Julien Dairou

Guanine glycation repair by DJ-1/Park7 and its bacterial homologs

Not-so-sweet DNA damage repaired Glyoxal and methylglyoxal, by-products of sugar metabolism that are present in all cells, can react with, and thus damage, DNA. Indeed, glycation of guanine (G) is as prevalent as the major product of oxidative damage in DNA, 8-oxo-dG. Richarme et al. show that both prokaryotes and eukaryotes have dedicated systems that specifically repair glycation damage (see the Perspective by Dingler and Patel). The parkinsonism-associated protein DJ-1/Park7 and its bacterial homologs Hsp31, YhbO, and YajL direct the enzymatic repair of damaged glycated bases in DNA. The proteins also clean up the more vulnerable pool of free nucleotides in the cell, which are more susceptible to glycation than the nucleotides within DNA. Science, this issue p. 208; see also p. 130 A DNA repair system acts specifically on bases damaged by reaction with by-products of sugar metabolism in the cell. DNA damage induced by reactive carbonyls (mainly methylglyoxal and glyoxal), called DNA glycation, is quantitatively as important as oxidative damage. DNA glycation is associated with increased mutation frequency, DNA strand breaks, and cytotoxicity. However, in contrast to guanine oxidation repair, how glycated DNA is repaired remains undetermined. Here, we found that the parkinsonism-associated protein DJ-1 and its bacterial homologs Hsp31, YhbO, and YajL could repair methylglyoxal- and glyoxal-glycated nucleotides and nucleic acids. DJ-1–depleted cells displayed increased levels of glycated DNA, DNA strand breaks, and phosphorylated p53. Deglycase-deficient bacterial mutants displayed increased levels of glycated DNA and RNA and exhibited strong mutator phenotypes. Thus, DJ-1 and its prokaryotic homologs constitute a major nucleotide repair system that we name guanine glycation repair.

Inhibition of central de novo ceramide synthesis restores insulin signaling in hypothalamus and enhances β-cell function of obese Zucker rats

Objectives Hypothalamic lipotoxicity has been shown to induce central insulin resistance and dysregulation of glucose homeostasis; nevertheless, elucidation of the regulatory mechanisms remains incomplete. Here, we aimed to determine the role of de novo ceramide synthesis in hypothalamus on the onset of central insulin resistance and the dysregulation of glucose homeostasis induced by obesity. Methods Hypothalamic GT1-7 neuronal cells were treated with palmitate. De novo ceramide synthesis was inhibited either by pharmacological (myriocin) or molecular (si-Serine Palmitoyl Transferase 2, siSPT2) approaches. Obese Zucker rats (OZR) were intracerebroventricularly infused with myriocin to inhibit de novo ceramide synthesis. Insulin resistance was determined by quantification of Akt phosphorylation. Ceramide levels were quantified either by a radioactive kinase assay or by mass spectrometry analysis. Glucose homeostasis were evaluated in myriocin-treated OZR. Basal and glucose-stimulated parasympathetic tonus was recorded in OZR. Insulin secretion from islets and β-cell mass was also determined. Results We show that palmitate impaired insulin signaling and increased ceramide levels in hypothalamic neuronal GT1-7 cells. In addition, the use of deuterated palmitic acid demonstrated that palmitate activated several enzymes of the de novo ceramide synthesis pathway in hypothalamic cells. Importantly, myriocin and siSPT2 treatment restored insulin signaling in palmitate-treated GT1-7 cells. Protein kinase C (PKC) inhibitor or a dominant-negative PKCζ also counteracted palmitate-induced insulin resistance. Interestingly, attenuating the increase in levels of hypothalamic ceramides with intracerebroventricular infusion of myriocin in OZR improved their hypothalamic insulin-sensitivity. Importantly, central myriocin treatment partially restored glucose tolerance in OZR. This latter effect is related to the restoration of glucose-stimulated insulin secretion and an increase in β-cell mass of OZR. Electrophysiological recordings also showed an improvement of glucose-stimulated parasympathetic nerve activity in OZR centrally treated with myriocin. Conclusion Our results highlight a key role of hypothalamic de novo ceramide synthesis in central insulin resistance installation and glucose homeostasis dysregulation associated with obesity.

