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1.
Plant J ; 119(2): 746-761, 2024 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-38733631

RESUMO

The jasmonic acid (JA) signaling pathway plays an important role in promoting the biosynthesis of tanshinones. While individual transcription factors have been extensively studied in the context of tanshinones biosynthesis regulation, the influence of methyl jasmonate (MeJA)-induced transcriptional complexes remains unexplored. This study elucidates the positive regulatory role of the basic helix-loop-helix protein SmMYC2 in tanshinones biosynthesis in Salvia miltiorrhiza. SmMYC2 not only binds to SmGGPPS1 promoters, activating their transcription, but also interacts with SmMYB36. This interaction enhances the transcriptional activity of SmMYC2 on SmGGPPS1, thereby promoting tanshinones biosynthesis. Furthermore, we identified three JA signaling repressors, SmJAZ3, SmJAZ4, and SmJAZ8, which interact with SmMYC2. These repressors hindered the transcriptional activity of SmMYC2 on SmGGPPS1 and disrupted the interaction between SmMYC2 and SmMYB36. MeJA treatment triggered the degradation of SmJAZ3 and SmJAZ4, allowing the SmMYC2-SmMYB36 complex to subsequently activate the expression of SmGGPPS1, whereas SmJAZ8 inhibited MeJA-mediated degradation due to the absence of the LPIARR motif. These results demonstrate that the SmJAZ-SmMYC2-SmMYB36 module dynamically regulates the JA-mediated accumulation of tanshinones. Our results reveal a new regulatory network for the biosynthesis of tanshinones. This study provides valuable insight for future research on MeJA-mediated modulation of tanshinones biosynthesis.


Assuntos
Abietanos , Acetatos , Ciclopentanos , Regulação da Expressão Gênica de Plantas , Oxilipinas , Proteínas de Plantas , Salvia miltiorrhiza , Ciclopentanos/metabolismo , Ciclopentanos/farmacologia , Oxilipinas/metabolismo , Oxilipinas/farmacologia , Salvia miltiorrhiza/genética , Salvia miltiorrhiza/metabolismo , Salvia miltiorrhiza/efeitos dos fármacos , Acetatos/farmacologia , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Reguladores de Crescimento de Plantas/metabolismo , Fatores de Transcrição/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Transdução de Sinais , Regiões Promotoras Genéticas/genética
2.
Mar Drugs ; 21(4)2023 Apr 17.
Artigo em Inglês | MEDLINE | ID: mdl-37103388

RESUMO

Precursor regulation has been an effective strategy to improve carotenoid production and the availability of novel precursor synthases facilitates engineering improvements. In this work, the putative geranylgeranyl pyrophosphate synthase encoding gene (AlGGPPS) and isopentenyl pyrophosphate isomerase encoding gene (AlIDI) from Aurantiochytrium limacinum MYA-1381 were isolated. We applied the excavated AlGGPPS and AlIDI to the de novo ß-carotene biosynthetic pathway in Escherichia coli for functional identification and engineering application. Results showed that the two novel genes both functioned in the synthesis of ß-carotene. Furthermore, AlGGPPS and AlIDI performed better than the original or endogenous one, with 39.7% and 80.9% increases in ß-carotene production, respectively. Due to the coordinated expression of the 2 functional genes, ß-carotene content of the modified carotenoid-producing E. coli accumulated a 2.99-fold yield of the initial EBIY strain in 12 h, reaching 10.99 mg/L in flask culture. This study helped to broaden current understanding of the carotenoid biosynthetic pathway in Aurantiochytrium and provided novel functional elements for carotenoid engineering improvements.


Assuntos
Escherichia coli , beta Caroteno , beta Caroteno/metabolismo , Escherichia coli/genética , Escherichia coli/metabolismo , Geranil-Geranildifosfato Geranil-Geraniltransferase/genética , Carotenoides/metabolismo
3.
Int J Mol Sci ; 24(2)2023 Jan 04.
Artigo em Inglês | MEDLINE | ID: mdl-36674507

RESUMO

As one of the most imperative antioxidants in higher plants, carotenoids serve as accessory pigments to harvest light for photosynthesis and photoprotectors for plants to adapt to high light stress. Here, we report a small subunit (SSU) of geranylgeranyl diphosphate synthase (GGPPS) in Nicotiana tabacum, NtSSU II, which takes part in the regulation carotenoid biosynthesis by forming multiple enzymatic components with NtGGPPS1 and downstream phytoene synthase (NtPSY1). NtSSU II transcript is widely distributed in various tissues and stimulated by low light and high light treatments. The confocal image revealed that NtSSU II was localized in the chloroplast. Bimolecular fluorescence complementation (BiFC) indicated that NtSSU II and NtGGPPS1 formed heterodimers, which were able to interact with phytoene synthase (NtPSY1) to channel GGPP into the carotenoid production. CRISPR/Cas9-induced ntssu II mutant exhibited decreased leaf area and biomass, along with a decline in carotenoid and chlorophyll accumulation. Moreover, the genes involved in carotenoid biosynthesis were also downregulated in transgenic plants of ntssu II mutant. Taken together, the newly identified NtSSU II could form multiple enzymatic components with NtGGPPS1 and NtPSY1 to regulate carotenoid biosynthesis in N. tabacum, in addition to the co-expression of genes in carotenoids biosynthetic pathways.


