RESUMEN
D-3-phosphoglycerate dehydrogenase (PHGDH) catalyzes the NAD+-dependent conversion of D-3-phospho-glycerate to 3-phosphohydroxypyruvate, the first step in the phosphorylated pathway for L-serine (L-Ser) biosynthesis. L-Ser plays different relevant metabolic roles in eukaryotic cells: alterations in L-Ser metabolism have been linked to serious neurological disorders. The human PHGDH (hPHGDH), showing a homotetrameric state in solution, is made of four domains, among which there are two regulatory domains at the C-terminus: the aspartate kinase-chorismate mutase-tyrA prephenate dehydrogenase (ACT) and allosteric substrate-binding (ASB) domains. The structure of hPHGDH was solved only for a truncated, dimeric form harboring the N-terminal end containing the substrate and the cofactor binding domains. A model ensemble of the tetrameric hPHGDH was generated using AlphaFold coupled with molecular dynamics refinement. By analyzing the inter-subunit interactions at the tetrameric interface, the residues F418, L478, P479, R454, and Y495 were selected and their role was studied by the alanine-scanning mutagenesis approach. The F418A variant modifies the putative ASB, slightly alters the activity, the fraction of protein in the tetrameric state, and the protein stability; it seems relevant in dimers' recognition to yield the tetrameric oligomer. On the contrary, the R454A, L478A, P479A, and Y495A variants (ACT domain) determine a loss of the tetrameric assembly, resulting in low stability and misfolding, triggering the aggregation and hampering the activity. The predicted tetrameric interface seems mediated by residues at the ACT domain, and the tetramer formation seems crucial for proper folding of hPHGDH, which, in turn, is essential for both stability and functionality.
Asunto(s)
Fosfoglicerato-Deshidrogenasa , Fosfoglicerato-Deshidrogenasa/química , Fosfoglicerato-Deshidrogenasa/metabolismo , Fosfoglicerato-Deshidrogenasa/genética , Humanos , Estructura Cuaternaria de Proteína , Modelos Moleculares , Multimerización de Proteína , Simulación de Dinámica Molecular , Dominios Proteicos , Cristalografía por Rayos XRESUMEN
Cancer cells are often addicted to serine synthesis to support growth. How serine synthesis is regulated in cancer is not well understood. We recently demonstrated protein arginine methyltransferase 1 (PRMT1) is upregulated in hepatocellular carcinoma (HCC) to methylate and activate phosphoglycerate dehydrogenase (PHGDH), thereby promoting serine synthesis. However, the mechanisms underlying PRMT1 upregulation and regulation of PRMT1-PHGDH axis remain unclear. Here, we show the E3 ubiquitin ligase F-box-only protein 7 (FBXO7) inhibits serine synthesis in HCC by binding PRMT1, inducing lysine 37 ubiquitination, and promoting proteosomal degradation of PRMT1. FBXO7-mediated PRMT1 downregulation cripples PHGDH arginine methylation and activation, resulting in impaired serine synthesis, accumulation of reactive oxygen species (ROS), and inhibition of HCC cell growth. Notably, FBXO7 is significantly downregulated in human HCC tissues, and inversely associated with PRMT1 protein and PHGDH methylation level. Overall, our study provides mechanistic insights into the regulation of cancer serine synthesis by FBXO7-PRMT1-PHGDH axis, and will facilitate the development of serine-targeting strategies for cancer therapy.
Asunto(s)
Carcinoma Hepatocelular , Proteínas F-Box , Neoplasias Hepáticas , Fosfoglicerato-Deshidrogenasa , Proteína-Arginina N-Metiltransferasas , Serina , Ubiquitinación , Proteína-Arginina N-Metiltransferasas/metabolismo , Proteína-Arginina N-Metiltransferasas/genética , Humanos , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patología , Carcinoma Hepatocelular/genética , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patología , Neoplasias Hepáticas/genética , Proteínas F-Box/metabolismo , Proteínas F-Box/genética , Serina/metabolismo , Serina/biosíntesis , Fosfoglicerato-Deshidrogenasa/metabolismo , Fosfoglicerato-Deshidrogenasa/genética , Línea Celular Tumoral , Animales , Proteínas Represoras/metabolismo , Proteínas Represoras/genética , Ratones , Proliferación Celular , Metilación , Regulación Neoplásica de la Expresión Génica , Ratones Desnudos , Masculino , Células HEK293 , Femenino , Células Hep G2RESUMEN
Serine is a key contributor to the generation of one-carbon units for DNA synthesis during cellular proliferation. In addition, it plays a crucial role in the production of antioxidants that prevent abnormal proliferation and stress in cancer cells. In recent studies, the relationship between cancer metabolism and the serine biosynthesis pathway has been highlighted. In this context, 3-phosphoglycerate dehydrogenase (PHGDH) is notable as a key enzyme that functions as the primary rate-limiting enzyme in the serine biosynthesis pathway, facilitating the conversion of 3-phosphoglycerate to 3-phosphohydroxypyruvate. Elevated PHGDH activity in diverse cancer cells is mediated through genetic amplification, posttranslational modification, increased transcription, and allosteric regulation. Ultimately, these characteristics allow PHGDH to not only influence the growth and progression of cancer but also play an important role in metastasis and drug resistance. Consequently, PHGDH has emerged as a crucial focal point in cancer research. In this review, the structural aspects of PHGDH and its involvement in one-carbon metabolism are investigated, and PHGDH is proposed as a potential therapeutic target in diverse cancers. By elucidating how PHGDH expression promotes cancer growth, the goal of this review is to provide insight into innovative treatment strategies. This paper aims to reveal how PHGDH inhibitors can overcome resistance mechanisms, contributing to the development of effective cancer treatments.
