Homeostasis-related Genes Research Articles

Expression level of Na+ homeostasis-related genes and salt-tolerant abilities in backcross introgression lines of rice crop under salt stress at reproductive stage.

Salt stress in the rice field is one of the most common abiotic stresses, reducing crop productivity, especially at reproductive stage, which is very sensitive to salt stress. The aim of this investigation was to study mRNA-related Na+ uptake/translocation and Na+ enrichment in the cellular level, leading to physiological changes, growth characteristics, and yield attributes in FL530 [salt-tolerant genotype; carrying SKC1 (in relation to high-affinity potassium transporters controlling Na+ and K+ translocation) and qSt1b (linking to salt injury score) QTLs] and KDML105 (salt-sensitive cultivar; lacking both QTLs) parental lines and 221-48 (carrying SKC1 and qSt1b QTLs) derived from BILs (backcross introgression lines) at 50% flowering of rice, under 150-mM NaCl until harvesting process. The upregulation of OsHKT1;5 (mediating Na+ exclusion into xylem parenchyma cells) and OsNHX1 (Na+/H+ exchanger to secrete Na+ into vacuole) and downregulation of OsHKT2;1 and OsHKT2;2 (mediating Na+ restriction in the roots, leaf sheath and older leaves) in cvs. FL530 and 221-48 (+ SKC1; + qSt1b) under salt stress were observed. It restricted Na+ level in flag leaf, thereby preventing salt toxicity, as indicated by maintenance of photon yield of PSII (ΦPSII), net photosynthetic rate (Pn), transpiration rate (E) and overall growth performances. In contrast, Na+ enrichment in flag leaf of cv. KDML105 (-SKC1;-qSt1b) caused the reduction in ΦPSII by 30.5% over the control, leading to the reduction in Pn by 62.3%, in seed sterility by 88.2%, and yield loss by 85.1%. Moreover, the negative relationships between Na+ enrichment in flag leaf, physiological changes, and yield traits in rice crop grown under salt stress were demonstrated. Based on this investigation, rice genotype 221-48 was found to possess salt-tolerant traits at reproductive stage and thus could prove to be a potential candidate for future breeding programs.

Transcriptional analysis reveals sodium nitroprusside affects alfalfa in response to PEG-induced osmotic stress at germination stage.

Drought is one of the most common environmental factors that affect alfalfa germination and development. Nitric oxide (NO) could mediate stress tolerance in plants. The goal of this study was to determine exogenous NO donor-mediated drought adaption molecular mechanisms during the alfalfa germination stage. In this study, physiological and transcriptome analyses were performed on 7days of the growth period seedlings by sodium nitroprusside (SNP) and polyethylene glycol (PEG) treatment. The results showed that SNP supplementation alleviated malondialdehyde accumulation, increased levels of proline and soluble sugars, and enhanced antioxidant enzyme activity under osmotic stress conditions. RNA-Seq experiments identified 5828 genes exhibiting differential expression in seedlings treated with PEG, SNP, or SNP+PEG relative to seedlings treated with distilled water. Of these DEGs, 3235 were upregulated, and 2593 were downregulated relative to the controls. Fifteen DEGs were amplified by qRT-PCR to verify the changes in expression determined by RNA-Seq, revealing that PIF3, glnA, PLCG1, and RP-S11e exhibited enhanced expression under the SNP+PEG treatment. SNP was found to modulate redox homeostasis-related genes such as GSTs, SOD2, GPX, and RBOH, and triggered calcium signaling transduction. It also induced some key genes relating to the abscisic acid, ethylene, and auxin signaling transduction in response to PEG stress. Conversely, genes associated with secondary metabolite biosynthesis and the metabolism of starch and sucrose during osmotic stress were downregulated by SNP. These results provide new insights into SNP-mediated drought adaption mechanisms at transcriptome-wide in alfalfa and reveal key drought tolerance pathways in this species.