Genetic deficiency of indoleamine 2,3-dioxygenase promotes gut microbiota-mediated metabolic health

The association between altered gut microbiota, intestinal permeability, inflammation and cardiometabolic diseases is becoming increasingly clear but remains poorly understood1,2. Indoleamine 2,3-dioxygenase is an enzyme induced in many types of immune cells, including macrophages in response to inflammatory stimuli, and catalyzes the degradation of tryptophan along the kynurenine pathway. Indoleamine 2,3-dioxygenase activity is better known for its suppression of effector T cell immunity and its activation of regulatory T cells3,4. However, high indoleamine 2,3-dioxygenase activity predicts worse cardiovascular outcome5–9 and may promote atherosclerosis and vascular inflammation6, suggesting a more complex role in chronic inflammatory settings. Indoleamine 2,3-dioxygenase activity is also increased in obesity10–13, yet its role in metabolic disease is still unexplored. Here, we show that obesity is associated with an increase of intestinal indoleamine 2,3-dioxygenase activity, which shifts tryptophan metabolism from indole derivative and interleukin-22 production toward kynurenine production. Indoleamine 2,3-dioxygenase deletion or inhibition improves insulin sensitivity, preserves the gut mucosal barrier, decreases endotoxemia and chronic inflammation, and regulates lipid metabolism in liver and adipose tissues. These beneficial effects are due to rewiring of tryptophan metabolism toward a microbiota-dependent production of interleukin-22 and are abrogated after treatment with a neutralizing anti-interleukin-22 antibody. In summary, we identify an unexpected function of indoleamine 2,3-dioxygenase in the fine tuning of intestinal tryptophan metabolism with major consequences on microbiota-dependent control of metabolic disease, which suggests indoleamine 2,3-dioxygenase as a potential therapeutic target.Indoleamine 2,3-dioxygenase normally suppresses inflammation, but knockout of its gene is metabolically beneficial as its depletion reshapes the gut microbiota.

DYRK1A inhibition and cognitive rescue in a Down syndrome mouse model are induced by new fluoro-DANDY derivatives

Inhibition of DYRK1A kinase, produced by chromosome 21 and consequently overproduced in trisomy 21 subjects, has been suggested as a therapeutic approach to treating the cognitive deficiencies observed in Down syndrome (DS). We now report the synthesis and potent DYRK1A inhibitory activities of fluoro derivatives of 3,5-di(polyhydroxyaryl)-7-azaindoles (F-DANDYs). One of these compounds (3-(4-fluorophenyl)-5-(3,4-dihydroxyphenyl)-1H-pyrrolo[2,3-b]pyridine, 5a) was selected for in vivo studies of cognitive rescuing effects in a standard mouse model of DS (Ts65Dn line). Using the Morris water maze task, Ts65Dn mice treated i.p. with 20 mg/kg of 5a performed significantly better than Ts65Dn mice treated with placebo, confirming the promnesiant effect of 5a in the trisomic mice. Overall, these results demonstrate for the first time that selective and competitive inhibition of DYRK1A kinase by the F-DANDY derivative 5a may provide a viable treatment strategy for combating the memory and learning deficiencies encountered in DS.

Overexpression of the DYRK1A Gene (Dual-Specificity Tyrosine Phosphorylation-Regulated Kinase 1A) Induces Alterations of the Serotoninergic and Dopaminergic Processing in Murine Brain Tissues

Trisomy 21 (T21) or Down syndrome (DS) is the most common genetic disorder associated with intellectual disability and affects around 5 million persons worldwide. Neuroanatomical phenotypes associated with T21 include slight reduction of brain size and weight, abnormalities in several brain areas including spines dysgenesis, dendritic morphogenesis, and early neuroanatomical characteristics of Alzheimer’s disease. Monoamine neurotransmitters are involved in dendrites development, functioning of synapses, memory consolidation, and their levels measured in the cerebrospinal fluid, blood, or brain areas that are modified in individuals with T21. DYRK1A is one of the recognized key genes that could explain some of the deficits present in individuals with T21. We investigated by high-performance liquid chromatography with electrochemical detection the contents and processing of monoamines neurotransmitters in four brain areas of female and male transgenic mice for the Dyrk1a gene (mBactgDyrk1a). DYRK1A overexpression induced dramatic deficits in the serotonin contents of the four brain areas tested and major deficits in dopamine and adrenaline contents especially in the hypothalamus. These results suggest that DYRK1A overexpression might be associated with the modification of monoamines content found in individuals with T21 and reinforce the interest to target the level of DYRK1A expression as a therapeutic approach for persons with T21.

