Endothelial PHACTR1 Promotes Endothelial Activation and Atherosclerosis by Repressing PPARγ Activity Under Disturbed Flow in Mice.

BACKGROUND: Numerous genome-wide association studies revealed that SNPs (single nucleotide polymorphisms) at the PHACTR1 (phosphatase and actin regulator 1) locus strongly correlate with coronary artery disease. However, the biological function of PHACTR1 remains poorly understood. Here, we identified the proatherosclerotic effect of endothelial PHACTR1, contrary to macrophage PHACTR1. METHODS: We generated global (Phactr1-/-) and endothelial cell (EC)-specific (Phactr1ECKO) Phactr1 KO (knockout) mice and crossed these mice with apolipoprotein E-deficient (Apoe-/-) mice. Atherosclerosis was induced by feeding the high-fat/high-cholesterol diet for 12 weeks or partially ligating carotid arteries combined with a 2-week high-fat/high-cholesterol diet. PHACTR1 localization was identified by immunostaining of overexpressed PHACTR1 in human umbilical vein ECs exposed to different types of flow. The molecular function of endothelial PHACTR1 was explored by RNA sequencing using EC-enriched mRNA from global or EC-specific Phactr1 KO mice. Endothelial activation was evaluated in human umbilical vein ECs transfected with siRNA targeting PHACTR1 and in Phactr1ECKO mice after partial carotid ligation. RESULTS: Global or EC-specific Phactr1 deficiency significantly inhibited atherosclerosis in regions of disturbed flow. PHACTR1 was enriched in ECs and located in the nucleus of disturbed flow areas but shuttled to cytoplasm under laminar flow in vitro. RNA sequencing showed that endothelial Phactr1 depletion affected vascular function, and PPARγ (peroxisome proliferator-activated receptor gamma) was the top transcription factor regulating differentially expressed genes. PHACTR1 functioned as a PPARγ transcriptional corepressor by binding to PPARγ through the corepressor motifs. PPARγ activation protects against atherosclerosis by inhibiting endothelial activation. Consistently, PHACTR1 deficiency remarkably reduced endothelial activation induced by disturbed flow in vivo and in vitro. PPARγ antagonist GW9662 abolished the protective effects of Phactr1 KO on EC activation and atherosclerosis in vivo. CONCLUSIONS: Our results identified endothelial PHACTR1 as a novel PPARγ corepressor to promote atherosclerosis in disturbed flow regions. Endothelial PHACTR1 is a potential therapeutic target for atherosclerosis treatment.

Knockdown of Sly-miR164a Enhanced Plant Salt Tolerance and Improved Preharvest and Postharvest Fruit Nutrition of Tomato.

Salinity stress is a serious limitation to tomato growth and development. The aim of this study was to investigate the effects of Sly-miR164a on tomato growth and fruit nutritional quality under salt stress. The results showed that the root length, fresh weight, plant height, stem diameter and ABA content of miR164a#STTM (knockdown of Sly-miR164a) lines were higher than those of WT and miR164a#OE (overexpression of Sly-miR164a) lines under salt stress. Compared with WT, miR164a#STTM tomato lines exhibited lower ROS accumulation under salt stress. In addition, the fruits of miR164a#STTM tomato lines had higher soluble solids, lycopene, ascorbic acid (ASA) and carotenoid content compared with WT. The study indicated that tomato plants were more sensitive to salt when Sly-miR164a was overexpressed, while knockdown of Sly-miR164a enhanced plant salt tolerance and improved fruit nutritional value.

Vinpocetine Attenuates Pathological Cardiac Remodeling by Inhibiting Cardiac Hypertrophy and Fibrosis.