Transcriptional expression of Na+ homeostasis-related genes and physiological responses of rice seedlings under salt stress

Being a salt-sensitive crop, the growth, development and productivity of rice is severely affected by saline soil. In the present study, Na+ homeostasis, physiological and morphological responses in both the root and leaf tissues of rice (Oryza sativa) seedlings of backcross introgression lines of KDML105 (KD; −/−) × FL530 (+/+) with or without SKC1 and qSt1b QTLs grown under 200 mM NaCl were investigated. Expression levels of OsHKT1;5 in FL530 (+/+) were very low, causing to low Na+ in both the root and leaf tissues, whereas it was up-regulated in KD (−/−), relating to Na+ enrichment. Expression levels of OsHKT2;1 in rice lines, 221-44 (+/+) and 221-48 (+/+) were down-regulated, thus limiting Na+ xylem upload, and store Na+ in the root tissues. OsSOS1 expression level in the leaves of 221-44 (+/+) and 221-58 (+/−) was up-regulated in relation to Na+ efflux. Na+ secretion into vacuoles via OsNHX1 in the leaf tissues was evidently demonstrated in 221-48 (+/+) as a major defense mechanism adopted by the plants to maintain photosynthetic abilities and growth performances, when plants were subjected to salt stress. In addition, the high Fv/Fm ratio in the leaves of 221-58 (+/−) was associated with increased expression level of OsNHX3, in comparison to KD (−/−) seedlings grown under salt stress (diminution by 22.8% over control). Moreover, reduction on growth characters and physiological parameters, viz., shoot fresh weight, shoot height, number of leaves, leaf area, Pn, and accumulation of Na+ in the roots under salt stress were applied as effective indices; thereby classifying FL530 (+/+) and 221-48 (+/+) as salt-tolerant, and 221-44 (+/+), 221-58 (+/−), KD (−/−), 221-3 (+/−), and 221-54 (−/−) as salt-susceptible.

Neuronal deficiency of p38α-MAPK ameliorates symptoms and pathology of APP or Tau-transgenic Alzheimer's mouse models.

Alzheimer's disease (AD) is the leading cause of dementia with very limited therapeutic options. Amyloid β (Aβ) and phosphorylated Tau (p-Tau) are key pathogenic molecules in AD. P38α-MAPK is specifically activated in AD lesion sites. However, its effects on AD pathogenesis, especially on p-Tau-associated brain pathology, and the underlying molecular mechanisms remain unclear. We mated human APP-transgenic mice and human P301S Tau-transgenic mice with mapk14-floxed and neuron-specific Cre-knock-in mice. We observed that deletion of p38α-MAPK specifically in neurons improves the cognitive function of both 9-month-old APP and Tau-transgenic AD mice, which is associated with decreased Aβ and p-Tau load in the brain. We further used next-generation sequencing to analyze the gene transcription in brains of p38α-MAPK deficient and wild-type APP-transgenic mice, which indicated that deletion of p38α-MAPK regulates the transcription of calcium homeostasis-related genes, especially downregulates the expression of grin2a, a gene encoding NMDAR subunit NR2A. Cell culture experiments further verified that deletion of p38α-MAPK inhibits NMDA-triggered calcium influx and neuronal apoptosis. Our systemic studies of AD pathogenic mechanisms using both APP- and Tau-transgenic mice suggested that deletion of neuronal p38α-MAPK attenuates AD-associated brain pathology and protects neurons in AD pathogenesis. This study supports p38α-MAPK as a novel target for AD therapy.

Genome-Wide Dissection of the CRF Gene Family in Brassica napus Indicates that BnaCRF8s Specifically Regulate Root Architecture and Phosphate Homeostasis against Phosphate Fluctuation in Plants.

Phosphorus (P) is an essential macronutrient required for plant growth and development. The involvement of cytokinin response factors (CRFs) in phosphate (Pi) homeostasis and lateral root (LR) initiation in Arabidopsis has been revealed. However, little is known in oil crops. Here, we performed genome-wide dissection of the CRF family in Brassica napus to identify 44 members, which were evolutionally classified into 6 subgroups. Among them, four BnaCRF8 genes were strongly upregulated by P deprivation, and were selected to be further investigated. Time course qRT-PCR analyses showed that four BnaCRF8 genes were enhanced dramatically after 12 h of P stress. Analyses of the subcellular localization in tobacco leaves indicated that BnaA7.CRF8 and BnaC2.CRF8 were localized in the nucleus. The expression of BnaCRF8 genes had constant negative effects on primary root growth and LR initiation and growth, and it reduced Pi acquisition and plant growth in Arabidopsis. Moreover, the expression of Pi homeostasis-related genes was modulated in BnaA7.CRF8 overexpression plants. These results suggest that BnaCRF8 genes might negatively regulate root architecture and plant growth through transcriptional modification of Pi homeostasis-related components. Overall, this study suggests that upregulation of BnaCRF8 genes might be a smart adaptive strategy to cope with continuous Pi deficiency in the environment.

