Je-Hyun Yoon

Long noncoding RNAs (lncRNAs) have emerged as critical regulators of cancer development and progression by modulating enzymatic reactions involved in cellular metabolism and protein modifications. These noncoding transcripts exert their roles through molecular interactions with target proteins. Here we explore the diverse mechanisms through which lncRNAs optimize enzymatic reactions in metabolic processes and protein modifications, both of which are fundamental to the growth, adaptation, and survival of cancer cells. These processes, often dysregulated during cancer progression, are modulated by lncRNAs through direct interactions with specific proteins, underscoring their critical role in oncogenesis. Key research findings are summarized to provide a comprehensive understanding of the roles of lncRNAs in modulating these cellular processes. This Review offers an in-depth overview of the complex regulatory networks through which lncRNAs influence cancer pathogenesis and provides essential insights for the development of future therapeutic strategies.

Uridine is the ubiquitous nucleoside form of the RNA base uracil. It occupies a prominent ‘hub’ position in energy metabolism; for example, it is metabolically linked to de novo pyrimidine biosynthesis and glycolysis and biologically linked to diverse processes, such as RNA synthesis/degradation and glycosylation. It is a vital interorgan ‘currency’ nutrient readily imported by mammalian cells, and its supplementation can exert both cytoprotective and toxic effects, for which the underlying mechanisms are poorly understood. Importantly, it is a route by which the decay of RNA can be repurposed as an alternative fuel source under nutrient-limiting conditions to aid in tumor initiation, development and metastasis. Here we explain how the upstream inputs and downstream metabolic fates of uridine influence cancer traits and illustrate both established and hypothetical strategies targeting uridine metabolism for cancer therapy.

Signaling pathways and transcriptional regulation during cellular senescence have been investigated; however, energy metabolism is one of the understudied areas. Senescent cells secrete pro-inflammatory cytokines and release proteins and RNAs via exosomes that contribute to organismal aging. Although senescent fibroblasts in solid organs are in a low oxygen environment, these fibroblasts have more active glucose metabolism and consume more oxygen than proliferating ones. A critical gap in our knowledge is how senescent fibroblasts facilitate glucose metabolism and organismal aging by creating a distinct microenvironment. Our high throughput profiling of mRNAs and proteins from Human Diploid Fibroblasts (HDFs) revealed that the expression of pyruvate metabolic enzymes is inhibited by the anti-senescent RNA-binding protein (RBP) AUF1 (AU-binding Factor 1). Our studies revealed that AUF1 promotes the decay of mRNAs encoding two enzymes: PGAM1 (phosphoglycerate mutase 1), a glycolytic enzyme involved in the pyruvate synthetic pathway, and PDP2 (Pyruvate Dehydrogenase Phosphatase 2), which activates Pyruvate Dehydrogenase. We also demonstrated that AUF1 is phosphorylated by a Serine/Threonine kinase, MST1 (Mammalian Ste20-like kinase 1; encoded by STK4), resulting in the inactivation of AUF1, which leads to target mRNA stabilization and senescence. Overexpression of PGAM1 and PDP2 predicts an acceleration of pyruvate production and subsequent citrate metabolism, leading to cellular senescence and aging. Thus, our studies revealed regulatory mechanisms of glycolysis-driven cellular senescence by AUF1-mediated decay of PGAM1 and PDP2 mRNAs.

SARS-CoV-2 is the betacoronavirus causing the COVID-19 pandemic. Although the SARS-CoV-2 genome and transcriptome were reported previously, the function of individual viral proteins is largely unknown. Utilizing biochemical and molecular biology methods, we identified that four SARS-CoV-2 RNA-binding proteins (RBPs) regulate the host RNA metabolism by direct interaction with mature miRNA let-7b revealed by Nuclear Magnetic Resonance spectroscopy (NMR). SARS-CoV-2 RBP Nsp9 primarily binds mature miRNA let-7b, a direct ligand of the Toll-like Receptor 7 (TLR7), one of the potential SARS-CoV-2 therapeutics. Nsp9 suppresses host gene expression possibly by promoting let-7b-mediated silencing of a cellular RNA polymerase, POLR2D. In addition, Nsp9 inhibits extracellular release of let-7b and subsequent antiviral activity via TLR7. These results demonstrate that SARS-CoV-2 hijacks the host RNA metabolism to suppress antiviral responses and to shut down cellular transcription. Our findings of how a natural ligand of TLR7, miRNA let-7b, is suppressed by SARS-CoV-2 RBPs will advance our understanding of COVID-19 and SARS-CoV-2 therapeutics.

