Hexanoic acid (Hx) is a short natural monocarboxylic acid present in some fruits and plants. Previous studies reported that soil drench application of this acid induces effective resistance in tomato plants against Botrytis cinerea and Pseudomonas syringae and in citrus against Alternaria alternata and Xanthomonas citri. In this work, we performed an in deep study of the metabolic changes produced in citrus by the application of Hx in response to the challenge pathogen A. alternata, focusing on the response of the plant. Moreover, we used (13)C labeled hexanoic to analyze its behavior inside the plants. Finally, we studied the volatile emission of the treated plants after the challenge inoculation. Drench application of (13)C labeled hexanoic demonstrated that this molecule stays in the roots and is not mobilized to the leaves, suggesting long distance induction of resistance. Moreover, the study of the metabolic profile showed an alteration of more than 200 molecules differentially induced by the application of the compound and the inoculation with the fungus. Bioinformatics analysis of data showed that most of these altered molecules could be related with the mevalonic and linolenic pathways suggesting the implication of these pathways in the induced resistance mediated by Hx. Finally, the application of this compound showed an enhancement of the emission of 17 volatile metabolites. Taken together, this study indicates that after the application of Hx this compound remains in the roots, provoking molecular changes that may trigger the defensive response in the rest of the plant mediated by changes in the mevalonic and linolenic pathways and enhancing the emission of volatile compounds, suggesting for the first time the implication of mevalonic pathway in response to hexanoic application.

Prevention of triple-negative breast cancer (TNBC) is hampered by lack of knowledge about the drivers of tumorigenesis.

The yeast Saccharomyces cerevisiae can be a useful model for studying cellular mechanisms related to sterol synthesis in humans due to the high similarity of the mevalonate pathway between these organisms. This metabolic pathway plays a key role in multiple cellular processes by synthesizing sterol and nonsterol isoprenoids. Statins are well-known inhibitors of 3-hydroxy-3-methylglutaryl-CoA reductase (HMGR), the key enzyme of the cholesterol synthesis pathway. However, the effects of statins extend beyond their cholesterol-lowering action, since inhibition of HMGR decreases the synthesis of all products downstream in the mevalonate pathway. Using transgenic yeast expressing human HMGR or either yeast HMGR isoenzyme we studied the effects of simvastatin, atorvastatin, fluvastatin and rosuvastatin on the cell metabolism.

The role of hepatic free cholesterol (FC) in nonalcoholic steatohepatitis (NASH) is raised up and the intervention with cholesterol synthesis will be a potential therapeutic target. This study investigated the hepatoprotective effect of mevalonic acid pathway inhibition by Zoledronic acid (ZA) on the hepatic changes associated with high fat diet (HFD) in rats.Thirty two male Wistar rats were used. They were divided into 2 groups: (I) control naïve (II) NASH: induced by HFD for 12 weeks, this group is subdivided into (A) NASH untreated (B)NASH + ZA (50ug/kg/week) i.p. for 12 weeks (C) NASH + ZA (100μg/kg/week) i.p. for 12 weeks. Portal pressure (PP), liver enzymes AST and ALT, serum glucose, lipid profile, hepatic levels of tumor necrosis factor alpha (TNF-α), vascular endothelial growth factor (VEGF), FC and triglyceride (TG), histopathological changes and expression of both hepatic alpha smooth muscle actin (α-SMA) and Caspase-3 were measured. ZA significantly prevented portal hypertension, worsening in liver function, and dyslipidemia. The hepatic levels of TNF-α, VEGF, FC and TG were significantly decreased in comparison to NASH untreated group. ZA hindered the histopathological changes induced by HFD. ZA inhibited the expression of hepatic α-SMA and Caspase-3 with significant difference favor the high dose intervention. ZA in a dose related manner prevents the hepatic pathological effects of chronic HFD ingestion in rats. This may be largely mediated by its ability to reduce TNF-α and hepatic FC.