PURPOSE: Pathological cardiac remodeling, characterized by cardiac hypertrophy and fibrosis, is a pathological feature of many cardiac disorders that leads to heart failure and cardiac arrest. Vinpocetine, a derivative of the alkaloid vincamine, has been used for enhancing cerebral blood flow to treat cognitive impairment. However, its role in pathological cardiac remodeling remains unknown. The aim of this study is to examine the effect of vinpocetine on pathological cardiac remodeling induced by chronic stimulation with angiotensin II (Ang II). METHODS: Mice received Ang II infusion via osmotic pumps in the presence of vehicle or vinpocetine. Cardiac hypertrophy and fibrosis were assessed by morphological, histological, and biochemical analyses. Mechanistic studies were carried out in vitro with isolated mouse adult cardiac myocytes and fibroblasts. RESULTS: We showed that chronic Ang II infusion caused cardiac hypertrophy and fibrosis, which were all significantly attenuated by systemic administration of vinpocetine. In isolated adult mouse cardiomyocytes, vinpocetine suppressed Ang II-stimulated myocyte hypertrophic growth. In cultured cardiac fibroblasts, vinpocetine suppressed TGFß-induced fibroblast activation and matrix gene expression, consistent with its effect in attenuating cardiac fibrosis. The effects of vinpocetine on cardiac myocyte hypertrophy and fibroblast activation are likely mediated by targeting cyclic nucleotide phosphodiesterase 1 (PDE1). CONCLUSIONS: Our results reveal a novel protective effect of vinpocetine in attenuating pathological cardiac remodeling through suppressing cardiac myocyte hypertrophic growth and fibroblast activation and fibrotic gene expression. These studies may also shed light on developing novel therapeutic agents for antagonizing pathological cardiac remodeling.

G-protein-coupled receptor kinase interacting protein-1 mediates intima formation by regulating vascular smooth muscle proliferation, apoptosis, and migration.

OBJECTIVE: The G-protein-coupled receptor kinase interacting protein-1 (GIT1) is a scaffold protein that is important for phospholipase Cγ and extracellular signal-regulated kinase 1/2 signaling induced by angiotensin II and epidermal growth factor. Because GIT1 regulates signaling by several vascular smooth muscle cell (VSMC) growth factors, we hypothesized that intima formation would be inhibited by GIT1 depletion. APPROACH AND RESULTS: Complete carotid ligation was performed on GIT1 wild-type and knockout (KO) mice. We compared changes between GIT1 wild-type and KO mice in carotid vascular remodeling, VSMC proliferation, and apoptosis in vivo and in vitro. Our data demonstrated that GIT1 deficiency significantly decreased intima formation after carotid ligation as a result of both reduced VSMC proliferation and enhanced apoptosis. To confirm the effects of GIT1 in vitro, we performed proliferation and apoptosis assays in VSMC. In mouse aortic smooth muscle cells (MASM), we found that the growth rate and [3H]-thymidine incorporation of the GIT1 KO MASM were significantly decreased compared with the wild-type MASM. Cyclin D1, which is a key cell cycle regulator, was significantly decreased in GIT1 KO cells. Serum deprivation of GIT1 KO MASM increased apoptosis 3-fold compared with wild-type MASM. Treatment of rat aortic smooth muscle cells with GIT1 small interfering RNA impaired cell migration. Both phospholipase Cγ and extracellular signal-regulated kinase 1/2 signaling were required for GIT1-dependent VSMC proliferation and migration, whereas only phospholipase Cγ was involved in GIT1-mediated VSMC apoptosis. CONCLUSIONS: GIT1 is a novel mediator of vascular remodeling by regulating VSMC proliferation, migration, and apoptosis through phospholipase Cγ and extracellular signal-regulated kinase 1/2 signaling pathways.

EVI1 acts as an inducible negative-feedback regulator of NF-κB by inhibiting p65 acetylation.