Evaluation of the Epigenetic Demethylation of NRF2, a Master Transcription Factor for Antioxidant Enzymes, in Colorectal Cancer

Epigenetic changes in CpG islands of the promoter regions of homeostasis-related genes, including nuclear factor erythroid 2-related factor 2 (NRF2), have been shown to hold a significant role in the development of colorectal cancer. Therefore, we aimed to examine the DNA demethylation pattern of the NRF2 promoter region in cancerous lesions from patients with colorectal cancer and the association of methylation status with clinicopathological features in the Iranian population. In this cross-sectional study, 114 colorectal tissue samples were collected. These samples included: 34 tumour tissue samples, 60 precancerous polyps, and 20 normal tissue samples. The promoter methylation status of the NRF2 gene was examined using methylation-specific PCR. Additionally, the relationship between the methylation status and the clinicopathological features was investigated. The frequency of NRF2 demethylation in the tumour samples was significantly higher compared to the polyp tissues (p= 0.003) and normal tissue (p= 0.009), indicating that cancerous colorectal tissues exhibit increased demethylation of the NRF2 promoter. After examining the demethylation status of tissue samples, the clinicopathological features were compared to the demethylation results. No significant association was found between NRF2 promoter demethylation and the clinicopathological features of patient samples. Our findings suggest that the epigenetic modifications leading to NRF2 demethylation found in colorectal tumour samples may contribute to cancer progression from precancerous polyps to cancerous lesions.

Adipose tissue gene expression of long non-coding RNAs; MALAT1, TUG1 in obesity: is it associated with metabolic profile and lipid homeostasis-related genes expression?

BackgroundRecent studies point toward the possible regulatory roles of two lncRNAs; metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) and taurine upregulated gene 1 (TUG1) in the pathogenesis of obesity-related disorders and regulation of lipogenesis and adipogenesis. In an attempt to understand the molecules involved in human obesity pathogenesis, we aimed to evaluate the expression of MALAT1 and TUG1 in visceral adipose tissues (VAT) and subcutaneous adipose tissues (SAT) of obese women, as compared to normal-weight women. The mRNA expression of possible target genes including peroxisome proliferator-activated receptor gamma (PPARγ), PPARγ coactivator-1 alpha (PGC1α), sterol regulatory element-binding protein-1c (SREBP-1c), fatty acid synthase (FAS), and acetyl-CoA carboxylase (ACC) which are involved in adipogenesis and lipogenesis were also examined.MethodsThis study was conducted on 20 obese [body mass index (BMI) ≥ 30 kg/m 2] female participants and 19 normal-weight (BMI < 25 kg/m 2) female participants. Real-time PCR was performed to investigate the mRNA expression of the above-mentioned genes in VAT and SAT from all participants.ResultsThe results showed lower mRNA levels of TUG1 in both the VAT and SAT of obese women, compared to normal-weight women. Furthermore, TUG1 expression in SAT positively correlated with BMI, waist circumference (WC), hip circumference, HOMA-IR, and insulin levels, eGFR value, creatinine levels, and hs-CRP in all participants independent of age and HOMA-IR. However, VAT mRNA expression of TUG1 had a positive correlation with obesity indices and HOMA-IR and insulin levels in the whole population. Moreover, SAT mRNA level of TUG1 was positively correlated with SAT gene expression of PGC1α, SREBP-1c, FAS, and ACC independent of age and HOMA-IR. Although mRNA expression of MALAT1 did not differ between two groups for any tissue, it was positively correlated with SAT mRNA levels of SREBP-1c, PPARγ, and their targets; FAS and ACC, as well as with VAT mRNA levels of PGC1α.ConclusionsIt seems likely that TUG1 with distinct expression pattern in VAT and SAT are involved in the regulation of lipogenic and adipogenic genes and obesity-related parameters. However, more studies are necessary to establish this concept.

