Metabolic Enzymes Enjoying New Partnerships as RNA-Binding Proteins
2015; Elsevier BV; Volume: 26; Issue: 12 Linguagem: Inglês
10.1016/j.tem.2015.09.012
ISSN1879-3061
AutoresAlfredo Castelló, Matthias W. Hentze, Thomas Preiß,
Tópico(s)RNA and protein synthesis mechanisms
ResumoGenetic control of metabolism is currently best understood at the level of transcription and epigenetics. Only limited information is available on post-transcriptional regulation of metabolism. While a few metabolic enzymes were previously known to moonlight as RNA-binding proteins in physiologically relevant contexts, recent discoveries highlight that several dozen of metabolic enzymes belonging to a wide spectrum of pathways exhibit RNA-binding activity in living mammalian cells. Abundant RNA–enzyme interactions might suggest novel roles of RNA in affecting enzyme function, for instance, as competitive inhibitors or allosteric regulators. A function of RNA as assembly scaffold for enzyme complexes is also conceivable, with potentially wide-ranging implications for our understanding of how cells organize and control metabolic flux. Finally, enzymes can moonlight as regulators of (m)RNAs, as exemplified by aconitase/IRP1 and GAPDH. In the past century, few areas of biology advanced as much as our understanding of the pathways of intermediary metabolism. Initially considered unimportant in terms of gene regulation, crucial cellular fate changes, cell differentiation, or malignant transformation are now known to involve 'metabolic remodeling' with profound changes in the expression of many metabolic enzyme genes. This review focuses on the recent identification of RNA-binding activity of numerous metabolic enzymes. We discuss possible roles of this unexpected second activity in feedback gene regulation ('moonlighting') and/or in the control of enzymatic function. We also consider how metabolism-driven post-translational modifications could regulate enzyme–RNA interactions. Thus, RNA emerges as a new partner of metabolic enzymes with far-reaching possible consequences to be unraveled in the future. In the past century, few areas of biology advanced as much as our understanding of the pathways of intermediary metabolism. Initially considered unimportant in terms of gene regulation, crucial cellular fate changes, cell differentiation, or malignant transformation are now known to involve 'metabolic remodeling' with profound changes in the expression of many metabolic enzyme genes. This review focuses on the recent identification of RNA-binding activity of numerous metabolic enzymes. We discuss possible roles of this unexpected second activity in feedback gene regulation ('moonlighting') and/or in the control of enzymatic function. We also consider how metabolism-driven post-translational modifications could regulate enzyme–RNA interactions. Thus, RNA emerges as a new partner of metabolic enzymes with far-reaching possible consequences to be unraveled in the future. Metabolic enzymes were long considered to be constitutively expressed housekeeping proteins, and even nowadays glyceraldehyde-3-phosphate dehydrogenase (GAPDH) mRNA continues to be broadly used for normalization of real-time quantitative PCR experiments. However, this traditional view is challenged by advances in many areas, including developmental, cancer, and stem cell biology. The expression profiles of metabolic enzymes are controlled by cell identity, which enables tissue metabolic specialization. Furthermore, metabolic enzyme expression is also subject to fine-tuning temporal regulation in response to feast/famine and to day/night cycles (reviewed in [1Hong S.H. et al.Nuclear receptors and metabolism: from feast to famine.Diabetologia. 