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1.
Microbiol Spectr ; 12(2): e0259423, 2024 Feb 06.
Artigo em Inglês | MEDLINE | ID: mdl-38230926

RESUMO

Fungal infections are a growing global health concern due to the limited number of available antifungal therapies as well as the emergence of fungi that are resistant to first-line antimicrobials, particularly azoles and echinocandins. Development of novel, selective antifungal therapies is challenging due to similarities between fungal and mammalian cells. An attractive source of potential antifungal treatments is provided by ecological niches co-inhabited by bacteria, fungi, and multicellular organisms, where complex relationships between multiple organisms have resulted in evolution of a wide variety of selective antimicrobials. Here, we characterized several analogs of one such natural compound, collismycin A. We show that NR-6226C has antifungal activity against several pathogenic Candida species, including C. albicans and C. glabrata, whereas it only has little toxicity against mammalian cells. Mechanistically, NR-6226C selectively chelates iron, which is a limiting factor for pathogenic fungi during infection. As a result, NR-6226C treatment causes severe mitochondrial dysfunction, leading to formation of reactive oxygen species, metabolic reprogramming, and a severe reduction in ATP levels. Using an in vivo model for fungal infections, we show that NR-6226C significantly increases survival of Candida-infected Galleria mellonella larvae. Finally, our data indicate that NR-6226C synergizes strongly with fluconazole in inhibition of C. albicans. Taken together, NR-6226C is a promising antifungal compound that acts by chelating iron and disrupting mitochondrial functions.IMPORTANCEDrug-resistant fungal infections are an emerging global threat, and pan-resistance to current antifungal therapies is an increasing problem. Clearly, there is a need for new antifungal drugs. In this study, we characterized a novel antifungal agent, the collismycin analog NR-6226C. NR-6226C has a favorable toxicity profile for human cells, which is essential for further clinical development. We unraveled the mechanism of action of NR-6226C and found that it disrupts iron homeostasis and thereby depletes fungal cells of energy. Importantly, NR-6226C strongly potentiates the antifungal activity of fluconazole, thereby providing inroads for combination therapy that may reduce or prevent azole resistance. Thus, NR-6226C is a promising compound for further development into antifungal treatment.


Assuntos
Anti-Infecciosos , Micoses , Animais , Humanos , Antifúngicos/farmacologia , Fluconazol/farmacologia , Ferro , Candida , Micoses/microbiologia , Candida albicans , Anti-Infecciosos/farmacologia , Azóis/farmacologia , Candida glabrata , Quelantes de Ferro/farmacologia , Farmacorresistência Fúngica , Testes de Sensibilidade Microbiana , Mamíferos
2.
Transcription ; : 1-21, 2023 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-37655806

RESUMO

The preservation of gene expression patterns that define cellular identity throughout the cell division cycle is essential to perpetuate cellular lineages. However, the progression of cells through different phases of the cell cycle severely disrupts chromatin accessibility, epigenetic marks, and the recruitment of transcriptional regulators. Notably, chromatin is transiently disassembled during S-phase and undergoes drastic condensation during mitosis, which is a significant challenge to the preservation of gene expression patterns between cell generations. This article delves into the specific gene expression and chromatin regulatory mechanisms that facilitate the preservation of transcriptional identity during replication and mitosis. Furthermore, we emphasize our recent findings revealing the unconventional role of yeast centromeres and mitotic chromosomes in maintaining transcriptional fidelity beyond mitosis.

3.
Proc Natl Acad Sci U S A ; 120(4): e2210593120, 2023 01 24.
Artigo em Inglês | MEDLINE | ID: mdl-36656860

RESUMO

Mitotic entry correlates with the condensation of the chromosomes, changes in histone modifications, exclusion of transcription factors from DNA, and the broad downregulation of transcription. However, whether mitotic condensation influences transcription in the subsequent interphase is unknown. Here, we show that preventing one chromosome to condense during mitosis causes it to fail resetting of transcription. Rather, in the following interphase, the affected chromosome contains unusually high levels of the transcription machinery, resulting in abnormally high expression levels of genes in cis, including various transcription factors. This subsequently causes the activation of inducible transcriptional programs in trans, such as the GAL genes, even in the absence of the relevant stimuli. Thus, mitotic chromosome condensation exerts stringent control on interphase gene expression to ensure the maintenance of basic cellular functions and cell identity across cell divisions. Together, our study identifies the maintenance of transcriptional homeostasis during interphase as an unexpected function of mitosis and mitotic chromosome condensation.