Asunto(s)
Neoplasias , Fosfoglicerato-Deshidrogenasa , Fosfoglicerato-Deshidrogenasa/metabolismo , Fosfoglicerato-Deshidrogenasa/antagonistas & inhibidores , Fosfoglicerato-Deshidrogenasa/genética , Humanos , Neoplasias/metabolismo , Neoplasias/tratamiento farmacológico , Neoplasias/patología , Animales , Terapia Molecular Dirigida , Antineoplásicos/uso terapéutico , Antineoplásicos/farmacología , Serina/metabolismoRESUMEN
Mitochondrial dysfunction is the pivotal driving factor of multiple inflammatory diseases, and targeting mitochondrial biogenesis represents an efficacious approach to ameliorate such dysfunction in inflammatory diseases. Here, we demonstrated that phosphoglycerate dehydrogenase (PHGDH) deficiency promotes mitochondrial biogenesis in inflammatory macrophages. Mechanistically, PHGDH deficiency boosts mitochondrial reactive oxygen species (mtROS) by suppressing cytoplasmic glutathione synthesis. mtROS provokes hypoxia-inducible factor-1α signaling to direct nuclear specificity protein 1 and nuclear respiratory factor 1 transcription. Moreover, myeloid Phgdh deficiency reverses diet-induced obesity. Collectively, this study reveals that a mechanism involving de novo serine synthesis orchestrates mitochondrial biogenesis via mitochondrial-to-nuclear communication, and provides a potential therapeutic target for tackling inflammatory diseases and mitochondria-mediated diseases.
Asunto(s)
Macrófagos , Mitocondrias , Biogénesis de Organelos , Fosfoglicerato-Deshidrogenasa , Especies Reactivas de Oxígeno , Serina , Macrófagos/metabolismo , Animales , Mitocondrias/metabolismo , Fosfoglicerato-Deshidrogenasa/metabolismo , Fosfoglicerato-Deshidrogenasa/genética , Serina/metabolismo , Ratones , Especies Reactivas de Oxígeno/metabolismo , Transducción de Señal , Ratones Noqueados , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Subunidad alfa del Factor 1 Inducible por Hipoxia/genética , Inflamación/metabolismo , Inflamación/patología , Obesidad/metabolismo , Obesidad/patología , Obesidad/genética , Ratones Endogámicos C57BLRESUMEN
Argininosuccinate synthase (ASS1), a critical enzyme in the urea cycle, acts as a tumor suppressor in many cancers. To date, the anticancer mechanism of ASS1 has not been fully elucidated. Here, we found that phosphoglycerate dehydrogenase (PHGDH), a key rate-limiting enzyme in serine synthesis, is a pivotal protein that interacts with ASS1. Our results showed that ASS1 directly binds to PHGDH and promotes its ubiquitination-mediated degradation to inhibit serine synthesis, consequently suppressing tumorigenesis. Importantly, the tumor suppressive effects of ASS1 were strongly abrogated by PHGDH knockout. In addition, ASS1 knockout and knockdown partially rescued cell proliferation when serine and glycine were depleted, while the inhibitory effect of ASS1 overexpression on cell proliferation was restored by the addition of serine and glycine. These findings unveil a novel role of ASS1 and suggest that the ASS1/PHGDH serine synthesis pathway is a promising target for cancer therapy.
