The rDNA gene is organized into tandem repeats that are spread throughout the mammalian genome with somatic cells containing approximately 200 copies of the rDNA gene (Fig. 6.1); however, not all rDNA copies are actively transcribed [30]. The regulation of the rDNA repeats’ conformation is important for ribosome biogenesis [19]. The nucleolus is an extremely prominent region within the nucleus where rDNA transcription occurs and is heavily packed with proteins responsible for synthesis of the 45S pre-rRNA and ribosome assembly (Fig. 6.2).

The Pol I enzyme requires the formation of a protein complex – the preinitiation complex (PIC) – at the rDNA promoter region. The upstream binding factor (UBF) is the first protein attaching to the rDNA. UBF binds physically to the promoter region (both UPE and the core promoters) via high mobility group (HMG) box domains, which confers DNA-binding activity of UBF. This association is the basic mechanism for the formation of the enhanceosome at the rRNA gene. The ribosomal enhanceosome is an almost 360° loop on the promoter elements allowing PIC formation for subsequent Pol I transcription [53]. Following enhanceosome formation, selectivity factor 1 (SL1) complex (formed by several TATA-binding proteins) associates with UBF and the rDNA [13]. The SL1 complex binds to the promoter region alongside UBF and recruits TIF-IA (also known as RRN3) and Pol I to initiate transcription [45]. TIF-IA, via protein-protein interactions, associates with SL1 and Pol I enzyme complex [5, 6], serving as a bridge between PIC and PolI[8, 18, 38].

Following synthesis, the 45S pre-rRNA will be processed into the mature rRNAs for maturation with ribosomal protein into the 40S and 60S subunits. First, the ETS and ITS regions are removed by nucleases and then degraded [20]. The majority of the processing occurs while pre-rRNA is still in the nucleolus, although some processing also occurs in the nucleoplasm and cytoplasm [20]. In addition to the processing of the 45S, the 5S rRNA transcribed by the Pol III and the ribosomal proteins translated in the cytoplasm will also join the rRNA processing and maturation to form the ribosomal subunits [29].

The majority of total cellular RNA is by far rRNA. Depending on the cell, as much as 80% of extracted RNA in tissue or cells can be rRNAs [61]. In proliferating cells, the majority of new transcripts are rRNA to make new ribosomes and support growth [61]. Due to the high cost of production, the process of making ribosomes is tightly regulated at many levels by nutritional, hormonal, and energy-sensing cues. It is widely known that many growth-related pathways, such as PI3K-Akt-mTOR axis and MAPK/ERK, trigger cell growth via increased rates of translation, especially at initiation and elongation steps. Those same pathways can also alter the rate of pre-rRNA synthesis. The transcription of rDNA is the central process governing ribosome biogenesis, in which the formation of the preinitiation complex (PIC) at the rDNA promoter is regulated by those pathways, such as mTOR and MAPK (Fig.6.3).

The mammalian target of rapamycin (mTOR) is a master kinase controlling ribosome activity and its biogenesis. mTOR complex 1 (mTORC1) controls protein synthesis by phosphorylating many targets, especially the 4E-BP1 and p70S6K at the TOS motif sequence [47]. Hyperphosphorylation of 4E-BP1 releases the inhibitory effect of 4E-BP1 on translation initiation of 5’TOP mRNAs. Members of this selected group of mRNAs encode proteins involved in cellular growth. The majority of mRNA translation that 4E-BP protein inhibits encodes ribosomal proteins, mitochondrial ribosomal proteins, and proteins that associate with ribosomes to drive translation, the eukaryotic initiation, and elongation factors (eIFs and eEfs) [55]. mTORC1 also regulates the transcription of the RNA polymerases I, II, and III (Pol I, II, and III) [34, 56]. In addition to these effects, mTOR also promote the phosphorylation of TIF-IA [35]. Therefore, activation of mTORC1 increases translation efficiency and transcription of the RNA polymerases toward augmentation of translational capacity, supporting cell growth in addition to modulating PIC formation at the rDNA promoter (Fig. 6.4).

In addition to mTORC1, the MAPK pathway also promotes ribosome biogenesis. Different proteins of MAPK pathway also directly phosphorylate key proteins promoting assembly of the PIC components at the rDNA promoter. ERK and RSK proteins can phosphorylate TIF-IA and UBF proteins and promote rDNA transcription [52, 54, 66]. In addition, as ribosome biogenesis is also important in cell proliferation, cell cycle regulators, such as the cyclin-dependent kinases (CDK), have also been shown to modulate ribosome biogenesis. CDKs can phosphorylate UBF directly [57, 58] and TIF-IA [35]. In particular, CDK4, when complexed with cyclin D1, can directly phosphorylate UBF protein [58]. Moreover, the pro-oncogene protein c-Myc, known for its role in cell cycle and proliferation, also promotes ribosome biogenesis. The c-Myc protein has been shown to directly interact with PIC components at the rDNA promoter [1].

Ribosome biogenesis is a highly metabolic demanding cellular process, and as such, AMPK has been found to be an important factor linking energy balance with rDNA transcription. Not only can AMPK interfere with ribosome biogenesis via its regulation at mTORC1 and translation initiation level [31], but AMPK has also been found to interact with PIC components. AMPK can directly phosphorylate and destabilize TIF-IA association with SL1 [21] impacting negatively in the rate of pre-rRNA synthesis (Fig. 6.3).

Many recent studies have shown that changes in ribosome biogenesis and translational capacity correlate well with changes in muscle mass in both growth and wasting conditions [9, 10, 43, 50]. Thus, muscle ribosome biogenesis appears to be necessary for muscle hypertrophy [62]. However, more mechanistic and proof-of-principle evidences are necessary to stablish a causal relationship  between ribosome biogenesis and muscle growth.

The regulation of ribosome biogenesis and translational capacity is an open venue for muscle biology researchers with interest in both the fundamental understanding of the molecular mechanisms of muscle growth and also of great applicability for clinical research. Novel aspects of regulation of muscle ribosome biogenesis may help elucidate the mechanism of muscle adaptation to different stimuli, providing tools to better design interventions of clinical interest. In addition to resistance exercise, endurance training studies will also further our understanding regarding the different exercise stimuli on ribosome biogenesis and the potential influence on skeletal muscle phenotypes. Moreover, of clinical relevance, novel therapeutic approaches to ameliorate the loss of muscle mass during muscle disuse, chronic inflammatory conditions, cancer cachexia, and sarcopenia can be developed through the elucidation of the role of ribosome biogenesis in skeletal muscle size regulation.