SirT1 and rRNA in the nucleolus: regulating the regulator

The synthesis of ribosomal RNA (rRNA) is tightly coupled to nutrient conditions and growth factors. Murayama et al. described a novel nucleolar silencing complex eNoSC involved in rRNA transcription regulation in response to glucose deprivation [1]. This complex contains SirT1, SUV39H1 and the nucleolar protein nucleomethylin (NML). Upon glucose starvation, NML inhibits rRNA synthesis by recruiting SirT1 and SUV39H1 to rDNA promoter and spreading heterochromatin marks across the rDNA repeat. Unlike yeast Sir2, which localizes to the nucleolus and telomeres, mammalian SirT1 is predominantly located in the nucleoplasm. Therefore, SirT1 function in the nucleolus may require recruitment by the nucleolar NML. In a recent study, we investigated the dynamic regulation of eNoSC assembly by nutrient conditions and found that the interaction of SirT1 and NML was regulated by rRNA [2]. A disordered region adjacent to the methyltransferase domain of NML was found to bind RNA in a sequenceindependent manner in vitro. The RNA and SirT1 binding sites on NML overlap partially. Thus, RNA and SirT1 compete for interacting with NML. The NML complex purified from cells contains predominantly ribosomal rRNA (5S, 5.8S and 28S), presumably due to its nucleolar localization. The interaction of NML with rRNA in vivo was inhibited by glucose deprivation, which is known to down-regulate pre-rRNA transcription through the AMPK-mTOR-RNA Pol I signaling pathway. Therefore, glucose deprivation strongly stimulates NML-SirT1 binding, resulting in the recruitment of a minor fraction of SirT1 to the nucleolus. Assembly of eNoSC contributes to the down regulation of rRNA transcription in the absence of glucose. In fact, various stresses that down regulate rRNA transcription, such as DNA damage, inhibition of mTOR using rapamycin, or direct inhibition of RNA Pol I using actinomycin D or CX5461, all promote SirT1-NML binding. These findings led to a positive feedback model that coordinates SirT1 function in the nucleolus with nutrient signaling: Under nutrient-rich condition, mTOR and other factors stimulate RNA Pol I and Pol III synthesis of rRNA. Nascent rRNA is processed and assembled into pre-ribosomal particles at the nucleolus. The abundant nascent rRNA also interacts with NML and prevents the recruitment of SirT1. This mechanism provides a positive feedback loop to nutrient signaling by protecting the active nucleolar rDNA from repression by the NMLSirT1 complex (Figure 1a). During starvation, mTOR inactivation reduces Pol I and Pol III activity. Reduced level of nascent rRNA enables NML-SirT1 complex formation in the nucleolus, promoting heterochromatin formation and silencing of the rDNA (Figure 1b). Therefore, NML provides positive feedback on mTOR regulation of rRNA synthesis by sensing the level of the nascent rRNA output. SirT1 is well known for its regulation by both the substrate and the reaction products. NAD+ is required for SirT1-mediated deacetylation reaction, whereas the deacetylation byproduct nicotinamide is an inhibitor of SirT1 activity [3]. Assembly of the SIR complex in yeast is inhibited by another product of deacetylation, O-acetylADP-ribose [4]. Our study identified a novel mechanism of SirT1 regulation that involves the RNA product of transcription. Furthermore, the results suggest that the RNA-mediated regulation may not require sequence Editorial Material