Target Of Rapamycin Gene Expression Research Articles

Yeast in addition to pollen enhances the reproduction of the predatory mite Euseius nicholsi by increasing the target of rapamycin gene expression

Euseius species are good predators and can control infestations of non-web producing spider mites (e.g., Panonychus); they face difficulties, though, in controlling web producing spider mite species such as Tentranychus urticae. Euseius nicholsi has not yet been mass reared using an alternative diet. Here we investigated the effects of nutrient supplementation to a diet of pollen on the growth, development, and reproduction of Euseius nicholsi and the underlying molecular mechanisms. When Euseius nicholsi groups fed pollen were compared to groups fed pollen + yeast powder or pollen + sugar, yeast supplementation shortened the immature stages, but sugar supplementation lengthened the immature stages. The oviposition of Euseius nicholsi reared on pollen + yeast was higher than those in the other treatments. Based on these findings, we propose that the combination of pollen and yeast powder can enhance the reproduction in Euseius nicholsi females and could be used as an alternative diet. We also characterized the expression profile of the Euseius nicholsi target of rapamycin (EnTOR) gene, which was expressed in all developmental stages tested. The EnTOR expression levels were highest in adults when reared on pollen + yeast powder. When EnTOR was silenced using RNA interference, expression of the Euseius nicholsi vitellogenin (Vg) genes was significantly decreased, reducing the oviposition period, fecundity, and egg hatching rate. Based on these findings, we conclude that protein supplementation (yeast powder) in Euseius nicholsi enhances TOR gene expression, there by promoting the expression of the vitellogeningenes and thus improving fecundity. These data provide a foundation for further research on TOR function and provide a theoretical basis for exploring the reproduction mechanism of Euseius nicholsi.

Target of rapamycin (TOR) gene is vital for whitefly survival and reproduction

The whitefly Bemisia tabaci (Gennadius) is a devastating polyphagous pest that causes significant losses in several economically important crops worldwide. Although chemical control is the fastest management strategy, insecticide resistance development poses a greater constraint in whitefly management. In this study, we used RNA interference (RNAi) technique to understand the role of target of rapamycin (TOR) gene in whitefly. In insects, TOR plays a crucial role in cell growth, metabolism, proliferation, and reproduction. Although sequence analyses of TOR revealed 95 nucleotides different in TOR gene sequence compared to predicted sequence (GenBank accession no. XM019060193.1), but this led to synthesis of only two different amino acids at positions 1448 and 1768. Phylogenetic analysis of amino acid sequence revealed that TOR gene of B. tabaci, Acrythosiphum pisum and Nilaparvata lugens were more closely related to each other and were clustered together. Oral feeding of artificial diet containing dsRNA viz 2.0, 1.0, 0.5, 0.1 μg/μl corresponding to TOR gene to adult whitefly resulted in 42.5% adult mortality, reduced fecundity i.e. 36.57 eggs per female, and 62.50% total nymphal mortality in the next generation. The qRT-PCR analysis confirmed the downregulation of TOR gene expression in adult whiteflies upon dstor feeding. These results suggest that TOR gene significantly regulates the survival and reproduction in whitefly and can be a potential RNAi target for insect management.

Effects of Dietary Corn Gluten Meal on Growth, Immunity, Enzyme Activity, and Protein Metabolism in Relation to TOR Gene Expression in Juvenile Large Yellow Croaker Larimichthys crocea

A series of six isonitrogenous (45% crude protein) and isolipidic (10% crude lipid) diets were formulated by replacing 0% (control), 15%, 30%, 45%, 60% and 75% of fish meal (FM) with corn gluten meal (CGM), including 40% FM. In addition, all diets were supplemented with crystalline amino including lysine and methionine in all diets except the control. Results showed that there were no significant effects on survival, weight gain ratio (WGR), specific growth rate (SGR), and feed conversion ratio (FCR) among all dietary diet groups (P>0.05). There were also no significant differences (P > 0.05) among all diets in the serum immune enzyme, Alkaline Phosphatase (AKP), Lysozyme (LYZ) and Complement 3 (C3), but in Complement 4 (C4), there was a significantly higher content in C75 compared with the other diets (P < 0.05). There were no significant differences in alpha-amylase activities of digestive enzymes in hindgut among the dietary treatments (P > 0.05), while the activities of lipase were significantly lower in C0 than in the other groups (P < 0.05). There were no significant differences in trypsin activity in fish fed the C15, and C75 diets than the control group (P > 0.05). There were no significant differences (P > 0.05) among all the diets in aspartate amino transferase (AST), but in alanine aminotransferase (ALT) the activities of C75 diet were significantly lower compared with the other diets (P < 0.05). mRNA expression levels of target of rapamycin (TOR) in dietary groups were significantly lower than the control group (P < 0.05), however, there were no significant differences in C15 to C75 groups (P > 0.05). Results of the present study suggested that CGM could substitute up to 75% of FM without influencing the growth of juvenile Larimichthys Crocea.

