to the upstream control element and core promoter, inducing DNA bending that facilitates SL1 binding. SL1 then stabilizes the pre-initiation complex by recruiting RNA polymerase I and transcription initiation factors such as RRN3. The interaction between RRN3 and RNA polymerase I is essential for polymerase recruitment and promoter escape.

Once the pre-initiation complex is assembled, RNA polymerase I undergoes conformational changes that enable the transition from a closed to an open complex, allowing the unwinding of DNA at the transcription start site. This step is followed by the synthesis of short abortive transcripts before the polymerase successfully clears the promoter and enters elongation. The efficiency of initiation is influenced by chromatin modifications, including histone acetylation and DNA methylation, which regulate promoter accessibility.

Elongation

During elongation, RNA polymerase I moves along the rDNA template, synthesizing the 45S precursor rRNA. This process is facilitated by elongation factors such as SPT5 and PAF1C, which enhance polymerase processivity and prevent premature termination. The nascent transcript undergoes co-transcriptional modifications, including 2′-O-methylation and pseudouridylation, guided by small nucleolar RNAs (snoRNAs) and associated proteins. These modifications are essential for proper folding and stability of the pre-rRNA.

Histone chaperones such as FACT assist in nucleosome displacement ahead of the polymerase and reassembly behind it. Additionally, topoisomerases alleviate supercoiling stress generated by transcription, ensuring smooth progression of RNA polymerase I. The rate of elongation is tightly regulated to coordinate with downstream processing events, preventing the accumulation of defective rRNA precursors.

Termination

Termination occurs at specific sequences downstream of the 45S pre-rRNA coding region. This process is mediated by the transcription termination factor TTF-I, which binds to terminator elements and induces polymerase pausing. The paused polymerase is then released through the action of PTRF (polymerase I and transcript release factor) and other termination-associated factors. Efficient termination prevents readthrough transcription into adjacent rDNA repeats, maintaining rRNA synthesis integrity.

Following termination, the released pre-rRNA undergoes processing, while RNA polymerase I is recycled for subsequent rounds of transcription. The termination process is also linked to chromatin remodeling, as TTF-I recruits chromatin modifiers that help reset the rDNA locus for the next transcription cycle.

Role of RNA Polymerase I in rRNA Synthesis

RNA polymerase I (Pol I) is the dedicated enzyme for transcribing rDNA into precursor rRNA, a fundamental step in ribosome biogenesis. Unlike RNA polymerase II, which transcribes protein-coding genes, or RNA polymerase III, which synthesizes small non-coding RNAs, Pol I is specialized for the rapid and efficient production of rRNA. This specialization is reflected in its structure, which includes unique subunits such as RPA34 and RPA49 that enhance its processivity and resistance to transcriptional pausing.

Pol I transcription efficiency is dictated by its ability to navigate the repetitive rDNA landscape while maintaining fidelity. Unlike RNA polymerase II, which frequently encounters regulatory pauses, Pol I is optimized for continuous elongation, ensuring rapid synthesis of long pre-rRNA transcripts. Structural studies using cryo-electron microscopy have revealed that Pol I adopts a more closed and rigid conformation compared to Pol II, reducing the likelihood of premature dissociation from the DNA template.

Pol I activity is regulated by cellular metabolism and growth signals. Under high metabolic demand, Pol I transcription is upregulated to support ribosome production. During cellular stress or nutrient deprivation, Pol I activity is suppressed to conserve energy. This regulation occurs through post-translational modifications of Pol I subunits, interactions with transcription factors, and chromatin remodeling. Dysregulation of these mechanisms has been implicated in diseases such as cancer, where hyperactive Pol I transcription contributes to uncontrolled cell proliferation.

Epigenetic Modifications and Nucleolar Architecture

rDNA transcription is controlled by epigenetic modifications that regulate chromatin accessibility within the nucleolus. DNA methylation plays a key role, with hypermethylated rDNA copies forming repressive heterochromatin, while hypomethylated repeats remain accessible for transcription. Increased methylation of rDNA promoters correlates with reduced rRNA synthesis in aging and disease states.

Histone modifications further refine this control by influencing nucleosome positioning and chromatin compaction. Acetylation of histone H3 at lysine 9 (H3K9ac) and histone H4 at lysine 16 (H4K16ac) is associated with active rDNA, promoting an open chromatin conformation. Conversely, trimethylation of histone H3 at lysine 9 (H3K9me3) and lysine 27 (H3K27me3) marks repressive rDNA regions, leading to condensed chromatin domains.

Cellular Signaling Pathways Affecting Transcription

rRNA transcription is integrated with cellular signaling pathways that respond to metabolic conditions, growth signals, and stress. These pathways modulate RNA polymerase I activity through post-translational modifications, transcription factor recruitment, and chromatin remodeling.

The mTOR pathway promotes rRNA transcription under favorable growth conditions. Activation of mTORC1 leads to phosphorylation of UBF, enhancing RNA polymerase I recruitment. Conversely, energy stress activates AMPK, which phosphorylates TIF-IA, leading to its inactivation and reduced rRNA synthesis. The tumor suppressor p53 further modulates Pol I activity in response to DNA damage by repressing rRNA transcription.

Coordination With Ribosome Assembly

Efficient rRNA transcription must be coordinated with ribosome assembly to ensure functional ribosome production. As rRNA is transcribed, it undergoes co-transcriptional folding and modification, facilitated by snoRNAs and associated proteins. These modifications guide the processing of the 45S pre-rRNA into mature 18S, 5.8S, and 28S rRNAs, which are then assembled with ribosomal proteins.

Assembly factors such as nucleolin and fibrillarin assist in ribosome formation, ensuring that rRNA adopts the correct structure. Disruptions in this coordination, whether due to mutations in ribosomal proteins or defects in rRNA processing, can result in ribosomopathies—diseases characterized by impaired ribosome function, such as Diamond-Blackfan anemia and Treacher Collins syndrome.