2014; 57: 860-867Crossref PubMed Scopus (26) Google Scholar, 2Zhao X. et al.Nuclear receptors rock around the clock.EMBO Rep. 2014; 15: 518-528Crossref PubMed Scopus (49) Google Scholar], respectively). The discovery of the nuclear hormone receptors (NHRs) in the 1980s represented a breakthrough in the understanding of the transcriptional control of metabolic networks. NHRs represent an extended family of ligand-responsive DNA-binding proteins that, upon activation, can switch transcriptional programs in cooperation with coactivators or corepressors [3Privalsky M.L. The role of corepressors in transcriptional regulation by nuclear hormone receptors.Annu. Rev. Physiol. 2004; 66: 315-360Crossref PubMed Scopus (258) Google Scholar]. NHRs are transcriptional master regulators of metabolism by altering the metabolic enzyme profiles in response to feeding and fasting as well as circadian signaling. An illustrative example is the role of NHRs in liver metabolism. Secretion of cortisol from the adrenal gland during prolonged starvation induces the activation of the glucocorticoid receptor in the liver. This leads to the transcription of two master regulators of sugar metabolism, glucose-6-phosphatase (G6PC) and phosphoenolpyruvate carboxykinase (PECK), which promote the synthesis of glucose via gluconeogenesis [1Hong S.H. et al.Nuclear receptors and metabolism: from feast to famine.Diabetologia. 2014; 57: 860-867Crossref PubMed Scopus (26) Google Scholar, 4Liu Y. et al.Reduction of hepatic glucocorticoid receptor and hexose-6-phosphate dehydrogenase expression ameliorates diet-induced obesity and insulin resistance in mice.J. Mol. Endocrinol. 2008; 41: 53-64Crossref PubMed Scopus (47) Google Scholar]. By contrast, liver X receptors (LXRs) and farnesoid X receptor (FXR) are activated by feeding-induced synthesis of their respective ligands, oxysterols and bile acid. In antagonism to fasting-activated NHRs, both LXRs and FXR suppress gluconeogenesis by upregulating the expression of glucokinase, which promotes glucose utilization, and by increasing glycogen synthesis [5Laffitte B.A. et al.Activation of liver X receptor improves glucose tolerance through coordinate regulation of glucose metabolism in liver and adipose tissue.Proc. Natl. Acad. Sci. U.S.A. 2003; 100: 5419-5424Crossref PubMed Scopus (419) Google Scholar, 6Renga B. et al.Glucocorticoid receptor mediates the gluconeogenic activity of the farnesoid X receptor in the fasting condition.FASEB J. 2012; 26: 3021-3031Crossref PubMed Scopus (43) Google Scholar, 7Zhang Y. et al.Activation of the nuclear receptor FXR improves hyperglycemia and hyperlipidemia in diabetic mice.Proc. Natl. Acad. Sci. U.S.A. 2006; 103: 1006-1011Crossref PubMed Scopus (716) Google Scholar]. 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Metabolic enzymes that bind RNA: yet another level of cellular regulatory network?.Acta Biochim. Pol. 2006; 53: 11-32Crossref PubMed Scopus (86) Google Scholar]. For example, thymidine synthase (TYMS) can bind and inhibit the translation of its own RNA when the levels of its substrates are low, establishing a negative feedback loop [13Chu E. et al.Autoregulation of human thymidylate synthase messenger RNA translation by thymidylate synthase.Proc. Natl. Acad. Sci. U.S.A. 1991; 88: 8977-8981Crossref PubMed Scopus (331) Google Scholar, 14Chu E. et al.Regulation of thymidylate synthase in human colon cancer cells treated with 5-fluorouracil and interferon-gamma.Mol. Pharmacol. 1993; 43: 527-533PubMed Google Scholar, 15Chu E. et al.Identification of a thymidylate synthase ribonucleoprotein complex in human colon cancer cells.Mol. Cell. Biol. 1994; 14: 207-213Crossref PubMed Scopus (84) Google Scholar]. Conceptually, such a mechanism represents a simple yet effective way to adjust to conditions when the enzyme is not required. In this review, we discuss the emerging roles of protein–RNA interactions in controlling metabolism. Over the past three decades, sporadic reports have shown that metabolic enzymes can moonlight as RNA-binding proteins (RBPs) and, in some instances, regulate the expression of their target mRNAs [12Ciesla J. Metabolic enzymes that bind RNA: yet another level of cellular regulatory network?.Acta Biochim. Pol. 