Immunity decreases, antioxidant system damages and tight junction changes in the intestine of grass carp (Ctenopharyngodon idella) during folic acid deficiency: Regulation of NF-κB, Nrf2 and MLCK mRNA levels.

This investigation used the same growth trial as the previous study, which showed that folic acid deficiency retarded growth in young grass carp (the percent weight gain of Groups 1-6 were 102.32±3.41%, 137.25±10.48%, 179.78±3.95%, 164.33±3.21%, 143.35±8.12% and 115.28±2.66%) [1]. In the present study, we investigated the effects of dietary folic acid on the immune response, antioxidant status and tight junctions in the intestine of young grass carp (Ctenopharyngodon idella). A total of 540 young grass carp were fed diets containing graded levels of folic acid at 0.10, 0.47, 1.03, 1.48, 1.88 and 3.12mgkg(-1) diet for 8 weeks. The results indicated that acid phosphatase and lysozyme activities, and the complement component 3 content in the proximal intestine (PI), mid intestine (MI) and distal intestine (DI) were decreased with folic acid deficiency (0.1mgkg(-1))(P<0.05). Folic acid deficiency (0.1mgkg(-1)) up-regulated interleukin 1β, interleukin 8, tumor necrosis factor α, nuclear factor κB p65 (NF-κB p65), IκB kinase α (IKK-α), IKK-β and IKK-γ gene expression, meanwhile down-regulated interleukin 10, transforming growth factor β, IκB and target of rapamycin gene expression in the PI, MI and DI (P<0.05). These data suggested that folic acid deficiency decreased fish intestinal innate immune function may be partly contributed to the regulation of NF-κB p65 pathway. Moreover, the activities and corresponding gene expression of glutathione content, Cu/Zn superoxide dismutase, catalase, glutathione peroxidase, glutathione s-transferases and glutathione reductase in fish intestine were depressed by deficient folic acid diet (0.1mgkg(-1))(P<0.05). Furthermore, folic acid deficiency (0.1mgkg(-1)) down-regulated NF-E2-related factor 2 (Nrf2) gene expression, up-regulated Kelch-like-ECH-associated protein 1a (Keap1a) and Keap1b gene expression in fish intestine (P<0.05). These data indicated that deficient folic acid diet damaged fish intestinal antioxidant capacity partly by regulating Nrf2/Keap1 pathway. Additionally, folic acid deficiency (0.1mgkg(-1)) down-regulated claudin-b, claudin-c, claudin-3, occludin and zonula occludens 1 gene expression; whereas folic acid deficiency (0.1mgkg(-1)) up-regulated claudin-12, claudin-15, myosin light chain kinase (MLCK) and p38 mitogen activated protein kinase (p38 MAPK) gene expression in the PI, MI and DI (P<0.05), suggesting that folic acid deficiency may damage fish intestinal tight junctions associated with the mediation of MLCK and p38 MAPK gene expression. In conclusion, folic acid deficiency (0.1mgkg(-1)) impaired fish intestinal immunity, antioxidant capacity and tight junctions.

Folic acid deficiency impairs the gill health status associated with the NF-κB, MLCK and Nrf2 signaling pathways in the gills of young grass carp (Ctenopharyngodon idella).