Assuntos
Cromatina , Cromossomos , Cromatina/genética , Cromossomos/genética , Cromossomos/metabolismo , Interfase/genética , Mitose/genética , Fatores de Transcrição/metabolismo
4.
STAR Protoc ; 3(1): 101210, 2022 03 18.
Artigo em Inglês | MEDLINE | ID: mdl-35265859

RESUMO

FUS3 and STE2 expression levels can be used as reporters for signaling through the pheromone pathway in the budding yeast Saccharomyces cerevisiae. Here, we describe an optimized protocol to measure the expression levels of FUS3 and STE2 using quantitative reverse transcription PCR (RT-qPCR). We describe the steps for comparing untreated and pheromone-treated yeast cells and how to quantify the changes in various deletion strains. The protocol can be applied to determine potential regulators of the pheromone pathway. For complete details on the use and execution of this protocol, please refer to Garcia et al. (2021).


Assuntos
Proteínas de Saccharomyces cerevisiae , Fermento Seco , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Feromônios/metabolismo , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/genética , Transdução de Sinais/genética
5.
Int J Mol Sci ; 23(3)2022 Jan 24.
Artigo em Inglês | MEDLINE | ID: mdl-35163213

RESUMO

The cyclin-dependent kinase Cdk1 is best known for its function as master regulator of the cell cycle. It phosphorylates several key proteins to control progression through the different phases of the cell cycle. However, studies conducted several decades ago with mammalian cells revealed that Cdk1 also directly regulates the basal transcription machinery, most notably RNA polymerase II. More recent studies in the budding yeast Saccharomyces cerevisiae have revisited this function of Cdk1 and also revealed that Cdk1 directly controls RNA polymerase III activity. These studies have also provided novel insight into the physiological relevance of this process. For instance, cell cycle-stage-dependent activity of these complexes may be important for meeting the increased demand for various proteins involved in housekeeping, metabolism, and protein synthesis. Recent work also indicates that direct regulation of the RNA polymerase II machinery promotes cell cycle entry. Here, we provide an overview of the regulation of basal transcription by Cdk1, and we hypothesize that the original function of the primordial cell-cycle CDK was to regulate RNAPII and that it later evolved into specialized kinases that govern various aspects of the transcription machinery and the cell cycle.


Assuntos
Proteína Quinase CDC2/genética , Proteína Quinase CDC2/metabolismo , Transcrição Gênica/fisiologia , Animais , Proteína Quinase CDC2/fisiologia , Ciclo Celular/genética , Ciclo Celular/fisiologia , Proteínas de Ciclo Celular/metabolismo , Quinases Ciclina-Dependentes/genética , Quinases Ciclina-Dependentes/metabolismo , Humanos , Fosforilação , RNA Polimerase II/metabolismo , Transcrição Gênica/genética
6.
Nucleic Acids Res ; 50(3): 1351-1369, 2022 02 22.
Artigo em Inglês | MEDLINE | ID: mdl-35100417

RESUMO

Tight control of gene expression networks required for adipose tissue formation and plasticity is essential for adaptation to energy needs and environmental cues. However, the mechanisms that orchestrate the global and dramatic transcriptional changes leading to adipocyte differentiation remain to be fully unraveled. We investigated the regulation of nascent transcription by the sumoylation pathway during adipocyte differentiation using SLAMseq and ChIPseq. We discovered that the sumoylation pathway has a dual function in differentiation; it supports the initial downregulation of pre-adipocyte-specific genes, while it promotes the establishment of the mature adipocyte transcriptional program. By characterizing endogenous sumoylome dynamics in differentiating adipocytes by mass spectrometry, we found that sumoylation of specific transcription factors like PPARγ/RXR and their co-factors are associated with the transcription of adipogenic genes. Finally, using RXR as a model, we found that sumoylation may regulate adipogenic transcription by supporting the chromatin occurrence of transcription factors. Our data demonstrate that the sumoylation pathway supports the rewiring of transcriptional networks required for formation of functional adipocytes. This study also provides the scientists in the field of cellular differentiation and development with an in-depth resource of the dynamics of the SUMO-chromatin landscape, SUMO-regulated transcription and endogenous sumoylation sites during adipocyte differentiation.


