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Th17 cells favor glycolytic metabolism, and pyruvate dehydrogenase (PDH) is the key bifurcation enzyme, which in its active dephosphorylated form advances the oxidative phosphorylation from glycolytic pathway. The transcriptional factor, inducible cAMP early repressor/cAMP response element modulator (ICER/CREM), has been shown to be induced in Th17 cells and to be overexpressed in CD4⁺ T cells from the patients with systemic lupus erythematosus (SLE). We found that glycolysis and lactate production in in vitro Th17-polarized T cells was reduced and that the expression of pyruvate dehydrogenase phosphatase catalytic subunit 2 (PDP2), an enzyme that converts the inactive PDH to its active form, and PDH enzyme activity were increased in Th17 cells from ICER/CREM-deficient animals. ICER was found to bind to the Pdp2 promoter and suppress its expression. Furthermore, forced expression of PDP2 in CD4⁺ cells reduced the in vitro Th17 differentiation, whereas shRNA-based suppression of PDP2 expression increased in vitro Th17 differentiation and augmented experimental autoimmune encephalomyelitis. At the translational level, PDP2 expression was decreased in memory Th17 cells from patients with SLE and forced expression of PDP2 in CD4⁺ T cells from lupus-prone MRL/lpr mice and patients with SLE suppressed Th17 differentiation. These data demonstrate the direct control of energy production during Th17 differentiation in health and disease by the transcription factor ICER/CREM at the PDH metabolism bifurcation level.

Aging is a multifactorial process that affects most of the biological functions of the organism and increases susceptibility to disease and death. Recent studies with animal models of accelerated aging have unveiled some mechanisms that also operate in physiological aging. However, little is known about the role of microRNAs (miRNAs) in this process. To address this question, we have analysed miRNA levels in Zmpste24 ‐deficient mice, a model of Hutchinson–Gilford progeria syndrome. We have found that expression of the miR‐29 family of miRNAs is markedly upregulated in Zmpste24 −/− progeroid mice as well as during normal aging in mouse. Functional analysis revealed that this transcriptional activation of miR‐29 is triggered in response to DNA damage and occurs in a p53‐dependent manner since p53 −/− murine fibroblasts do not increase miR‐29 expression upon doxorubicin treatment. We have also found that miR‐29 represses Ppm1d phosphatase, which in turn enhances p53 activity. Based on these results, we propose the existence of a novel regulatory circuitry involving miR‐29, Ppm1d and p53, which is activated in aging and in response to DNA damage. Previous data from worms implicated miRNAs in lifespan regulation. Analysis of a mammalian progeria model now identifies a miRNA induced upon both premature and physiological aging, contributing to these processes via p53 and DNA damage response pathways.

Anxiety is a serious psychiatric disorder, with a higher incidence rate in women than in men. Protein phosphatase Mg 2+ /Mn 2+ -dependent 1F (PPM1F), a serine/threonine phosphatase, has been shown to have multiple biological and cellular functions. However, the effects of PPM1F and its neuronal substrates on anxiety remain largely unclear. In this study, we showed that chronic restraint stress (CRS) induced anxiety-related behaviors only in female mice, while acute restraint stress (ARS) produced anxiety-related behaviors in both male and female mice in light–dark and elevated plus maze tests and induced upregulation of PPM1F and downregulation of brain-derived neurotrophic factor (BDNF) expression in the hippocampus. Adeno-associated virus–mediated overexpression of PPM1F or conditional knockout of BDNF in dentate gyrus (DG) led to a more pronounced anxiety-related behavior in female than in male mice as indicated by the behavioral evaluations. Meanwhile, overexpression of PPM1F in the DG decreased total Bdnf exon–specific messenger RNA expression in the hippocampus with the decreased binding activity of phosphorylated H3S10 to its individual promoters in female mice. Furthermore, we identified that overexpression of PPM1F decreased the phosphorylation levels of AKT and JNK in the hippocampus of female mice. These results may suggest that PPM1F regulates anxiety-related behaviors by modulating BDNF expression and H3S10 phosphorylation–mediated epigenetic modification, which may be served as potentially pathological genes associated with anxiety or other mental diseases.

