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1.
BMC Cancer ; 21(1): 211, 2021 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-33648471

RESUMO

BACKGROUND: Meningiomas are common brain tumours that are usually defined by benign clinical course. However, some meningiomas undergo a malignant transformation and recur within a short time period regardless of their World Health Organization (WHO) grade. The current study aimed to identify potential markers that can discriminate between benign and malignant meningioma courses. METHODS: We profiled the metabolites from 43 patients with low- and high-grade meningiomas. Tumour specimens were analyzed by nuclear magnetic resonance analysis; 270 metabolites were identified and clustered with the AutoPipe algorithm. RESULTS: We observed two distinct clusters marked by alterations in glycine/serine and choline/tryptophan metabolism. Glycine/serine cluster showed significantly lower WHO grades and proliferation rates. Also progression-free survival was significantly longer in the glycine/serine cluster. CONCLUSION: Our findings suggest that alterations in glycine/serine metabolism are associated with lower proliferation and more recurrent tumours. Altered choline/tryptophan metabolism was associated with increases proliferation, and recurrence. Our results suggest that tumour malignancy can be reflected by metabolic alterations, which may support histological classifications to predict the clinical outcome of patients with meningiomas.


Assuntos
Biomarcadores Tumorais/análise , Neoplasias Meníngeas/metabolismo , Meningioma/metabolismo , Idoso , Algoritmos , Colina/metabolismo , Análise por Conglomerados , Progressão da Doença , Feminino , Glicina/metabolismo , Humanos , Masculino , Neoplasias Meníngeas/química , Neoplasias Meníngeas/mortalidade , Meningioma/química , Meningioma/mortalidade , Pessoa de Meia-Idade , Gradação de Tumores , Ressonância Magnética Nuclear Biomolecular , Intervalo Livre de Progressão , Serina/metabolismo , Resultado do Tratamento , Triptofano/metabolismo
2.
Nat Plants ; 7(3): 310-316, 2021 03.
Artigo em Inglês | MEDLINE | ID: mdl-33686225

RESUMO

The nitrate transporter NRT2.1, which plays a central role in high-affinity nitrate uptake in roots, is activated at the post-translational level in response to nitrogen (N) starvation1,2. However, the critical enzymes required for the post-translational activation of NRT2.1 remain to be identified. Here, we show that a type 2C protein phosphatase, designated CEPD-induced phosphatase (CEPH), activates high-affinity nitrate uptake by directly dephosphorylating Ser501 of NRT2.1, a residue that functions as a negative phospho-switch in Arabidopsis2. CEPH is predominantly expressed in epidermal and cortex cells in roots and is upregulated by N starvation via a CEPDL2/CEPD1/2-mediated long-distance signalling from shoots3,4. The loss of CEPH leads to marked decreases in high-affinity nitrate uptake, tissue nitrate content and plant biomass. Collectively, our results identify CEPH as a crucial enzyme in the N-starvation-dependent activation of NRT2.1 and provide molecular and mechanistic insights into how plants regulate high-affinity nitrate uptake at the post-translational level in response to the N environment.


Assuntos
Proteínas de Transporte de Ânions/metabolismo , Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Nitratos/metabolismo , Arabidopsis/enzimologia , Glutarredoxinas/metabolismo , Fosforilação , Serina/metabolismo
3.
Nat Commun ; 12(1): 1899, 2021 03 26.
Artigo em Inglês | MEDLINE | ID: mdl-33771996

RESUMO

Polo-like kinase 1 (Plk1) is instrumental for mitotic entry and progression. Plk1 is activated by phosphorylation on a conserved residue Thr210 in its activation segment by the Aurora A kinase (AURKA), a reaction that critically requires the co-factor Bora phosphorylated by a CyclinA/B-Cdk1 kinase. Here we show that phospho-Bora is a direct activator of AURKA kinase activity. We localize the key determinants of phospho-Bora function to a 100 amino acid region encompassing two short Tpx2-like motifs and a phosphoSerine-Proline motif at Serine 112, through which Bora binds AURKA. The latter substitutes in trans for the Thr288 phospho-regulatory site of AURKA, which is essential for an active conformation of the kinase domain. We demonstrate the importance of these determinants for Bora function in mitotic entry both in Xenopus egg extracts and in human cells. Our findings unveil the activation mechanism of AURKA that is critical for mitotic entry.