Assuntos
Carotenoides , Nicotiana , Farnesiltranstransferase/genética , Farnesiltranstransferase/metabolismo , Nicotiana/genética , Nicotiana/metabolismo , Carotenoides/metabolismo , Fotossíntese , Geranil-Geranildifosfato Geranil-Geraniltransferase/genética , Geranil-Geranildifosfato Geranil-Geraniltransferase/metabolismo
4.
Immunopharmacol Immunotoxicol ; 44(1): 58-66, 2022 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-34845949

RESUMO

AIM: Acute lung injury (ALI) and resultant acute respiratory distress syndrome (ARDS) are detrimental inflammatory disease associated with high rates of morbidity and mortality due to a lack of effective treatment options. Previous study has demonstrated that an inhibition of geranylgeranyl pyrophosphate synthase large subunit 1 (GGPPS1) show a protective effect against ALI. METHOD: In this study, by using connective map (CMAP), we identified catechin as a potential drug to exhibit similar effects to inhibit GGPPS1. Furthermore, we detected the protective effect of catechin on lipopolysaccharide (LPS)-induced ALI and delineated the underlying mechanism. RESULTS: We found that catechin effectively ameliorated LPS-induced lung inflammation and alleviated the release of cytokines into alveolar space. Notably, miR-182/GGPPS1 signaling pathway was reactivated upon catechin administration, which was essential for the catechin-induced protective effect against ALI. CONCLUSION: catechin regulates miR-182/GGPPS1 signaling pathway and efficaciously ameliorates LPS-induced acute lung injury in mice model, which provided a promising therapeutic strategy in ALI and ARDS.


Assuntos
Lesão Pulmonar Aguda , Catequina , MicroRNAs , Lesão Pulmonar Aguda/induzido quimicamente , Lesão Pulmonar Aguda/tratamento farmacológico , Lesão Pulmonar Aguda/metabolismo , Animais , Catequina/efeitos adversos , Catequina/metabolismo , Lipopolissacarídeos/toxicidade , Pulmão/metabolismo , Camundongos , MicroRNAs/genética , MicroRNAs/metabolismo , Transdução de Sinais
5.
J Biol Chem ; 295(47): 15988-16001, 2020 11 20.
Artigo em Inglês | MEDLINE | ID: mdl-32913122

RESUMO

Metaflammation is a primary inflammatory complication of metabolic disorders characterized by altered production of many inflammatory cytokines, adipokines, and lipid mediators. Whereas multiple inflammation networks have been identified, the mechanisms by which metaflammation is initiated have long been controversial. As the mevalonate pathway (MVA) produces abundant bioactive isoprenoids and abnormal MVA has a phenotypic association with inflammation/immunity, we speculate that isoprenoids from the MVA may provide a causal link between metaflammation and metabolic disorders. Using a line with the MVA isoprenoid producer geranylgeranyl diphosphate synthase (GGPPS) deleted, we find that geranylgeranyl pyrophosphate (GGPP) depletion causes an apparent metaflammation as evidenced by abnormal accumulation of fatty acids, eicosanoid intermediates, and proinflammatory cytokines. We also find that GGPP prenylate cytochrome b5 reductase 3 (CYB5R3) and the prenylated CYB5R3 then translocate from the mitochondrial to the endoplasmic reticulum (ER) pool. As CYB5R3 is a critical NADH-dependent reductase necessary for eicosanoid metabolism in ER, we thus suggest that GGPP-mediated CYB5R3 prenylation is necessary for metabolism. In addition, we observe that pharmacological inhibition of the MVA pathway by simvastatin is sufficient to inhibit CYB5R3 translocation and induces smooth muscle death. Therefore, we conclude that the dysregulation of MVA intermediates is an essential mechanism for metaflammation initiation, in which the imbalanced production of eicosanoid intermediates in the ER serve as an important pathogenic factor. Moreover, the interplay of MVA and eicosanoid metabolism as we reported here illustrates a model for the coordinating regulation among metabolite pathways.


Assuntos
Citocromo-B(5) Redutase/metabolismo , Eicosanoides/metabolismo , Retículo Endoplasmático/metabolismo , Mitocôndrias/metabolismo , Fosfatos de Poli-Isoprenil/metabolismo , Prenilação , Animais , Citocromo-B(5) Redutase/genética , Eicosanoides/genética , Retículo Endoplasmático/genética , Ácido Mevalônico/metabolismo , Camundongos , Camundongos Knockout , Mitocôndrias/genética , Transporte Proteico/efeitos dos fármacos , Transporte Proteico/genética , Sinvastatina/farmacologia
6.
J Biol Chem ; 295(15): 5152-5162, 2020 04 10.
Artigo em Inglês | MEDLINE | ID: mdl-32139507

RESUMO

Protein prenylation is an essential posttranslational modification and includes protein farnesylation and geranylgeranylation using farnesyl diphosphate or geranylgeranyl diphosphate as substrates, respectively. Geranylgeranyl diphosphate synthase is a branch point enzyme in the mevalonate pathway that affects the ratio of farnesyl diphosphate to geranylgeranyl diphosphate. Abnormal geranylgeranyl diphosphate synthase expression and activity can therefore disrupt the balance of farnesylation and geranylgeranylation and alter the ratio between farnesylated and geranylgeranylated proteins. This change is associated with the progression of nonalcoholic fatty liver disease (NAFLD), a condition characterized by hepatic fat overload. Of note, differential accumulation of farnesylated and geranylgeranylated proteins has been associated with differential stages of NAFLD and NAFLD-associated liver fibrosis. In this review, we summarize key aspects of protein prenylation as well as advances that have uncovered the regulation of associated metabolic patterns and signaling pathways, such as Ras GTPase signaling, involved in NAFLD progression. Additionally, we discuss unique opportunities for targeting prenylation in NAFLD/hepatocellular carcinoma with agents such as statins and bisphosphonates to improve clinical outcomes.


