RESUMO
Aberrantly accumulated metabolites elicit intra- and inter-cellular pro-oncogenic cascades, yet current measurement methods require sample perturbation/disruption and lack spatio-temporal resolution, limiting our ability to fully characterize their function and distribution. Here, we show that Raman spectroscopy (RS) can directly detect fumarate in living cells in vivo and animal tissues ex vivo, and that RS can distinguish between Fumarate hydratase (Fh1)-deficient and Fh1-proficient cells based on fumarate concentration. Moreover, RS reveals the spatial compartmentalization of fumarate within cellular organelles in Fh1-deficient cells: consistent with disruptive methods, we observe the highest fumarate concentration (37 ± 19 mM) in mitochondria, where the TCA cycle operates, followed by the cytoplasm (24 ± 13 mM) and then the nucleus (9 ± 6 mM). Finally, we apply RS to tissues from an inducible mouse model of FH loss in the kidney, demonstrating RS can classify FH status. These results suggest RS could be adopted as a valuable tool for small molecule metabolic imaging, enabling in situ non-destructive evaluation of fumarate compartmentalization.
Assuntos
Fumarato Hidratase , Fumaratos , Análise Espectral Raman , Análise Espectral Raman/métodos , Animais , Fumaratos/metabolismo , Camundongos , Fumarato Hidratase/metabolismo , Fumarato Hidratase/genética , Rim/metabolismo , Mitocôndrias/metabolismo , Humanos , Núcleo Celular/metabolismo , Citoplasma/metabolismoRESUMO
Mutations in metabolic enzymes are associated with hereditary and sporadic forms of cancer. For example, loss-of-function mutations affecting fumarate hydratase (FH), the tricarboxylic acid (TCA) cycle enzyme, result in the accumulation of millimolar levels of fumarate that cause an aggressive form of kidney cancer. A distinct feature of fumarate is its ability to spontaneously react with thiol groups of cysteines in a chemical reaction termed succination. Although succination of a few proteins has been causally implicated in the molecular features of FH-deficient cancers, the stoichiometry, wider functional consequences, and contribution of succination to disease development remain largely unexplored. We discuss the functional implications of fumarate-induced succination in FH-deficient cells, the available methodologies, and the current challenges in studying this post-translational modification.
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Xeroderma pigmentosum (XP) is caused by defective nucleotide excision repair of DNA damage. This results in hypersensitivity to ultraviolet light and increased skin cancer risk, as sunlight-induced photoproducts remain unrepaired. However, many XP patients also display early-onset neurodegeneration, which leads to premature death. The mechanism of neurodegeneration is unknown. Here, we investigate XP neurodegeneration using pluripotent stem cells derived from XP patients and healthy relatives, performing functional multi-omics on samples during neuronal differentiation. We show substantially increased levels of 5',8-cyclopurine and 8-oxopurine in XP neuronal DNA secondary to marked oxidative stress. Furthermore, we find that the endoplasmic reticulum stress response is upregulated and reversal of the mutant genotype is associated with phenotypic rescue. Critically, XP neurons exhibit inappropriate downregulation of the protein clearance ubiquitin-proteasome system (UPS). Chemical enhancement of UPS activity in XP neuronal models improves phenotypes, albeit inadequately. Although more work is required, this study presents insights with intervention potential.
Assuntos
Células-Tronco Pluripotentes Induzidas , Xeroderma Pigmentoso , Xeroderma Pigmentoso/patologia , Xeroderma Pigmentoso/metabolismo , Xeroderma Pigmentoso/genética , Células-Tronco Pluripotentes Induzidas/metabolismo , Humanos , Neurônios/metabolismo , Neurônios/patologia , Estresse Oxidativo , Estresse do Retículo Endoplasmático , Complexo de Endopeptidases do Proteassoma/metabolismo , Diferenciação Celular , Dano ao DNA , Modelos Biológicos , MultiômicaRESUMO
Polycystic kidney disease (PKD) is a genetic disorder characterized by bilateral cyst formation. We showed that PKD cells and kidneys display metabolic alterations, including the Warburg effect and glutaminolysis, sustained in vitro by the enzyme asparagine synthetase (ASNS). Here, we used antisense oligonucleotides (ASO) against Asns in orthologous and slowly progressive PKD murine models and show that treatment leads to a drastic reduction of total kidney volume (measured by MRI) and a prominent rescue of renal function in the mouse. Mechanistically, the upregulation of an ATF4-ASNS axis in PKD is driven by the amino acid response (AAR) branch of the integrated stress response (ISR). Metabolic profiling of PKD or control kidneys treated with Asns-ASO or Scr-ASO revealed major changes in the mutants, several of which are rescued by Asns silencing in vivo. Indeed, ASNS drives glutamine-dependent de novo pyrimidine synthesis and proliferation in cystic epithelia. Notably, while several metabolic pathways were completely corrected by Asns-ASO, glycolysis was only partially restored. Accordingly, combining the glycolytic inhibitor 2DG with Asns-ASO further improved efficacy. Our studies identify a new therapeutic target and novel metabolic vulnerabilities in PKD.