The aim of this study was to investigate the effect of dietary folic acid on fish growth, the immune and barrier functions of fish gills, and the potential mechanisms of these effects. Young grass carp (Ctenopharyngodon idella) were fed diets containing graded levels of folic acid at 0.10 (basal diet), 0.47, 1.03, 1.48, 1.88 and 3.12 mg kg(-1) diet for 8 weeks. The results showed that acid phosphatase and lysozyme activities and the complement component 3 content in fish gills decreased with folic acid deficiency (P < 0.05). Folic acid deficiency up-regulated liver-expressed antimicrobial peptide 1, interleukin 1β, interleukin 8, tumor necrosis factor α, nuclear factor κB p65, IκB kinase α (IKK-α), IKK-β and IKK-γ gene expression. Folic acid deficiency down-regulated interleukin 10, transforming growth factor β, IκB and target of rapamycin gene expression in fish gills (P < 0.05). These results showed that limited folic acid decreased fish gill immune status. Furthermore, folic acid deficiency down-regulated claudin-b, claudin-c, claudin-3, occludin and zonula occludens 1 gene expression, whereas folic acid deficiency up-regulated claudin-12, claudin-15, myosin light chain kinase and p38 mitogen activated protein kinase gene expression in fish gills (P < 0.05). These results suggested that folic acid deficiency disrupted tight junction-mediated fish gill barrier function. Additionally, folic acid deficiency increased the content of reactive oxygen species, protein carbonyl and malondialdehyde (MDA); Mn superoxide dismutase activity and gene expression; and Kelch-like-ECH-associated protein 1a (Keap1a) and Keap1b gene expression (P < 0.05). Conversely, folic acid deficiency decreased Cu/Zn superoxide dismutase, catalase, glutathione peroxidase, glutathione s-transferases and glutathione reductase activities and gene expression as well as NF-E2-related factor 2 gene expression in fish gills (P < 0.05). All of these results indicated that folic acid deficiency impaired fish gill health status via regulating gene expression of cytokines, tight junction proteins, antioxidant enzymes, NF-κB p65, MLCK and Nrf2. Based on percent weight gain, LZ activity and MDA content in the gills, the dietary folic acid requirements for young grass carp were 1.60, 2.07 and 2.08 mg kg(-1), respectively.

Optimal dietary alpha-linolenic acid/linoleic acid ratio improved digestive and absorptive capacities and target of rapamycin gene expression of juvenile grass carp (Ctenopharyngodon idellus)

Growth performance, digestive and absorptive capacities and target of rapamycin (TOR), ribosomal protein S6 kinase 1 (S6K1) and eIF4E-binding protein (4E-BP) gene expression in the hepatopancreas and intestine of juvenile grass carp (Ctenopharyngodon idellus) fed graded ratios of dietary alpha-linolenic acid/linoleic acid (ALA/LNA) (0.01, 0.34, 0.68, 1.03, 1.41, 1.76 and 2.15) for 60 days were investigated. The results showed that ALA/LNA ratio of 1.03 significantly improved (i) per cent weight gain (PWG) and feed efficiency, (ii) hepatopancreatic trypsin, chymotrypsin, lipase, amylase and intestinal creatine kinase (CK) activities, (iii) hepatopancreatic trypsinogen-2 and chymotrypsinogen mRNA levels. Meanwhile, fish fed with ALA/LNA ratio of 0.68 significantly enhanced, (iv) Na+/K+-ATPase and γ-glutamyl transpeptidase activities in whole intestine, and alkaline phosphatase activities in the proximal intestine (PI) and distal intestine, (v) amylase, intestinal Na+/K+-ATPase alpha-subunit isoform 1, Na+/K+-ATPase alpha-subunit isoform 8 and CK mRNA abundances, (vi) TOR and S6K1 gene expression in the hepatopancreas and intestine of juvenile grass carp. Based on the quadratic regression analysis of PWG, cholecystokinin and leptin contents in the PI, optimal dietary ALA/LNA ratio of juvenile grass carp (8.78–72.00 g) was estimated to be 1.08, 1.19 and 1.05, respectively.

Effect of tryptophan on growth, intestinal enzyme activities and TOR gene expression in juvenile Jian carp (Cyprinus carpio var. Jian): Studies in vivo and in vitro