Assuntos
Farnesiltranstransferase/metabolismo , Hepatopatia Gordurosa não Alcoólica/patologia , Fosfatos de Poli-Isoprenil/metabolismo , Prenilação de Proteína , Processamento de Proteína Pós-Traducional , Animais , Progressão da Doença , Humanos , Hepatopatia Gordurosa não Alcoólica/metabolismo
7.
Am J Physiol Lung Cell Mol Physiol ; 320(6): L1011-L1024, 2021 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-33729030

RESUMO

Macrophage activation is a key contributing factor for excessive inflammatory responses of acute lung injury (ALI)/acute respiratory distress syndrome (ARDS). Geranylgeranyl diphosphate synthase (GGPPS) plays a key role in the development of inflammatory diseases. Our group previously showed that GGPPS in alveolar epithelium have deleterious effects on acute lung injury induced by LPS or mechanical ventilation. Herein, we examined the role of GGPPS in modulating macrophage activation in ALI/ARDS. We found significant increased GGPPS expression in alveolar macrophages in patients with ARDS compared with healthy volunteers and in ALI mice induced by LPS. GGPPS-floxed control (GGPPSfl/fl) and myeloid-selective knockout (GGPPSfl/flLysMcre) mice were then generated. Interestingly, using an LPS-induced ALI mouse model, we showed that myeloid-specific GGPPS knockout significantly increased mortality, aggravated lung injury, and increased the accumulation of inflammatory cells, total protein, and inflammatory cytokines in BALF. In vitro, GGPPS deficiency upregulated the production of LPS-induced IL-6, IL-1ß, and TNF-α in alveolar macrophages, bone marrow-derived macrophages (BMDMs), and THP-1 cells. Mechanistically, GGPPS knockout increased phosphorylation and nuclear translocation of NF-κB p65 induced by LPS. In addition, GGPPS deficiency increased the level of GTP-Rac1, which was responsible for NF-κB activation. In conclusion, decreased expression of GGPPS in macrophages aggravates lung injury and inflammation in ARDS, at least partly by regulating Rac1-dependent NF-κB signaling. GGPPS in macrophages may represent a novel therapeutic target in ARDS.


Assuntos
Lesão Pulmonar Aguda/tratamento farmacológico , Lipopolissacarídeos/farmacologia , Ativação de Macrófagos/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Lesão Pulmonar Aguda/induzido quimicamente , Lesão Pulmonar Aguda/metabolismo , Animais , Citocinas/metabolismo , Modelos Animais de Doenças , Regulação da Expressão Gênica/efeitos dos fármacos , Inflamação/tratamento farmacológico , Inflamação/metabolismo , Pulmão/efeitos dos fármacos , Pulmão/metabolismo , Macrófagos/metabolismo , Camundongos
8.
J Inherit Metab Dis ; 43(1): 133-144, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-30942483

RESUMO

There are many metabolic disorders that present with bone phenotypes. In some cases, the pathological bone symptoms are the main features of the disease whereas in others they are a secondary characteristic. In general, the generation of the bone problems in these disorders is not well understood and the therapeutic options for them are scarce. Bone development occurs in the early stages of embryonic development where the bone formation, or osteogenesis, takes place. This osteogenesis can be produced through the direct transformation of the pre-existing mesenchymal cells into bone tissue (intramembranous ossification) or by the replacement of the cartilage by bone (endochondral ossification). In contrast, bone remodeling takes place during the bone's growth, after the bone development, and continues throughout the whole life. The remodeling involves the removal of mineralized bone by osteoclasts followed by the formation of bone matrix by the osteoblasts, which subsequently becomes mineralized. In some metabolic diseases, bone pathological features are associated with bone development problems but in others they are associated with bone remodeling. Here, we describe three examples of impaired bone development or remodeling in metabolic diseases, including work by others and the results from our research. In particular, we will focus on hereditary multiple exostosis (or osteochondromatosis), Gaucher disease, and the susceptibility to atypical femoral fracture in patients treated with bisphosphonates for several years.