Inflammation is a hallmark of many important human diseases. Appropriate inflammation is critical for host defense; however, an overactive response is detrimental to the host. Thus, inflammation must be tightly regulated. The molecular mechanisms underlying the tight regulation of inflammation remain largely unknown. Ecotropic viral integration site 1 (EVI1), a proto-oncogene and zinc finger transcription factor, plays important roles in normal development and leukemogenesis. However, its role in regulating NF-κB-dependent inflammation remains unknown. In this article, we show that EVI1 negatively regulates nontypeable Haemophilus influenzae- and TNF-α-induced NF-κB-dependent inflammation in vitro and in vivo. EVI1 directly binds to the NF-κB p65 subunit and inhibits its acetylation at lysine 310, thereby inhibiting its DNA-binding activity. Moreover, expression of EVI1 itself is induced by nontypeable Haemophilus influenzae and TNF-α in an NF-κB-dependent manner, thereby unveiling a novel inducible negative feedback loop to tightly control NF-κB-dependent inflammation. Thus, our study provides important insights into the novel role for EVI1 in negatively regulating NF-κB-dependent inflammation, and it may also shed light on the future development of novel anti-inflammatory strategies.

Vinpocetine inhibits NF-kappaB-dependent inflammation via an IKK-dependent but PDE-independent mechanism.

Inflammation is a hallmark of many diseases, such as atherosclerosis, chronic obstructive pulmonary disease, arthritis, infectious diseases, and cancer. Although steroids and cyclooxygenase inhibitors are effective antiinflammatory therapeutical agents, they may cause serious side effects. Therefore, developing unique antiinflammatory agents without significant adverse effects is urgently needed. Vinpocetine, a derivative of the alkaloid vincamine, has long been used for cerebrovascular disorders and cognitive impairment. Its role in inhibiting inflammation, however, remains unexplored. Here, we show that vinpocetine acts as an antiinflammatory agent in vitro and in vivo. In particular, vinpocetine inhibits TNF-alpha-induced NF-kappaB activation and the subsequent induction of proinflammatory mediators in multiple cell types, including vascular smooth muscle cells, endothelial cells, macrophages, and epithelial cells. We also show that vinpocetine inhibits monocyte adhesion and chemotaxis, which are critical processes during inflammation. Moreover, vinpocetine potently inhibits TNF-alpha- or LPS-induced up-regulation of proinflammatory mediators, including TNF-alpha, IL-1beta, and macrophage inflammatory protein-2, and decreases interstitial infiltration of polymorphonuclear leukocytes in a mouse model of TNF-alpha- or LPS-induced lung inflammation. Interestingly, vinpocetine inhibits NF-kappaB-dependent inflammatory responses by directly targeting IKK, independent of its well-known inhibitory effects on phosphodiesterase and Ca(2+) regulation. These studies thus identify vinpocetine as a unique antiinflammatory agent that may be repositioned for the treatment of many inflammatory diseases.

Investigations of Single-Subunit tRNA Methyltransferases from Yeast.

tRNA methylations, including base modification and 2'-O-methylation of ribose moiety, play critical roles in the structural stabilization of tRNAs and the fidelity and efficiency of protein translation. These modifications are catalyzed by tRNA methyltransferases (TRMs). Some of the TRMs from yeast can fully function only by a single subunit. In this study, after performing the primary bioinformatic analyses, the progress of the studies of yeast single-subunit TRMs, as well as the studies of their homologues from yeast and other types of eukaryotes and the corresponding TRMs from other types of organisms was systematically reviewed, which will facilitate the understanding of the evolutionary origin of functional diversity of eukaryotic single-subunit TRM.

The effect of BVOCs produced by Lysinibacillus fusiformis and LED irradiation on pigment metabolism in stored broccoli.

Volatile organic compounds produced by bacteria (BVOCs) have been proven to effect the postharvest metabolism of fruits and vegetables. The quality, color and antioxidant capacity of membrane lipids of broccoli in storage were effectively maintained by fumigation with BVOCs produced by Lysinibacillus fusiformis combined with white light emitting diode (LED) technology. An analysis of the transcriptome and metabolome of broccoli treated with the combined LED-BVOCs technology resulted in the identification of 49 differentially expressed genes (DEGs) and 13 differentially abundant metabolites (DAMs) involved in photosynthesis (32/0 DEGs upregulated/downregulated; 0/0 DAMs with increased/decreased abundance), chlorophyll (7/0; 1/2), carotenoid (5/0; 1/4) and flavonoid (3/3; 3/2) metabolism. The maintenance of green color in harvested broccoli treated by LED-BVOCs was associated with DEGs and DAMs that inhibited chlorophyll degradation and carotenoid accumulation. Our study provides a theoretical basis for understanding the delayed senescence of broccoli during storage using BVOCs-LED technology.