2006; 53: 11-32Crossref PubMed Scopus (86) Google Scholar, 16Hentze M.W. Enzymes as RNA-binding proteins: a role for (di)nucleotide-binding domains?.Trends Biochem. Sci. 1994; 19: 101-103Abstract Full Text PDF PubMed Scopus (123) Google Scholar, 17Hentze M.W. Preiss T. The REM phase of gene regulation.Trends Biochem. Sci. 2010; 35: 423-426Abstract Full Text Full Text PDF PubMed Scopus (82) Google Scholar] (Table 1). These moonlighting enzymes (see Glossary) participate in varied metabolic pathways, such as glycolysis, the tricarboxylic acid (TCA) cycle, lipid metabolism, and deoxynucleotide biosynthesis, and catalyze different reactions. In most cases, RNA binding was observed in vitro, using filter binding or electrophoretic mobility shift assays [12Ciesla J. Metabolic enzymes that bind RNA: yet another level of cellular regulatory network?.Acta Biochim. Pol. 2006; 53: 11-32Crossref PubMed Scopus (86) Google Scholar, 15Chu E. et al.Identification of a thymidylate synthase ribonucleoprotein complex in human colon cancer cells.Mol. Cell. Biol. 1994; 14: 207-213Crossref PubMed Scopus (84) Google Scholar, 18Chu E. et al.Identification of an RNA binding site for human thymidylate synthase.Proc. Natl. Acad. Sci. 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While most of the reported moonlighting metabolic enzymes still await validation in living cells and animals, the functions and modes of RNA binding of aconitase 1 (ACO1, also known as iron regulatory protein 1, IRP1), GAPDH, and TYMS have been explored by biophysical and structural approaches [21Walden W.E. et al.Structure of dual function iron regulatory protein 1 complexed with ferritin IRE–RNA.Science. 2006; 314: 1903-1908Crossref PubMed Scopus (229) Google Scholar], and investigated in cellular and animal models as described later [22Zhang Y. et al.Interaction between thymidylate synthase and its cognate mRNA in zebrafish embryos.PLoS ONE. 2010; 5: e10618Crossref PubMed Scopus (12) Google Scholar, 23Galy B. et al.Iron regulatory proteins are essential for intestinal function and control key iron absorption molecules in the duodenum.Cell Metab. 2008; 7: 79-85Abstract Full Text Full Text PDF PubMed Scopus (151) Google Scholar, 24Galy B. et al.Iron homeostasis in the brain: complete iron regulatory protein 2 deficiency without symptomatic neurodegeneration in the mouse.Nat. Genet. 2006; 38 (discussion 969–970): 967-969Crossref PubMed Scopus (54) Google Scholar, 25Chang C.H. et al.Posttranscriptional control of T cell effector function by aerobic glycolysis.Cell. 2013; 153: 1239-1251Abstract Full Text Full Text PDF PubMed Scopus (1323) Google Scholar]. Insights from these examples form the basis of the 'REM (RNA–enzyme–metabolite) hypothesis', which proposes the existence of regulatory links between gene expression and intermediary metabolism mediated by moonlighting RNA-binding metabolic enzymes [17Hentze M.W. Preiss T. The REM phase of gene regulation.Trends Biochem. Sci. 2010; 35: 423-426Abstract Full Text Full Text PDF PubMed Scopus (82) Google Scholar].Table 1Examples of Metabolic Enzymes Identified as RBPs in the RNA Interactome StudiesGene NameComplete NameFunctionDi/mononucleotide BindingHeLa RNA InteractomeHEK293 RNA InteractomemESC RNA InteractomeADKAdenylate kinaseAMP biosynthesisATP and adenosineYesALDH18A1Delta-1-pyrroline-5-carboxylate synthaseBiosynthesis of proline, ornithine, and arginineATP and NADPYesALDH6A1Methylmalonate-semialdehyde dehydrogenase (acylating), mitochondrialValine and pyrimidine metabolismNAD(P)/HYesALDOAFructose-bisphosphate aldolase AGlycolysisYesASS1Argininosuccinate synthasel-Arginine biosynthesisATPYesCCBL2Kynurenine–oxoglutarate transaminase 3Transaminase activity for several amino acidsYesCSCitrate synthase, mitochondrialTCA cycleYesDUTDeoxyuridine 5′-triphosphate nucleotidohydrolase, mitochondrialNucleotide metabolismdUTPYesYesENO1α-EnolaseGlycolysisYesYesFASNFatty acid synthaseFatty acid synthesisNADP/HYesYesFDPSFarnesyl