Caloric restriction reverses left ventricular hypertrophy through the regulation of cardiac iron homeostasis in impaired leptin signaling mice

Leptin-deficient and leptin-resistant mice manifest obesity, insulin resistance, and left ventricular hypertrophy (LVH); however, LVH’s mechanisms are not fully understood. Cardiac iron dysregulation has been recently implicated in cardiomyopathy. Here we investigated the protective effects of caloric restriction on cardiac remodeling in impaired leptin signaling obese mice. RNA-seq analysis was performed to assess the differential gene expressions in the heart of wild-type and ob/ob mice. In particular, to investigate the roles of caloric restriction on iron homeostasis-related gene expressions, 10-week-old ob/ob and db/db mice were assigned to ad libitum or calorie-restricted diets for 12 weeks. Male ob/ob mice exhibited LVH, cardiac inflammation, and oxidative stress. Using RNA-seq analysis, we identified that an iron uptake-associated gene, transferrin receptor, was upregulated in obese ob/ob mice with LVH. Caloric restriction attenuated myocyte hypertrophy, cardiac inflammation, fibrosis, and oxidative stress in ob/ob and db/db mice. Furthermore, we found that caloric restriction reversed iron homeostasis-related lipocalin 2, divalent metal transporter 1, transferrin receptor, ferritin, ferroportin, and hepcidin expressions in the heart of ob/ob and db/db mice. These findings demonstrate that the cardioprotective effects of caloric restriction result from the cellular regulation of iron homeostasis, thereby decreasing oxidative stress, inflammation, and cardiac remodeling. We suggest that decreasing iron-mediated oxidative stress and inflammation offers new therapeutic approaches for obesity-induced cardiomyopathy.

Epigenomic regulation of OTU5 in Arabidopsis thaliana

Epigenetic regulation by DNA methylation and histone marks is crucial to plant development. In Arabidopsis, the otu5 mutant exhibited altered root phenotypes resembling those of phosphate-deficient plants. In low phosphate (Pi) conditions, altered H3K4 and H3K27 trimethylation were associated with the expression of Pi homeostasis-related genes. However, the genetic effect of OTU5 on the epigenomes was left unexplored. We assessed genome-wide DNA methylation, gene expression and histone modifications of roots from both Col-0 and otu5 mutants. We found that OTU5 altered DNA methylation profile with a context-specific effect through targeting local genomic regions. Our analysis showed that in otu5 the abundance of H3K4me3 was clearly associated with the changes of DNA methylation, leading to the transcriptional difference from wildtype. We concluded that OTU5 induced cross-talks among epigenomes that altogether impacted the regulation of approximately 7060 genes. Of which 186 genes associated with root development were likely to be epigenetically regulated.

Subchronic continuous inhalation exposure to zinc oxide nanoparticles induces pulmonary cell response in mice

ObjectivesWe used mice as an animal model to investigate the entry of ZnO nanoparticles from the ambient air into the lungs and other organs, subsequent changes in Zn levels and the impact on the transcription of Zn homeostasis-related genes in the lungs. MethodsThe mice were exposed to two concentrations of ZnO nanoparticles; lower (6.46 × 104 particles/cm3) and higher (1.93 × 106 particles/cm3), allowed to breathe the nanoparticles in the air for 12 weeks and subjected to necropsy. Characterization of the ZnO nanoparticles was done using transmission electron microscopy (TEM). Energy-dispersive X-ray (EDX) spectroscopy was used to quantify ZnO nanoparticles in the lungs, brain, liver and kidney. The total zinc content in the lungs, brain, liver, kidney, red blood cells and plasma was estimated by inductively coupled plasma mass spectroscopy (ICP-MS). Transcription rate of the genes was evaluated by RealTime PCR. ResultsThe two concentration of ZnO nanoparticles in the ambient air produced two different outcomes. The lower concentration resulted in significant increases in Zn content of the liver while the higher concentration significantly increased Zn in the lungs (p < 0.05). Additionally, at the lower concentration, Zn content was found to be lower in brain tissue (p < 0.05). Using TEM/EDX we detected ZnO nanoparticles inside the cells in the lungs, kidney and liver. Inhaling ZnO NP at the higher concentration increased the levels of mRNA of the following genes in the lungs: Mt2 (2.56 fold), Slc30a1 (1.52 fold) and Slc30a5 (2.34 fold). At the lower ZnO nanoparticle concentration, only Slc30a7 mRNA levels in the lungs were up (1.74 fold). Thus the two air concentrations of ZnO nanoparticles produced distinct effects on the expression of the Zn-homeostasis related genes. ConclusionUntil adverse health effects of ZnO nanoparticles deposited in organs such as lungs are further investigated and/or ruled out, the exposure to ZnO nanoparticles in aerosols should be avoided or minimised.