Effects of spray drying and freeze drying on the structure and emulsifying properties of yam soluble protein: A study by experiment and molecular dynamics simulation.

This study focused on the effects of freeze drying (FD) and sprays drying (SD) on the structure and emulsifying properties of yam soluble protein (YSP). The results showed that the surface hydrophobicity (Ho) value, free sulfhydryl group (SH) content, turns content, denaturation temperature and enthalpy value of spray-dried YSP (SD-YSP) were higher than freeze-dried YSP (FD-YSP), but the apparent hydrodynamic diameter (Dh) value of SD-YSP was smaller. The smaller Dh, higher Ho and free SH led to higher percentage of adsorbed proteins and stronger binding between protein and oil droplet in emulsions. Thus, the emulsifying properties of SD-YSP were better, and the SD-YSP-stabilized emulsion had better dynamical rheological properties. Molecular dynamics (MD) simulations suggested that some intramolecular disulfide bonds and hydrogen bonds of dioscorin were broken, and some helices transformed into turns during the SD process. These structural changes resulted in better thermal stability and emulsification properties of SD-YSP.

Revealing the mechanism underlying the effects of γ-aminobutyric acid-dioscorin interactions on dioscorin structure and emulsifying properties by molecular dynamic simulations.

Many studies have shown that γ-aminobutyric acid (GABA) exhibits various beneficial biological activities, including gut-modulating, neuro-stimulating, and cardio-protecting activities. Naturally, GABA exists in small amounts in yam, which is primarily synthesized by the decarboxylation of L-glutamic acid in the presence of glutamate decarboxylase. Dioscorin, the major tuber storage protein of yam, has been shown to have good solubility and emulsifying activity. However, how GABA interacts with dioscorin and affects their properties has yet to be clarified. In this research, the physicochemical and emulsifying properties of GABA-fortified dioscorin, which was dried by spray drying and freeze drying, were studied. As results, the freeze-dried (FD) dioscorin produced more stable emulsions, while the spray-dried (SD) dioscorin adsorbed more rapidly to oil/water (O/W) interface. The fluorescence spectroscopy, ultraviolet spectroscopy and circular dichroism spectroscopy showed that GABA changed the structure of dioscorin, by exposing its hydrophobic groups. The addition of GABA significantly promoted the adsorption of dioscorin to the O/W interface and prevented droplets coalescence. The results of molecular dynamics simulation (MD) showed that GABA destroyed the H-bond network between dioscorin and water, increased surface hydrophobicity and finally improved the emulsifying properties of dioscorin.

Chronic administration of hexarelin attenuates cardiac fibrosis in the spontaneously hypertensive rat.

Cardiac fibrosis is a hallmark of heart disease and plays a vital role in cardiac remodeling during heart diseases, including hypertensive heart disease. Hexarelin is one of a series of synthetic growth hormone secretagogues (GHSs) possessing a variety of cardiovascular effects via action on GHS receptors (GHS-Rs). However, the role of hexarelin in cardiac fibrosis in vivo has not yet been investigated. In the present study, spontaneously hypertensive rats (SHRs) were treated with hexarelin alone or in combination with a GHS-R antagonist for 5 wk from an age of 16 wk. Hexarelin treatment significantly reduced cardiac fibrosis in SHRs by decreasing interstitial and perivascular myocardial collagen deposition and myocardial hydroxyproline content and reducing mRNA and protein expression of collagen I and III in SHR hearts. Hexarelin treatment also increased matrix metalloproteinase (MMP)-2 and MMP-9 activities and decreased myocardial mRNA expression of tissue inhibitor of metalloproteinase (TIMP)-1 in SHRs. In addition, hexarelin treatment significantly attenuated left ventricular (LV) hypertrophy, LV diastolic dysfunction, and high blood pressure in SHRs. The effect of hexarelin on cardiac fibrosis, blood pressure, and cardiac function was mediated by its receptor, GHS-R, since a selective GHS-R antagonist abolished these effects and expression of GHS-Rs was upregulated by hexarelin treatment. In summary, our data demonstrate that hexarelin reduces cardiac fibrosis in SHRs, perhaps by decreasing collagen synthesis and accelerating collagen degradation via regulation of MMPs/TIMP. Hexarelin-reduced systolic blood pressure may also contribute to this reduced cardiac fibrosis in SHRs. The present findings provided novel insights and underscore the therapeutic potential of hexarelin as an antifibrotic agent for the treatment of cardiac fibrosis.