The metazoan lifespan is determined in part by a complex signaling network that regulates energy metabolism and stress responses. Key signaling hubs in this network include insulin/IGF‐1, AMPK, mTOR, and sirtuins. The Hippo/Mammalian Ste20‐like Kinase1 (MST1) pathway has been reported to maintain lifespan in Caenorhabditis elegans , but its role has not been studied in higher metazoans. In this study, we report that overexpression of Hpo, the MST1 homolog in Drosophila melanogaster , decreased lifespan with concomitant changes in lipid metabolism and aging‐associated gene expression, while RNAi Hpo depletion increased lifespan. These effects were mediated primarily by Hpo‐induced transcriptional activation of the RNA‐binding protein maternal expression at 31B (Me31b)/RCK, resulting in stabilization of mRNA‐encoding a lipolytic hormone, Akh. In mouse adipocytes, Hpo/Mst1 mediated adipocyte differentiation, phosphorylation of RNA‐binding proteins such as Rck, decapping MRNA 2 (Dcp2), enhancer Of MRNA decapping 3 (Edc3), nucleolin (NCL), and glucagon mRNA stability by interacting with Rck. Decreased lifespan in Hpo‐overexpressing Drosophila lines required expression of Me31b, but not DCP2, which was potentially mediated by recovering expression of lipid metabolic genes and formation of lipid droplets. Taken together, our findings suggest that Hpo/Mst1 plays a conserved role in longevity by regulating adipogenesis and fatty acid metabolism.

Eukaryotic RNA synthesis and degradation are intricately regulated, impacting on gene expression dynamics. RNA stability varies in individual transcripts and is modulated by trans-acting factors such as microRNAs, long noncoding RNAs, and RNA-binding proteins, which determine protein output and subsequent cellular processes. To measure RNA decay rate, accurate and reliable methodologies are essential in the field of RNA biology. Transcription inhibition and metabolic labeling enable comprehensive investigations on RNA decay, offering critical insights into dynamic regulation of RNA decay. Transcription shut-off has been employed widely by using various approaches, such as treatment with chemical inhibitors or generation of temperature-sensitive mutants of RNA polymerases. However, it has limitations, providing a static view and lacking real-time dynamics as well as precise measurement of decay rate. Metabolic labeling, a method of incorporating modified nucleotides into RNA transcripts, complements shut-off approaches, allowing selective monitoring of newly synthesized RNA and tracing decay intermediates. The purpose of the protocol described in this chapter is to assess the kinetics and statics of newly synthesized RNA and its decay by 5-ethynyl uridine labeling.

Argonaute (AGO), a component of RNA-induced silencing complexes (RISCs), is a representative RNA-binding protein (RBP) known to bind with mature microRNAs (miRNAs) and is directly involved in post-transcriptional gene silencing. However, despite the biological significance of miRNAs, the roles of other miRNA-binding proteins (miRBPs) remain unclear in the regulation of miRNA loading, dissociation from RISCs and extracellular release. In this study, we performed protein arrays to profile miRBPs and identify 118 RBPs that directly bind to miRNAs. Among those proteins, the RBP quaking (QKI) inhibits extracellular release of the mature microRNA let-7b by controlling the loading of let-7b into extracellular vesicles via additional miRBPs such as AUF1 (also known as hnRNPD) and hnRNPK. The enhanced extracellular release of let-7b after QKI depletion activates Toll-like receptor 7 (TLR7) and promotes the production of proinflammatory cytokines in recipient cells, leading to brain inflammation in the mouse cortex. Thus, this study reveals the contribution of QKI to the inhibition of brain inflammation via regulation of extracellular let-7b release.