Assuntos
Aurora Quinase A/metabolismo , Proteínas de Ciclo Celular/metabolismo , Mitose , Treonina/metabolismo , Motivos de Aminoácidos/genética , Animais , Aurora Quinase A/genética , Proteínas de Ciclo Celular/genética , Linhagem Celular Tumoral , Ciclina A2/genética , Ciclina A2/metabolismo , Quinase 2 Dependente de Ciclina/genética , Quinase 2 Dependente de Ciclina/metabolismo , Ativação Enzimática , Feminino , Humanos , Oócitos/metabolismo , Fosforilação , Prolina/genética , Prolina/metabolismo , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , Serina/genética , Serina/metabolismo , Treonina/genética , Xenopus laevis
4.
Nat Commun ; 12(1): 1392, 2021 03 02.
Artigo em Inglês | MEDLINE | ID: mdl-33654102

RESUMO

Rice grains typically contain high levels of toxic arsenic but low levels of the essential micronutrient selenium. Anthropogenic arsenic contamination of paddy soils exacerbates arsenic toxicity in rice crops resulting in substantial yield losses. Here, we report the identification of the gain-of-function arsenite tolerant 1 (astol1) mutant of rice that benefits from enhanced sulfur and selenium assimilation, arsenic tolerance, and decreased arsenic accumulation in grains. The astol1 mutation promotes the physical interaction of the chloroplast-localized O-acetylserine (thiol) lyase protein with its interaction partner serine-acetyltransferase in the cysteine synthase complex. Activation of the serine-acetyltransferase in this complex promotes the uptake of sulfate and selenium and enhances the production of cysteine, glutathione, and phytochelatins, resulting in increased tolerance and decreased translocation of arsenic to grains. Our findings uncover the pivotal sensing-function of the cysteine synthase complex in plastids for optimizing stress resilience and grain quality by regulating a fundamental macronutrient assimilation pathway.


Assuntos
Arsênico/metabolismo , Oryza/metabolismo , Sementes/metabolismo , Selênio/metabolismo , Enxofre/metabolismo , Alelos , Cloroplastos/metabolismo , Cisteína Sintase/metabolismo , Redes e Vias Metabólicas , Modelos Biológicos , Mutação/genética , Fenótipo , Fitoquelatinas/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Serina/metabolismo , Frações Subcelulares/metabolismo
5.
Int J Mol Sci ; 22(5)2021 Feb 25.
Artigo em Inglês | MEDLINE | ID: mdl-33669046

RESUMO

The phosphorylation of serine 10 in histone 3 (p-S10H3) has recently been demonstrated to participate in spinal nociceptive processing. However, superficial dorsal horn (SDH) neurons involved in p-S10H3-mediated nociception have not been fully characterized. In the present work, we combined immunohistochemistry, in situ hybridization with the retrograde labeling of projection neurons to reveal the subset of dorsal horn neurons presenting an elevated level of p-S10H3 in response to noxious heat (60 °C), causing burn injury. Projection neurons only represented a small percentage (5%) of p-S10H3-positive cells, while the greater part of them belonged to excitatory SDH interneurons. The combined immunolabeling of p-S10H3 with markers of already established interneuronal classes of the SDH revealed that the largest subset of neurons with burn injury-induced p-S10H3 expression was dynorphin immunopositive in mice. Furthermore, the majority of p-S10H3-expressing dynorphinergic neurons proved to be excitatory, as they lacked Pax-2 and showed Lmx1b-immunopositivity. Thus, we showed that neurochemically heterogeneous SDH neurons exhibit the upregulation of p-S10H3 shortly after noxious heat-induced burn injury and consequential tissue damage, and that a dedicated subset of excitatory dynorphinergic neurons is likely a key player in the development of central sensitization via the p-S10H3 mediated pathway.


Assuntos
Queimaduras/metabolismo , Histonas/metabolismo , Nociceptividade/fisiologia , Dor/metabolismo , Células do Corno Posterior/metabolismo , Serina/metabolismo , Medula Espinal/metabolismo , Animais , Epigênese Genética , Feminino , Imuno-Histoquímica , Hibridização In Situ , Proteínas com Homeodomínio LIM/metabolismo , Masculino , Camundongos , Camundongos Transgênicos , Fator de Transcrição PAX2/metabolismo , Fosforilação , Ratos , Ratos Wistar , Medula Espinal/citologia , Medula Espinal/fisiologia , Fatores de Transcrição/metabolismo
6.
Nat Commun ; 12(1): 1676, 2021 03 15.
Artigo em Inglês | MEDLINE | ID: mdl-33723254

RESUMO

The recently identified Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is the cause of the COVID-19 pandemic. How this novel beta-coronavirus virus, and coronaviruses more generally, alter cellular metabolism to support massive production of ~30 kB viral genomes and subgenomic viral RNAs remains largely unknown. To gain insights, transcriptional and metabolomic analyses are performed 8 hours after SARS-CoV-2 infection, an early timepoint where the viral lifecycle is completed but prior to overt effects on host cell growth or survival. Here, we show that SARS-CoV-2 remodels host folate and one-carbon metabolism at the post-transcriptional level to support de novo purine synthesis, bypassing viral shutoff of host translation. Intracellular glucose and folate are depleted in SARS-CoV-2-infected cells, and viral replication is exquisitely sensitive to inhibitors of folate and one-carbon metabolism, notably methotrexate. Host metabolism targeted therapy could add to the armamentarium against future coronavirus outbreaks.