Assuntos
Aspartato-Amônia Ligase , Modelos Animais de Doenças , Doenças Renais Policísticas , Animais , Humanos , Camundongos , Aspartato-Amônia Ligase/metabolismo , Aspartato-Amônia Ligase/genética , Aspartato-Amônia Ligase/antagonistas & inibidores , Progressão da Doença , Rim/patologia , Rim/metabolismo , Oligonucleotídeos Antissenso/farmacologia , Oligonucleotídeos Antissenso/uso terapêutico , Doenças Renais Policísticas/metabolismo , Doenças Renais Policísticas/tratamento farmacológico , Doenças Renais Policísticas/patologia , Doenças Renais Policísticas/genéticaRESUMO
Knudson's "two-hit" paradigm posits that carcinogenesis requires inactivation of both copies of an autosomal tumor suppressor gene. Here, we report that the glycolytic metabolite methylglyoxal (MGO) transiently bypasses Knudson's paradigm by inactivating the breast cancer suppressor protein BRCA2 to elicit a cancer-associated, mutational single-base substitution (SBS) signature in nonmalignant mammary cells or patient-derived organoids. Germline monoallelic BRCA2 mutations predispose to these changes. An analogous SBS signature, again without biallelic BRCA2 inactivation, accompanies MGO accumulation and DNA damage in Kras-driven, Brca2-mutant murine pancreatic cancers and human breast cancers. MGO triggers BRCA2 proteolysis, temporarily disabling BRCA2's tumor suppressive functions in DNA repair and replication, causing functional haploinsufficiency. Intermittent MGO exposure incites episodic SBS mutations without permanent BRCA2 inactivation. Thus, a metabolic mechanism wherein MGO-induced BRCA2 haploinsufficiency transiently bypasses Knudson's two-hit requirement could link glycolysis activation by oncogenes, metabolic disorders, or dietary challenges to mutational signatures implicated in cancer evolution.
Assuntos
Proteína BRCA2 , Neoplasias da Mama , Glicólise , Aldeído Pirúvico , Animais , Proteína BRCA2/metabolismo , Proteína BRCA2/genética , Camundongos , Humanos , Feminino , Aldeído Pirúvico/metabolismo , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Haploinsuficiência , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patologia , Mutação , Dano ao DNA , Reparo do DNA , Linhagem Celular TumoralRESUMO
The in vitro oxygen microenvironment profoundly affects the capacity of cell cultures to model physiological and pathophysiological states. Cell culture is often considered to be hyperoxic, but pericellular oxygen levels, which are affected by oxygen diffusivity and consumption, are rarely reported. Here, we provide evidence that several cell types in culture actually experience local hypoxia, with important implications for cell metabolism and function. We focused initially on adipocytes, as adipose tissue hypoxia is frequently observed in obesity and precedes diminished adipocyte function. Under standard conditions, cultured adipocytes are highly glycolytic and exhibit a transcriptional profile indicative of physiological hypoxia. Increasing pericellular oxygen diverted glucose flux toward mitochondria, lowered HIF1α activity, and resulted in widespread transcriptional rewiring. Functionally, adipocytes increased adipokine secretion and sensitivity to insulin and lipolytic stimuli, recapitulating a healthier adipocyte model. The functional benefits of increasing pericellular oxygen were also observed in macrophages, hPSC-derived hepatocytes and cardiac organoids. Our findings demonstrate that oxygen is limiting in many terminally-differentiated cell types, and that considering pericellular oxygen improves the quality, reproducibility and translatability of culture models.