Assuntos
Desenvolvimento Ósseo/fisiologia , Remodelação Óssea/fisiologia , Cartilagem/crescimento & desenvolvimento , Doenças Metabólicas/metabolismo , Osteogênese/fisiologia , Animais , Cartilagem/citologia , Condrócitos/ultraestrutura , Difosfonatos/uso terapêutico , Exostose Múltipla Hereditária/metabolismo , Fraturas do Fêmur/tratamento farmacológico , Fraturas do Fêmur/metabolismo , Doença de Gaucher/metabolismo , Humanos , Osteoclastos/metabolismo
9.
J Pathol ; 247(3): 283-286, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30374976

RESUMO

Non-alcoholic fatty liver disease (NAFLD) often develops in concert with related metabolic diseases, such as obesity, dyslipidemia and insulin resistance. Prolonged lipid accumulation and inflammation can progress to non-alcoholic steatohepatitis (NASH). Although factors associated with the development of NAFLD are known, triggers for the progression of NAFLD to NASH are poorly understood. Recent findings published in The Journal of Pathology reveal the possible regulation of NASH progression by metabolites of the mevalonate pathway. Mevalonate can be converted into the isoprenoids farnesyldiphosphate (FPP) and geranylgeranyl diphosphate (GGPP). GGPP synthase (GGPPS), the enzyme that converts FPP to GGPP, is dysregulated in humans and mice during NASH. Both FPP and GGPP can be conjugated to proteins through prenylation, modifying protein function and localization. Deletion or knockdown of GGPPS favors FPP prenylation (farnesylation) and augments the function of liver kinase B1, an upstream kinase of AMP-activated protein kinase (AMPK). Despite increased AMPK activation, livers in Ggpps-deficient mice on a high-fat diet poorly oxidize lipids due to mitochondrial dysfunction. Although work from Liu et al provides evidence as to the potential importance of the prenylation portion of the mevalonate pathway during NAFLD, future studies are necessary to fully grasp any therapeutic or diagnostic potential. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.


Assuntos
Hepatopatia Gordurosa não Alcoólica , Animais , Dieta Hiperlipídica , Farnesiltranstransferase , Fibrose , Glucose , Humanos , Fígado , Camundongos , Prenilação , Reino Unido
10.
Biochem Biophys Res Commun ; 518(3): 479-485, 2019 10 20.
Artigo em Inglês | MEDLINE | ID: mdl-31427080

RESUMO

Isoprenoids comprise a diverse group of natural products with a broad range of metabolic functions. Isoprenoids are synthesized from prenyl pyrophosphates by prenyltransferases that catalyze the isoprenoid chain-elongation process to different chain lengths. We hereby present the crystal structure of geranylgeranyl pyrophosphate synthase from the marine flavobacterium Nonlabens dokdonensis DSW-6 (NdGGPPS). NdGGPPS forms a hexamer composed of homodimeric trimer, and the monomeric structure is composed of 15 α-helices (α1-α15). In this structure, we observed the binding of one pyrophosphate molecule and two glycerol molecules that mimicked substrate binding to the enzyme. The substrate binding site of NdGGPPS contains large hydrophobic residues such as Phe, His and Tyr, and structural and amino acids sequence analyses thereof suggest that the protein belongs to the short-chain prenyltransferase family.


Assuntos
Proteínas de Bactérias/química , Flavobacteriaceae/química , Geranil-Geranildifosfato Geranil-Geraniltransferase/química , Sequência de Aminoácidos , Cristalografia por Raios X , Modelos Moleculares , Conformação Proteica , Conformação Proteica em alfa-Hélice , Multimerização Proteica , Alinhamento de Sequência
11.
J Pathol ; 246(3): 277-288, 2018 11.
Artigo em Inglês | MEDLINE | ID: mdl-29971772

RESUMO

Patients with obesity have a high prevalence of non-alcoholic fatty liver disease (NAFLD) and, in parallel, increased susceptibility to fibrosis/cirrhosis/hepatocellular carcinoma (HCC). Herein, we report that a high-fat diet (HFD) can augment glycolysis and then accelerate NAFLD-fibrosis progression by downregulating the expression of geranylgeranyl diphosphate synthase (GGPPS), which is a critical enzyme in the mevalonate pathway. Long-term HFD overloading decreases GGPPS expression in mice, which shifts the fuel preference from fatty acids towards glucose. Liver-specific Ggpps deficiency drives the Warburg effect by impairing mitochondrial function, and then induces hepatic inflammation, thus exacerbating fibrosis. Ggpps deficiency also enhances the hyperfarnesylation of liver kinase B1, and promotes metabolic reprogramming by regulating 5'-AMP-activated protein kinase activity. Clinical data further imply that GGPPS expression can predict the stage of NAFLD and recurrence of NAFLD-associated HCC. We conclude that the level of GGPPS is a susceptibility factor for NAFLD-fibrosis progression, and requires more stringent surveillance to ensure early prediction and precision of treatment of NAFLD-related HCC. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.