Assuntos
/metabolismo , Carbono/metabolismo , Ácido Fólico/metabolismo , Replicação Viral , Células A549 , Animais , Chlorocebus aethiops , Efeito Citopatogênico Viral , Antagonistas do Ácido Fólico/farmacologia , Glucose/metabolismo , Humanos , Metotrexato/farmacologia , RNA Viral/biossíntese , Serina/metabolismo , Transcrição Genética , Células Vero , Proteínas Virais/genética , Replicação Viral/efeitos dos fármacos
7.
Nat Commun ; 12(1): 1028, 2021 02 15.
Artigo em Inglês | MEDLINE | ID: mdl-33589610

RESUMO

Upon binding to DNA breaks, poly(ADP-ribose) polymerase 1 (PARP1) ADP-ribosylates itself and other factors to initiate DNA repair. Serine is the major residue for ADP-ribosylation upon DNA damage, which strictly depends on HPF1. Here, we report the crystal structures of human HPF1/PARP1-CAT ΔHD complex at 1.98 Å resolution, and mouse and human HPF1 at 1.71 Å and 1.57 Å resolution, respectively. Our structures and mutagenesis data confirm that the structural insights obtained in a recent HPF1/PARP2 study by Suskiewicz et al. apply to PARP1. Moreover, we quantitatively characterize the key residues necessary for HPF1/PARP1 binding. Our data show that through salt-bridging to Glu284/Asp286, Arg239 positions Glu284 to catalyze serine ADP-ribosylation, maintains the local conformation of HPF1 to limit PARP1 automodification, and facilitates HPF1/PARP1 binding by neutralizing the negative charge of Glu284. These findings, along with the high-resolution structural data, may facilitate drug discovery targeting PARP1.


Assuntos
Proteínas de Transporte/química , DNA/química , Histonas/química , Proteínas Nucleares/química , Poli(ADP-Ribose) Polimerase-1/química , Serina/metabolismo , ADP-Ribosilação , Sequência de Aminoácidos , Animais , Sítios de Ligação , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Clonagem Molecular , Cristalografia por Raios X , DNA/genética , DNA/metabolismo , Escherichia coli/genética , Escherichia coli/metabolismo , Expressão Gênica , Vetores Genéticos/química , Vetores Genéticos/metabolismo , Glutamina/metabolismo , Histonas/genética , Histonas/metabolismo , Humanos , Camundongos , Modelos Moleculares , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Poli(ADP-Ribose) Polimerase-1/genética , Poli(ADP-Ribose) Polimerase-1/metabolismo , Ligação Proteica , Conformação Proteica em alfa-Hélice , Conformação Proteica em Folha beta , Domínios e Motivos de Interação entre Proteínas , Isoformas de Proteínas/química , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Eletricidade Estática
8.
Nat Struct Mol Biol ; 28(3): 249-257, 2021 03.
Artigo em Inglês | MEDLINE | ID: mdl-33558762

RESUMO

Human serine palmitoyltransferase (SPT) complex catalyzes the initial and rate-limiting step in the de novo biosynthesis of all sphingolipids. ORMDLs regulate SPT function, with human ORMDL3 being related to asthma. Here we report three high-resolution cryo-EM structures: the human SPT complex, composed of SPTLC1, SPTLC2 and SPTssa; the SPT-ORMDL3 complex; and the SPT-ORMDL3 complex bound to two substrates, PLP-L-serine (PLS) and a non-reactive palmitoyl-CoA analogue. SPTLC1 and SPTLC2 form a dimer of heterodimers as the catalytic core. SPTssa participates in acyl-CoA coordination, thereby stimulating the SPT activity and regulating the substrate selectivity. ORMDL3 is located in the center of the complex, serving to stabilize the SPT assembly. Our structural and biochemical analyses provide a molecular basis for the assembly and substrate selectivity of the SPT and SPT-ORMDL3 complexes, and lay a foundation for mechanistic understanding of sphingolipid homeostasis and for related therapeutic drug development.