There is a growing interest in the use of microbial fermentation for the generation of high-demand, high-purity chemicals using cheap feedstocks in an environmentally friendly manner. One example explored here is the production of isoprene (C5H8), a hemiterpene, which is primarily polymerized to polyisoprene in synthetic rubber in tires but which can also be converted to C10 and C15 biofuels. The strictly anaerobic, acetogenic bacterium Clostridium ljungdahlii, used in all of the work described here, is capable of glycolysis using the Embden-Meyerhof-Parnas pathway and of carbon fixation using the Wood-Ljungdahl pathway. Clostridium-Escherichia coli shuttle plasmids, each bearing either 2 or 3 different heterologous genes of the eukaryotic mevalonic acid (MVA) pathway or eukaryotic isopentenyl pyrophosphate isomerase (Idi) and isoprene synthase (IspS), were constructed and electroporated into C. ljungdahlii These plasmids, one or two of which were introduced into the host cells, enabled the synthesis of mevalonate and of isoprene from fructose and from syngas (H2, CO2, and CO) and the conversion of mevalonate to isoprene. All of the heterologous enzymes of the MVA pathway, as well as Idi and IspS, were shown to be synthesized at high levels in C. ljungdahlii, as demonstrated by Western blotting, and were enzymatically active, as demonstrated by in vivo product synthesis. The quantities of mevalonate and isoprene produced here are far below what would be required of a commercial production strain. However, proposals are made that could enable a substantial increase in the mass yield of product formation.IMPORTANCE This study demonstrates the ability to synthesize a heterologous metabolic pathway in C. ljungdahlii, an organism capable of metabolizing either simple sugars or syngas or both together (mixotrophy). Syngas, an inexpensive source of carbon and reducing equivalents, is produced as a major component of some industrial waste gas, and it can be generated by gasification of cellulosic biowaste and of municipal solid waste. Its conversion to useful products therefore offers potential cost and environmental benefits. The ability of C. ljungdahlii to grow mixotrophically also enables the recapture, should there be sufficient reducing equivalents available, of the CO2 released upon glycolysis, potentially increasing the mass yield of product formation. Isoprene is the simplest of the terpenoids, and so the demonstration of its production is a first step toward the synthesis of higher-value products of the terpenoid pathway.

The spontaneous fermentation of alcoholic beverage is a bioprocess donated by microbiota with complex stress environments. Among various microbes, non-Saccharomyces yeasts have high stress tolerance and significantly affect the taste and quality of products in process. Although many researchers have focused on the influence of acid stress, the mechanism of non-Saccharomyces yeasts to tolerant stress remains unclear in microbiota. To bridge the gap, we constructed in situ and in vitro studies to explore the reduction pathway of acetic acid in non-Saccharomyces yeasts. In this study, we found Schizosaccharomyces pombe has special capacities to resist 10 g/L acetic acid in laboratory cultures and decrease the average concentration of acetic acid from 9.62 to 6.55 g/kg fermented grains in Chinese Maotai-flavor liquor (Baijiu) production. Moreover, Schi. pombe promoted metabolic level of mevalonate pathway (high expressions of gene ACCAT1, HMGCS1, and HMGCR1) to degrade a high concentration of acetic acid. Meanwhile, Schi. pombe also improved the concentration of mevalonic acid that is the precursor of terpenes to enhance the taste and quality of Baijiu. Overall, the synchronicity of reduction and generation in Schi. pombe advances the current knowledge to guide more suitable strategies for mechanism studies of non-Saccharomyces yeasts in fermented industries of alcoholic beverages.

Geranylgeranoic acid (GGA) has been developed as a preventive agent against second primary hepatoma. Recently, GGA was reported to induce cell death in human hepatoma cells via TLR4-mediated pyroptosis. We have reported that GGA is enzymatically biosynthesized from mevalonic acid in human hepatoma-derived cells and that endogenous GGA is found in most organs of rats. In addition, we found that upregulation of endogenous GGA levels by zaragozic acid A (ZAA) induced cell death in human hepatoma-derived cells. Therefore, we investigated the age-related changes in hepatic GGA and the possibility of suppressing hepatocarcinogenesis by GGA supplementation using male C3H/HeN mice that spontaneously develop hepatoma. We measured endogenous GGA and mRNA of monoamine oxidase (BMAOB), a key enzyme of GGA biosynthesis, in the liver of male C3H/HeN mice aged 6-93 weeks. We also tried suppressing spontaneous hepatocarcinogenesis by a single administration of GGA to C3H/HeN mice. Hepatic GGA content and Maob mRNA expression level age-dependently decreased in male C3H/HeN mice; some of which produced spontaneous hepatoma in 2 years. A single oral administration of GGA at 11 months of age significantly prevented hepatoma in terms of the number and weight of tumors per mouse at 24 months. Oral supplementation with GGA or geranylgeraniol significantly increased endogenous hepatic GGA contents dose-dependently; and ZAA dramatically upregulated hepatic GGA. In this study; we found an age-dependent decrease in hepatic endogenous GGA in male C3H/HeN mice and efficient prevention of spontaneous hepatoma by a single administration of GGA at 11 months of age.