Assuntos
Adipócitos , Diferenciação Celular , Oxigênio , Oxigênio/metabolismo , Adipócitos/metabolismo , Adipócitos/citologia , Humanos , Técnicas de Cultura de Células/métodos , Animais , Glicólise , Hepatócitos/metabolismo , Hipóxia Celular , Mitocôndrias/metabolismo , Camundongos , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Células Cultivadas , Glucose/metabolismo , Macrófagos/metabolismoRESUMO
Digested dietary fats are taken up by enterocytes where they are assembled into pre-chylomicrons in the endoplasmic reticulum followed by transport to the Golgi for maturation and subsequent secretion to the circulation1. The role of mitochondria in dietary lipid processing is unclear. Here we show that mitochondrial dysfunction in enterocytes inhibits chylomicron production and the transport of dietary lipids to peripheral organs. Mice with specific ablation of the mitochondrial aspartyl-tRNA synthetase DARS2 (ref. 2), the respiratory chain subunit SDHA3 or the assembly factor COX10 (ref. 4) in intestinal epithelial cells showed accumulation of large lipid droplets (LDs) in enterocytes of the proximal small intestine and failed to thrive. Feeding a fat-free diet suppressed the build-up of LDs in DARS2-deficient enterocytes, which shows that the accumulating lipids derive mostly from digested fat. Furthermore, metabolic tracing studies revealed an impaired transport of dietary lipids to peripheral organs in mice lacking DARS2 in intestinal epithelial cells. DARS2 deficiency caused a distinct lack of mature chylomicrons concomitant with a progressive dispersal of the Golgi apparatus in proximal enterocytes. This finding suggests that mitochondrial dysfunction results in impaired trafficking of chylomicrons from the endoplasmic reticulum to the Golgi, which in turn leads to storage of dietary lipids in large cytoplasmic LDs. Taken together, these results reveal a role for mitochondria in dietary lipid transport in enterocytes, which might be relevant for understanding the intestinal defects observed in patients with mitochondrial disorders5.
Assuntos
Gorduras na Dieta , Enterócitos , Metabolismo dos Lipídeos , Mitocôndrias , Animais , Camundongos , Aspartato-tRNA Ligase/metabolismo , Quilomícrons/metabolismo , Gorduras na Dieta/metabolismo , Complexo II de Transporte de Elétrons/metabolismo , Retículo Endoplasmático/metabolismo , Enterócitos/metabolismo , Enterócitos/patologia , Células Epiteliais/metabolismo , Complexo de Golgi/metabolismo , Intestinos , Gotículas Lipídicas/metabolismo , Mitocôndrias/metabolismo , Mitocôndrias/patologiaRESUMO
Background: Branched-chain aminotransferase 1 (BCAT1) has been proposed to drive proliferation and invasion of isocitrate dehydrogenase (IDH) wild-type glioblastoma cells. However, the Cancer Genome Atlas (TCGA) dataset shows considerable variation in the expression of this enzyme in glioblastoma. The aim of this study was to determine the role of BCAT1 in driving the proliferation and invasion of glioblastoma cells and xenografts that have widely differing levels of BCAT1 expression and the mechanism responsible. Methods: The activity of BCAT1 was modulated in IDH wild-type patient-derived glioblastoma cell lines, and in orthotopically implanted tumors derived from these cells, to examine the effects of BCAT1 expression on tumor phenotype. Results: In cells with constitutively high BCAT1 expression and a glycolytic metabolic phenotype, inducible shRNA knockdown of the enzyme resulted in reduced proliferation and invasion by increasing the concentration of α-ketoglutarate, leading to reduced DNA methylation, HIF-1α destabilization, and reduced expression of the transcription factor Forkhead box protein M1 (FOXM1). Conversely, overexpression of the enzyme increased HIF-1α expression and promoted proliferation and invasion. However, in cells with an oxidative phenotype and very low constitutive expression of BCAT1 increased expression of the enzyme had no effect on invasion and reduced cell proliferation. This occurred despite an increase in HIF-1α levels and could be explained by decreased TCA cycle flux. Conclusions: There is a wide variation in BCAT1 expression in glioblastoma and its role in proliferation and invasion is dependent on tumor subtype.