Assuntos
Microscopia Crioeletrônica , Proteínas de Membrana/química , Proteínas de Membrana/metabolismo , Complexos Multiproteicos , Serina C-Palmitoiltransferase/química , Serina C-Palmitoiltransferase/metabolismo , Acil Coenzima A/química , Acil Coenzima A/metabolismo , Acil Coenzima A/ultraestrutura , Sítios de Ligação , Biocatálise , Humanos , Proteínas de Membrana/genética , Proteínas de Membrana/ultraestrutura , Modelos Moleculares , Complexos Multiproteicos/química , Complexos Multiproteicos/genética , Complexos Multiproteicos/metabolismo , Complexos Multiproteicos/ultraestrutura , Mutação , Fosfato de Piridoxal/química , Fosfato de Piridoxal/metabolismo , Reprodutibilidade dos Testes , Serina/química , Serina/metabolismo , Serina C-Palmitoiltransferase/genética , Serina C-Palmitoiltransferase/ultraestrutura , Especificidade por Substrato
9.
Nat Genet ; 53(3): 342-353, 2021 03.
Artigo em Inglês | MEDLINE | ID: mdl-33558758

RESUMO

Technological and computational advances in genomics and interactomics have made it possible to identify how disease mutations perturb protein-protein interaction (PPI) networks within human cells. Here, we show that disease-associated germline variants are significantly enriched in sequences encoding PPI interfaces compared to variants identified in healthy participants from the projects 1000 Genomes and ExAC. Somatic missense mutations are also significantly enriched in PPI interfaces compared to noninterfaces in 10,861 tumor exomes. We computationally identified 470 putative oncoPPIs in a pan-cancer analysis and demonstrate that oncoPPIs are highly correlated with patient survival and drug resistance/sensitivity. We experimentally validate the network effects of 13 oncoPPIs using a systematic binary interaction assay, and also demonstrate the functional consequences of two of these on tumor cell growth. In summary, this human interactome network framework provides a powerful tool for prioritization of alleles with PPI-perturbing mutations to inform pathobiological mechanism- and genotype-based therapeutic discovery.


Assuntos
Biologia Computacional/métodos , Mutação , Neoplasias/genética , Mapas de Interação de Proteínas/genética , Araquidonato 5-Lipoxigenase/genética , Araquidonato 5-Lipoxigenase/metabolismo , Arginina/genética , Arginina/metabolismo , Doença/genética , Genoma Humano , Histonas/genética , Histonas/metabolismo , Humanos , Testes Farmacogenômicos , Pró-Proteína Convertase 9/genética , Pró-Proteína Convertase 9/metabolismo , Receptores de LDL/genética , Receptores de LDL/metabolismo , Reprodutibilidade dos Testes , Serina/genética , Serina/metabolismo , Inibidor alfa de Dissociação do Nucleotídeo Guanina rho/genética , Inibidor alfa de Dissociação do Nucleotídeo Guanina rho/metabolismo , Proteína rhoA de Ligação ao GTP/genética , Proteína rhoA de Ligação ao GTP/metabolismo
10.
Cancer Sci ; 112(5): 1723-1734, 2021 May.
Artigo em Inglês | MEDLINE | ID: mdl-33609296

RESUMO

T cells could be engineered to overcome the aberrant metabolic milieu of solid tumors and tip the balance in favor of a long-lasting clinical response. Here, we explored the therapeutic potential of stably overexpressing cystathionine-gamma-lyase (CTH, CSE, or cystathionase), a pivotal enzyme of the transsulfuration pathway, in antitumor CD8+ T cells with the initial aim to boost intrinsic cysteine metabolism. Using a mouse model of adoptive cell transfer (ACT), we found that CTH-expressing T cells showed a superior control of tumor growth compared to control T cells. However, contrary to our hypothesis, this effect was not associated with increased T cell expansion in vivo or proliferation rescue in the absence of cysteine/cystine in vitro. Rather than impacting methionine or cysteine, ACT with CTH overexpression unexpectedly reduced glycine, serine, and proline concentration within the tumor interstitial fluid. Interestingly, in vitro tumor cell growth was mostly impacted by the combination of serine/proline or serine/glycine deprivation. These results suggest that metabolic gene engineering of T cells could be further investigated to locally modulate amino acid availability within the tumor environment while avoiding systemic toxicity.


Assuntos
Transferência Adotiva/métodos , Linfócitos T CD8-Positivos/metabolismo , Cistationina gama-Liase/metabolismo , Cisteína/biossíntese , Animais , Engenharia Celular , Linhagem Celular Tumoral , Proliferação de Células , Líquido Extracelular/metabolismo , Feminino , Glicina/metabolismo , Metionina/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Modelos Animais , Neoplasias Ovarianas/terapia , Prolina/metabolismo , Serina/metabolismo , Microambiente Tumoral/imunologia
11.
Nat Metab ; 3(2): 131-141, 2021 02.
Artigo em Inglês | MEDLINE | ID: mdl-33510397