pyrophosphate synthaseFormation of farnesyl diphosphateYesGOT2Aspartate aminotransferase, mitochondrialAmino acid metabolismYesHADHBTrifunctional enzyme subunit beta, mitochondrialbeta-Oxidation of fatty acidsYesHK2Hexokinase-2GlycolysisATPYesHSD17B103-Hydroxyacyl-CoA dehydrogenase type-2β-Oxidation at position 17 of androgens and estrogensNAD/NAD(P)YesLTA4HLeukotriene A4 hydrolaseBiosynthesis of leukotriene B4YesMDH2Malate dehydrogenase 2, mitochondrialTCA cycleNAD/HYesYesNME1Nucleoside diphosphate kinase ASynthesis of nucleoside triphosphatesATPYesNQO1NAD(P)H dehydrogenase (quinone) 1Detoxification pathways and vitamin K-dependent γ-carboxylation of glutamate residuesNAD(P)HYesPKM2Pyruvate kinaseGlycolysisATPYesYesPPP1CCSerine/threonine–protein phosphatase 1–γ catalytic subunitGlycogen metabolism, muscle contractility, and protein synthesisYesSUCLG1Succinyl-CoA ligase (ADP/GDP-forming) subunit α, mitochondrialTCA cycleATP/GTPYesTPI1Triosephosphate isomeraseGlycolysis and gluconeogenesisYes Open table in a new tab Recent system-wide approaches have been developed to identify a (near) complete compendium of RBPs. Initially, two parallel works used Saccharomyces cerevisiae proteome-wide protein arrays to interrogate protein binding to RNA in vitro. These studies catalogued 180 [26Scherrer T. et al.A screen for RNA-binding proteins in yeast indicates dual functions for many enzymes.PLoS ONE. 2010; 5: e15499Crossref PubMed Scopus (108) Google Scholar] and 42 proteins [27Tsvetanova N.G. et al.Proteome-wide search reveals unexpected RNA-binding proteins in Saccharomyces cerevisiae.PLoS ONE. 2010; 5: e12671Crossref PubMed Scopus (131) Google Scholar], respectively, as putative RBPs, including many not previously known to interact with RNA. Among the dozen metabolic enzymes reliably associated with RNA in vitro, oxidoreductases and proteins involved in lipid metabolism were the most prominent classes of putative moonlighting metabolic enzymes. The peroxisomal malate dehydrogenase (MDH3) was identified in both studies as an RBP; immunoprecipitation followed by microarray (RIP-Chip) showed modest RNA-binding capacity towards a limited pool of target RNAs [26Scherrer T. et al.A screen for RNA-binding proteins in yeast indicates dual functions for many enzymes.PLoS ONE. 2010; 5: e15499Crossref PubMed Scopus (108) Google Scholar]. Because the peroxisome is not an organelle classically associated with RNA biology, these results called for further experimental validation in cellular models. To address the technical limitations of in vitro RBP identification screens, two groups developed in parallel a new approach named RNA interactome capture (Figure 1). Applying UV crosslinking to proliferative cell monolayers, followed by stringent denaturing oligo(dT) isolation of protein–RNA complexes and quantitative mass spectrometry, these studies identified a total of 1106 high-confidence RBPs in HeLa and HEK293 cells [28Baltz A.G. et al.The mRNA-bound proteome and its global occupancy profile on protein-coding transcripts.Mol. Cell. 2012; 46: 674-690Abstract Full Text Full Text PDF PubMed Scopus (800) Google Scholar, 29Castello A. et al.Insights into RNA biology from an atlas of mammalian mRNA-binding proteins.Cell. 2012; 149: 1393-1406Abstract Full Text Full Text PDF PubMed Scopus (1353) Google Scholar]. Notably, hundreds of them were novel RNA interactors and lacked known RNA-binding domains (RBDs). This method offers several advantages over previous approaches: (i) UV light promotes free radical formation at the nucleotide base that can establish covalent bonds only with amino acids placed at 'zero distance' (≤2 Å); (ii) UV crosslinking does not promote protein–protein crosslinks; (iii) because UV is applied directly to living cells, hybridization with oligo(dT) captures native protein–RNA complexes; (iv) nucleic acid hybridization is compatible with high salt and denaturing agents including chaotropic detergents, thus allowing stringent removal of noncovalent binders; and (v) to qualify as high-confidence RBP, quantitative information and rigorous statistic methods are applied [29Castello A. et al.Insights into RNA biology from an atlas of mammalian mRNA-binding proteins.Cell. 2012; 149: 1393-1406Abstract Full Text Full Text PDF PubMed Scopus (1353) Google Scholar, 30Castello A. et al.System-wide identification of RNA-binding proteins by interactome capture.Nat. Protoc. 