Assuntos
Adipogenia , Sumoilação , Adipócitos/metabolismo , Adipogenia/genética , Diferenciação Celular/genética , Cromatina/genética , Cromatina/metabolismo , Fatores de Transcrição/metabolismo
7.
Cell Rep ; 37(13): 110186, 2021 12 28.
Artigo em Inglês | MEDLINE | ID: mdl-34965431

RESUMO

Mechanisms have evolved that allow cells to detect signals and generate an appropriate response. The accuracy of these responses relies on the ability of cells to discriminate between signal and noise. How cells filter noise in signaling pathways is not well understood. Here, we analyze noise suppression in the yeast pheromone signaling pathway and show that the poorly characterized protein Kel1 serves as a major noise suppressor and prevents cell death. At the molecular level, Kel1 prevents spontaneous activation of the pheromone response by inhibiting membrane recruitment of Ste5 and Far1. Only a hypophosphorylated form of Kel1 suppresses signaling, reduces noise, and prevents pheromone-associated cell death, and our data indicate that the MAPK Fus3 contributes to Kel1 phosphorylation. Taken together, Kel1 serves as a phospho-regulated suppressor of the pheromone pathway to reduce noise, inhibit spontaneous activation of the pathway, regulate mating efficiency, and prevent pheromone-associated cell death.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Ruído , Feromônios/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Inibidoras de Quinase Dependente de Ciclina/genética , Proteínas Inibidoras de Quinase Dependente de Ciclina/metabolismo , Proteínas Quinases Ativadas por Mitógeno/genética , Fosforilação , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/crescimento & desenvolvimento , Proteínas de Saccharomyces cerevisiae/genética , Transdução de Sinais
8.
J Biol Chem ; 294(49): 18784-18795, 2019 12 06.
Artigo em Inglês | MEDLINE | ID: mdl-31676685

RESUMO

Post-translational modification by small ubiquitin-like modifier (Sumo) regulates many cellular processes, including the adaptive response to various types of stress, referred to as the Sumo stress response (SSR). However, it remains unclear whether the SSR involves a common set of core proteins regardless of the type of stress or whether each particular type of stress induces a stress-specific SSR that targets a unique, largely nonoverlapping set of Sumo substrates. In this study, we used MS and a Gene Ontology approach to identify differentially sumoylated proteins during heat stress, hyperosmotic stress, oxidative stress, nitrogen starvation, and DNA alkylation in Saccharomyces cerevisiae cells. Our results indicate that each stress triggers a specific SSR signature centered on proteins involved in transcription, translation, and chromatin regulation. Strikingly, whereas the various stress-specific SSRs were largely nonoverlapping, all types of stress tested here resulted in desumoylation of subunits of RNA polymerase III, which correlated with a decrease in tRNA synthesis. We conclude that desumoylation and subsequent inhibition of RNA polymerase III constitutes the core of all stress-specific SSRs in yeast.


Assuntos
RNA Polimerase III/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/enzimologia , Saccharomyces cerevisiae/metabolismo , Espectrometria de Massas , Estresse Oxidativo , Processamento de Proteína Pós-Traducional
10.
Cell ; 175(3): 780-795.e15, 2018 10 18.
Artigo em Inglês | MEDLINE | ID: mdl-30318142

RESUMO

During mitosis, chromatin condensation shapes chromosomes as separate, rigid, and compact sister chromatids to facilitate their segregation. Here, we show that, unlike wild-type yeast chromosomes, non-chromosomal DNA circles and chromosomes lacking a centromere fail to condense during mitosis. The centromere promotes chromosome condensation strictly in cis through recruiting the kinases Aurora B and Bub1, which trigger the autonomous condensation of the entire chromosome. Shugoshin and the deacetylase Hst2 facilitated spreading the condensation signal to the chromosome arms. Targeting Aurora B to DNA circles or centromere-ablated chromosomes or releasing Shugoshin from PP2A-dependent inhibition bypassed the centromere requirement for condensation and enhanced the mitotic stability of DNA circles. Our data indicate that yeast cells license the chromosome-autonomous condensation of their chromatin in a centromere-dependent manner, excluding from this process non-centromeric DNA and thereby inhibiting their propagation.