RESUMO

Cancer cells reprogramme their metabolism to support unrestrained proliferation and survival in nutrient-poor conditions. Whereas non-transformed cells often have lower demands for serine and glycine, several cancer subtypes hyperactivate intracellular serine and glycine synthesis and become addicted to de novo production. Copy-number amplifications of serine- and glycine-synthesis genes and genetic alterations in common oncogenes and tumour-suppressor genes enhance serine and glycine synthesis, resulting in high production and secretion of these oncogenesis-supportive metabolites. In this Review, we discuss the contribution of serine and glycine synthesis to cancer progression. By relying on de novo synthesis pathways, cancer cells are able to enhance macromolecule synthesis, neutralize high levels of oxidative stress and regulate methylation and tRNA formylation. Furthermore, we discuss the immunosuppressive potential of serine and glycine, and the essentiality of both amino acids to promoting survival of non-transformed neighbouring cells. Finally, we point to the emerging data proposing moonlighting functions of serine- and glycine-synthesis enzymes and examine promising small molecules targeting serine and glycine synthesis.


Assuntos
Glicina/metabolismo , Neoplasias/metabolismo , Serina/metabolismo , Animais , Humanos
12.
J Virol ; 95(6)2021 02 24.
Artigo em Inglês | MEDLINE | ID: mdl-33408177

RESUMO

Influenza A virus (IAV) nonstructural protein 1 (NS1) is a protein with multiple functions that are regulated by phosphorylation. Phosphoproteomic screening of H1N1 virus-infected cells revealed that NS1 was phosphorylated at serine 205 in intermediate stages of the viral life cycle. Interestingly, S205 is one of six amino acid changes in NS1 of post-pandemic H1N1 viruses currently circulating in humans compared to the original swine-origin 2009 pandemic (H1N1pdm09) virus, suggesting a role in host adaptation. To identify NS1 functions regulated by S205 phosphorylation, we generated recombinant PR8 H1N1 NS1 mutants with S205G (nonphosphorylatable) or S205N (H1N1pdm09 signature), as well as H1N1pdm09 viruses harboring the reverse mutation NS1 N205S or N205D (phosphomimetic). Replication of PR8 NS1 mutants was attenuated relative to wild-type (WT) virus replication in a porcine cell line. However, PR8 NS1 S205N showed remarkably higher attenuation than PR8 NS1 S205G in a human cell line, highlighting a potential host-independent advantage of phosphorylatable S205, while an asparagine at this position led to a potential host-specific attenuation. Interestingly, PR8 NS1 S205G did not show polymerase activity-enhancing functions, in contrast to the WT, which can be attributed to diminished interaction with cellular restriction factor DDX21. Analysis of the respective kinase mediating S205 phosphorylation indicated an involvement of casein kinase 2 (CK2). CK2 inhibition significantly reduced the replication of WT viruses and decreased NS1-DDX21 interaction, as observed for NS1 S205G. In summary, NS1 S205 is required for efficient NS1-DDX21 binding, resulting in enhanced viral polymerase activity, which is likely to be regulated by transient phosphorylation.IMPORTANCE Influenza A viruses (IAVs) still pose a major threat to human health worldwide. As a zoonotic virus, IAV can spontaneously overcome species barriers and even reside in new hosts after efficient adaptation. Investigation of the functions of specific adaptational mutations can lead to a deeper understanding of viral replication in specific hosts and can probably help to find new targets for antiviral intervention. In the present study, we analyzed the role of NS1 S205, a phosphorylation site that was reacquired during the circulation of pandemic H1N1pdm09 "swine flu" in the human host. We found that phosphorylation of human H1N1 virus NS1 S205 is mediated by the cellular kinase CK2 and is needed for efficient interaction with human host restriction factor DDX21, mediating NS1-induced enhancement of viral polymerase activity. Therefore, targeting CK2 activity might be an efficient strategy for limiting the replication of IAVs circulating in the human population.


Assuntos
Vírus da Influenza A/fisiologia , Serina/metabolismo , Proteínas não Estruturais Virais/metabolismo , Adaptação Fisiológica/genética , Animais , Caseína Quinase II/metabolismo , Linhagem Celular , RNA Helicases DEAD-box/metabolismo , Interações Hospedeiro-Patógeno , Humanos , Vírus da Influenza A Subtipo H1N1/genética , Vírus da Influenza A Subtipo H1N1/metabolismo , Vírus da Influenza A Subtipo H1N1/fisiologia , Vírus da Influenza A/genética , Vírus da Influenza A/metabolismo , Mutação , Fosforilação , Ligação Proteica , Suínos , Proteínas não Estruturais Virais/genética , Replicação Viral
13.
Nat Commun ; 12(1): 704, 2021 01 29.
Artigo em Inglês | MEDLINE | ID: mdl-33514736