Assuntos
Metabolismo Energético , Farnesiltranstransferase/metabolismo , Ácidos Graxos/metabolismo , Glucose/metabolismo , Hepatócitos/enzimologia , Cirrose Hepática/enzimologia , Fígado/enzimologia , Hepatopatia Gordurosa não Alcoólica/enzimologia , Proteínas Quinases Ativadas por AMP/metabolismo , Animais , Células Cultivadas , Dieta Hiperlipídica , Modelos Animais de Doenças , Farnesiltranstransferase/deficiência , Farnesiltranstransferase/genética , Glicólise , Hepatócitos/patologia , Humanos , Fígado/patologia , Cirrose Hepática/genética , Cirrose Hepática/patologia , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mitocôndrias Hepáticas/enzimologia , Mitocôndrias Hepáticas/patologia , Hepatopatia Gordurosa não Alcoólica/genética , Hepatopatia Gordurosa não Alcoólica/patologia , Oxirredução , Prenilação de Proteína , Proteínas Serina-Treonina Quinases/metabolismo , Transdução de Sinais
12.
Plant Cell Rep ; 38(1): 117-128, 2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-30448883

RESUMO

KEY MESSAGE: We found that ApGGPPS1, ApGGPPS2, and ApGGPPS3 can convert IPP and DMAPP to GGPP. ApGGPPS2 is probably involved in andrographolide biosynthesis. ApGGPPS3 may be responsible for the synthesis of the cytosolic GGPP. Andrographis paniculata is a traditional herb for the treatment of sore throat, flu, upper respiratory tract infections and other disorders. In A. paniculata, GGPP is not only the precursor of andrographolide and its primary bioactive compounds, but also the precursor of chlorophylls, carotenoids, gibberellins, and abscisic acid, which are the biomolecules of photosynthesis, growth regulation and other physiological and ecological processes. In this study, four cDNAs (named ApGGPPS1, ApGGPPS2, ApGGPPS3 and ApGGPPS4) encoding geranylgeranyl pyrophosphate synthases from A. paniculata were putatively isolated. Bioinformatic and phylogenetic analyses suggested that these ApGGPPS are highly similar to the geranylgeranyl pyrophosphate synthases in other plants. Prokaryotic expression showed that ApGGPPS1, ApGGPPS2 and ApGGPPS3 could convert IPP and DMAPP to GGPP, although ApGGPPS4 lacks a similar function. The expression of ApGGPPS2 was similar as ApCPS2 under MeJA treatment, ApCPS2 involved in the biosynthesis pathway of andrographolide (Shen et al., Biotechnol Lett 38:131-137, 2016a), has been proven through Virus-induced Gene Siliencing (VIGS) (Shen et al., Acta Bot Boreal 36:17-22, 2016b), and the subcellular localization of ApGGPPS2 was shown to localize in the plastid, suggested that ApGGPPS2 could be the key synthase in the biosynthesis pathway of andrographolide. In addition, ApGGPPS3 was shown to localize in the cytoplasm, suggested that ApGGPPS3 may be responsible for the synthesis of cytosolic GGPP, which may participate in the synthesis of cytosolic oligoprenols as side chains to produce ubiquinone, dolichols or other isoprenoids, in the synthesis of polyisoprenoids, and in protein prenylation.


Assuntos
Andrographis/metabolismo , Clonagem Molecular , Geranil-Geranildifosfato Geranil-Geraniltransferase/metabolismo , Andrographis/genética , Regulação da Expressão Gênica de Plantas/genética , Regulação da Expressão Gênica de Plantas/fisiologia , Inativação Gênica/fisiologia , Geranil-Geranildifosfato Geranil-Geraniltransferase/genética
13.
Mar Drugs ; 17(12)2019 Dec 12.
Artigo em Inglês | MEDLINE | ID: mdl-31842293

RESUMO

Haematococcus pluvialis is widely distributed in the world and well known as the richest natural source of astaxanthin that is a strong antioxidant with excellent commercial value. The pathway of astaxanthin biosynthesis in H. pluvialis has been documented as an enzymatic reaction. Several enzymes have been reported, but their isoforms or homologs have not been investigated genome-wide. To better understand the astaxanthin biosynthesis pathway in H. pluvialis, eight candidates of the geranylgeranyl pyrophosphate synthase gene (HpGGPPS) predicted from Iso-seq data were isolated in this study. The length of coding region of these candidates varied from 960 bp to 1272 bp, composing of 7-9 exons. The putative amino acids of all candidates composed the signature domain of GGPPS gene. However, the motifs in the domain region are varied, indicating different bio-functions. Phylogenetic analysis revealed eight candidates can be clustered into three groups. Only two candidates in Group1 encode the synthase participating in the astaxanthin formation. The yield of astaxanthin from these two candidates, 7.1 mg/g (DW) and 6.5 mg/g (DW) respectively, is significant higher than that from CrtE (2.4 mg/g DW), a GGPPS gene from Pantoea ananatis. This study provides a potential productive pathway for astaxanthin synthesis.


Assuntos
Antioxidantes/isolamento & purificação , Clorofíceas/química , Geranil-Geranildifosfato Geranil-Geraniltransferase/genética , Genoma , Filogenia , Xantofilas/isolamento & purificação
14.
J Pathol ; 238(1): 109-19, 2016 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-26434932

RESUMO

Loss of first-phase insulin secretion associated with ß cell dysfunction is an independent predictor of type 2 diabetes mellitus (T2DM) onset. Here we found that a critical enzyme involved in protein prenylation, geranylgeranyl pyrophosphate synthase (GGPPS), is required to maintain first-phase insulin secretion. GGPPS shows a biphasic expression pattern in islets of db/db mice during the progression of T2DM: GGPPS is increased during the insulin compensatory period, followed by a decrease during ß cell dysfunction. Ggpps deletion in ß cells results in typical T2DM ß cell dysfunction, with blunted glucose-stimulated insulin secretion and consequent insulin secretion insufficiency. However, the number and size of islets and insulin biosynthesis are unaltered. Transmission electron microscopy shows a reduced number of insulin granules adjacent to the cellular membrane, suggesting a defect in docked granule pool formation, while the reserve pool is unaffected. Ggpps ablation depletes GGPP and impairs Rab27A geranylgeranylation, which is responsible for the docked pool deficiency in Ggpps-null mice. Moreover, GGPPS re-expression or GGPP administration restore glucose-stimulated insulin secretion in Ggpps-null islets. These results suggest that GGPPS-controlled protein geranylgeranylation, which regulates formation of the insulin granule docked pool, is critical for ß cell function and insulin release during the development of T2DM.