Assuntos
Centrômero/genética , Cromossomos Fúngicos/genética , Mitose , Saccharomyces cerevisiae/genética , Aurora Quinase B/genética , Aurora Quinase B/metabolismo , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Proteína Fosfatase 2/genética , Proteína Fosfatase 2/metabolismo , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Sirtuína 2/genética , Sirtuína 2/metabolismo
11.
Nucleic Acids Res ; 46(22): 11698-11711, 2018 12 14.
Artigo em Inglês | MEDLINE | ID: mdl-30247619

RESUMO

tRNA genes are transcribed by RNA polymerase III (RNAPIII). During recent years it has become clear that RNAPIII activity is strictly regulated by the cell in response to environmental cues and the homeostatic status of the cell. However, the molecular mechanisms that control RNAPIII activity to regulate the amplitude of tDNA transcription in normally cycling cells are not well understood. Here, we show that tRNA levels fluctuate during the cell cycle and reveal an underlying molecular mechanism. The cyclin Clb5 recruits the cyclin dependent kinase Cdk1 to tRNA genes to boost tDNA transcription during late S phase. At tDNA genes, Cdk1 promotes the recruitment of TFIIIC, stimulates the interaction between TFIIIB and TFIIIC, and increases the dynamics of RNA polymerase III in vivo. Furthermore, we identified Bdp1 as a putative Cdk1 substrate in this process. Preventing Bdp1 phosphorylation prevented cell cycle-dependent recruitment of TFIIIC and abolished the cell cycle-dependent increase in tDNA transcription. Our findings demonstrate that under optimal growth conditions Cdk1 gates tRNA synthesis in S phase by regulating the RNAPIII machinery, revealing a direct link between the cell cycle and RNAPIII activity.


Assuntos
Proteína Quinase CDC2/genética , Proteína Quinase CDC28 de Saccharomyces cerevisiae/genética , Ciclo Celular/genética , RNA Polimerase III/genética , RNA de Transferência/genética , Proteína Quinase CDC2/metabolismo , Proteína Quinase CDC28 de Saccharomyces cerevisiae/metabolismo , Ciclina B/genética , Ciclina B/metabolismo , Perfilação da Expressão Gênica , Regulação Fúngica da Expressão Gênica , Fosforilação , Ligação Proteica , RNA Polimerase III/metabolismo , RNA de Transferência/metabolismo , Fase S/genética , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Fator de Transcrição TFIIIB/genética , Fator de Transcrição TFIIIB/metabolismo , Fatores de Transcrição TFIII/genética , Fatores de Transcrição TFIII/metabolismo
12.
Biochim Biophys Acta Gene Regul Mech ; 1861(4): 310-319, 2018 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-29127063

RESUMO

RNA polymerase III (RNAPIII) transcribes tRNA genes, 5S RNA as well as a number of other non-coding RNAs. Because transcription by RNAPIII is an energy-demanding process, its activity is tightly linked to the stress levels and nutrient status of the cell. Multiple signaling pathways control RNAPIII activity in response to environmental cues, but exactly how these pathways regulate RNAPIII is still poorly understood. One major target of these pathways is the transcriptional repressor Maf1, which inhibits RNAPIII activity under conditions that are detrimental to cell growth. However, recent studies have found that the cell can also directly regulate the RNAPIII machinery through phosphorylation and sumoylation of RNAPIII subunits. In this review we summarize post-translational modifications of RNAPIII subunits that mainly have been identified in large-scale proteomics studies, and we highlight several examples to discuss their relevance for regulation of RNAPIII.