Geranylgeranoic acid (GGA) was developed as a preventative agent against second primary hepatoma, and was reported to induce cell death in human hepatoma cells via Toll-like receptor 4 (TLR4)-mediated pyroptosis. We recently reported that GGA is enzymatically biosynthesized from mevalonic acid in human hepatoma-derived HuH-7 cells and that endogenous GGA is found in most rat organs including the liver. An unbiased metabolomics analysis of ice-cold 50% acetonitrile extracts from control and GGA-treated cells was performed in this study to characterize the intracellular metabolic changes in GGA-induced pyroptosis and to analyze their relationship with the mechanism of GGA-induced cell death. The total positive ion chromatograms of the cellular extracts in ultra-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry were apparently unchanged after GGA treatment, but an orthogonal partial least squares-discriminant analysis score plot clearly discriminated the intracellular metabolite profiles of GGA-treated cells from that of control cells. S-plot analysis revealed 15 potential biomarkers up-regulated by 24-h GGA treatment according to their variable importance in the projection value of more than 1, and the subsequent metabolomics analysis identified nine of these metabolites as a group of lysophospholipids containing lysophosphatidylcholine with C16:0, C20:4, or C20:3 fatty acids. The possible roles of these lysophospholipids in GGA-induced pyroptosis are discussed.

Human studies have shown diurnal rhythms of cholesterol and bile acid synthesis, but a better understanding of the role of the circadian system in cholesterol homeostasis is needed for the development of targeted interventions to improve metabolic health. Therefore, we performed a systematic literature search on the diurnal rhythms of cholesterol synthesis and absorption markers and of bile acid synthesis markers. We also examined the diurnal rhythms of the cholesterol synthesis markers lathosterol and desmosterol, and of the cholesterol absorption markers cholestanol, campesterol, and sitosterol in serum samples from the Bispebjerg study. These samples were collected every three hours over a 24-hour period in healthy males (n = 24) who consumed low-fat meals. The systematic search identified sixteen papers that had examined the diurnal rhythms of the cholesterol synthesis markers lathosterol (n = 3), mevalonate (n = 9), squalene (n = 2), or the bile acid synthesis marker 7α-hydroxy-4-cholesten-3-one (C4) (n = 4). Results showed that lathosterol, mevalonate, and squalene had a diurnal rhythm with nocturnal peaks, while C4 had a diurnal rhythm with daytime peaks. Furthermore, cosinor analyses of the serum samples showed a significant diurnal rhythm for lathosterol (cosinor p < 0.001), but not for desmosterol, campesterol, sitosterol, and cholestanol (cosinor p > 0.05). In conclusion, cholesterol synthesis and bile acid synthesis have a diurnal rhythm, though no evidence for a diurnal rhythm of cholesterol absorption was found under highly standardised conditions. More work is needed to further explore the influence of external factors on the diurnal rhythms regulating cholesterol homeostasis.

Statins are cholesterol-lowering drugs with pleiotropic activities including inhibition of isoprenylation and reduction of signals driving cell proliferation and survival responses.

Ent-homocyclopiamine B (1), a new prenylated indole alkaloid bearing an alicyclic nitro group along with 2-methylbutane-1,2,4-triol (2) were isolated from an endophytic fungus Penicillium concentricum of the liverwort Trichocolea tomentella (Trichocoleaceae). The structure of 1 was elucidated through extensive spectroscopic analyses and comparison with data reported for a structurally related nitro-bearing Penicillium metabolite, clopiamine C (3), which contain an indolizidine ring instead of the quinolizine ring in 1. The new compound, ent-homocyclopiamine B, exhibited slight growth inhibition against Gram-positive bacteria. Based on the reported biosynthesis of related compounds and the isolation of the mevalonic acid derived compound 2-methyl-1,2,4-butanetriol (2), we proposed that ent-homocylopiamine B (1) was biosynthesized from lysine and prenyl group-producing mevalonic pathway.