Silicon reduces the iron uptake in rice and induces iron homeostasis related genes

Gramineous plants take up silicon (Si) that enhances the formation of exodermal Casparian bands (CBs) in the roots of rice (Oryza sativa L.). Furthermore, it is known that Si supply reduces the concentration of Fe in rice shoots. We hypothesized that the Si-enhanced CB formation in the exodermis reduces in the flux of Fe in the apoplast and the uptake of Fe loaded deoxymugineic acid. Thus, the effect of silicic acid supply at varied Fe concentrations and Fe forms was investigated in nutrient solution. The Fe concentrations in the shoot and apoplastic Fe concentrations in the root were determined and an Affymetrix GeneChip experiment was carried out together with qRT-PCR measurements for observation of transcriptomic reactions. Additionally, the Fe uptake of an overexpression mutant of OsABCG25 with an enhanced exodermal CB formation was investigated. The application of silicic acid reduced the Fe concentrations in shoot DM independently of the supplied Fe concentration and Fe form. As a reaction to the Fe shortage, the full cascade of Fe-homeostasis-related genes in the roots was upregulated. Silicic acid supply also decreased the apoplastic Fe concentrations in roots. In addition, an overexpression mutant of OsABCG25 with an enhanced CB formation showed a reduced uptake of Fe in excess Fe conditions. The results suggest that the Si-induced CB formation in the exodermis hampers the flux of Fe into the apoplast of the cortex and, thus, Fe uptake of rice grown in nutrient solution which is reflected in the upregulation of Fe homeostasis-related genes.

Lipidomics-based study on the neuroprotective effect of geissoschizine methyl ether against oxidative stress-induced cytotoxicity

Ethnopharmacological relevanceLipid homoeostasis is important for neurodevelopment, cell signaling and neurotransmission. Alteration of lipid metabolism has been demonstrated in many neurological disorders and neurodegenerative diseases. Geissoschizine methyl ether (GM) is an active alkaloid ingredient in the traditional Chinese medicine Uncaria hook. It has been shown that GM has strong potency in neuroprotective activity and GM reduces the production of reactive oxygen species by regulating glucose metabolism, which protects neurons against oxidative stress-induced cell death. However, it is unknown whether GM could regulate neuronal lipid metabolism during oxidative challenge. Aim of the studyThe current study aimed to explore whether GM regulates lipid metabolism in oxidative damaged neurons and to determine the underlying mechanism involved in this neuro-protection. Materials and methodsUsing a glutamate-induced oxidative toxicity model in mouse hippocampal neuronal cell line (HT-22 cells), we investigated the effect of GM on glutamate-induced lipid peroxidation, lipotoxicity and mitochondrial dysfunction. In order to clarify the mechanism underlying the neuroprotection by GM, lipid metabolomics was performed to investigate whether GM prevent oxidative stress-induced lipid metabolism disruption. Furthermore, the expression of lipid metabolism-related genes was measured. ResultsThe results show the protective effect of GM against oxidative stress through blocking glutamate-induced lipid peroxidation and lipotoxicity. Overall, lipidomics analysis revealed that glutamate treatment resulted in different extents of changes in a wide range of lipid classes such as fatty acids (FA), triacylglycerol (TG), sphingomyelin (SM), cardiolipin (CL), lysophosphatidylcholines (LPC). However, GM treatment can significantly reverse glutamate-induced lipids disorder to the homeostasis level. GM prevented the disruption of lipid metabolism by regulating the expression of lipid homeostasis related genes, which contributes to preserve mitochondrial function under oxidative damage. ConclusionThese findings clearly demonstrated a novel protective mechanism of GM against glutamate-induced oxidative toxicity in neurons via regulating lipid metabolism. GM may provide an effective approach for the prevention and treatment of oxidative damaged neurons.

High-carbohydrate diet promotes the adaptation to acute hypoxia in zebrafish.