Chilling injury of tomato fruit was alleviated under low-temperature storage by silencing Sly-miR171e with short tandem target mimic technology.

In this study, the role of Sly-miR171e on post-harvest cold tolerance of tomato fruit was researched. The results showed that overexpression of Sly-miR171e (miR171e-OE) promoted postharvest chilling injury (CI) of tomato fruit at the mature red (MR) and mature green (MG) stage. Contrasted with the wild type (WT) and miR171e-OE fruit, the knockdown of Sly-miR171e (miR171e-STTM) showed a lower CI index, lower hydrogen peroxide (H2O2) content, and higher fruit firmness after harvest. In the fruit of miR171e-STTM, the expression level of GRAS24, CBF1, GA2ox1, and COR, and the GA3 content were ascended, while the expression levels of GA20ox1 and GA3ox1 were descended. The research demonstrated that CI in tomato fruit was alleviated at low temperature storage by silencing Sly-miR171e with short tandem target mimic (STTM) technology. Furthermore, it also provided helpful information for genetic modification of miR171e and control of CI in the postharvest fruit.

Melatonin maintained higher contents of unsaturated fatty acid and cell membrane structure integrity in banana peel and alleviated postharvest chilling injury.

Exogenous melatonin confers the chilling tolerance of banana fruit by promoting reactive oxygen species (ROS) scavenging system and inducing the unsaturated fatty acid synthesis. The results showed that melatonin treatment increased the contents of phospholipids, promoted the ROS scavenging enzyme, and restrained the activities of lipoxygenase (LOX), and thus reduced the lipid peroxidation of banana peel. In addition, melatonin treatment increased the flavonoids and proline contents, which was conducive to antioxidant capacity. Interestingly, the enhanced antioxidant capacity is conducive to the stability of unsaturated fatty acids and reduce the enzymatic browning reaction. Moreover, melatonin treatment induced the expression of omega-3/6 fatty acid desaturase and triggered the fatty acid metabolism activity, by which maintained higher contents of unsaturated fatty acid in banana peel. Moreover, melatonin treatment stimulated the accumulation of fatty acids in banana peel, and was involved in alleviating fruit chilling injury.

Silencing of Sly-miR171d increased the expression of GRAS24 and enhanced postharvest chilling tolerance of tomato fruit.

The role of Sly-miR171d on tomato fruit chilling injury (CI) was investigated. The results showed that silencing the endogenous Sly-miR171d effectively delayed the increase of CI and electrolyte leakage (EL) in tomato fruit, and maintained fruit firmness and quality. After low temperature storage, the expression of target gene GRAS24 increased in STTM-miR171d tomato fruit, the level of GA3 anabolism and the expression of CBF1, an important regulator of cold resistance, both increased in STTM-miR171d tomato fruit, indicated that silencing the Sly-miR171d can improve the resistance ability of postharvest tomato fruit to chilling tolerance.

Physiological and Transcriptome Analyses of CaCl2 Treatment to Alleviate Chilling Injury in Pineapple.