Asunto(s)
Argininosuccinato Sintasa , Proliferación Celular , Fosfoglicerato-Deshidrogenasa , Serina , Neoplasias de la Mama Triple Negativas , Fosfoglicerato-Deshidrogenasa/metabolismo , Fosfoglicerato-Deshidrogenasa/genética , Serina/metabolismo , Serina/biosíntesis , Humanos , Femenino , Neoplasias de la Mama Triple Negativas/metabolismo , Neoplasias de la Mama Triple Negativas/patología , Neoplasias de la Mama Triple Negativas/genética , Animales , Argininosuccinato Sintasa/metabolismo , Argininosuccinato Sintasa/genética , Línea Celular Tumoral , Ratones Desnudos , Ubiquitinación , Ratones , Glicina/metabolismoRESUMEN
The increased de novo serine biosynthesis confers many advantages for tumorigenesis and metastasis. Phosphoglycerate dehydrogenase (PHGDH), a rate-limiting enzyme in serine biogenesis, exhibits hyperactivity across multiple tumors and emerges as a promising target for cancer treatment. Through screening our in-house compound library, we identified compound Stattic as a potent PHGDH inhibitor (IC50 = 1.98 ± 0.66 µM). Subsequent exploration in structural activity relationships led to the discovery of compound B12 that demonstrated the increased enzymatic inhibitory activity (IC50 = 0.29 ± 0.02 µM). Furthermore, B12 exhibited robust inhibitory effects on the proliferation of MDA-MB-468, NCI-H1975, HT1080 and PC9 cells that overexpress PHGDH. Additionally, using a [U-13C6]-glucose tracing assay, B12 was found to reduce the production of glucose-derived serine in MDA-MB-468 cells. Finally, mass spectrometry-based peptide profiling, mutagenesis experiment and molecular docking study collectively suggested that B12 formed a covalent bond with Cys421 of PHGDH.
Asunto(s)
Inhibidores Enzimáticos , Fosfoglicerato-Deshidrogenasa , Simulación del Acoplamiento Molecular , Inhibidores Enzimáticos/farmacología , Inhibidores Enzimáticos/química , Serina , Glucosa , Línea Celular TumoralRESUMEN
BACKGROUND: Although papillary thyroid carcinoma (PTC) has a favorable prognosis, it could affect patient life quality and become a serious threat because of invasion and metastasis. Many investigations have suggested that circular RNAs (circRNAs) are involved in different cancer regulations. Nevertheless, circRNAs role in invasive PTC remains unclear. METHODS: In the present investigation, next-generation sequencing was applied to explore abnormal circRNA expression. The expression of circRNA phosphoglycerate dehydrogenase (circPHGDH) in PTC cell lines and tissues were examined. Then, we investigated regulatory mechanism and circPHGDH downstream targets using bioinformatics analysis and luciferase reporting analysis. Then transwell migration, Cell Counting Kit-8 (CCK8) and 5-ethynyl-2'-deoxyuridine (EdU) assays were used for cells migration and proliferation analysis. In vivo metastasis and tumorigenesis assays were also employed to evaluate the circPHGDH role in PTC. RESULTS: The data showcased that circPHGDH expression increased in both PTC cell lines and tissues, which suggested that circPHGDH functions in PTC progression. circPHGDH downregulation suppressed PTC invasion and proliferation in both in vivo and in vitro experiments. Bioinformatics and luciferase reporter results confirmed that both microRNA (miR)-122-5p and pyruvate kinase M2 subtype (PKM2) were downstream targets of circPHGDH. PKM2 overexpression or miR-122-5p suppression reversed PTC cell invasion and proliferation post silencing circPHGDH by restoring aerobic glycolysis. CONCLUSION: Taken together, our research found that circPHGDH downregulation reduced PTC progression via miR-122-5p/PKM2 axis regulation mediated by aerobic glycolysis.
Asunto(s)
Proliferación Celular , Progresión de la Enfermedad , Regulación hacia Abajo , Proteínas de la Membrana , MicroARNs , Fosfoglicerato-Deshidrogenasa , ARN Circular , Cáncer Papilar Tiroideo , Neoplasias de la Tiroides , Animales , Femenino , Humanos , Masculino , Ratones , Línea Celular Tumoral , Movimiento Celular/genética , Proliferación Celular/genética , Regulación Neoplásica de la Expresión Génica , Proteínas de la Membrana/metabolismo , Proteínas de la Membrana/genética , Ratones Desnudos , MicroARNs/genética , MicroARNs/metabolismo , Invasividad Neoplásica , Fosfoglicerato-Deshidrogenasa/genética , ARN Circular/genética , ARN Circular/metabolismo , Cáncer Papilar Tiroideo/genética , Cáncer Papilar Tiroideo/patología , Cáncer Papilar Tiroideo/metabolismo , Neoplasias de la Tiroides/genética , Neoplasias de la Tiroides/patología , Neoplasias de la Tiroides/metabolismo , Piruvato Quinasa/genética , Piruvato Quinasa/metabolismoRESUMEN
Circular RNAs are a class of covalently closed single-stranded loop RNAs that have been implicated to play a functional role in almost all types of cancers. Previous studies have revealed that circMYBL2 acts as a tumor-promoting circular RNA. In this study, we found that circMYBL2 in colorectal cancer encodes a 185-amino acid protein, p185. Functionally, circMYBL2-encoded p185 suppressed the growth and aggressiveness of colorectal cancer cells in vitro and in vivo. Mechanistically, p185 counteracted ubiquitin C-terminal hydrolase L3 (UCHL3)-mediated deubiquitination of phosphoglycerate dehydrogenase (PHGDH) by competitively binding to the C1 domain of UCHL3, resulting in PHGDH degradation and a subsequent reduction in serine and glycine biosynthesis. These data revealed that the circMYBL2-encoded p185 isoform serves as a tumor suppressor to inhibit the progression of colorectal cancer by reducing serine biosynthesis. Significance: A p185 protein encoded by circMYBL2 functions as a tumor suppressor that inhibits the progression of colorectal cancer by increasing the degradation of PHGDH to reduce serine biosynthesis.