Oxygen deprivation (hypoxia) is a common challenge in water environment, which causes lack of energy and oxidative damage in organisms. Many studies have indicated a number of physiological and metabolic changes under hypoxia, but the effects of dietary nutrients on hypoxia tolerance have not been well evaluated. In the present 7-week feeding trial, we fed zebrafish with low-protein diet (LP), high-protein diet (HP), low-fat diet (LF), high-fat diet (HF), low-carbohydrate diet (LC), and high-carbohydrate diet (HC), respectively. Afterward, the resistance to acute hypoxia challenge, growth, body composition, activities of metabolic enzymes, and expressions of energy homeostasis-related genes and six hifαs genes were measured. The results indicated that only the HC diet could significantly improve the resistance to hypoxia challenge. Moreover, the HC diet feeding caused higher glycogen deposition in the liver and muscle, and these glycogens were significantly reduced after 6-h acute hypoxia challenge. Meanwhile, the lactate content in the liver and blood was increased in the HC groups. At hypoxia status, the relative mRNA expressions of the genes related to glycolysis, ATP production, insulin signaling pathway, and hif-3a (hif1al) were all significantly increased in the muscle of the HC diet-fed fish. This study revealed that high-carbohydrate diet could improve the resistance to hypoxia by activating glycolysis and hif/insulin signaling pathway in zebrafish, mainly in the muscle, to efficiently supply energy. Therefore, our results highlight the importance of dietary carbohydrate in resisting hypoxia in fish.

Expression levels of vacuolar ion homeostasis-related genes, Na+ enrichment, and their physiological responses to salt stress in sugarcane genotypes.

Sugarcane is a sugar-producing crop widely grown in tropical regions in over 120 countries of the world. Salt-affected soil is one of the most significant abiotic constraints that inhibit growth and crop productivity, and, consequently, reduce sucrose concentration in the stalk. The present study investigated vacuolar ion homeostasis, Na+ accumulation, and physiological and morphological adaptations under salt stress in two different sugarcane genotypes (salt-tolerant K88-92 and salt-sensitive K92-80) under greenhouse conditions. Na+ was rapidly absorbed by the root tissues of both sugarcane genotypes within 3-7 days of 150 mM NaCl treatment, as confirmed by the results of CoroNa Green fluorescence staining. In addition, the rate of Na+ translocation from roots to shoots was evidently reduced, leading to lower amount of Na+ in the leaf tissues. At the cellular level, expression of ShNHX1 (vacuolar Na+/H+ antiporter), ShV-PPase (vacuolar H+-pyrophosphatase), and ShV-ATPase (vacuolar H+-ATPase) was upregulated in salt-stressed plants for the compartmentation of Na+ into the vacuoles of root cells. Interestingly, sucrose, glucose, and fructose in root tissues of salt-stressed sugarcane cv. K88-92 were increased by 10.61, 5.58, and 1.81 folds, respectively, over the control. Total soluble sugars in the roots and free proline in the leaves of sugarcane cv. K88-92 (salt-tolerant) were enriched by 3.08 and 1.99 folds, respectively, when plants were exposed to 150 mM NaCl, leading to maintain better photosynthetic abilities, net photosynthetic rate (Pn), stomatal conductance (gs), transpiration rate (E), and water use efficiency (WUE) in sugarcane cv. K88-92 than those in cv. K92-80. The study concludes that Na+ compartmentation in the root tissue acts as a major defense mechanism in sugarcane, especially in salt-tolerant genotype.

Quantum Blue Reduces the Severity of Woody Breast Myopathy via Modulation of Oxygen Homeostasis-Related Genes in Broiler Chickens.

The incidence of woody breast (WB) is increasing on a global scale representing a significant welfare problem and economic burden to the poultry industry and for which there is no effective treatment due to its unknown etiology. In this study, using diffuse reflectance spectroscopy (DRS) coupled with iSTAT portable clinical analyzer, we provide evidence that the circulatory- and breast muscle-oxygen homeostasis is dysregulated [low oxygen and hemoglobin (HB) levels] in chickens with WB myopathy compared to healthy counterparts. Molecular analysis showed that blood HB subunit Mu (HBM), Zeta (HBZ), and hephaestin (HEPH) expression were significantly down regulated; however, the expression of the subunit rho of HB beta (HBBR) was upregulated in chicken with WB compared to healthy counterparts. The breast muscle HBBR, HBE, HBZ, and hypoxia-inducible factor prolyl hydroxylase 2 (PHD2) mRNA abundances were significantly down regulated in WB-affected compared to normal birds. The expression of HIF-1α at mRNA and protein levels was significantly induced in breasts of WB-affected compared to unaffected birds confirming a local hypoxic status. The phosphorylated levels of the upstream mediators AKT at Ser473 site, mTOR at Ser2481 site, and PI3K P85 at Tyr458 site, as well as their mRNA levels were significantly increased in breasts of WB-affected birds. In attempt to identify a nutritional strategy to reduce WB incidence, male broiler chicks (Cobb 500, n = 576) were randomly distributed into 48 floor pens and subjected to six treatments (12 birds/pen; 8 pens/treatment): a nutrient adequate control group (PC), the PC supplemented with 0.3% myo-inositol (PC + MI), a negative control (NC) deficient in available P and Ca by 0.15 and 0.16%, respectively, the NC fed with quantum blue (QB) at 500 (NC + 500 FTU), 1,000 (NC + 1,000 FTU), or 2,000 FTU/kg of feed (NC + 2,000 FTU). Although QB-enriched diets did not affect growth performances (FCR and FE), it did reduce the severity of WB by 5% compared to the PC diet. This effect is mediated by reversing the expression profile of oxygen homeostasis-related genes; i.e., significant down regulation of HBBR and upregulation of HBM, HBZ, and HEPH in blood, as well as a significant upregulation of HBA1, HBBR, HBE, HBZ, and PHD2 in breast muscle compared to the positive control.