A spontaneous late-ripening mutant of 'Jincheng' (C. sinensis L. Osbeck) sweet orange exhibited a delay of fruit pigmentation and harvesting. In this work, we studied the processes of orange fruit ripening through the comparative analysis between the Jincheng mutant and its wild type. This study revealed that the fruit quality began to differ on 166th days after anthesis. At this stage, fruits were subjected to transcriptome analysis by RNA sequencing. 13,412 differentially expressed unigenes (DEGs) were found. Of these unigenes, 75.8% were down-regulated in the wild type, suggesting that the transcription level of wild type was lower than that of the mutant during this stage. These DEGs were mainly clustered into five pathways: metabolic pathways, plant-pathogen interaction, spliceosome, biosynthesis of plant hormones and biosynthesis of phenylpropanoids. Therefore, the expression profiles of the genes that are involved in abscisic acid, sucrose, and jasmonic acid metabolism and signal transduction pathways were analyzed during the six fruit ripening stages. The results revealed the regulation mechanism of sweet orange fruit ripening metabolism in the following four aspects: First, the more mature orange fruits were, the lower the transcription levels were. Second, the expression level of PME boosted with the maturity of the citrus fruit. Therefore, the expression level of PME might represent the degree of the orange fruit ripeness. Third, the interaction of PP2C, PYR/PYL, and SnRK2 was peculiar to the orange fruit ripening process. Fourth, abscisic acid, sucrose, and jasmonic acid all took part in orange fruit ripening process and might interact with each other. These findings provide an insight into the intricate process of sweet orange fruit ripening.

For developmental processes, we know most of the gene networks controlling specific cell responses. We still have to determine how these networks cooperate and how signals become integrated. The JNK pathway is one of the key elements modulating cellular responses during development. Yet, we still know little about how the core components of the pathway interact with additional regulators or how this network modulates cellular responses in the whole organism in homeostasis or during tissue morphogenesis. We have performed a promoter analysis, searching for potential regulatory sequences of puckered (puc) and identified different specific enhancers directing gene expression in different tissues and at different developmental times. Remarkably, some of these domains respond to the JNK activity, but not all. Altogether, these analyses show that puc expression regulation is very complex and that JNK activities participate in non-previously known processes during the development of Drosophila.

Phosphoglycerate mutase 5 (PGAM5) is a mitochondrial membrane protein that plays crucial roles in necroptosis and apoptosis. Though PGAM5 is known to be required for inducing intrinsic apoptosis through interacting with BCL2 associated X protein (Bax) and dynamin-related protein 1 (Drp1), the expression and role of PGAM5 in cardiomyocyte apoptosis driven by myocardial ischemia/reperfusion injury(MIRI) has not been studied. The present study shows that PGAM5 expression decreased after MIRI in vivo, positively correlated with Bcl-xL expression, negatively correlated with Kelch-ECH associating protein 1 (Keap1) expression. Furthermore, PGAM5 expression also decreased in cardiomyocytes after hypoxia/reoxygenation (H/R) treatment in vitro. PGAM5 silence promoted cardiomyocyte apoptosis and inhibited Bcl-xL expression, but with no effect on Keap1 expression. Accordingly, Keap1 overexpression further inhibited Bcl-xL and PGAM5 expression. Additionally, PGAM5-Bcl-xL-Keap1 interaction was identified, suggesting that PGAM5 might participate in the degradation of Bcl-xL mediated by Keap1. In summary, PGAM5 controls cardiomyocyte apoptosis induced by MIRI through regulating Keap1-mediated Bcl-xL degradation, which may supply a novel molecular target for acute myocardial infarction (AMI) therapy.

The tetronate ring appears in several natural products, but the biosynthetic path to this structure has proven elusive. Reconstitution of a polyketide assembly line and in vitro assays with a chemically synthesized intermediate now point to a single enzyme as catalyzing ring formation. The protein phosphatase inhibitor RK-682 is one of a number of potentially valuable tetronate polyketide natural products. Understanding how the tetronate ring is formed has been frustrated by the inaccessibility of the putative substrates. We report the heterologous expression of rk genes in Saccharopolyspora erythraea and reconstitution of the RK-682 pathway using recombinant enzymes, and we show that RkD is the enzyme required for RK-682 formation from acyl carrier protein–bound substrates.