Asunto(s)
Neoplasias Colorrectales , Progresión de la Enfermedad , Fosfoglicerato-Deshidrogenasa , ARN Circular , Serina , Humanos , Neoplasias Colorrectales/patología , Neoplasias Colorrectales/genética , Neoplasias Colorrectales/metabolismo , Serina/metabolismo , Serina/biosíntesis , Ratones , Animales , ARN Circular/genética , Fosfoglicerato-Deshidrogenasa/metabolismo , Fosfoglicerato-Deshidrogenasa/genética , Ratones Desnudos , Proliferación Celular , Línea Celular Tumoral , Masculino , Ratones Endogámicos BALB C , Femenino , Regulación Neoplásica de la Expresión Génica , Ensayos Antitumor por Modelo de Xenoinjerto , Células HCT116RESUMEN
Objective. Glucose and glutamine supply as well as serine synthesis and endoplasmic reticulum (ER) stress are important factors of glioblastoma growth. Previous studies showed that the knockdown of ERN1 (ER to nucleus signaling 1) suppressed glioblastoma cell proliferation and modified the sensitivity of numerous gene expressions to nutrient deprivations. The present study is aimed to investigate the impact of glucose and glutamine deprivations on the expression of serine synthesis genes in U87MG glioblastoma cells in relation to ERN1 knockdown with the intent to reveal the role of ERN1 signaling pathway on the ER stress-dependent regulation of these gene expressions. Clarification of the regulatory mechanisms of serine synthesis is a great significance for glioblastoma therapy. Methods. The control U87MG glioblastoma cells (transfected by empty vector) and ERN1 knockdown cells (transfected by dominant-negative ERN1) were exposed under glucose and glutamine deprivation conditions for 16 h. RNA was extracted from cells and reverse transcribed. The expression level of PHGDH (phosphoglycerate dehydrogenase), PSAT1 (phosphoserine amino-transferase 1), PSPH (phosphoserine phosphatase), ATF4 (activating transcription factor 4), and SHMT1 (serine hydroxymethyltransferase 1) genes was studied by real-time qPCR and normalized to ACTB. Results. It was found that the expression level of genes responsible for serine synthesis such as PHGDH, PSAT1, PSPH, and transcription factor ATF4 was up-regulated in U87MG glioblastoma cells under glucose and glutamine deprivations. Furthermore, inhibition of ERN1 significantly enhances the impact of glucose and especially glutamine deprivations on these gene expressions. At the same time, the expression of the SHMT1 gene, which is responsible for serine conversion to glycine, was down-regulated in both nutrient deprivation conditions with more significant changes in ERN1 knockdown glioblastoma cells. Conclusion. Taken together, the results of present study indicate that the expression of genes responsible for serine synthesis is sensitive to glucose and glutamine deprivations in gene-specific manner and that suppression of ERN1 signaling significantly modifies the impact of both glucose and glutamine deprivations on PHGDH, PSAT1, PSPH, ATF4, and SHMT1 gene expressions and reflects the ERN1-mediated genome reprograming introduced by nutrient deprivation condition.
Asunto(s)
Endorribonucleasas , Regulación Neoplásica de la Expresión Génica , Glioblastoma , Glucosa , Glutamina , Fosfoglicerato-Deshidrogenasa , Monoéster Fosfórico Hidrolasas , Proteínas Serina-Treonina Quinasas , Serina , Transaminasas , Humanos , Factor de Transcripción Activador 4/genética , Factor de Transcripción Activador 4/metabolismo , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/metabolismo , Línea Celular Tumoral , Estrés del Retículo Endoplásmico/genética , Estrés del Retículo Endoplásmico/efectos de los fármacos , Endorribonucleasas/genética , Endorribonucleasas/metabolismo , Técnicas de Silenciamiento del Gen , Glioblastoma/genética , Glioblastoma/metabolismo , Glucosa/metabolismo , Glutamina/metabolismo , Glicina Hidroximetiltransferasa/genética , Glicina Hidroximetiltransferasa/metabolismo , Fosfoglicerato-Deshidrogenasa/genética , Fosfoglicerato-Deshidrogenasa/metabolismo , Monoéster Fosfórico Hidrolasas/genética , Monoéster Fosfórico Hidrolasas/metabolismo , Proteínas Serina-Treonina Quinasas/genética , Proteínas Serina-Treonina Quinasas/metabolismo , Serina/metabolismo , Serina/biosíntesis , Transducción de SeñalRESUMEN
Colorectal cancer (CRC) has emerged as the third most prevalent and second deadliest cancer worldwide. Metabolic reprogramming is a key hallmark of cancer cells. Phosphoglycerate dehydrogenase (PHGDH) is over-expressed in multiple cancers, including CRC. Although the role of PHGDH in metabolism has been extensively investigated, its effects on CRC development remains to be elucidated. In the present study, it was demonstrated that PHGDH expression was significantly up-regulated in colorectal cancer. PHGDH expression was positively correlated with that of the aryl hydrocarbon receptor (AhR) and its target genes, CYP1A1 and CYP1B1, in CRC cells. Knockdown of PHGDH reduced AhR levels and activity, as well as the ratio of reduced to oxidized glutathione. The selective AhR antagonist stemregenin 1 induced cell death through reactive oxygen species-dependent autophagy in CRC cells. PHGDH knockdown induced CRC cell sensitivity to stemregenin 1 via the autophagy pathway. Our findings suggest that PHGDH modulates AhR signaling and the redox-dependent autophagy pathway in CRC, and that the combination of inhibition of both PHGDH and AhR may be a novel therapeutic strategy for CRC.