The post-harvest ripening of pineapples can be effectively postponed by refrigerated storage. Nevertheless, internal browning (IB) frequently appears in pineapples after refrigerated storage during the course of the shelf life at room temperature, which is known as chilling injury (CI). In this study, the chilling injury of pineapple fruit was induced by a low temperature (6 °C) and transferred to normal-temperature storage; the best concentration of 50 µmol/L of CaCl2 was selected by the IB appearance and electrical conductivity. Fruit quality, reactive oxygen species (ROS), antioxidants, and transcription factors were investigated. The physiological data results indicated that pineapples treated with 50 µmol/L of CaCl2 maintained fruit quality, decreased reactive oxygen species (ROS), and enhanced the antioxidant activity of fruits, alleviating internal browning (IB) symptoms in pineapple fruit. The expressions of related genes were also consistent with the physiological changes by the transcriptome data analysis. In addition, we focused on some related metabolic pathways, including phenylpropanoid biosynthesis, MAPK pathway, plant hormone, plant-pathogen interaction, tricarboxylic acid cycle (TAC), and fatty acid biosynthesis. We performed integrative analyses of transcriptome data combined with a series of physiology and experimental analyses on the internal browning of pineapples, which will be of great significance to extending the shelf life of pineapples through molecular biology in the future.

SlBEL11 affects tomato carotenoid accumulation by regulating SlLCY-b2.

Extensive data have demonstrated that carotenoid accumulation in tomato fruit is influenced by environmental cues and hormonal signals. However, there is insufficient information on the mechanism of its transcriptional regulation, as many molecular roles of carotenoid biosynthetic pathways remain unknown. In this work, we found that the silence of the BEL1-like family transcription factor (TF) BEL1-LIKE HOMEODOMAIN 11 (SlBEL11) enhanced carotenoid accumulation in virus induced gene silencing (VIGS) analysis. In its RNA interference (RNAi) transgenic lines, a significant increase in the transcription level for the lycopene beta cyclase 2 (SlLCY-b2) gene was detected, which encoded a key enzyme located at the downstream branch of the carotenoid biosynthetic pathway. In Electrophoretic mobility shift assay (EMSA), SlBEL11 protein was confirmed to bind to the promoter of SlLCY-b2 gene. In addition, the dual-luciferase reporter assay showed its intrinsic transcriptional repression activity. Collectively, our findings added a new member to the carotenoid transcriptional regulatory network and expanded the functions of the SlBEL11 transcription factor.

Discovery of SHR5133, a Highly Potent and Novel HBV Capsid Assembly Modulator.

Capsid assembly modulators (CpAMs) represent a new class of antivirals targeting hepatitis B virus (HBV) core protein to disrupt the assembly process. In this work, a novel chemotype featuring a fused heterocycle amide was discovered through pharmacophore exploration. Lead optimization resulted in compound 8 with an EC50 value of 511 nM, and then methyl substitution on the piperazine was found to improve the in vitro potency remarkably. Further SAR studies established the key compound SHR5133, which showed high in vitro antiviral potency, favorable pharmacokinetic profiles across species, and robust in vivo efficacy.

G protein coupled receptor kinase 2 interacting protein 1 (GIT1) is a novel regulator of mitochondrial biogenesis in heart.

G-protein-coupled receptor (GPCR)-kinase interacting protein-1 (GIT1) is a multi-function scaffold protein. However, little is known about its physiological role in the heart. Here we sought to identify the cardiac function of GIT1. Global GIT1 knockout (KO) mice were generated and exhibited significant cardiac hypertrophy that progressed to heart failure. Electron microscopy revealed that the hearts of GIT1 KO mice demonstrated significant morphological abnormities in mitochondria, including decreased mitochondrial volume density, cristae density and increased vacuoles. Moreover, mitochondrial biogenesis-related gene peroxisome proliferator-activated receptor γ (PPARγ) co-activator-1α (PGC-1α), PGC-1ß, mitochondrial transcription factor A (Tfam) expression, and total mitochondrial DNA were remarkably decreased in hearts of GIT1 KO mice. These animals also had impaired mitochondrial function, as evidenced by reduced ATP production and dissipated mitochondrial membrane potential (Ψ(m)) in adult cardiomyocytes. Concordant with these mitochondrial observations, GIT1 KO mice showed enhanced cardiomyocyte apoptosis and cardiac dysfunction. In conclusion, our findings identify GIT1 as a new regulator of mitochondrial biogenesis and function, which is necessary for postnatal cardiac maturation.