Due to its ability to be rapidly generated and propagated over long distances, H2O2 is an important second messenger for biotic and abiotic stress signaling in plants. In response to low water potential and high salt concentrations sensed in the roots of plants, the stress hormone abscisic acid (ABA) activates NADPH oxidase to generate H2O2, which is propagated in guard cells in leaves to induce stomatal closure and prevent water loss from transpiration. Using a reconstituted system, we demonstrate that H2O2 reversibly prevents the protein phosphatase HAB1, a key component of the core ABA-signaling pathway, from inhibiting its main target in guard cells, SnRK2.6/OST1 kinase. We have identified HAB1 C186 and C274 as H2O2-sensitive thiols and demonstrate that their oxidation inhibits both HAB1 catalytic activity and its ability to physically associate with SnRK2.6 by formation of intermolecular dimers.

The objective of this study was to investigate the effect of silencing gene protein phosphatase 1H (PPM1H) on malignant phenotype of human pancreatic cancer cell line BxPC-3. In order to explore the function of PPM1H in pancreatic cancer cells, real-time PCR and western blotting were used to detect the expression of PPM1H in different pancreatic cancer cell lines. Human pancreatic cancer cell line BxPC-3 was treated with 10 ng/ml TGF-β1 and 200 ng/ml BMP2 for 72 h, respectively, and the mRNA and protein expression levels of PPM1H and EMT-related markers (E-cadherin, vimentin) were detected by real-time PCR and western blotting, respectively. Using exogenous RNA interference technology to silence the PPM1H gene, the expression of PPM1H and EMT-related markers at mRNA and protein levels were detected by real-time PCR and western blotting. The cell migration and invasion were measured using Transwell assays. Finally, cell counting kit-8 (CCK-8) and flow cytometry were used to determine the effect of PPM1H on cell proliferation and apoptosis of BxPC-3 cells. The expression levels of PPM1H in all of the examined pancreatic cancer cell lines (BxPC-3, MIA-PACA2, PANC-1, SW1990, PANC-03.27) were lower than that of normal pancreatic ductal epithelial cells (HPDE6-C7) at both mRNA and protein levels. Both TGF-β1 and BMP2 treatment induced EMT and downregulation of PPM1H in BxPC-3 cells. By using RNA interference to transiently knock down PPM1H expression in BxPC-3 cells, we found that the expression of E-cadherin was downregulated while vimentin was up regulated. The data suggested that silencing PPM1H gene can induce EMT in BxPC-3 cells. In addition, Transwell migration assays showed that silencing PPM1H gene can promote the invasion and metastasis of BxPC-3 cells. Cell proliferation and apotosis detection demonstrated that silencing PPM1H gene can promote the proliferation and inhibit apoptosis of BxPC-3 cells. In conclusion, PPM1H is aberrantly expressed in human pancreatic cancer cell lines and can be downregulated when EMT is induced by cytokine stimulation. Silencing PPM1H gene can induce EMT in BxPC-3 cells, and promote the invasion and metastasis of BxPC-3 cells. Moreover, silencing PPM1H gene can promote the proliferation and inhibit apoptosis of BxPC-3 cells. PPM1H may be a new tumor-suppressor factor for pancreatic cancer and provides new insight into molecular targets for gene therapy of pancreatic cancer.

Decreased IL-2 production in systemic lupus erythematosus (SLE) represents a central component of the disease immunopathology. We report that the message, protein, and enzymatic activity of the catalytic subunit of protein phosphatase 2A (PP2Ac), but not PP1, are increased in patients with SLE regardless of disease activity and treatment and in a disease-specific manner. Treatment of SLE T cells with PP2Ac-siRNA decreased the protein levels and activity of PP2Ac in a specific manner and increased the levels of phosphorylated cAMP response element-binding protein and its binding to the IL2 and c-fos promoters, as well as increased activator protein 1 activity, causing normalization of IL-2 production. Our data document increased activity of PP2A as a novel SLE disease-specific abnormality and define a distinct mechanism whereby it represses IL-2 production. We propose the use of PP2Ac-siRNA as a novel tool to correct T cell IL-2 production in SLE patients.