Plants synthesize a great variety of isoprenoid products that are required not only for normal growth and development but also for their adaptive responses to environmental challenges. However, despite the remarkable diversity in the structure and function of plant isoprenoids, they all originate from a single metabolic precursor, mevalonic acid. The synthesis of mevalonic acid is catalysed by the enzyme, 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMG- CoA reductase). The analysis of the amino acid sequence of HMG-CoA reductase from Artemisia annua L. plant showed that it belongs to class I HMG-CoA reductase family. The three dimensional structure of HMG-CoA reductase of Artemisia annua has been generated from amino acid sequence using homology modelling with backbone structure of human HMG-CoA reductase as template. The model was generated using the SWISS MODEL SERVER. The generated 3-D structure of HMG-CoA reductase was evaluated at various web interfaced servers to checks the stereo interfaced quality of the structure in terms of bonds, bond angles, dihedral angles and non-bonded atom-atom distances, structural as well as functional domains etc. The generated model was visualized using the RASMOL. Structural analysis of HMG-CoA reductase from Artemisia annua L. plant hypothesize that the N and C-terminals are positioned in cytosol by the two membrane spanning helices and the C-terminals domain shows similarity to the human HMG-CoA reductase enzyme indicating that they both had potential catalytic similarities.

Terpenes are industrially relevant natural compounds the biosynthesis of which relies on two well-established-mevalonic acid (MVA) and methyl erythritol phosphate (MEP)-pathways. Both pathways are widely distributed in all domains of life, the former is predominantly found in eukaryotes and archaea and the latter in eubacteria and chloroplasts. These two pathways supply isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP), the universal building blocks of terpenes.

Whether bisphosphonates and RANKL inhibitors play a novel role in delaying cardiovascular calcification is unknown. Their action on regulatory enzymes in the mevalonic acid pathway, which is implicated in both bone and lipid metabolism, may be a novel therapeutic target to manage coronary artery disease (CAD). Such therapies may particularly be relevant in those for whom traditional cardiovascular therapies are no longer sufficient to control disease progression.

Statin derivatives can inhibit the replication of a range of viruses, including hepatitis C virus (HCV, Hepacivirus), dengue virus (Flavivirus), African swine fever virus (Asfarviridae) and poliovirus (Picornaviridae). We assess the antiviral effect of fluvastatin in cells infected with orbiviruses (bluetongue virus (BTV) and Great Island virus (GIV)). The synthesis of orbivirus outer-capsid protein VP2 (detected by confocal immunofluorescence imaging) was used to assess levels of virus replication, showing a reduction in fluvastatin-treated cells. A reduction in virus titres of ~1.7 log (98%) in fluvastatin-treated cells was detected by a plaque assay. We have previously identified a fourth non-structural protein (NS4) of BTV and GIV, showing that it interacts with lipid droplets in infected cells. Fluvastatin, which inhibits 3-hydroxy 3-methyl glutaryl CoA reductase in the mevalonic acid pathway, disrupts these NS4 interactions. These findings highlight the role of the lipid pathways in orbivirus replication and suggest a greater role for the membrane-enveloped orbivirus particles than previously recognised. Chemical intermediates of the mevalonic acid pathway were used to assess their potential to rescue orbivirus replication. Pre-treatment of IFNAR(-/-) mice with fluvastatin promoted their survival upon challenge with live BTV, although only limited protection was observed.

Statins are widely used cholesterol-lowering drugs. Their potential application in anti-cancer treatment is also under investigation. The individual variance in statin response has been observed, which may be caused by the variation in human HMG-CoA reductase (hHMGR)-the inhibition target of statin drugs. Herein, we reported the design and construction of two Escherichia coli whole-cell biosensors. The first one is statin-efficacy testing sensor, which is composed of two separate modules: a hybrid mevalonate (MVA) pathway and a HMG-CoA sensing system. A truncated hHMGR was used as the key enzyme of the MVA pathway and a promiscuous transcription factor (TF) BsFapR was used as the HMG-CoA sensor. When hHMGR was inhibited by statins, HMG-CoA accumulated intracellularly and was sensed by BsFapR, which subsequently turned on its cognate promoter. This biosensor has the potential to be used as a "precision medicine" tool-selecting potent statin drugs for individual patients. The second one is a statin-production testing sensor, which is based on another promiscuous TF AraCM that can sense statins. This biosensor can be used in optimization of statin-producing strains. The prototypes of these two biosensors were successfully constructed and their further optimization is highly expected.