RESUMO

p53 mutations with single amino acid changes in cancer often lead to dominant oncogenic changes. Here, we have developed a mouse model of gain-of-function (GOF) p53-driven lung cancer utilizing conditionally active LSL p53-R172H and LSL K-Ras-G12D knock-in alleles that can be activated by Cre in lung club cells. Mutation of the p53 transactivation domain (TAD) (p53-L25Q/W26S/R172H) eliminating significant transactivation activity resulted in loss of tumorigenicity, demonstrating that transactivation mediated by or dependent on TAD is required for oncogenicity by GOF p53. GOF p53 TAD mutations significantly reduce phosphorylation of nearby p53 serine 20 (S20), which is a target for PLK3 phosphorylation. Knocking out PLK3 attenuated S20 phosphorylation along with transactivation and oncogenicity by GOF p53, indicating that GOF p53 exploits PLK3 to trigger its transactivation capability and exert oncogenic functions. Our data show a mechanistic involvement of PLK3 in mutant p53 pathway of oncogenesis.


Assuntos
Carcinogênese/genética , Neoplasias Pulmonares/genética , Proteínas Serina-Treonina Quinases/metabolismo , Proteína Supressora de Tumor p53/genética , Animais , Carcinogênese/patologia , Linhagem Celular Tumoral , Modelos Animais de Doenças , Feminino , Mutação com Ganho de Função , Técnicas de Introdução de Genes , Técnicas de Inativação de Genes , Humanos , Pulmão/patologia , Neoplasias Pulmonares/patologia , Camundongos , Camundongos Transgênicos , Fosforilação/genética , Domínios Proteicos/genética , Proteínas Serina-Treonina Quinases/genética , Serina/metabolismo , Esferoides Celulares , Ativação Transcricional , Proteína Supressora de Tumor p53/metabolismo
14.
Nat Genet ; 53(1): 54-64, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-33414548

RESUMO

In cross-platform analyses of 174 metabolites, we identify 499 associations (P < 4.9 × 10-10) characterized by pleiotropy, allelic heterogeneity, large and nonlinear effects and enrichment for nonsynonymous variation. We identify a signal at GLP2R (p.Asp470Asn) shared among higher citrulline levels, body mass index, fasting glucose-dependent insulinotropic peptide and type 2 diabetes, with ß-arrestin signaling as the underlying mechanism. Genetically higher serine levels are shown to reduce the likelihood (by 95%) and predict development of macular telangiectasia type 2, a rare degenerative retinal disease. Integration of genomic and small molecule data across platforms enables the discovery of regulators of human metabolism and translation into clinical insights.


Assuntos
Saúde , Metabolismo/genética , Diabetes Mellitus Tipo 2/genética , Oftalmopatias/genética , Frequência do Gene/genética , Loci Gênicos , Pleiotropia Genética , Genoma Humano , Receptor do Peptídeo Semelhante ao Glucagon 2/genética , Glicina/metabolismo , Humanos , Modelos Lineares , Análise da Randomização Mendeliana , Erros Inatos do Metabolismo/genética , Metaboloma/genética , Mutação de Sentido Incorreto/genética , Fenótipo , Polimorfismo de Nucleotídeo Único/genética , Telangiectasia Retiniana/genética , Tamanho da Amostra , Serina/metabolismo
15.
Nat Commun ; 12(1): 594, 2021 01 26.
Artigo em Inglês | MEDLINE | ID: mdl-33500413

RESUMO

Telomeres are organized into a heterochromatin structure and maintenance of silent heterochromatin is required for chromosome stability. How telomere heterochromatin is dynamically regulated in response to stimuli remains unknown. Pyruvate kinase Pyk1 forms a complex named SESAME (Serine-responsive SAM-containing Metabolic Enzyme complex) to regulate gene expression by phosphorylating histone H3T11 (H3pT11). Here, we identify a function of SESAME in regulating telomere heterochromatin structure. SESAME phosphorylates H3T11 at telomeres, which maintains SIR (silent information regulator) complex occupancy at telomeres and protects Sir2 from degradation by autophagy. Moreover, SESAME-catalyzed H3pT11 directly represses autophagy-related gene expression to further prevent autophagy-mediated Sir2 degradation. By promoting H3pT11, serine increases Sir2 protein levels and enhances telomere silencing. Loss of H3pT11 leads to reduced Sir2 and compromised telomere silencing during chronological aging. Together, our study provides insights into dynamic regulation of silent heterochromatin by histone modifications and autophagy in response to cell metabolism and aging.