Ion accumulation and expression of ion homeostasis-related genes associated with halophilism, NaCl-promoted growth in a halophyte Mesembryanthemum crystallinum L.

ABSTRACTA halophyte, the common ice plant (Mesembryanthemum crystallinum L.), shows the maximal growth under salinity, in which almost all crops die. The NaCl-stimulated growth, which is referred to as halophilism, is an important trait for adaptation to salinity, but the mechanism is still unclear. To elucidate factors contributing to the halophilism, we tested the effects of NaCl on growth, ion accumulation, and expression of ion homeostasis-related genes in suspension-cultured cells. Among nine ions analyzed, Na+, Cl−, K+, and NO3− were accumulated significantly in the cells showing halophilism than that in the salt-untreated cells, and the accumulation of these ions was positively correlated with the growth. Also, the expression of ion homeostasis-related genes for plasma membrane transporters and channels for incorporation of nitrate (McNRT1), sodium (McHKT1), potassium (McKmt1), cations/Cl− (McCCC1), for tonoplast antiporters H+/Cl− (McCLC1) and Na+/H+ (McNHX1), and V-ATPase subunit c (McVmac1) for sequestration of Cl− and Na+ into the vacuole, and for enzymes catalyzing biosynthesis of proline (McP5CS) and ononitol (McImt1) was higher in the cells showing halophilism than that in the salt-untreated cells. These results indicate that the ion accumulation and the expression of ion homeostasis-related genes contribute to the NaCl-stimulated growth enhancement in the halophyte, the common ice plant.Abbreviations: CCC: cation/Cl- cotransporter; CLC: H+/Cl–antiporter; DW: dry weight; FW: fresh weight; HAK: high affinity potassium; HKT: high potassium transporter; Imt: myo-inositol O-methyl transferase; MIP: major intrinsic protein; NHX: Sodium/proton antiporter; P5CS: delta 1-pyrroline-5-carboxylate synthase; PCV: Packed cell volume; PEG: polyethylene glycol; XTH: xyloglucan endotransglucosylase/hydrolase

Abstract LB-295: Therapeutic implications of altered cholesterol homeostasis mediated by loss of CYP46A1 in human glioblastoma