Assuntos
Linfócitos B/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Farnesiltranstransferase/metabolismo , Insulina/metabolismo , Complexos Multienzimáticos/metabolismo , Proteínas rab de Ligação ao GTP/metabolismo , Animais , Western Blotting , Imunoprecipitação , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Microscopia Eletrônica de Transmissão , Prenilação de Proteína/fisiologia , RNA Interferente Pequeno , Reação em Cadeia da Polimerase em Tempo Real , Vesículas Secretórias/metabolismo , Vesículas Secretórias/ultraestrutura , Transfecção , Proteínas rab27 de Ligação ao GTP
15.
Appl Microbiol Biotechnol ; 101(20): 7523-7533, 2017 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-28918530

RESUMO

Taxol is an anticancer identified in both endophytic fungus and its host plant. Plant Taxol is a diterpenoid with geranylgeranyl diphosphate (GGPP) mediates the biosynthesis of its terpenoid moiety. Previous report has suggested that fungal Taxol may require terpenoid pathway for its biosynthesis. Here in this study, feeding a Taxol-producing endophytic fungus (Paraconiothyrium SSM001) with terpenoid precursors including isopentenyl pyrophosphate (IPP, isoprene) and GGPP enhanced Taxol production threefold and fivefold, respectively, compared to the control. Thus, we assumed that increasing the terpenoid pool size in particular GGPP by introducing a new copy number of GGPPS particularly from a Taxol-producing plant might increase the production level of fungal Taxol. Agrobacterium-mediated integration of Taxus canadensis geranylgeranyl diphosphate synthase (GGPPS) gene into the Paraconiothyrium SSM001 genome was successful and increased the terpenoid pool size indicated by an increase in carotenoid level and orange to red coloration of some GGPPS-transformed SSM001 colonies. Furthermore, the integration improved the level of Taxol production threefold. Feeding a GGPPS-transformed SSM001 fungus with a GGPP precursor increased the expression level of GGPPS transcript and Taxol production. The successful increase in both terpenoid and Taxol production levels due to GGPPS gene integration into the fungal genome might be a step forward in manipulating Taxol-producing endophytic fungi. Future control of the transformation time and the manipulation of the phenolic pathway could maximize the production level.


Assuntos
Antineoplásicos/metabolismo , Ascomicetos/metabolismo , Farnesiltranstransferase/metabolismo , Engenharia Metabólica , Paclitaxel/metabolismo , Fosfatos de Poli-Isoprenil/metabolismo , Ascomicetos/genética , Butadienos/metabolismo , Farnesiltranstransferase/genética , Hemiterpenos/metabolismo , Pentanos/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Taxus/enzimologia , Taxus/genética
16.
J Biol Chem ; 290(33): 20086-97, 2015 Aug 14.
Artigo em Inglês | MEDLINE | ID: mdl-26112408

RESUMO

Elevated circulating free fatty acid levels are important contributors to insulin resistance in the muscle and liver, but the underlying mechanisms require further elucidation. Here, we show that geranylgeranyl diphosphate synthase 1 (GGPPS), which is a branch point enzyme in the mevalonic acid pathway, promotes lipid-induced muscle insulin resistance through activation of the RhoA/Rho kinase signaling pathway. We have found that metabolic perturbation would increase GGPPS expression in the skeletal muscles of db/db mice and high fat diet-fed mice. To address the metabolic effects of GGPPS activity in skeletal muscle, we generated mice with specific GGPPS deletions in their skeletal muscle tissue. Heterozygous knock-out of GGPPS in the skeletal muscle improved systemic insulin sensitivity and glucose homeostasis in mice fed both normal chow and high fat diets. These metabolic alterations were accompanied by activated PI3K/Akt signaling and enhanced glucose uptake in the skeletal muscle. Further investigation showed that the free fatty acid-stimulated GGPPS expression in the skeletal muscle was able to enhance the geranylgeranylation of RhoA, which further induced the inhibitory phosphorylation of IRS-1 (Ser-307) by increasing Rho kinase activity. These results implicate a crucial role of the GGPPS/RhoA/Rho kinase/IRS-1 pathway in skeletal muscle, in which it mediates lipid-induced systemic insulin resistance in obese mice. Therefore, skeletal muscle GGPPS may represent a potential pharmacological target for the prevention and treatment of obesity-related type 2 diabetes.