This study was conducted both in vivo and in vitro to investigate the effects of tryptophan on growth performance, digestive and absorptive function and protein synthesis of juvenile Jian carp (Cyprinus carpio var. Jian). 1050 juvenile Jian carp (initial weight 7.73±0.03g) were fed seven isonitrogenous diets with graded concentrations of tryptophan (1.1, 1.7, 2.5, 3.8, 4.9, 6.0, 6.9g/kg diet) for 8weeks. Percent weight gain, feed intake and protein retention value were markedly improved, with increases in dietary tryptophan up to 3.8g/kg diet. Similar trend was found in glutamate–oxaloacetate transaminase (GOT) and glutamate–pyruvate transaminase (GPT) activities, trypsin, lipase and α-amylase activities, Na+/K+-ATPase, alkaline phosphatase (AKP), γ-glutamyl transpeptidase and creatine kinase activities, and relative expression of eIF4E-binding protein (4E-BP). On the other hand, feed conversion ratio, plasma ammonia concentration and the relative expression of target of rapamycin (TOR) in different tissue showed an opposite pattern. A series of experiments in vitro were then carried out. Compared with the control group, tryptophan supplementation increased 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide (MTT) OD value, protein content and activity of AKP, GOT, GPT and Na+/K+-ATPase in enterocytes and decreased lactate dehydrogenase activity and ammonia concentration in the culture medium. Protein synthesis rate was 17% higher and relative expression of TOR was 28% lower in tryptophan-supplemented than in control carp enterocytes. In conclusion, our results indicate that tryptophan improved fish growth, digestive and absorptive function as well as protein synthesis, which may be partly related to the TOR signaling pathway.

Differences in Mammalian target of rapamycin gene expression in the peripheral blood and articular cartilages of osteoarthritic patients and disease activity.

The gene expression of mTOR, autophagy-related ULK1, caspase 3, CDK-inhibitor p21, and TNF α was measured in the peripheral blood of osteoarthritic (OA) patients at different stages of the disease aiming to establish a gene expression profile that might indicate the activity of the disease and joint destruction. Whole blood of 65 OA outpatients, 27 end-stage OA patients, 27 healthy volunteers, and knee articular cartilages of 28 end-stage OA patients and 26 healthy subjects were examined. OA outpatients were subjected to clinical testing, ultrasonography, and radiographic and WOMAC scoring. Protein levels of p70-S6K, p21, and caspase 3 were quantified by ELISA. Gene expression was measured using real-time RT-PCR. Upregulation of mTOR gene expression was observed in PBMCs of 42 OA outpatients (“High mTOR expression subset”) and in PBMCs and articular cartilages of all end-stage OA patients. A positive correlation between mTOR gene expression in PBMCs and cartilage was observed in the end-stage OA patients. 23 OA outpatients in the “Low mTOR expression subset” exhibited significantly lower mTOR gene expression in PBMCs compared to healthy controls. These “Low mTOR” subset subjects experienced significantly more pain upon walking, and standing and increased total joint stiffness versus “High mTOR” subset, while the latter more often exhibited synovitis. The protein concentrations of p70-S6K, p21, and caspase 3 in PBMCs were significantly lower in the “Low” subset versus “High” subset and end-stage subjects. Increases in the expression of mTOR in PBMCs of OA patients are related to disease activity, being associated with synovitis more than with pain.

Effects of dietary amino acid patterns on growth and protein metabolism of large yellow croaker (Larimichthys crocea) larvae

This study was conducted to evaluate the effects of dietary amino acids (AA) patterns on growth, survival, activities of digestive enzymes and aminotransferases and target of rapamycin (TOR) gene expression of large yellow croaker (Larimichthys crocea) larvae. The control diet was produced using intact protein as the only protein source. Four isonitrogenous and isolipidic semi-purified diets were formulated with crystalline-AA replacing approximately 40% fish meal protein-bound nitrogen. The AA patterns of these diets were adjusted according to the overall AA pattern of large yellow croaker egg protein (LEP), large yellow croaker larvae whole-body protein (LLP), large yellow croaker muscle protein (LMP) and white fishmeal protein (WFP), respectively. The test diets and live copepod were fed to triplicate groups of larvae (initial body weight 3.15±0.15mg) five times (6:00, 8:30, 12:30, 14:30, and 17:00) daily for 30days. The results showed that specific growth rate (SGR) of fish fed the WFP diet was significantly higher than LEP or LLP diet (P<0.05). At the end of the growth trial, there was no significant difference in survival rate among larvae fed LEP, LLP, WFP and the control diet (P>0.05), whereas the highest value was recorded in larvae fed the LLP diet, followed by WFP, the control, LEP and LMP diet, respectively. Whole-body crude protein content was significantly higher in larvae fed the WFP diet compared to the LEP or LLP diet (P<0.05). Larvae fed the live copepod had significantly higher whole-body crude protein and lipid contents than that in larvae fed artificial microdiets (P<0.05). Whole-body moisture content was not significantly affected by dietary treatments (P>0.05). The specific activities of digestive enzymes and the ratio “pancreatic enzyme in intestinal segment/pancreatic enzyme in pancreatic segment” were significantly higher in fish fed the WFP diet than fish fed LLP and LEP diet (P<0.05). Specific activities of alanine and aspartate aminotransferases were significantly higher in fish fed the WFP diet compared to the other treatments (P<0.05). No significant differences were observed in larvae body TOR gene expression among dietary treatments (P>0.05). Results of this study indicated that white fishmeal protein amino acid pattern was a more suitable amino acid pattern in diets of large yellow croaker larvae compared to the amino acid pattern of LEP, LLP, and LMP.