Assuntos
Instabilidade Cromossômica , Histonas/metabolismo , Complexos Multienzimáticos/metabolismo , Saccharomyces cerevisiae/genética , Telômero/metabolismo , Autofagia , Regulação Fúngica da Expressão Gênica , Heterocromatina/metabolismo , Fosforilação , Proteólise , Piruvato Quinase/metabolismo , Saccharomyces cerevisiae/enzimologia , Serina/metabolismo , Proteínas Reguladoras de Informação Silenciosa de Saccharomyces cerevisiae/metabolismo , Sirtuína 2/metabolismo
16.
Biochem J ; 478(2): 299-326, 2021 01 29.
Artigo em Inglês | MEDLINE | ID: mdl-33367571

RESUMO

Mutations that increase the protein kinase activity of LRRK2 are one of the most common causes of familial Parkinson's disease. LRRK2 phosphorylates a subset of Rab GTPases within their Switch-II motif, impacting interaction with effectors. We describe and validate a new, multiplexed targeted mass spectrometry assay to quantify endogenous levels of LRRK2-phosphorylated Rab substrates (Rab1, Rab3, Rab8, Rab10, Rab35 and Rab43) as well as total levels of Rabs, LRRK2 and LRRK2-phosphorylated at the Ser910 and Ser935 biomarker sites. Exploiting this assay, we quantify for the first time the relative levels of each of the pRab proteins in different cells (mouse embryonic fibroblasts, human neutrophils) and mouse tissues (brain, kidney, lung and spleen). We define how these components are impacted by Parkinson's pathogenic mutations (LRRK2[R1441C] and VPS35[D620N]) and LRRK2 inhibitors. We find that the VPS35[D620N], but not LRRK2[R1441C] mutation, enhances Rab1 phosphorylation in a manner blocked by administration of an LRRK2 inhibitor, providing the first evidence that endogenous Rab1 is a physiological substrate for LRRK2. We exploit this assay to demonstrate that in Parkinson's patients with VPS35[D620N] mutations, phosphorylation of multiple Rab proteins (Rab1, Rab3, Rab8, Rab10 and Rab43) is elevated. We highlight the benefits of this assay over immunoblotting approaches currently deployed to assess LRRK2 Rab signalling pathway.


Assuntos
Biomarcadores/análise , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/metabolismo , Espectrometria de Massas em Tandem/métodos , Proteínas rab de Ligação ao GTP/metabolismo , Animais , Biomarcadores/metabolismo , Fibroblastos/metabolismo , Humanos , Imunoprecipitação/métodos , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/antagonistas & inibidores , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/genética , Limite de Detecção , Camundongos Mutantes , Mutação , Doença de Parkinson/genética , Fosforilação , Serina/metabolismo , Proteínas de Transporte Vesicular/genética , Proteínas de Transporte Vesicular/metabolismo
17.
Biochim Biophys Acta Proteins Proteom ; 1869(1): 140542, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-32950692

RESUMO

Among aging-induced impairments, those affecting cognitive functions certainly represent one the most major challenge to face to improve elderly quality of life. In last decades, our knowledge on changes in the morphology and function of neuronal networks associated with normal and pathological brain aging has rapidly progressed, initiating the development of different pharmacological and behavioural strategies to alleviate cognitive aging. In particular, experimental evidences have accumulated indicating that the communication between neurons and its plasticity gradually weakens with aging. Because of its pivotal role for brain functional plasticity, the N-Methyl­d-Aspartate receptor subtype of glutamate receptors (NMDAr) has gathered much of the experimental interest. NMDAr activation is regulated by many mechanisms. Among is the mandatory binding of a co-agonist, such as the amino acid d-serine, in order to activate NMDAr. This mini-review presents the most recent information indicating how d-serine could contribute to mechanisms of physiological cognitive aging and also considers the divergent views relative of the role of the NMDAr co-agonist in Alzheimer's disease.


Assuntos
Envelhecimento/metabolismo , Doença de Alzheimer/metabolismo , Plasticidade Neuronal/fisiologia , Receptores de N-Metil-D-Aspartato/metabolismo , Serina/metabolismo , Envelhecimento/patologia , Doença de Alzheimer/genética , Doença de Alzheimer/patologia , Animais , Astrócitos/metabolismo , Astrócitos/patologia , Cognição/fisiologia , Expressão Gênica , Glucose/metabolismo , Ácido Glutâmico/metabolismo , Hipocampo/metabolismo , Hipocampo/patologia , Humanos , Memória/fisiologia , Neurônios/metabolismo , Neurônios/patologia , Racemases e Epimerases/genética , Racemases e Epimerases/metabolismo , Receptores de N-Metil-D-Aspartato/genética
18.
Nat Commun ; 11(1): 5003, 2020 10 06.
Artigo em Inglês | MEDLINE | ID: mdl-33024099

RESUMO

Recognition of a start codon by the initiator aminoacyl-tRNA determines the reading frame of messenger RNA (mRNA) translation by the ribosome. In eukaryotes, the GTPase eIF5B collaborates in the correct positioning of the initiator Met-tRNAiMet on the ribosome in the later stages of translation initiation, gating entrance into elongation. Leveraging the long residence time of eIF5B on the ribosome recently identified by single-molecule fluorescence measurements, we determine the cryoEM structure of the naturally long-lived ribosome complex with eIF5B and Met-tRNAiMet immediately before transition into elongation. The structure uncovers an unexpected, eukaryotic specific and dynamic fidelity checkpoint implemented by eIF5B in concert with components of the large ribosomal subunit.