Asunto(s)
Neoplasias Colorrectales , Receptores de Hidrocarburo de Aril , Humanos , Autofagia/genética , Línea Celular Tumoral , Neoplasias Colorrectales/metabolismo , Fosfoglicerato-Deshidrogenasa/deficiencia , Fosfoglicerato-Deshidrogenasa/genética , Receptores de Hidrocarburo de Aril/antagonistas & inhibidores , Receptores de Hidrocarburo de Aril/genética , Receptores de Hidrocarburo de Aril/metabolismoRESUMEN
Phosphoglycerate dehydrogenase (PHGDH) has emerged as a crucial factor in macromolecule synthesis, neutralizing oxidative stress, and regulating methylation reactions in cancer cells, lymphocytes, and endothelial cells. However, the role of PHGDH in tumor-associated macrophages (TAMs) is poorly understood. Here, we found that the T helper 2 (Th2) cytokine interleukin-4 and tumor-conditioned media upregulate the expression of PHGDH in macrophages and promote immunosuppressive M2 macrophage activation and proliferation. Loss of PHGDH disrupts cellular metabolism and mitochondrial respiration, which are essential for immunosuppressive macrophages. Mechanistically, PHGDH-mediated serine biosynthesis promotes α-ketoglutarate production, which activates mTORC1 signaling and contributes to the maintenance of an M2-like macrophage phenotype in the tumor microenvironment. Genetic ablation of PHGDH in macrophages from tumor-bearing mice results in attenuated tumor growth, reduced TAM infiltration, a phenotypic shift of M2-like TAMs toward an M1-like phenotype, downregulated PD-L1 expression and enhanced antitumor T-cell immunity. Our study provides a strong basis for further exploration of PHGDH as a potential target to counteract TAM-mediated immunosuppression and hinder tumor progression.
Asunto(s)
Ácidos Cetoglutáricos , Diana Mecanicista del Complejo 1 de la Rapamicina , Fosfoglicerato-Deshidrogenasa , Transducción de Señal , Microambiente Tumoral , Macrófagos Asociados a Tumores , Animales , Macrófagos Asociados a Tumores/inmunología , Macrófagos Asociados a Tumores/metabolismo , Diana Mecanicista del Complejo 1 de la Rapamicina/metabolismo , Fosfoglicerato-Deshidrogenasa/metabolismo , Ratones , Ácidos Cetoglutáricos/metabolismo , Humanos , Ratones Endogámicos C57BL , Fenotipo , Línea Celular Tumoral , Activación de MacrófagosRESUMEN
Serine metabolism plays a pivotal role in cancer, making it an appealing therapeutic target. Two recent studies published in Nature Metabolism and Science Translational Medicine uncovered novel players and therapeutic opportunities within this crucial metabolic pathway. Papalazarou and colleagues employed genetic tools coupled with metabolomics and high-throughput imaging to identify and characterize membrane transporters involved in serine uptake and mitochondrial import in colorectal cancer. Notably, they showed that dual inhibition of these transporters in combination with impaired serine biosynthesis reduced tumor growth in xenograft models. In a parallel study, Zhang and colleagues identified isocitrate dehydrogenase I (IDH1) as a novel regulator of serine biosynthesis in non-small cell lung cancer. Through extensive mechanistic studies, they demonstrated that IDH1 enhances the expression of the key enzymes phosphoglycerate dehydrogenase and phosphoserine aminotransferase 1 via a noncanonical function independent of its enzymatic activity. Strikingly, pharmacologic disruption of this novel function of IDH1 not only diminished tumor growth but also enhanced the anticancer efficacy of dietary serine restriction in mouse models of lung cancer. Together, these studies advance our mechanistic understanding of how cancer cells fulfill their serine requirements and reveal innovative therapeutic avenues to deprive tumors of this vital nutrient.