Original Chemical Series of Pyrimidine Biosynthesis Inhibitors That Boost the Antiviral Interferon Response

ABSTRACT De novo pyrimidine biosynthesis is a key metabolic pathway involved in multiple biosynthetic processes. Here, we identified an original series of 3-(1H-indol-3-yl)-2,3-dihydro-4H-furo[3,2-c]chromen-4-one derivatives as a new class of pyrimidine biosynthesis inhibitors formed by two edge-fused polycyclic moieties. We show that identified compounds exhibit broad-spectrum antiviral activity and immunostimulatory properties, in line with recent reports linking de novo pyrimidine biosynthesis with innate defense mechanisms against viruses. Most importantly, we establish that pyrimidine deprivation can amplify the production of both type I and type III interferons by cells stimulated with retinoic acid-inducible gene 1 (RIG-I) ligands. Altogether, our results further expand the current panel of pyrimidine biosynthesis inhibitors and illustrate how the production of antiviral interferons is tightly coupled to this metabolic pathway. Functional and structural similarities between this new chemical series and dicoumarol, which was reported before to inhibit pyrimidine biosynthesis at the dihydroorotate dehydrogenase (DHODH) step, are discussed.

Enhancement of 5-Fluorouracil cytotoxicity by Pyridoxal 5'-phosphate and Folinic acid in tandem.

The current study originates from the assumption that, in tumors, levels of naturally occurring pyridoxal 5′-phosphate (PLP) are too small to allow conversion of tetra hydro pteroylglutamate (H4PteGlu) into methylene tetra hydro pteroylglutamate (CH2-H4PteGlu) in amounts required to improve inhibition of thymidylate synthase by 5-fluorouracil (FUra) through ternary complex stabilization. The hypothesis relates to the low affinity for cofactor of the PLP-dependent serine hydroxymethyl transferase (SHMT), the enzyme that catalyzes formation of CH2-H4PteGlu by transfer of the Cβ of serine to H4PteGlu. Intracellular concentrations of PLP are smaller than the dissociation constant of SHMT for cofactor, which suggests that enzyme activity should be sensitive to PLP level changes. Three cancer cell lines were supplemented with PLP to investigate the influence of this cofactor on FUra cytotoxicity. Cells were exposed to FUra, FUra and folinic acid (FA), FUra and PLP, and FUra combined with both FA and PLP. The median–effect principle for concentration–effect analysis and combination indices were used to determine interactions on cytotoxicity. FUra cytotoxicity in vitro was enhanced by FA and PLP in tandem. Synergistic cytotoxic interaction of FUra with FA and PLP was demonstrated in HT29 and L1210 cells. Summation was found in HCT116 cells. Parenteral pyridoxamine was administered in mice to explore erythrocyte production of PLP in vivo. Cofactor attained levels in the range of the KD for binding to SHMT, and it was rapidly cleared from cells. Pharmacokinetics of pyridoxamine suggests that modulation of FUra by vitamin B6 could be achieved in vivo.

Further characterization of the Maillard deglycase DJ-1 and its prokaryotic homologs, deglycase 1/Hsp31, deglycase 2/YhbO, and deglycase 3/YajL

We reported recently that the Parkinsonism-associated protein DJ-1 and its bacterial homologs Hsp31, YhbO and YajL function as deglycases that repair proteins and nucleotides from endogeneous glycation by glyoxal and methylglyoxal, two reactive by-products of glucose metabolism responsible for up to 60% of glycation damage. Here, we show that DJ-1, deglycase 1 and deglycase 2 repair glyoxal- and methylglyoxal-glycated substrates, whereas deglycase 3 principally repairs glyoxal-glycated substrates. Moreover, deglycase 1 and 2 are overexpressed in stationary phase, whereas deglycase 3 is steadily expressed throughout bacterial growth. Finally, deglycase mutants overexpress glyoxalases, aldoketoreductases, glutathione-S-transferase and efflux pumps to alleviate carbonyl stress. In the discussion, we present an overview of the multiple functions of DJ-1 proteins. Our thourough work on deglycases provides compelling evidence that their previously reported glyoxalase III activity merely reflects their deglycase activity. Moreover, for their deglycase activity the Maillard deglycases likely recruit: i) their chaperone activity to interact with glycated proteins, ii) glyoxalase 1 activity to catalyze the rearrangement of Maillard products (aminocarbinols and hemithioacetals) into amides and thioesters, respectively, iii) their protease activity to cleave amide bonds of glycated arginine, lysine and guanine, and iv) glyoxalase 2 activity to cleave thioester bonds of glycated cysteine. Finally, because glycation affects many cellular processes, the discovery of the Maillard deglycases, awaited since 1912, likely constitutes a major advance for medical research, including ageing, cancer, atherosclerosis, neurodegenerative, post-diabetic, renal and autoimmune diseases.