Assuntos
Fatores de Iniciação em Eucariotos/química , Fatores de Iniciação em Eucariotos/metabolismo , Elongação Traducional da Cadeia Peptídica , Iniciação Traducional da Cadeia Peptídica , Subunidades Ribossômicas Maiores/metabolismo , Acilação , Anticódon , Microscopia Crioeletrônica , Fatores de Iniciação em Eucariotos/genética , Guanosina Difosfato/metabolismo , Modelos Moleculares , Conformação de Ácido Nucleico , RNA de Transferência de Metionina/química , RNA de Transferência de Metionina/metabolismo , Subunidades Ribossômicas Maiores/química , Subunidades Ribossômicas Maiores/genética , Subunidades Ribossômicas Maiores de Eucariotos , Subunidades Ribossômicas Menores de Eucariotos , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/química , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Serina/metabolismo
19.
Nat Commun ; 11(1): 5007, 2020 10 06.
Artigo em Inglês | MEDLINE | ID: mdl-33024116

RESUMO

p50, the mature product of NFKB1, is constitutively produced from its precursor, p105. Here, we identify BARD1 as a p50-interacting factor. p50 directly associates with the BARD1 BRCT domains via a C-terminal phospho-serine motif. This interaction is induced by ATR and results in mono-ubiquitination of p50 by the BARD1/BRCA1 complex. During the cell cycle, p50 is mono-ubiquitinated in S phase and loss of this post-translational modification increases S phase progression and chromosomal breakage. Genome-wide studies reveal a substantial decrease in p50 chromatin enrichment in S phase and Cycln E is identified as a factor regulated by p50 during the G1 to S transition. Functionally, interaction with BARD1 promotes p50 protein stability and consistent with this, in human cancer specimens, low nuclear BARD1 protein strongly correlates with low nuclear p50. These data indicate that p50 mono-ubiquitination by BARD1/BRCA1 during the cell cycle regulates S phase progression to maintain genome integrity.


Assuntos
Neoplasias da Mama/metabolismo , Ciclo Celular/fisiologia , Instabilidade Genômica , Subunidade p50 de NF-kappa B/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Animais , Sítios de Ligação , Neoplasias da Mama/mortalidade , Linhagem Celular Tumoral , Feminino , Fibroblastos , Humanos , Lisina/metabolismo , Camundongos , Subunidade p50 de NF-kappa B/genética , Neuroblastoma/metabolismo , Domínios Proteicos , Processamento de Proteína Pós-Traducional , Serina/metabolismo , Proteínas Supressoras de Tumor/genética , Ubiquitina-Proteína Ligases/genética , Ubiquitinação
20.
Nat Commun ; 11(1): 5052, 2020 10 07.
Artigo em Inglês | MEDLINE | ID: mdl-33028817

RESUMO

The mechanism and function of autophagy as a highly-conserved bulk degradation pathway are well studied, but the physiological role of autophagy remains poorly understood. We show that autophagy is involved in the adaptation of Saccharomyces cerevisiae to respiratory growth through its recycling of serine. On respiratory media, growth onset, mitochondrial initiator tRNA modification and mitochondrial protein expression are delayed in autophagy defective cells, suggesting that mitochondrial one-carbon metabolism is perturbed in these cells. The supplementation of serine, which is a key one-carbon metabolite, is able to restore mitochondrial protein expression and alleviate delayed respiratory growth. These results indicate that autophagy-derived serine feeds into mitochondrial one-carbon metabolism, supporting the initiation of mitochondrial protein synthesis and allowing rapid adaptation to respiratory growth.


Assuntos
Adaptação Fisiológica , Autofagia/fisiologia , Proteínas Mitocondriais/biossíntese , Saccharomyces cerevisiae/fisiologia , Carbono/metabolismo , Respiração Celular/fisiologia , Mitocôndrias/metabolismo , Biossíntese de Proteínas/fisiologia , RNA de Transferência/metabolismo , Saccharomyces cerevisiae/citologia , Proteínas de Saccharomyces cerevisiae/metabolismo , Serina/metabolismo
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