Asunto(s)
Carcinoma de Pulmón de Células no Pequeñas , Neoplasias Pulmonares , Animales , Ratones , Humanos , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Carcinoma de Pulmón de Células no Pequeñas/genética , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/genética , Serina/metabolismo , Línea Celular Tumoral , Fosfoglicerato-DeshidrogenasaRESUMEN
Triple-negative breast cancer (TNBC) chemoresistance hampers the ability to effectively treat patients. Identification of mechanisms driving chemoresistance can lead to strategies to improve treatment. Here, we revealed that protein arginine methyltransferase-1 (PRMT1) simultaneously methylates D-3-phosphoglycerate dehydrogenase (PHGDH), a critical enzyme in serine synthesis, and the glycolytic enzymes PFKFB3 and PKM2 in TNBC cells. 13C metabolic flux analyses showed that PRMT1-dependent methylation of these three enzymes diverts glucose toward intermediates in the serine-synthesizing and serine/glycine cleavage pathways, thereby accelerating the production of methyl donors in TNBC cells. Mechanistically, PRMT1-dependent methylation of PHGDH at R54 or R20 activated its enzymatic activity by stabilizing 3-phosphoglycerate binding and suppressing polyubiquitination. PRMT1-mediated PHGDH methylation drove chemoresistance independently of glutathione synthesis. Rather, activation of the serine synthesis pathway supplied α-ketoglutarate and citrate to increase palmitate levels through activation of fatty acid synthase (FASN). Increased palmitate induced protein S-palmitoylation of PHGDH and FASN to further enhance fatty acid synthesis in a PRMT1-dependent manner. Loss of PRMT1 or pharmacologic inhibition of FASN or protein S-palmitoyltransferase reversed chemoresistance in TNBC. Furthermore, IHC coupled with imaging MS in clinical TNBC specimens substantiated that PRMT1-mediated methylation of PHGDH, PFKFB3, and PKM2 correlates with chemoresistance and that metabolites required for methylation and fatty acid synthesis are enriched in TNBC. Together, these results suggest that enhanced de novo fatty acid synthesis mediated by coordinated protein arginine methylation and protein S-palmitoylation is a therapeutic target for overcoming chemoresistance in TNBC. SIGNIFICANCE: PRMT1 promotes chemoresistance in TNBC by methylating metabolic enzymes PFKFB3, PKM2, and PHGDH to augment de novo fatty acid synthesis, indicating that targeting this axis is a potential treatment strategy.
Asunto(s)
Fosfoglicerato-Deshidrogenasa , Neoplasias de la Mama Triple Negativas , Humanos , Neoplasias de la Mama Triple Negativas/tratamiento farmacológico , Neoplasias de la Mama Triple Negativas/genética , Resistencia a Antineoplásicos , Serina/metabolismo , Palmitatos , Ácidos Grasos , Línea Celular Tumoral , Proteína-Arginina N-Metiltransferasas/genética , Proteínas RepresorasRESUMEN
In skeletal muscle, the Hippo effector Yap promotes satellite cell, myoblast, and rhabdomyoblast proliferation but prevents myogenic differentiation into multinucleated muscle fibres. We previously noted that Yap drives expression of the first enzyme of the serine biosynthesis pathway, phosphoglycerate dehydrogenase (Phgdh). Here, we examined the regulation and function of Phgdh in satellite cells and myoblasts and found that Phgdh protein increased during satellite cell activation. Analysis of published data reveal that Phgdh mRNA in mouse tibialis anterior muscle was highly expressed at day 3 of regeneration after cardiotoxin injection, when markers of proliferation are also robustly expressed and in the first week of synergist-ablated muscle. Finally, siRNA-mediated knockdown of PHGDH significantly reduced myoblast numbers and the proliferation rate. Collectively, our data suggest that Phgdh is a proliferation-enhancing metabolic enzyme that is induced when quiescent satellite cells become activated.
Asunto(s)
Fosfoglicerato-Deshidrogenasa , Células Satélite del Músculo Esquelético , Ratones , Animales , Fosfoglicerato-Deshidrogenasa/genética , Fosfoglicerato-Deshidrogenasa/metabolismo , Proliferación Celular/fisiología , Músculo Esquelético/metabolismo , Fibras Musculares Esqueléticas/metabolismo , Células Satélite del Músculo Esquelético/metabolismoRESUMEN
Bone-resorbing osteoclasts are vital for postnatal bone health, as increased differentiation or activity results in skeletal pathologies such as osteoporosis. The metabolism of mature osteoclasts differs from their progenitor cells, but whether the observed metabolic changes are secondary to the altered cell state or actively drive the process of cell differentiation is unknown. Here, we show that transient activation of the serine synthesis pathway (SSP) is essential for osteoclastogenesis, as deletion of the rate-limiting enzyme phosphoglycerate dehydrogenase in osteoclast progenitors impairs their differentiation and results in increased bone mass. In addition, pharmacological phosphoglycerate dehydrogenase inhibition abrogated bone loss in a mouse model of postmenopausal osteoporosis by blocking bone resorption. Mechanistically, SSP-derived α-ketoglutarate is necessary for histone demethylases that remove repressive histone methylation marks at the nuclear factor of activated T cells, cytoplasmic 1 (Nfatc1) gene locus, thereby inducing NFATc1 expression and consequent osteoclast maturation. Taken together, this study reveals a metabolic-epigenetic coupling mechanism that directs osteoclast differentiation and suggests that the SSP can be therapeutically targeted to prevent osteoporotic bone loss.