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Fumarate hydratase (FH) is an enzyme of the Tricarboxylic Acid (TCA) cycle whose mutations lead to hereditary and sporadic forms of cancer. Although more than twenty years have passed since its discovery as the leading cause of the cancer syndrome Hereditary leiomyomatosis and Renal Cell Carcinoma (HLRCC), it is still unclear how the loss of FH causes cancer in a tissue-specific manner and with such aggressive behaviour. It has been shown that FH loss, via the accumulation of FH substrate fumarate, activates a series of oncogenic cascades whose contribution to transformation is still under investigation. In this review, we will summarise these recent findings in an integrated fashion and put forward the case that understanding the biology of FH and how its mutations promote transformation will be vital to establish novel paradigms of oncometabolism.
Assuntos
Carcinoma de Células Renais , Neoplasias Renais , Síndromes Neoplásicas Hereditárias , Neoplasias Cutâneas , Neoplasias Uterinas , Feminino , Humanos , Fumarato Hidratase/genética , Neoplasias Uterinas/genética , Neoplasias Cutâneas/genética , Síndromes Neoplásicas Hereditárias/genética , Carcinoma de Células Renais/genética , Carcinoma de Células Renais/patologia , Neoplasias Renais/genética , Neoplasias Renais/patologiaRESUMO
Metabolic competition within the tumor microenvironment (TME) shapes the efficacy of anticancer immunity. In the August 3rd issue of Nature, Guo et al.1 show that glutamine is an intercellular metabolic checkpoint between cancer and immune cells. Targeting glutamine metabolism in the TME is a promising strategy to improve anti-cancer therapy.
Assuntos
Glutamina , Microambiente Tumoral , Células DendríticasRESUMO
Mitochondrial metabolite reduces melanoma growth by boosting antigen presentation.
Assuntos
Apresentação de Antígeno , Melanoma , Mitocôndrias , Humanos , Melanoma/imunologia , Melanoma/patologia , Mitocôndrias/metabolismo , Carcinogênese/imunologia , Carcinogênese/patologiaRESUMO
DNA single-strand breaks (SSBs) disrupt DNA replication and induce chromosome breakage. However, whether SSBs induce chromosome breakage when present behind replication forks or ahead of replication forks is unclear. To address this question, we exploited an exquisite sensitivity of SSB repair-defective human cells lacking PARP activity or XRCC1 to the thymidine analogue 5-chloro-2'-deoxyuridine (CldU). We show that incubation with CldU in these cells results in chromosome breakage, sister chromatid exchange, and cytotoxicity by a mechanism that depends on the S phase activity of uracil DNA glycosylase (UNG). Importantly, we show that CldU incorporation in one cell cycle is cytotoxic only during the following cell cycle, when it is present in template DNA. In agreement with this, while UNG induces SSBs both in nascent strands behind replication forks and in template strands ahead of replication forks, only the latter trigger fork collapse and chromosome breakage. Finally, we show that BRCA-defective cells are hypersensitive to CldU, either alone and/or in combination with PARP inhibitor, suggesting that CldU may have clinical utility.
Assuntos
Antineoplásicos , Inibidores de Poli(ADP-Ribose) Polimerases , Humanos , Inibidores de Poli(ADP-Ribose) Polimerases/farmacologia , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Quebra Cromossômica , Reparo do DNA , Replicação do DNA , DNA , Proteína 1 Complementadora Cruzada de Reparo de Raio-X/metabolismoRESUMO
Hereditary leiomyomatosis and renal cell cancer (HLRCC) is a cancer syndrome caused by inactivating germline mutations in fumarate hydratase (FH) and subsequent accumulation of fumarate. Fumarate accumulation leads to profound epigenetic changes and the activation of an anti-oxidant response via nuclear translocation of the transcription factor NRF2. The extent to which chromatin remodeling shapes this anti-oxidant response is currently unknown. Here, we explored the effects of FH loss on the chromatin landscape to identify transcription factor networks involved in the remodeled chromatin landscape of FH-deficient cells. We identify FOXA2 as a key transcription factor that regulates anti-oxidant response genes and subsequent metabolic rewiring cooperating without direct interaction with the anti-oxidant regulator NRF2. The identification of FOXA2 as an anti-oxidant regulator provides additional insights into the molecular mechanisms behind cell responses to fumarate accumulation and potentially provides further avenues for therapeutic intervention for HLRCC.