2013; 8: 491-500Crossref PubMed Scopus (137) Google Scholar]. Among the newly identified RBP classes, the RNA interactome studies reported 23 distinct metabolic enzymes associated with polyadenylated RNAs [28Baltz A.G. et al.The mRNA-bound proteome and its global occupancy profile on protein-coding transcripts.Mol. Cell. 2012; 46: 674-690Abstract Full Text Full Text PDF PubMed Scopus (800) Google Scholar, 29Castello A. et al.Insights into RNA biology from an atlas of mammalian mRNA-binding proteins.Cell. 2012; 149: 1393-1406Abstract Full Text Full Text PDF PubMed Scopus (1353) Google Scholar, 31Kwon S.C. et al.The RNA-binding protein repertoire of embryonic stem cells.Nat. Struct. Mol. Biol. 2013; 20: 1122-1130Crossref PubMed Scopus (332) Google Scholar] (Table 1), suggesting that the interplay between RNA and metabolism is broader than previously realized and supporting the REM network hypothesis. Among these moonlighters, aldolase and trifunctional enzyme subunit β (HADHB) had previously been recognized to bind RNA in vitro [12Ciesla J. Metabolic enzymes that bind RNA: yet another level of cellular regulatory network?.Acta Biochim. Pol. 2006; 53: 11-32Crossref PubMed Scopus (86) Google Scholar] and the interaction of enolase 1 (ENO1), hydroxymethyltransferase (SHMT1), and pyruvate kinase M2 (PKM2) with RNA was validated in cells by an independent approach [29Castello A. et al.Insights into RNA biology from an atlas of mammalian mRNA-binding proteins.Cell. 2012; 149: 1393-1406Abstract Full Text Full Text PDF PubMed Scopus (1353) Google Scholar, 30Castello A. et al.System-wide identification of RNA-binding proteins by interactome capture.Nat. Protoc. 2013; 8: 491-500Crossref PubMed Scopus (137) Google Scholar]. Applying UV crosslinking, immunoprecipitation, and RNA sequencing (CLIPseq), it was shown that ENO1 and SHMT2 associate with hundreds of different mRNAs in HeLa cells, but display distinct binding patterns from each other, suggesting selectivity of binding [29Castello A. et al.Insights into RNA biology from an atlas of mammalian mRNA-binding proteins.Cell. 2012; 149: 1393-1406Abstract Full Text Full Text PDF PubMed Scopus (1353) Google Scholar]. In agreement, bacterial enolase has been recently identified, together with the RNase E, as a part of the degradosome complex, which suggests that the relationship of this enzyme with RNA is already observable in prokaryotes [32Ait-Bara S. Carpousis A.J. RNA degradosomes in bacteria and chloroplasts: classification, distribution and evolution of RNase E homologs.Mol. Microbiol. 2015; 97: 1021-1035Crossref PubMed Scopus (68) Google Scholar]. Although the 23 moonlighting enzymes identified by the RNA interactome studies belong to different metabolic pathways and catalyze distinct reactions, 13 of them bind either dinucleotides or mononucleotides (Table 1). This suggests that protein domains commonly involved in nucleotide binding, such as the Rossmann fold, may represent suitable protein surfaces to interact with RNA, as discussed in more detail later. Interestingly, some of the already known and newly discovered moonlighting RBPs are linked to hereditary diseases. Mutations in inosine 5′-monophosphate dehydrogenase 1 (IMPDH1), a dual RNA-binding and dinucleotide-binding enzyme [33McLean J.E. et al.Inosine 5′-monophosphate dehydrogenase binds nucleic acids in vitro and in vivo.Biochem. J. 2004; 379: 243-251Crossref PubMed Scopus (76) Google Scholar], cause retinitis pigmentosa [34Hedstrom L. 