Asunto(s)
Epigénesis Genética , Factores de Transcripción NFATC , Osteoclastos , Animales , Ratones , Factores de Transcripción NFATC/genética , Factores de Transcripción NFATC/metabolismo , Fosfoglicerato-Deshidrogenasa/genética , Serina/genética , Serina/metabolismoRESUMEN
A "switch" in the metabolic pattern of microglia is considered to be required to meet the metabolic demands of cell survival and functions. However, how metabolic switches regulate microglial function remains controversial. We found here that exposure to amyloid-ß triggers microglial inflammation accompanied by increasing GAPDH levels. The increase of GAPDH, a glycolysis enzyme, leads to the reduced release of interferon-γ (IFN-γ) from inflammatory microglia. Such alternation is translational and is regulated by the binding of glycolysis enzyme glyceraldehyde-3-phosphate dehydrogenase (GAPDH) to IFN-γ mRNA. GAPDH, by engaging/disengaging glycolysis and through influencing IFN-γ expression, regulates microglia functions, including phagocytosis and cytokine production. Phosphoglycerate dehydrogenase (PHGDH), screened from different state microglia by metabolomics combined with METARECON analysis, is a metabolic enzyme adjacent downstream of GAPDH and synthesizes serine on the collateral pathway derived from glycolysis. Polarization of microglial with PHGDH as a metabolic checkpoint can be bidirectionally regulated by adding IL-4 or giving PHGDH inhibitors. Therefore, regulation of metabolic enzymes not only reprograms metabolic patterns, but also manipulates microglia functions. Further study should be performed to explore the mechanism of metabolic checkpoints in human microglia or more in vivo animal experiments, and may expand to the effects of various metabolic substrates or enzyme, such as lipids and amino acids, on the functions of microglia.
Asunto(s)
Microglía , Fosfoglicerato-Deshidrogenasa , Animales , Humanos , Fosfoglicerato-Deshidrogenasa/genética , Interferón gamma , Multiómica , Gliceraldehído-3-Fosfato Deshidrogenasas/genéticaRESUMEN
Cells use glycolytic intermediates for anabolism, e.g., via the serine synthesis and pentose phosphate pathways. However, we still understand poorly how these metabolic pathways contribute to skeletal muscle cell biomass generation. The first aim of this study was therefore to identify enzymes that limit protein synthesis, myotube size, and proliferation in skeletal muscle cells. We inhibited key enzymes of glycolysis, the pentose phosphate pathway, and the serine synthesis pathway to evaluate their importance in C2C12 myotube protein synthesis. Based on the results of this first screen, we then focused on the serine synthesis pathway enzyme phosphoglycerate dehydrogenase (PHGDH). We used two different PHGDH inhibitors and mouse C2C12 and human primary muscle cells to study the importance and function of PHGDH. Both myoblasts and myotubes incorporated glucose-derived carbon into proteins, RNA, and lipids, and we showed that PHGDH is essential in these processes. PHGDH inhibition decreased protein synthesis, myotube size, and myoblast proliferation without cytotoxic effects. The decreased protein synthesis in response to PHGDH inhibition appears to occur mainly mechanistic target of rapamycin complex 1 (mTORC1)-dependently, as was evident from experiments with insulin-like growth factor 1 and rapamycin. Further metabolomics analyses revealed that PHGDH inhibition accelerated glycolysis and altered amino acid, nucleotide, and lipid metabolism. Finally, we found that supplementing an antioxidant and redox modulator, N-acetylcysteine, partially rescued the decreased protein synthesis and mTORC1 signaling during PHGDH inhibition. The data suggest that PHGDH activity is critical for skeletal muscle cell biomass generation from glucose and that it regulates protein synthesis and mTORC1 signaling.NEW & NOTEWORTHY The use of glycolytic intermediates for anabolism was demonstrated in both myoblasts and myotubes, which incorporate glucose-derived carbon into proteins, RNA, and lipids. We identify phosphoglycerate dehydrogenase (PHGDH) as a critical enzyme in those processes and also for muscle cell hypertrophy, proliferation, protein synthesis, and mTORC1 signaling. Our results thus suggest that PHGDH in skeletal muscle is more than just a serine-synthesizing enzyme.