Assuntos
Regulação da Expressão Gênica , Processamento de Proteína Pós-Traducional , RNA Polimerase III/metabolismo , RNA de Transferência/biossíntese , Animais , Caseína Quinases/genética , Proteína Coatomer/genética , Regulação Fúngica da Expressão Gênica , Fosforilação , Subunidades Proteicas , RNA Polimerase III/química , RNA Polimerase III/genética , RNA Fúngico/biossíntese , RNA Fúngico/genética , RNA de Transferência/genética , Proteína SUMO-1/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , Transdução de Sinais , Sumoilação , Proteína de Ligação a TATA-Box/genética , Fator de Transcrição TFIIIB/genética , Fatores de Transcrição/genética
13.
G3 (Bethesda) ; 7(6): 1753-1766, 2017 06 07.
Artigo em Inglês | MEDLINE | ID: mdl-28428242

RESUMO

Cdk1 (Cdc28 in yeast) is a cyclin-dependent kinase (CDK) essential for cell cycle progression and cell division in normal cells. However, CDK activity also underpins proliferation of tumor cells, making it a relevant study subject. While numerous targets and processes regulated by Cdc28 have been identified, the exact functions of Cdc28 are only partially understood. To further explore the functions of Cdc28, we systematically overexpressed ∼4800 genes in wild-type (WT) cells and in cells with artificially reduced Cdc28 activity. This screen identified 366 genes that, when overexpressed, specifically compromised cell viability under conditions of reduced Cdc28 activity. Consistent with the crucial functions of Cdc28 in cell cycle regulation and chromosome metabolism, most of these genes have functions in the cell cycle, DNA replication, and transcription. However, a substantial number of genes control processes not directly associated with the cell cycle, indicating that Cdc28 may also regulate these processes. Finally, because the dataset was enriched for direct Cdc28 targets, the results from this screen will aid in identifying novel targets and process regulated by Cdc28.


Assuntos
Quinases relacionadas a CDC2 e CDC28/genética , Mapeamento Cromossômico , Epistasia Genética , Mutações Sintéticas Letais , Quinases relacionadas a CDC2 e CDC28/metabolismo , Ciclo Celular/genética , Biologia Computacional/métodos , Análise Mutacional de DNA , Replicação do DNA , Regulação Fúngica da Expressão Gênica , Redes Reguladoras de Genes , Genômica/métodos , Fenótipo , Proteínas Recombinantes de Fusão , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo
17.
Proc Natl Acad Sci U S A ; 114(5): 1039-1044, 2017 01 31.
Artigo em Inglês | MEDLINE | ID: mdl-28096404

RESUMO

Maintaining cellular homeostasis under changing nutrient conditions is essential for the growth and development of all organisms. The mechanisms that maintain homeostasis upon loss of nutrient supply are not well understood. By mapping the SUMO proteome in Saccharomyces cerevisiae, we discovered a specific set of differentially sumoylated proteins mainly involved in transcription. RNA polymerase III (RNAPIII) components, including Rpc53, Rpc82, and Ret1, are particularly prominent nutrient-dependent SUMO targets. Nitrogen starvation, as well as direct inhibition of the master nutrient response regulator target of rapamycin complex 1 (TORC1), results in rapid desumoylation of these proteins, which is reflected by loss of SUMO at tRNA genes. TORC1-dependent sumoylation of Rpc82 in particular is required for robust tRNA transcription. Mechanistically, sumoylation of Rpc82 is important for assembly of the RNAPIII holoenzyme and recruitment of Rpc82 to tRNA genes. In conclusion, our data show that TORC1-dependent sumoylation of Rpc82 bolsters the transcriptional capacity of RNAPIII under optimal growth conditions.


Assuntos
Regulação Fúngica da Expressão Gênica , Processamento de Proteína Pós-Traducional , RNA Polimerase III/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/metabolismo , Fatores de Transcrição/metabolismo , Transcrição Gênica , Substituição de Aminoácidos , Ontologia Genética , Nitrogênio/metabolismo , Subunidades Proteicas , RNA Fúngico/biossíntese , RNA Fúngico/genética , RNA de Transferência/biossíntese , RNA de Transferência/genética , Saccharomyces cerevisiae/efeitos dos fármacos , Saccharomyces cerevisiae/crescimento & desenvolvimento , Proteínas de Saccharomyces cerevisiae/efeitos dos fármacos , Sirolimo/farmacologia , Sumoilação , Fatores de Transcrição/efeitos dos fármacos , Enzimas de Conjugação de Ubiquitina/genética
18.
Bioessays ; 37(10): 1095-105, 2015 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-26354225

RESUMO

The small ubiquitin-like modifier SUMO regulates many aspects of cellular physiology to maintain cell homeostasis, both under normal conditions and during cell stress. Components of the transcriptional apparatus and chromatin are among the most prominent SUMO substrates. The prevailing view is that SUMO serves to repress transcription. However, as we will discuss in this review, this model needs to be refined, because recent studies have revealed that SUMO can also have profound positive effects on transcription.


