Je-Hyun Yoon

Posttranscriptional Gene Regulation by Long Noncoding RNA

Eukaryotic cells transcribe a vast number of noncoding RNA species. Among them, long noncoding RNAs (lncRNAs) have been widely implicated in the regulation of gene transcription. However, examples of posttranscriptional gene regulation by lncRNAs are emerging. Through extended base-pairing, lncRNAs can stabilize or promote the translation of target mRNAs, while partial base-pairing facilitates mRNA decay or inhibits target mRNA translation. In the absence of complementarity, lncRNAs can suppress precursor mRNA splicing and translation by acting as decoys of RNA-binding proteins or microRNAs and can compete for microRNA-mediated inhibition leading to increased expression of the mRNA. Through these regulatory mechanisms, lncRNAs can elicit differentiation, proliferation, and cytoprotective programs, underscoring the rising recognition of lncRNA roles in human disease. In this review, we summarize the mechanisms of posttranscriptional gene regulation by lncRNAs identified until now.

Functional interactions among microRNAs and long noncoding RNAs.

In mammals, the vast majority of transcripts expressed are noncoding RNAs, ranging from short RNAs (including microRNAs) to long RNAs spanning up to hundreds of kb. While the actions of microRNAs as destabilizers and repressors of the translation of protein-coding transcripts (mRNAs) have been studied in detail, the influence of microRNAs on long noncoding RNA (lncRNA) function is only now coming into view. Conversely, the influence of lncRNAs upon microRNA function is also rapidly emerging. In some cases, lncRNA stability is reduced through the interaction of specific miRNAs. In other cases, lncRNAs can act as microRNA decoys, with the sequestration of microRNAs favoring expression of repressed target mRNAs. Other lncRNAs derepress gene expression by competing with miRNAs for interaction with shared target mRNAs. Finally, some lncRNAs can produce miRNAs, leading to repression of target mRNAs. These microRNA-lncRNA regulatory paradigms modulate gene expression patterns that drive major cellular processes (such as cell differentiation, proliferation, and cell death) which are central to mammalian physiologic and pathologic processes. We review and summarize the types of microRNA-lncRNA crosstalk identified to-date and discuss their influence on gene expression programs.

Scaffold function of long non-coding RNA HOTAIR in protein ubiquitination

Although mammalian long non-coding (lnc)RNAs are best known for modulating transcription, their post-transcriptional influence on mRNA splicing, stability and translation is emerging. Here we report a post-translational function for the lncRNA HOTAIR as an inducer of ubiquitin-mediated proteolysis. HOTAIR associates with E3 ubiquitin ligases bearing RNA-binding domains, Dzip3 and Mex3b, as well as with their respective ubiquitination substrates, Ataxin-1 and Snurportin-1. In this manner, HOTAIR facilitates the ubiquitination of Ataxin-1 by Dzip3 and Snurportin-1 by Mex3b in cells and in vitro, and accelerates their degradation. HOTAIR levels are highly upregulated in senescent cells, causing rapid decay of targets Ataxin-1 and Snurportin-1, and preventing premature senescence. These results uncover a role for a lncRNA, HOTAIR, as a platform for protein ubiquitination.

Senescence‐associated lncRNAs: senescence‐associated long noncoding RNAs

Noncoding RNAs include small transcripts, such as microRNAs and piwi‐interacting RNAs, and a wide range of long noncoding RNAs (lncRNAs). Although many lncRNAs have been identified, only a small number of lncRNAs have been characterized functionally. Here, we sought to identify lncRNAs differentially expressed during replicative senescence. We compared lncRNAs expressed in proliferating, early‐passage, ‘young’ human diploid WI‐38 fibroblasts [population doubling (PDL) 20] with those expressed in senescent, late‐passage, ‘old’ fibroblasts (PDL 52) by RNA sequencing (RNA‐Seq). Numerous transcripts in all lncRNA groups (antisense lncRNAs, pseudogene‐encoded lncRNAs, previously described lncRNAs and novel lncRNAs) were validated using reverse transcription (RT) and real‐time, quantitative (q)PCR. Among the novel senescence‐associated lncRNAs (SAL‐RNAs) showing lower abundance in senescent cells, SAL‐RNA1 (XLOC_023166) was found to delay senescence, because reducing SAL‐RNA1 levels enhanced the appearance of phenotypic traits of senescence, including an enlarged morphology, positive β‐galactosidase activity, and heightened p53 levels. Our results reveal that the expression of known and novel lncRNAs changes with senescence and suggests that SAL‐RNAs play direct regulatory roles in this important cellular process.

PAR-CLIP analysis uncovers AUF1 impact on target RNA fate and genome integrity

Post-transcriptional gene regulation is robustly regulated by RNA-binding proteins (RBPs). Here we describe the collection of RNAs regulated by AUF1 (AU-binding factor 1), an RBP linked to cancer, inflammation and aging. Photoactivatable ribonucleoside-enhanced crosslinking and immunoprecipitation (PAR-CLIP) analysis reveals that AUF1 primarily recognizes U-/GU-rich sequences in mRNAs and noncoding RNAs and influences target transcript fate in three main directions. First, AUF1 lowers the steady-state levels of numerous target RNAs, including long noncoding RNA NEAT1, in turn affecting the organization of nuclear paraspeckles. Second, AUF1 does not change the abundance of many target RNAs, but ribosome profiling reveals that AUF1 promotes the translation of numerous mRNAs in this group. Third, AUF1 unexpectedly enhances the steady-state levels of several target mRNAs encoding DNA-maintenance proteins. Through its actions on target RNAs, AUF1 preserves genomic integrity, in agreement with the AUF1-elicited prevention of premature cellular senescence.