Assuntos
Farnesiltranstransferase/metabolismo , Resistência à Insulina , Metabolismo dos Lipídeos , Complexos Multienzimáticos/metabolismo , Músculo Esquelético/metabolismo , Quinases Associadas a rho/metabolismo , Proteína rhoA de Ligação ao GTP/metabolismo , Animais , Diabetes Mellitus Experimental/complicações , Diabetes Mellitus Experimental/enzimologia , Farnesiltranstransferase/genética , Camundongos , Complexos Multienzimáticos/genética , Obesidade/complicações , Obesidade/enzimologia
17.
New Phytol ; 209(1): 252-64, 2016 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-26224411

RESUMO

Most plastid isoprenoids, including photosynthesis-related metabolites such as carotenoids and the side chain of chlorophylls, tocopherols (vitamin E), phylloquinones (vitamin K), and plastoquinones, derive from geranylgeranyl diphosphate (GGPP) synthesized by GGPP synthase (GGPPS) enzymes. Seven out of 10 functional GGPPS isozymes in Arabidopsis thaliana reside in plastids. We aimed to address the function of different GGPPS paralogues for plastid isoprenoid biosynthesis. We constructed a gene co-expression network (GCN) using GGPPS paralogues as guide genes and genes from the upstream and downstream pathways as query genes. Furthermore, knock-out and/or knock-down ggpps mutants were generated and their growth and metabolic phenotypes were analyzed. Also, interacting protein partners of GGPPS11 were searched for. Our data showed that GGPPS11, encoding the only plastid isozyme essential for plant development, functions as a hub gene among GGPPS paralogues and is required for the production of all major groups of plastid isoprenoids. Furthermore, we showed that the GGPPS11 protein physically interacts with enzymes that use GGPP for the production of carotenoids, chlorophylls, tocopherols, phylloquinone, and plastoquinone. GGPPS11 is a hub isozyme required for the production of most photosynthesis-related isoprenoids. Both gene co-expression and protein-protein interaction likely contribute to the channeling of GGPP by GGPPS11.


Assuntos
Alquil e Aril Transferases/metabolismo , Proteínas de Arabidopsis/metabolismo , Arabidopsis/enzimologia , Terpenos/metabolismo , Alquil e Aril Transferases/genética , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Carotenoides/metabolismo , Clorofila/metabolismo , Isoenzimas , Fenótipo , Fotossíntese , Plastídeos/enzimologia , Fosfatos de Poli-Isoprenil/metabolismo , Mapeamento de Interação de Proteínas
18.
J Pathol ; 235(5): 672-85, 2015 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-25385233

RESUMO

G protein-regulated cell function is crucial for cardiomyocytes, and any deregulation of its gene expression or protein modification can lead to pathological cardiac hypertrophy. Herein, we report that protein prenylation, a lipidic modification of G proteins that facilitates their association with the cell membrane, might control the process of cardiomyocyte hypertrophy. We found that geranylgeranyl diphosphate synthase (GGPPS), a key enzyme involved in protein prenylation, played a critical role in postnatal heart growth by regulating cardiomyocyte size. Cardiac-specific knockout of GGPPS in mice led to spontaneous cardiac hypertrophy, beginning from week 4, accompanied by the persistent enlargement of cardiomyocytes. This hypertrophic effect occurred by altered prenylation of G proteins. Evaluation of the prenylation, membrane association and hydrophobicity showed that Rheb was hyperactivated and increased mTORC1 signalling pathway after GGPPS deletion. Protein farnesylation or mTORC1 inhibition blocked GGPPS knockdown-induced mTORC1 activation and suppressed the larger neonatal rat ventricle myocyte size and cardiomyocyte hypertrophy in vivo, demonstrating a central role of the FPP-Rheb-mTORC1 axis for GGPPS deficiency-induced cardiomyocyte hypertrophy. The sustained cardiomyocyte hypertrophy progressively provoked cardiac decompensation and dysfunction, ultimately causing heart failure and adult death. Importantly, GGPPS was down-regulated in the hypertrophic hearts of mice subjected to transverse aortic constriction (TAC) and in failing human hearts. Moreover, HPLC-MS/MS detection revealed that the myocardial farnesyl diphosphate (FPP):geranylgeranyl diphosphate (GGPP) ratio was enhanced after pressure overload. Our observations conclude that the alteration of protein prenylation promotes cardiomyocyte hypertrophic growth, which acts as a potential cause for pathogenesis of heart failure and may provide a new molecular target for hypertrophic heart disease clinical therapy.


Assuntos
Cardiomegalia/enzimologia , Farnesiltranstransferase/deficiência , Insuficiência Cardíaca/enzimologia , Proteínas Monoméricas de Ligação ao GTP/metabolismo , Complexos Multiproteicos/metabolismo , Miócitos Cardíacos/enzimologia , Neuropeptídeos/metabolismo , Prenilação de Proteína , Transdução de Sinais , Serina-Treonina Quinases TOR/metabolismo , Animais , Cardiomegalia/tratamento farmacológico , Cardiomegalia/genética , Cardiomegalia/patologia , Cardiomegalia/fisiopatologia , Linhagem Celular , Modelos Animais de Doenças , Progressão da Doença , Farnesiltranstransferase/genética , Feminino , Insuficiência Cardíaca/tratamento farmacológico , Insuficiência Cardíaca/genética , Insuficiência Cardíaca/patologia , Insuficiência Cardíaca/fisiopatologia , Masculino , Alvo Mecanístico do Complexo 1 de Rapamicina , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas Monoméricas de Ligação ao GTP/genética , Complexos Multiproteicos/antagonistas & inibidores , Miócitos Cardíacos/efeitos dos fármacos , Miócitos Cardíacos/patologia , Neuropeptídeos/genética , Inibidores de Proteínas Quinases/farmacologia , Prenilação de Proteína/efeitos dos fármacos , Interferência de RNA , Proteína Enriquecida em Homólogo de Ras do Encéfalo , Ratos , Ratos Sprague-Dawley , Transdução de Sinais/efeitos dos fármacos , Sirolimo/farmacologia , Serina-Treonina Quinases TOR/antagonistas & inibidores , Fatores de Tempo , Transfecção , Função Ventricular Esquerda
19.
Plant Cell Rep ; 34(12): 2179-88, 2015 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-26449416

