USP2a alters chemotherapeutic response by modulating redox

Cancer cells are characterized by altered ubiquitination of many proteins. The ubiquitin-specific protease 2a (USP2a) is a deubiquitinating enzyme overexpressed in prostate adenocarcinomas, where it exhibits oncogenic behavior in a variety of ways including targeting c-Myc via the miR-34b/c cluster. Here we demonstrate that USP2a induces drug resistance in both immortalized and transformed prostate cells. Specifically, it confers resistance to typically pro-oxidant agents, such as cisplatin (CDDP) and doxorubicin (Doxo), and to taxanes. USP2a overexpression protects from drug-induced oxidative stress by reducing reactive oxygen species (ROS) production and stabilizing the mitochondrial membrane potential (ΔΨ), thus impairing downstream p38 activation and triggering of apoptosis. The molecular mediator of the USP2a protective function is the glutathione (GSH). Through miR-34b/c-driven c-Myc regulation, USP2a increases intracellular GSH content, thus interfering with the oxidative cascade triggered by chemotherapeutic agents. In light of these findings, targeting Myc and/or miR-34b/c might revert chemo-resistance

Effect of nitrogen fertilization and fungicide application at heading on the gluten protein composition and rheological quality of wheat

Optimizing the bread-making quality properties of flour is currently one of the main aims of the bakery industry. Therefore, this study has investigated the effects of N fertilization and fungicide application at wheat heading on the protein content (GPC), gluten composition and rheological properties of wheat. Field experiments were carried out in North-West Italy over a 3 year period, on a high protein cultivar of soft winter wheat. Grain samples were collected for each agronomic treatment at four ripening timings, from the milk stage to the final combine harvesting, and the contents of the different gluten fractions were quantified. The late N fertilization increased the GPC (+1.2%) and dough strength (W) (+22%) as a result of a similar enhancement of all the gluten protein fractions, while the fungicide application slightly reduced the GPC (0.3%) and W(4%), mainly because of a dilution of the gliadin content, due to the significantly higher grain yield (+8.6%) and thousand kernel weight (+5.5%). These agronomic practices did not modify the gluten composition significantly, expressed as the relative ratio between the gliadins (glia) and the high (HMW) and low (LMW) molecular weight glutenins (gs), and confirmed by the accumulation trend of the different protein fractions during ripening. The year resulted to have the most marked effect on the gluten protein fraction ratios and alveographic parameters. The lowest W was observed in 2015, and although the highest GPC was recorded for the same year, the lowest gs/glia ratio was also observed. Instead, 2016 showed the highest gs/glia and HMW-gs/LMW-gs (H/L) ratios, and also the highest P/L value (2.2). In 2015, a slightly higher temperature during the ripening stage resulted in a greater increase in the -gliadin enriched fraction than the / gliadin ones, and marked differences were noted in the rheological traits. This field experiment has highlighted the interactive role of environmental and agronomic factors on the content and quality of gluten proteins and their bread-making ability, thus making a further contribution to the development of an integrated crop strategy for the cultivation of high protein wheat in humid Mediterranean areas

Development and comparison of two multiresidue methods for the determination of 17 Aspergillus and Fusarium mycotoxins in cereals using HPLC-ESI-TQ-MS/MS

Cereals can be contaminated by several mycotoxins, whose co-presence may represent an undervalued risk for humans and animals. Maize and wheat are the most contaminated cereals and in temperate areas could be affected in field conditions by several Fusarium and Aspergillus infections. To date, only B-fumonisins (FBs), aflatoxins (AFs), zearalenone (ZEA), deoxynivalenol (DON) and T-2 and HT-2 toxins have been regulated in cereals in European Union. The other fungal metabolites, are commonly referred to as “emerging” and “masked” mycotoxins, and more information on their occurrence in combination with the regulated mycotoxins, are needed to design combined toxicological and exposure assessments.This research intends to develop and compare two multiresidue HPLC-ESI-TQ-MS/MS methods for the simultaneous determination of the main regulated, emerging and masked mycotoxins in maize and wheat, among which: FB1, FB2, DON, ZEA, AFB1, AFB2, AFG1, AFG2, moniliformin (MON), deoxynivalenol-3-glucoside (DON-3-G), 3-acetyldeoxynivalenol (3-ADON), 15-acetyldeoxynivalenol (15-ADON), nivalenol (NIV), enniatins A, A1, B, B1 (ENNA, ENNA1, ENNB, ENNB1). The extraction was performed for both methods using a mixture of CH3CN/H2O/CH3COOH (79/20/1, v/v/v), while the dilution/purification was carried out through two different procedures: (1) by the “dilute-and-shoot” technique diluting 1:2 the filtered extract with CH3CN/H2O/CH3COOH (20/79/1, v/v/v) to reduce the matrix effect; (2) using the Oasis® PRiME HLB clean-up columns. The analysis was carried out using CH3OH and H2O both acidified with 0.1% of CH3COOH as eluents. The injection volume was 20 μL and the flow rate 200 μL min-1. The analysis of two reference material (maize and wheat), was performed to evaluate the trueness and precision of the two methods by matrix-matched calibration curves. For all the regulated mycotoxins analyzed by both methods, the range of recovery percentage established by the Regulation (EC) No. 401/2006 was respected, except for ZEA by using the Oasis® PRiME HLB clean-up columns. Nevertheless, the results suggest that the Oasis® PRiME HLB clean-up columns, could be a valid alternative to the dilute-and-shoot method, although an additional cost for the clean-up has to be considered. In conclusion, both two analytical methods considerably reduce the analytical time and costs and therefore result to be promising and applicable for high-throughput routine multi-mycotoxins analysis by the use of a TQ

Influence of agronomic conditions on the efficacy of different fungicides applied to wheat at heading: effect on flag leaf senescence, Fusarium head blight attack, grain yield and deoxynivalenol contamination

A series of field experiments has been conducted in North West Italy over a period of 3 years to evaluate the effect of fungicide applications on common wheat yield and safety, combined with different agronomic conditions (high: a susceptible variety to FHB planted in an untilled field; low: a medium tolerant variety to FHB planted in a ploughed field) for Fusarium head blight (FHB) infection risk. A azole mixture (prochloraz + epoxiconazole), applied at heading, was compared in each year and for each agronomic condition with a triazole with high activity against F. graminearum and F. culmorum (metconazole), a strobilurin-azole mixture with elevated action to control leaf diseases and delay leaf senescence (azoxystrobin + prochloraz) and an untreated control. The following parameters were analyzed: flag leaf greenness, grain yield, test weight, thousand kernel weight (TKW), FHB incidence and severity and deoxynivalenol (DON) contamination. The results of this research, conducted over three years with different meteorological conditions, underline the important link between fungicide efficacy and environmental and agronomic conditions that can influence fungal disease pressure. The fungicide effect on the control of FHB and the increase in flag leaf longevity and grain yield were greater with an increase in the disease pressure. On the other hand, the DON contamination was reduced by the fungicide to a greater extent in the low risk agronomic and environmental conditions compared to the high risk ones. Prochloraz + epoxiconazole showed a lower efficacy in reducing DON contamination compared to metconazole, particularly in the higher FHB pressure conditions. No significant differences were observed between the azole mixture and the strobilurin-azole mixture for flag leaf greenness, grain yield, test weight or TKW. This study provides useful information that can be used to evaluate, in naturally-infected field conditions, the effect of a fungicide application at heading on wheat yield and safety in distinct scenarios for FHB and foliar disease pressure