Role of Ca2+/calmodulin-stimulated cyclic nucleotide phosphodiesterase 1 in mediating cardiomyocyte hypertrophy.

RATIONALE: Cyclic nucleotide phosphodiesterases (PDEs) through the degradation of cGMP play critical roles in maintaining cardiomyocyte homeostasis. Ca(2+)/calmodulin (CaM)-activated cGMP-hydrolyzing PDE1 family may play a pivotal role in balancing intracellular Ca(2+)/CaM and cGMP signaling; however, its function in cardiomyocytes is unknown. OBJECTIVE: Herein, we investigate the role of Ca(2+)/CaM-stimulated PDE1 in regulating pathological cardiomyocyte hypertrophy in neonatal and adult rat ventricular myocytes and in the heart in vivo. METHODS AND RESULTS: Inhibition of PDE1 activity using a PDE1-selective inhibitor, IC86340, or downregulation of PDE1A using siRNA prevented phenylephrine induced pathological myocyte hypertrophy and hypertrophic marker expression in neonatal and adult rat ventricular myocytes. Importantly, administration of the PDE1 inhibitor IC86340 attenuated cardiac hypertrophy induced by chronic isoproterenol infusion in vivo. Both PDE1A and PDE1C mRNA and protein were detected in human hearts; however, PDE1A expression was conserved in rodent hearts. Moreover, PDE1A expression was significantly upregulated in vivo in the heart and myocytes from various pathological hypertrophy animal models and in vitro in isolated neonatal and adult rat ventricular myocytes treated with neurohumoral stimuli such as angiotensin II (Ang II) and isoproterenol. Furthermore, PDE1A plays a critical role in phenylephrine-induced reduction of intracellular cGMP- and cGMP-dependent protein kinase (PKG) activity and thereby cardiomyocyte hypertrophy in vitro. CONCLUSIONS: These results elucidate a novel role for Ca(2+)/CaM-stimulated PDE1, particularly PDE1A, in regulating pathological cardiomyocyte hypertrophy via a cGMP/PKG-dependent mechanism, thereby demonstrating Ca(2+) and cGMP signaling cross-talk during cardiac hypertrophy.

Nitrate reductase-mediated nitric oxide generation is essential for fungal elicitor-induced camptothecin accumulation of Camptotheca acuminata suspension cell cultures.

Secondary metabolite accumulation and nitric oxide (NO) generation are two common responses of plant cells to fungal elicitors, and NO has been reported to play important roles in elicitor-induced secondary metabolite production. However, the source of elicitor-triggered NO generation in plant cells remains largely unknown. To investigate the origin of elicitor-triggered NO, we examined nitrate reductase (NR) activities and the expression levels of NIA1 and NIA2 genes of Camptotheca acuminata cells treated with PB90, a protein elicitor from Phytophthora boehmeriae. The data show that PB90 treatment stimulates NR activity and induces upregulation of NIA1 but does not affect NIA2 expression in the cells. Pretreatment of the cells with NR inhibitors tungstate and Gln abolishes not only the fungal elicitor-triggered NR activities but also the PB90-induced NO generation. Treatment of PB90 enhances camptothecin contents of the cells, suggesting that the fungal elicitor might stimulate camptothecin biosynthesis. Furthermore, application of tungstate and Gln suppresses the fungal elicitor-induced camptothecin accumulation of the cells and the suppression of NR inhibitors on PB90-induced camptothecin production can be reversed by NO via its donor sodium nitroprusside. Together, the results suggest that NIA1 is sensitive to PB90 and the fungal elicitor-induced upregulation of NIA1 may lead to higher NR activity. Furthermore, our data demonstrate that NR is involved in the fungal elicitor-triggered NO generation and the fungal elicitor induces camptothecin production of C. acuminata cells dependently on NR-mediated NO generation.