We have established methodology for the isolation and characterization of a novel endophytic fungus from the inner bark of medicinal plant Nothapodytes foetida, which produced camptothecin in Sabouraud broth (SB) under shake flask conditions. Camptothecin and its related compounds are at present obtained by extraction from intact plants, but fungal endopytes may be an alternative source of production. In present study we have observed the effect of different nutrient combinations and precursors (tryptophan, tryptamine, geraniol, citral, mevalonic acid and leucine) on the accumulation of camptothecin by endophytic fungus Entrophospora infrequens. The precursors were fed either alone or in combinations (tryptophan and geraniol, tryptophan and citral, tryptophan and mevalonic acid, tryptophan and leucine). The highest camptothecin content was observed in the range of 503 ± 25µg/100g dry cell mass in Sabouraud medium. Camptothecin content in the medium was increased by 2.5 folds by the presence of tryptophan and leucine whereas the production with trytophan was also significantly different from other treatments. Furthermore, the effect of fungal camptothecin on the morphology of human cancer cell lines was also studied. The treated cells showed reduction in size, condensation of nucleus and the protoplasmic extensions were reduced. All these characteristics are found in apoptotic cells.

Juvenile hormones (JH) regulate development and reproductive maturation in insects. JHs are synthesized through the mevalonate pathway (MVAP), an ancient metabolic pathway present in the three domains of life. Mevalonate kinase (MVK) is a key enzyme in the MVAP. MVK catalyzes the synthesis of phosphomevalonate (PM) by transferring the γ-phosphoryl group from ATP to the C5 hydroxyl oxygen of mevalonic acid (MA). Despite the importance of MVKs, these enzymes have been poorly characterized in insects.

Mevalonate kinase deficiency (MKD) is a monogenic auto-inflammatory disease. Its manifestations range from partial MKD to mevalonic aciduria (MVA). All patients display a periodic fever, and MVA patients additionally exhibit severe neurological involvement. The objective of this work was to describe neurological manifestations of MKD.

Statins and fibrates (weak PPARalpha agonists) are prescribed for the treatment of lipid disorders. Both drugs cause myopathy, but with a low incidence, 0.1-0.5%. However, combined statin and fibrate therapy can enhance myopathy risk. We tested the myotoxic potential of PPAR subtype selective agonists alone and in combination with statins in a differentiated rat myotube model. A pharmacologically potent experimental PPARalpha agonist, Compound A, induced myotoxicity as assessed by TUNEL staining at a minimum concentration of 1 nM, while other weaker PPARalpha compounds, for example, WY-14643, Gemfibrozil and Bezafibrate increased the percentage of TUNEL-positive nuclei at micromolar concentrations. In contrast, the PPARgamma agonist Rosiglitazone caused little or no cell death at up to 10 muM and the PPARdelta ligand GW-501516 exhibited comparatively less myotoxicity than that seen with Compound A. An experimental statin (Compound B) and Atorvastatin also increased the percentage of TUNEL-positive nuclei and co-treatment with WY-14643, Gemfibrozil or Bezafibrate had less than a full additive effect on statin-induced cell killing. The mechanism of PPARalpha agonist-induced cell death was different from that of statins. Unlike statins, Compound A and WY-14643 did not activate caspase 3/7. In addition, mevalonate and geranylgeraniol reversed the toxicity caused by statins, but did not prevent the cell killing induced by WY-14643. Furthermore, unlike statins, Compound A did not inhibit the isoprenylation of rab4 or rap1a. Interestingly, Compound A and Compound B had differential effects on ATP levels. Taken together, these observations support the hypothesis that in rat myotube cultures, PPARalpha agonism mediates in part the toxicity response to PPARalpha compounds. Furthermore, PPARalpha agonists and statins cause myotoxicity through distinct and independent pathways.