Assuntos
Carcinoma de Células Renais , Neoplasias Renais , Leiomiomatose , Síndromes Neoplásicas Hereditárias , Neoplasias Cutâneas , Neoplasias Uterinas , Feminino , Humanos , Fumarato Hidratase/genética , Antioxidantes , Fator 2 Relacionado a NF-E2/genética , Leiomiomatose/genética , Neoplasias Uterinas/genética , Neoplasias Cutâneas/genética , Síndromes Neoplásicas Hereditárias/genética , Cromatina , Neoplasias Renais/genética , Carcinoma de Células Renais/genética , Fator 3-beta Nuclear de Hepatócito/genéticaRESUMO
Myocardial ischemia-reperfusion (IR) injury may result in cardiomyocyte dysfunction. Mitochondria play a critical role in cardiomyocyte recovery after IR injury. The mitochondrial uncoupling protein 3 (UCP3) has been proposed to reduce mitochondrial reactive oxygen species (ROS) production and to facilitate fatty acid oxidation. As both mechanisms might be protective following IR injury, we investigated functional, mitochondrial structural, and metabolic cardiac remodeling in wild-type mice and in mice lacking UCP3 (UCP3-KO) after IR. Results showed that infarct size in isolated perfused hearts subjected to IR ex vivo was larger in adult and old UCP3-KO mice than in equivalent wild-type mice, and was accompanied by higher levels of creatine kinase in the effluent and by more pronounced mitochondrial structural changes. The greater myocardial damage in UCP3-KO hearts was confirmed in vivo after coronary artery occlusion followed by reperfusion. S1QEL, a suppressor of superoxide generation from site IQ in complex I, limited infarct size in UCP3-KO hearts, pointing to exacerbated superoxide production as a possible cause of the damage. Metabolomics analysis of isolated perfused hearts confirmed the reported accumulation of succinate, xanthine and hypoxanthine during ischemia, and a shift to anaerobic glucose utilization, which all recovered upon reoxygenation. The metabolic response to ischemia and IR was similar in UCP3-KO and wild-type hearts, being lipid and energy metabolism the most affected pathways. Fatty acid oxidation and complex I (but not complex II) activity were equally impaired after IR. Overall, our results indicate that UCP3 deficiency promotes enhanced superoxide generation and mitochondrial structural changes that increase the vulnerability of the myocardium to IR injury.
Assuntos
Doença da Artéria Coronariana , Isquemia Miocárdica , Traumatismo por Reperfusão Miocárdica , Camundongos , Animais , Superóxidos/metabolismo , Isquemia Miocárdica/metabolismo , Miócitos Cardíacos/metabolismo , Mitocôndrias/metabolismo , Estresse Oxidativo , Traumatismo por Reperfusão Miocárdica/genética , Traumatismo por Reperfusão Miocárdica/metabolismo , Doença da Artéria Coronariana/metabolismo , Metabolismo Energético , Isquemia/metabolismo , Reperfusão , Ácidos Graxos/metabolismo , Infarto/complicações , Infarto/metabolismoRESUMO
Metabolic rewiring underlies the effector functions of macrophages1-3, but the mechanisms involved remain incompletely defined. Here, using unbiased metabolomics and stable isotope-assisted tracing, we show that an inflammatory aspartate-argininosuccinate shunt is induced following lipopolysaccharide stimulation. The shunt, supported by increased argininosuccinate synthase (ASS1) expression, also leads to increased cytosolic fumarate levels and fumarate-mediated protein succination. Pharmacological inhibition and genetic ablation of the tricarboxylic acid cycle enzyme fumarate hydratase (FH) further increases intracellular fumarate levels. Mitochondrial respiration is also suppressed and mitochondrial membrane potential increased. RNA sequencing and proteomics analyses demonstrate that there are strong inflammatory effects resulting from FH inhibition. Notably, acute FH inhibition suppresses interleukin-10 expression, which leads to increased tumour necrosis factor secretion, an effect recapitulated by fumarate esters. Moreover, FH inhibition, but not fumarate esters, increases interferon-ß production through mechanisms that are driven by mitochondrial RNA (mtRNA) release and activation of the RNA sensors TLR7, RIG-I and MDA5. This effect is recapitulated endogenously when FH is suppressed following prolonged lipopolysaccharide stimulation. Furthermore, cells from patients with systemic lupus erythematosus also exhibit FH suppression, which indicates a potential pathogenic role for this process in human disease. We therefore identify a protective role for FH in maintaining appropriate macrophage cytokine and interferon responses.