Abstract BACKGROUND Tumor cells display higher cholesterol levels compared to normal cells and emerging evidence indicates a pivotal role of cholesterol content in tumor cell proliferation and growth mediated by a dysregulation of key regulators involved in cholesterol homeostasis has been implicated in various cancers. In this study, we assessed the dependency of glioblastoma (GBM) cells to extracellular cholesterol levels for growth. Through analysis of whole-transcriptome databases, we identified the cholesterol homeostasis-related genes whose expression are dysregulated in gliomas. One of the most dramatically down-regulated genes identified encodes cholesterol 24-hydroxylase (CYP46A1), a brain-specific enzyme responsible for elimination of cholesterol via conversion of cholesterol to 24(S)-hydroxycholesterol (24OHC). METHODS Molecular and clinical data was obtained from publicly genomic databases. Immunohistochemistry was applied to assess the protein level of CYP46A1 in tissue microarrays. Functionally, CYP46A1 overexpression was determined using lentiviral constructs in LN229 and a primary glioblastoma cell line (GBM#P3). Cells were characterized in vitro and implanted in vivo to generate orthotopic xenografts in order to assess their molecular status, cell survival, and self-renewal by western blotting, flow cytometry, and extreme limiting dilution assay (ELDA), respectively. RESULTS Down-regulation of CYP46A1 was found in malignant glioma samples compared to normal brain tissue. Importantly, reduced CYP46A1 expression was associated with aggressive clinicopathological features and poor prognosis of GBM patients. In functional assays, restoration of CYP46A1 suppressed cell proliferation and in vivo tumour growth by increasing 24OHC levels. Consistent, treatment of GBM cells with 24OHC inhibited tumour growth and induced apoptosis in a concentration dependent manner. RNA-seq data showed that 24OHC regulated LXR and sterol regulatory-element binding protein 1 (SREBP1) signalling activity. Intriguingly, efavirenz (EFV), a widespread used anti-HIV medication that effectively penetrates the blood–brain barrier, has recently been reported as a potent activator of the CYP46A1 enzyme. The therapeutic potential of EFV is currently being assessed in vivo and the data will be presented. CONCLUSIONS Collectively, our findings demonstrate that CYP46A1 is a critical cellular cholesterol regulator in GBM and that the CYP46A1/24OHC axis is a potential therapeutic target in GBM. Citation Format: Mingzhi Han, Shuai Wang, Xingang Li, Jian Wang, Rolf Bjerkvig. Therapeutic implications of altered cholesterol homeostasis mediated by loss of CYP46A1 in human glioblastoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr LB-295.

Metabolomics and Lipidomics Reveal the Effect of Hepatic Vps33b Deficiency on Bile Acids and Lipids Metabolism.

Vascular protein sorting-associated protein 33B (VPS33B) plays important roles in hepatic polarity, which directly maintains the functional structure of the liver. It has reported that VPS33B has close association with arthrogryposis, renal dysfunction and cholestasis (ARC) syndrome. Unfortunately, no further studies were conducted to reveal the role of Vps33b in the homeostasis of bile acids. In the current study, hepatic Vps33b-depleted male mice were used to investigate the metabolomics and lipidomics profiles of hepatic Vps33b deficiency based on ultrahigh-performance liquid chromatography coupled with an electrospray ionization high-resolution mass spectrometry (UHPLC-ESI-HRMS) system. Hepatic Vps33b-depleted male mice displayed cholestasis and slight liver damage with increased serum levels of ALT, AST, ALP and T-Bili compared to wild-type mice. Targeted metabolomics analysis of bile acids revealed that increased taurine-conjugated bile acids accumulated in the serum of hepatic Vps33b-depleted mice, while unconjugated bile acids were prone to decrease, accompanied by the regulation of bile acid homeostasis-related genes. In addition, lipid profiles were significantly altered with the lack of Vps33b in the liver. A variety of lipids, such as triglycerides and sphingomyelins, were significantly decreased in the liver and increased in the serum of hepatic Vps33b-depleted mice compared to those in wild-type mice. Our study demonstrated that Vps33b influences the progress of liver metabolism both in bile acid circulation and lipid metabolism, which is involved in the progression of liver cholestasis in mice.

The protein kinase Cmk2 negatively regulates the calcium/calcineurin signalling pathway and expression of calcium pump genes PMR1 and PMC1 in budding yeast

Through a genome-wide screen we have identified calcium-tolerant deletion mutants for five genes in the budding yeast Saccharomyces cerevisiae. In addition to CNB1 and RCN1 that are known to play a role in the calcium signalling pathway, the protein kinase gene CMK2, the sphingolipid homeostasis-related gene ORM2 and the gene SIF2 encoding the WD40 repeat-containing subunit of Set3C histone deacetylase complex are involved in the calcium sensitivity of yeast cells to extracellular calcium. Cmk2 and the transcription factor Crz1 have opposite functions in the response of yeast cells to calcium stress. Deletion of CMK2 elevates the level of calcium/calcineurin signalling and increases the expression level of PMR1 and PMC1, which is dependent on Crz1. Effects of Cmk2 on calcium sensitivity and calcium/calcineurin signalling are dependent on its kinase activity. Therefore, Cmk2 is a negative feedback controller of the calcium/calcineurin signalling pathway. Furthermore, the cmk2 crz1 double deletion mutant is more resistant than the crz1 deletion mutant, suggesting that Cmk2 has an additional Crz1-independent role in promoting calcium tolerance.

CYP46A1 expression modulates mitochondrial function and integrity in Niemann-Pick type C disease

CYP46A1 expression modulates mitochondrial function and integrity in Niemann-Pick type C disease