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Fosfoglicerato-Deshidrogenasa , Serina , Animales , Humanos , Ratones , Biomasa , Carbono/metabolismo , Proliferación Celular , Glucosa/metabolismo , Lípidos , Diana Mecanicista del Complejo 1 de la Rapamicina/metabolismo , Fibras Musculares Esqueléticas/metabolismo , Fosfoglicerato-Deshidrogenasa/genética , Fosfoglicerato-Deshidrogenasa/metabolismo , ARN/metabolismo , Serina/metabolismoRESUMEN
In the brain, the non-essential amino acid L-serine is produced through the phosphorylated pathway (PP) starting from the glycolytic intermediate 3-phosphoglycerate: among the different roles played by this amino acid, it can be converted into D-serine and glycine, the two main co-agonists of NMDA receptors. In humans, the enzymes of the PP, namely phosphoglycerate dehydrogenase (hPHGDH, which catalyzes the first and rate-limiting step of this pathway), 3-phosphoserine aminotransferase, and 3-phosphoserine phosphatase are likely organized in the cytosol as a metabolic assembly (a "serinosome"). The hPHGDH deficiency is a pathological condition biochemically characterized by reduced levels of L-serine in plasma and cerebrospinal fluid and clinically identified by severe neurological impairment. Here, three single-point variants responsible for hPHGDH deficiency and Neu-Laxova syndrome have been studied. Their biochemical characterization shows that V261M, V425M, and V490M substitutions alter either the kinetic (both maximal activity and Km for 3-phosphoglycerate in the physiological direction) and the structural properties (secondary, tertiary, and quaternary structure, favoring aggregation) of hPHGDH. All the three variants have been successfully ectopically expressed in U251 cells, thus the pathological effect is not due to hindered expression level. At the cellular level, mistargeting and aggregation phenomena have been observed in cells transiently expressing the pathological protein variants, as well as a reduced L-serine cellular level. Previous studies demonstrated that the pharmacological supplementation of L-serine in hPHGDH deficiencies could ameliorate some of the related symptoms: our results now suggest the use of additional and alternative therapeutic approaches.
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Encefalopatías , Ácidos Glicéricos , Serina , Humanos , Serina/genética , Fosfoglicerato-Deshidrogenasa/genética , Fosfoglicerato-Deshidrogenasa/química , Encefalopatías/metabolismo , AminoácidosRESUMEN
Clinical studies indicated that Serum Amyloid A (SAA) might be a promising biomarker for forecasting the activity, severity, and adverse prognosis of systemic lupus erythematosus (SLE). Simultaneously, a positive correlation has been observed between macrophages, Th17 cells, and SLE disease activity, with both these immune cells being affected by SAA. Presently, the relationship between SAA and the aforementioned immune cell types in SLE remains to be elucidated. To discern the immune cell type most closely associated with SAA, we undertook a single-cell RNA sequencing data analysis via the GEO database. Subsequent results revealed a strong association between macrophages and SAA, a relationship further validated through flow cytometry of spleen macrophages in the MRL/lpr model. We discovered that SAA stimulate M1 macrophage differentiation along with the upregulation of pro-inflammatory cytokines such as IL-6 and IL-1ß. Our findings suggest that SAA may promote M1 macrophage differentiation via the downregulation of phosphoglycerate dehydrogenase (PHGDH). Artesunate (ART), primarily utilized for malaria treatment, was shown to inhibit M1 macrophage differentiation and pro-inflammatory cytokine levels via upregulating the PHGDH expression, thereby attenuating the disease activity in SLE.
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Lupus Eritematoso Sistémico , Proteína Amiloide A Sérica , Humanos , Animales , Ratones , Artesunato/farmacología , Artesunato/uso terapéutico , Proteína Amiloide A Sérica/metabolismo , Fosfoglicerato-Deshidrogenasa/metabolismo , Fosfoglicerato-Deshidrogenasa/uso terapéutico , Macrófagos , Citocinas/metabolismo , Ratones Endogámicos MRL lprRESUMEN
The Warburg effect links growth and glycolysis in cancer. A key purpose of the Warburg effect is to generate glycolytic intermediates for anabolic reactions, such as nucleotides â RNA/DNA and amino acids â protein synthesis. The aim of this study was to investigate whether a similar 'glycolysis-for-anabolism' metabolic reprogramming also occurs in hypertrophying skeletal muscle. To interrogate this, we first induced C2C12 myotube hypertrophy with IGF-1. We then added 14C glucose to the differentiation medium and measured radioactivity in isolated protein and RNA to establish whether 14C had entered anabolism. We found that especially protein became radioactive, suggesting a glucose â glycolytic intermediates â non-essential amino acid(s) â protein series of reactions, the rate of which was increased by IGF-1. Next, to investigate the importance of glycolytic flux and non-essential amino acid synthesis for myotube hypertrophy, we exposed C2C12 and primary mouse myotubes to the glycolysis inhibitor 2-Deoxy-d-glucose (2DG). We found that inhibiting glycolysis lowered C2C12 and primary myotube size. Similarly, siRNA silencing of PHGDH, the key enzyme of the serine biosynthesis pathway, decreased C2C12 and primary myotube size; whereas retroviral PHGDH overexpression increased C2C12 myotube size. Together these results suggest that glycolysis is important for hypertrophying myotubes, which reprogram their metabolism to facilitate anabolism, similar to cancer cells.