MicroRNA 33 Regulates Glucose Metabolism

ABSTRACT Metabolic diseases are characterized by the failure of regulatory genes or proteins to effectively orchestrate specific pathways involved in the control of many biological processes. In addition to the classical regulators, recent discoveries have shown the remarkable role of small noncoding RNAs (microRNAs [miRNAs]) in the posttranscriptional regulation of gene expression. In this regard, we have recently demonstrated that miR-33a and miR33b, intronic miRNAs located within the sterol regulatory element-binding protein (SREBP) genes, regulate lipid metabolism in concert with their host genes. Here, we show that miR-33b also cooperates with SREBP1 in regulating glucose metabolism by targeting phosphoenolpyruvate carboxykinase (PCK1) and glucose-6-phosphatase (G6PC), key regulatory enzymes of hepatic gluconeogenesis. Overexpression of miR-33b in human hepatic cells inhibits PCK1 and G6PC expression, leading to a significant reduction of glucose production. Importantly, hepatic SREBP1c/miR-33b levels correlate inversely with the expression of PCK1 and G6PC upon glucose infusion in rhesus monkeys. Taken together, these results suggest that miR-33b works in concert with its host gene to ensure a fine-tuned regulation of lipid and glucose homeostasis, highlighting the clinical potential of miR-33a/b as novel therapeutic targets for a range of metabolic diseases.

7SL RNA represses p53 translation by competing with HuR

Noncoding RNAs (ncRNAs) and RNA-binding proteins are potent post-transcriptional regulators of gene expression. The ncRNA 7SL is upregulated in cancer cells, but its impact upon the phenotype of cancer cells is unknown. Here, we present evidence that 7SL forms a partial hybrid with the 3′-untranslated region (UTR) of TP53 mRNA, which encodes the tumor suppressor p53. The interaction of 7SL with TP53 mRNA reduced p53 translation, as determined by analyzing p53 expression levels, nascent p53 translation and TP53 mRNA association with polysomes. Silencing 7SL led to increased binding of HuR to TP53 mRNA, an interaction that led to the promotion of p53 translation and increased p53 abundance. We propose that the competition between 7SL and HuR for binding to TP53 3′UTR contributes to determining the magnitude of p53 translation, in turn affecting p53 levels and the growth-suppressive function of p53. Our findings suggest that targeting 7SL may be effective in the treatment of cancers with reduced p53 levels.

HuR and GRSF1 modulate the nuclear export and mitochondrial localization of the lncRNA RMRP

Some mitochondrial long noncoding RNAs (lncRNAs) are encoded by nuclear DNA, but the mechanisms that mediate their transport to mitochondria are poorly characterized. Using affinity RNA pull-down followed by mass spectrometry analysis, we found two RNA-binding proteins (RBPs), HuR (human antigen R) and GRSF1 (G-rich RNA sequence-binding factor 1), that associated with the nuclear DNA-encoded lncRNA RMRP and mobilized it to mitochondria. In cultured human cells, HuR bound RMRP in the nucleus and mediated its CRM1 (chromosome region maintenance 1)-dependent export to the cytosol. After RMRP was imported into mitochondria, GRSF1 bound RMRP and increased its abundance in the matrix. Loss of GRSF1 lowered the mitochondrial levels of RMRP, in turn suppressing oxygen consumption rates and modestly reducing mitochondrial DNA replication priming. Our findings indicate that RBPs HuR and GRSF1 govern the cytoplasmic and mitochondrial localization of the lncRNA RMRP, which is encoded by nuclear DNA but has key functions in mitochondria.

Long noncoding RNAs in diseases of aging

Aging is a process during which progressive deteriorating of cells, tissues, and organs over time lead to loss of function, disease, and death. Towards the goal of extending human health span, there is escalating interest in understanding the mechanisms that govern aging-associated pathologies. Adequate regulation of expression of coding and noncoding genes is critical for maintaining organism homeostasis and preventing disease processes. Long noncoding RNAs (lncRNAs) are increasingly recognized as key regulators of gene expression at all levels--transcriptional, post-transcriptional and post-translational. In this review, we discuss our emerging understanding of lncRNAs implicated in aging illnesses. We focus on diseases arising from age-driven impairment in energy metabolism (obesity, diabetes), the declining capacity to respond homeostatically to proliferative and damaging stimuli (cancer, immune dysfunction), and neurodegeneration. We identify the lncRNAs involved in these ailments and discuss the rising interest in lncRNAs as diagnostic and therapeutic targets to ameliorate age-associated pathologies and prolong health. This article is part of a Special Issue entitled: Clues to long noncoding RNA taxonomy1, edited by Dr. Tetsuro Hirose and Dr. Shinichi Nakagawa.

Tyrosine phosphorylation of HuR by JAK3 triggers dissociation and degradation of HuR target mRNAs

In response to stress conditions, many mammalian mRNAs accumulate in stress granules (SGs) together with numerous RNA-binding proteins that control mRNA turnover and translation. However, the signaling cascades that modulate the presence of ribonucleoprotein (RNP) complexes in SGs are poorly understood. Here, we investigated the localization of human antigen R (HuR), an mRNA-stabilizing RNA-binding protein, in SGs following exposure to the stress agent arsenite. Unexpectedly, the mobilization of HuR to SGs was prevented through the activation of Janus kinase 3 (JAK3) by the vitamin K3 analog menadione. JAK3 phosphorylated HuR at tyrosine 200, in turn inhibiting HuR localization in SGs, reducing HuR interaction with targets SIRT1 and VHL mRNAs, and accelerating target mRNA decay. Our findings indicate that HuR is tyrosine-phosphorylated by JAK3, and link this modification to HuR subcytoplasmic localization and to the fate of HuR target mRNAs.