RESUMO

KEY MESSAGE: We found triptolide synthesis is correlated with the expressions of TwGGPPS1 and TwGGPPS4 . This lays the foundation for future studies of biosynthetic pathways for triptolide and other diterpenoids in T. wilfordii. Tripterygium wilfordii is a traditional Chinese medical plant commonly used to treat rheumatoid arthritis. One of its main bioactive compounds is triptolide, which is identified as an abietane-type diterpenoid natural product. Geranylgeranyl diphosphate synthase (GGPPS) catalyses the synthesis of GGPP (geranylgeranyl diphosphate), the common precursor of diterpenes, and is therefore a crucial enzyme in diterpene biosynthesis. A previous study showed that GGPP could be catalyzed by copalyl diphosphate synthase and kaurene synthase like of Salvia miltiorrhiza (SmCPS, SmKSL) to miltiradiene, a key intermediate in tanshinone biosynthesis. In this paper, five new full-length cDNAs (TwGGPPS) encoding GGPP synthases were cloned from T. wilfordii. Sequence comparisons revealed that all six TwGGPPSs (including TwGGPPS2 cloned previously) exhibit similarities to GGPPSs of other plants. Subsequent functional complement assays demonstrated that TwGGPPS1, TwGGPPS4 and TwGGPPS5 can participate in miltiradiene biosynthesis in the recombinant E. coli. Correlation analysis of gene expressions and secondary metabolite accumulation indicated that TwGGPPS1 and TwGGPPS4 are likely involved in the biosynthesis of triptolide. These findings lay the foundation for future studies of the biosynthetic pathways for triptolide and other diterpenoids in T. wilfordii.


Assuntos
Diterpenos/metabolismo , Farnesiltranstransferase/genética , Fenantrenos/metabolismo , Tripterygium/enzimologia , Acetatos/metabolismo , Sequência de Aminoácidos , Vias Biossintéticas , Clonagem Molecular , Ciclopentanos/metabolismo , Diterpenos/química , Compostos de Epóxi/química , Compostos de Epóxi/metabolismo , Escherichia coli/genética , Escherichia coli/metabolismo , Farnesiltranstransferase/metabolismo , Dados de Sequência Molecular , Oxilipinas/metabolismo , Fenantrenos/química , Filogenia , Reguladores de Crescimento de Plantas/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Fosfatos de Poli-Isoprenil/química , Fosfatos de Poli-Isoprenil/metabolismo , Alinhamento de Sequência , Tripterygium/genética
20.
Inflammation ; 2024 Jul 25.
Artigo em Inglês | MEDLINE | ID: mdl-39052180

RESUMO

Acute respiratory distress syndrome (ARDS) and acute lung injury (ALI) are life-threatening diseases. Neutrophil extracellular traps (NETs) play a key role in lung damage. Geranylgeranyl diphosphate synthase (GGPPS) is associated with the development of inflammatory diseases. We aimed to explore the role of GGPPS in NETs formation in ARDS/ALI. First, lung pathological changes in lipopolysaccharide (LPS)-induced ALI mice after myeloid-specific GGPPS deletion were evaluated. The level of NETs formation was analyzed by immunofluorescence, PicoGreen assay and Western blotting. Next, we determined the role of GGPPS in NETs formation and underlying mechanisms using immunofluorescence, flow cytometry, DCFH-DA, and SYTOX GREEN staining in vitro. Finally, the correlation between GGPPS expression incirculating neutrophils and dsDNA levels in plasma was evaluated. Myeloid-specific GGPPS deletion mice showed increased NETs deposition in lung tissue and aggravated histopathological damage of lung tissue. In vitro, GGPPS deficiency in neutrophils resulted in increased NETs formation by Phorbol-12-myristate-13-acetate (PMA), which was reversed by Geranylgeranyl diphosphate (GGPP). In addition, inhibitors blocking protein kinase C (PKC) and NADPH-oxidase (NOX) decreased NETs formation induced by GGPPS deletion. Importantly, GGPPS expression in circulating neutrophils was decreased in ARDS patients compared with the healthy control, and the level of dsDNA in plasma of ARDS patients was negatively correlated with the GGPPS expression. Taken together, GGPPS deficiency in neutrophils aggravates LPS-induced lung injury by promoting NETs formation via PKC/NOX signaling. Thus, neutrophil GGPPS could be a key therapeutic target for ARDS.

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