Effects of dietary corn gluten meal on growth performance and protein metabolism in relation to IGF-I and TOR gene expression of juvenile cobia (Rachycentron canadum)

A growth experiment was conducted on cobia (Rachycentron canadum, initial weight 108.2 g ± 3.0 g) to investigate the effects of dietary corn gluten meal (CGM) levels on the fish growth, whole body composition and protein metabolism in relation to specific gene expression. Five isonitrogenous (crude protein 45%) and isoenergetic (gross energy 20 kJ g−1) practical diets were formulated by replacing 0% (the control), 17.5%, 35.0%, 52.5%, and 70.0% of fish meal (FM) protein with CGM protein. No significant differences were observed in the survival, feed intake (FI), specific growth rate (SGR), feed efficiency (FE) and protein productive value (PPV) among fish fed diets with 0%, 17.5%, 35.0%, and 52.5% of CGM protein. However, these indices were significantly lower in fish fed the diet with 70.0% of CGM protein than those in fish fed the control diet (P < 0.05). The whole-body crude protein and lipid contents were significantly lower while the whole-body moisture content was significantly higher in fish fed the diet with 70.0% of CGM protein compared with the control group (P < 0.05). When 70.0% of FM protein was replaced by CGM, plasma total protein and cholesterol contents were significantly lower than those in the control group (P < 0.05). Fish fed the diet with 70.0% of CGM protein had significantly lower hepatic insulin-like growth factor I (IGF-I) expression levels than those in the control group (P < 0.05). However, no significant differences were observed in hepatic target of rapamycin (TOR), dorsal muscle IGF-I and TOR expression levels among dietary treatments. Results of the present study indicated that 52.5% of FM protein could be replaced by CGM in the diets without significant influences on the growth, feed utilization and protein metabolism of juvenile cobia. The present results might be useful for developing cost effective and sustainable cobia dietary formulations.

Effect of dietary choline on growth, intestinal enzyme activities and relative expressions of target of rapamycin and eIF4E-binding protein2 gene in muscle, hepatopancreas and intestine of juvenile Jian carp ( Cyprinus carpio var. Jian)

The effects of dietary choline on growth, digestive and absorptive capacities, and target of rapamycin (TOR) and eIF4E-binding protein2 (4E-BP2) gene expression in muscle, hepatopancreas and intestine of juvenile Jian carp ( Cyprinus carpio var. Jian) were assessed. A total of 1200 juvenile Jian carp with an average initial weight of 7.94 ± 0.03 g were fed six semi-purified diets containing 165 (unsupplemented control group), 310, 607, 896, 1167 and 1820 mg choline kg − 1 diet for 65 days. Specific growth rate (SGR), feed intake, feed efficiency, protein retention value, protein content and lipid content of fish carcasses were significantly improved by dietary choline supplementation; whereas, protein efficiency ratio and body ash content were not significantly different among dietary groups. Patterns of differences in intestine length, relative gut length, intestine weight, hepatopancreas protein content, intestinal folds height in three intestinal segments, trypsin and chymotrypsin activities in intestine and hepatopancreas, α-amylase and creatine kinase activities in intestine, Na +/K +-ATPase activity in distal intestine and relative expression of TOR gene in distal intestine and 4E-BP2 gene in muscle responded similar to SGR; whereas, the trends of hepatosomatic index, lipase activity in intestine and hepatopancreas, Na +/K +-ATPase activity in proximal intestine and mid intestine, TOR gene expression in muscle and hepatopancreas, and 4E-BP2 gene expression in proximal intestine, mid intestine and distal intestine were opposite. Alkaline phosphatase and γ-glutamyl transpeptidase activities in intestinal segments were not significantly different among dietary groups. These results indicated that dietary choline could improve fish growth, enhance digestive and absorptive ability and regulate TOR and 4E-BP2 gene expression in tissues. The dietary choline requirement of Jian carp estimated by the broken-line model based on SGR was 566 mg choline kg − 1 diet in the form of choline chloride.