Assuntos
Fumarato Hidratase , Interferon beta , Macrófagos , Mitocôndrias , RNA Mitocondrial , Humanos , Argininossuccinato Sintase/metabolismo , Ácido Argininossuccínico/metabolismo , Ácido Aspártico/metabolismo , Respiração Celular , Citosol/metabolismo , Fumarato Hidratase/antagonistas & inibidores , Fumarato Hidratase/genética , Fumarato Hidratase/metabolismo , Fumaratos/metabolismo , Interferon beta/biossíntese , Interferon beta/imunologia , Lipopolissacarídeos/farmacologia , Lipopolissacarídeos/metabolismo , Lúpus Eritematoso Sistêmico/enzimologia , Macrófagos/enzimologia , Macrófagos/imunologia , Macrófagos/metabolismo , Potencial da Membrana Mitocondrial , Metabolômica , Mitocôndrias/genética , Mitocôndrias/metabolismo , RNA Mitocondrial/metabolismoRESUMO
Mutations in fumarate hydratase (FH) cause hereditary leiomyomatosis and renal cell carcinoma1. Loss of FH in the kidney elicits several oncogenic signalling cascades through the accumulation of the oncometabolite fumarate2. However, although the long-term consequences of FH loss have been described, the acute response has not so far been investigated. Here we generated an inducible mouse model to study the chronology of FH loss in the kidney. We show that loss of FH leads to early alterations of mitochondrial morphology and the release of mitochondrial DNA (mtDNA) into the cytosol, where it triggers the activation of the cyclic GMP-AMP synthase (cGAS)-stimulator of interferon genes (STING)-TANK-binding kinase 1 (TBK1) pathway and stimulates an inflammatory response that is also partially dependent on retinoic-acid-inducible gene I (RIG-I). Mechanistically, we show that this phenotype is mediated by fumarate and occurs selectively through mitochondrial-derived vesicles in a manner that depends on sorting nexin 9 (SNX9). These results reveal that increased levels of intracellular fumarate induce a remodelling of the mitochondrial network and the generation of mitochondrial-derived vesicles, which allows the release of mtDNAin the cytosol and subsequent activation of the innate immune response.
Assuntos
DNA Mitocondrial , Fumaratos , Imunidade Inata , Mitocôndrias , Animais , Camundongos , DNA Mitocondrial/metabolismo , Fumarato Hidratase/genética , Fumarato Hidratase/metabolismo , Fumaratos/metabolismo , Mitocôndrias/enzimologia , Mitocôndrias/genética , Mitocôndrias/metabolismo , Mitocôndrias/patologia , Rim/enzimologia , Rim/metabolismo , Rim/patologia , Citosol/metabolismoRESUMO
Depriving cells of nutrients triggers an energetic crisis, which is resolved by metabolic rewiring and organelle reorganization. Primary cilia are microtubule-based organelles at the cell surface, capable of integrating multiple metabolic and signalling cues, but their precise sensory function is not fully understood. Here we show that primary cilia respond to nutrient availability and adjust their length via glutamine-mediated anaplerosis facilitated by asparagine synthetase (ASNS). Nutrient deprivation causes cilia elongation, mediated by reduced mitochondrial function, ATP availability and AMPK activation independently of mTORC1. Of note, glutamine removal and replenishment is necessary and sufficient to induce ciliary elongation or retraction, respectively, under nutrient stress conditions both in vivo and in vitro by restoring mitochondrial anaplerosis via ASNS-dependent glutamate generation. Ift88-mutant cells lacking cilia show reduced glutamine-dependent mitochondrial anaplerosis during metabolic stress, due to reduced expression and activity of ASNS at the base of cilia. Our data indicate a role for cilia in responding to, and possibly sensing, cellular glutamine levels via ASNS during metabolic stress.