Assuntos
Regulação da Expressão Gênica , Proteína SUMO-1/fisiologia , Transcrição Gênica , Animais , Humanos , Lisina/metabolismo , Ligação Proteica/genética , Conformação Proteica , Sumoilação/fisiologia
19.
Genome Res ; 25(6): 897-906, 2015 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-25800674

RESUMO

Transcription factors are abundant Sumo targets, yet the global distribution of Sumo along the chromatin and its physiological relevance in transcription are poorly understood. Using Saccharomyces cerevisiae, we determined the genome-wide localization of Sumo along the chromatin. We discovered that Sumo-enriched genes are almost exclusively involved in translation, such as tRNA genes and ribosomal protein genes (RPGs). Genome-wide expression analysis showed that Sumo positively regulates their transcription. We also discovered that the Sumo consensus motif at RPG promoters is identical to the DNA binding motif of the transcription factor Rap1. We demonstrate that Rap1 is a molecular target of Sumo and that sumoylation of Rap1 is important for cell viability. Furthermore, Rap1 sumoylation promotes recruitment of the basal transcription machinery, and sumoylation of Rap1 cooperates with the target of rapamycin kinase complex 1 (TORC1) pathway to promote RPG transcription. Strikingly, our data reveal that sumoylation of Rap1 functions in a homeostatic feedback loop that sustains RPG transcription during translational stress. Taken together, Sumo regulates the cellular translational capacity by promoting transcription of tRNA genes and RPGs.


Assuntos
RNA Fúngico/isolamento & purificação , Proteínas de Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/genética , Sumoilação , Fator de Transcrição TFIID/genética , Fatores de Transcrição/genética , Proteínas rap1 de Ligação ao GTP/genética , Cromatina/genética , Cromatina/metabolismo , Estudos de Associação Genética , Regiões Promotoras Genéticas , RNA Fúngico/genética , Proteína SUMO-1/genética , Proteína SUMO-1/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Análise de Sequência de RNA , Transdução de Sinais , Fator de Transcrição TFIID/metabolismo , Fatores de Transcrição/metabolismo , Proteínas rap1 de Ligação ao GTP/metabolismo
20.
Cell Rep ; 5(4): 1036-46, 2013 Nov 27.
Artigo em Inglês | MEDLINE | ID: mdl-24239358

RESUMO

Very long chain fatty acids (VLCFAs) are essential fatty acids with multiple functions, including ceramide synthesis. Although the components of the VLCFA biosynthetic machinery have been elucidated, how their activity is regulated to meet the cell's metabolic demand remains unknown. The goal of this study was to identify mechanisms that regulate the rate of VLCFA synthesis, and we discovered that the fatty acid elongase Elo2 is regulated by phosphorylation. Elo2 phosphorylation is induced upon inhibition of TORC1 and requires GSK3. Expression of nonphosphorylatable Elo2 profoundly alters the ceramide spectrum, reflecting aberrant VLCFA synthesis. Furthermore, VLCFA depletion results in constitutive activation of autophagy, which requires sphingoid base phosphorylation. This constitutive activation of autophagy diminishes cell survival, indicating that VLCFAs serve to dampen the amplitude of autophagy. Together, our data reveal a function for TORC1 and GSK3 in the regulation of VLCFA synthesis that has important implications for autophagy and cell homeostasis.


Assuntos
Acetiltransferases/metabolismo , Ácidos Graxos Essenciais/biossíntese , Quinase 3 da Glicogênio Sintase/metabolismo , Proteínas de Membrana/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/metabolismo , Fatores de Transcrição/metabolismo , Acetiltransferases/biossíntese , Autofagia , Sobrevivência Celular , Ceramidas/biossíntese , Proteínas de Membrana/biossíntese , Fosforilação , Proteínas de Saccharomyces cerevisiae/antagonistas & inibidores , Proteínas de Saccharomyces cerevisiae/biossíntese , Fatores de Transcrição/antagonistas & inibidores
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