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IMPDH is involved in the post-transcriptional regulation of rhodopsin mRNA and this disease-associated mutation reduces its association with polysomes and thus its translation efficiency [37McGrew D.A. Hedstrom L. Towards a pathological mechanism for IMPDH1-linked retinitis pigmentosa.Adv. Exp. Med. Biol. 2012; 723: 539-545Crossref PubMed Scopus (14) Google Scholar]. Retinitis pigmentosa is also caused by mutations in components of the splicing machinery, such as U4/U6 small nuclear ribonucleoprotein Prp3 (PRPF3), PRPF8, and PRPF31, suggesting a considerable role of RNA biology in this disease [38Castello A. et al.RNA-binding proteins in Mendelian disease.Trends Genet. 2013; 29: 318-327Abstract Full Text Full Text PDF PubMed Scopus (174) Google Scholar]. The mitochondrial enzyme 17β-hydroxysteroid dehydrogenase 10 (HSD17B10; also known as MRPP2), catalyzes the dehydrogenation of 17-hydroxysteroids in steroidogenesis. However, it was catalogued as an RBP in HEK293 cells [28Baltz A.G. et al.The mRNA-bound proteome and its global occupancy profile on protein-coding transcripts.Mol. Cell. 2012; 46: 674-690Abstract Full Text Full Text PDF PubMed Scopus (800) Google Scholar] and also moonlights as a component of mitochondrial ribonuclease P, which is involved in the processing of the mitochondrial tRNAs [39Holzmann J. et al.RNase P without RNA: identification and functional reconstitution of the human mitochondrial tRNA processing enzyme.Cell. 2008; 135: 462-474Abstract Full Text Full Text PDF PubMed Scopus (432) Google Scholar]. HSD17B10 deficiency causes neurodegeneration in humans and has been associated with Alzheimer's disease. Curiously, there is no correlation between the degree of catalytic activity of the disease-associated mutant enzymes and the severity of the disease [40Rauschenberger K. et al.A non-enzymatic function of 17beta-hydroxysteroid dehydrogenase type 10 is required for mitochondrial integrity and cell survival.EMBO Mol. Med. 2010; 2: 51-62Crossref PubMed Scopus (61) Google Scholar], suggesting that the molecular mechanism underlying this pathology does not primarily derive from the catalytic activity of HSD17B10. Indeed, a recent study revealed that knock-down or mutation of HSD17D10 induces a defect in the processing of the heavy strand of the mitochondrial RNA [41Deutschmann A.J. et al.Mutation or knock-down of 17beta-hydroxysteroid dehydrogenase type 10 cause loss of MRPP1 and impaired processing of mitochondrial heavy strand transcripts.Hum. Mol. Genet. 2014; 23: 3618-3628Crossref PubMed Scopus (45) Google Scholar]. 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RNA stem loop structures termed iron-responsive elements (IREs) were first found in the 5′ untranslated regions (UTRs) of ferritin mRNAs [46Hentze M.W. et al.Identification of the iron-responsive element for the translational regulation of human ferritin mRNA.Science. 1987; 238: 1570-1573Crossref PubMed Scopus (390) Google Scholar] and in the 3′ UTR of transferrin receptor mRNA [47Casey J.L. et al.Iron-responsive elements: regulatory RNA sequences that control mRNA levels and translation.Science. 1988; 240: 924-928Crossref PubMed Scopus (507) Google Scholar, 48Mullner E.W. Kuhn L.C. A stem-loop in the 3′ untranslated region mediates iron-dependent regulation of transferrin receptor mRNA stability in the cytoplasm.Cell. 1988; 53: 815-825Abstract Full Text PDF PubMed Scopus (375) Google Scholar] (Figure 2). Specific IRE-binding proteins were identified [49Leibold E.A. Munro H.N. 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Both proteins are broadly expressed across tissues and single knockout mice are viable, while simultaneous knockout of both IRPs is early embryonic lethal, indicating essential but largely redundant functions. Nevertheless, the single knockout phenotypes also demonstrate specific roles for IRP1 in erythropoiesis and the cardiovascular system, while IRP2 is of particular importa
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