Effect of pH on MHC class II-peptide interactions.

The effect of pH on class II-peptide interactions has been analyzed using several mouse (IAd, IAk, IEd, IEk) and human (DR1, DR5, DR7) MHC specificities, and eight different class II-restricted determinants. In direct binding assays, acidic conditions led to increased binding capacity for many class II-peptide combinations. IE molecules seemed to bind optimally around pH 4.5, whereas IA molecules displayed binding optima in the 5.5 to 6.5 range. In contrast, the DR molecules studied were, in most cases, affected only marginally by pH changes in the 4.5 to 7.0 range. Despite these apparent isotype-specific trends, no general rule could be formulated, because even for the same class II molecules, the binding capacity could be increased for many peptides when the binding was performed under acidic conditions, was unaffected for some, and even decreased for others. The mechanisms responsible for this complex behavior were analyzed in more detail by kinetic and equilibrium analysis of three different class II-peptide combinations (IAd/OVA 323-339, IAk/HEL 46-61, and DR1/HA 307-319). It was found that acidic pH conditions could affect both on and off rates for class II-peptide complexes. Depending on the net balance of these effects, either increases, decreases, or no effect on overall affinities at equilibrium were detected. In the case of IAd/OVA 323-339, it was also found that acidic conditions influenced the binding capacity of class II molecules by increasing the fraction of sites available for peptide binding, presumably by favoring dissociation of endogenously bound, acid-sensitive peptides.     

Association of genetic variants rs641153 (CFB), rs2230199 (C3), and rs1410996 (CFH) with age-related macular degeneration in a Brazilian population

This study aimed to investigate the association among genetic variants of the complement pathway CFB R32Q (rs641153), C3 R102G (rs2230199), and CFH (rs1410996) with age-related macular degeneration (AMD) in a sample of the Brazilian population. In a case-control study, 484 AMD patients were classified according to the clinical age-related maculopathy grading system (CARMS) and compared to 479 unrelated controls. The genetic variants rs1410996 of complement H (CFH), rs641153 of complement factor B (CFB), and rs2230199 of complement 3 (C3) were evaluated through polymerase chain reaction (PCR) and direct sequencing. The associations between single nucleotide polymorphisms (SNPs) and AMD, adjusted by age, were assessed by using logistic regression models. A statistically significant association was observed between AMD risk and rs2230199 variant with an OR of 2.01 (P  = 0.0002) for CG individuals compared to CC individuals. Regarding the comparison of advanced AMD versus the control group, the OR was 2.12 (P = 0.0036) for GG versus AA genotypes for rs1410996 variant. Similarly, the OR for rs2230199 polymorphism was 2.3034 (P  = 5.47^e-05) when comparing CG individuals to CC carriers. In contrast, the rs641153 variant showed a significant protective effect against advanced AMD for GA versus GG genotype (OR = 0.4406; P  = 0.0019). When comparing wet AMD versus controls, a significant association was detected for rs1410996 variant (OR = 2.16; P  = 0.0039) comparing carriers of the homozygous GG versus AA genotype, as well as in the comparisons of GG (OR = 3.0713; P  = 0.0046) and CG genotypes (OR = 2.2249; P  = 0.0002) versus CC genotype for rs2230199 variant, respectively. The rs641153 variant granted a significant protective effect against wet AMD for GA versus GG genotypes (OR = 0.4601; P  = 0.0044). Our study confirmed the risk association between rs2230199 and rs1410996 variants and AMD, and the protective role against AMD for rs641153 variant.     

Genetic dissection of Al tolerance QTLs in the maize genome by high density SNP scan

Background

Aluminum (Al) toxicity is an important limitation to food security in tropical and subtropical regions. High Al saturation on acid soils limits root development, reducing water and nutrient uptake. In addition to naturally occurring acid soils, agricultural practices may decrease soil pH, leading to yield losses due to Al toxicity. Elucidating the genetic and molecular mechanisms underlying maize Al tolerance is expected to accelerate the development of Al-tolerant cultivars.

Results

Five genomic regions were significantly associated with Al tolerance, using 54,455 SNP markers in a recombinant inbred line population derived from Cateto Al237. Candidate genes co-localized with Al tolerance QTLs were further investigated. Near-isogenic lines (NILs) developed for ZmMATE2 were as Al-sensitive as the recurrent line, indicating that this candidate gene was not responsible for the Al tolerance QTL on chromosome 5, qALT5. However, ZmNrat1, a maize homolog to OsNrat1, which encodes an Al³⁺ specific transporter previously implicated in rice Al tolerance, was mapped at ~40 Mbp from qALT5. We demonstrate for the first time that ZmNrat1 is preferentially expressed in maize root tips and is up-regulated by Al, similarly to OsNrat1 in rice, suggesting a role of this gene in maize Al tolerance. The strongest-effect QTL was mapped on chromosome 6 (qALT6), within a 0.5 Mbp region where three copies of the Al tolerance gene, ZmMATE1, were found in tandem configuration. qALT6 was shown to increase Al tolerance in maize; the qALT6-NILs carrying three copies of ZmMATE1 exhibited a two-fold increase in Al tolerance, and higher expression of ZmMATE1 compared to the Al sensitive recurrent parent. Interestingly, a new source of Al tolerance via ZmMATE1 was identified in a Brazilian elite line that showed high expression of ZmMATE1 but carries a single copy of ZmMATE1.

Conclusions

High ZmMATE1 expression, controlled either by three copies of the target gene or by an unknown molecular mechanism, is responsible for Al tolerance mediated by qALT6. As Al tolerant alleles at qALT6 are rare in maize, marker-assisted introgression of this QTL is an important strategy to improve maize adaptation to acid soils worldwide.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2164-15-153) contains supplementary material, which is available to authorized users.     

Use of 2-[18F]fluoroethylazide for the Staudinger ligation – Preparation and characterisation of GABAA receptor binding 4-quinolones

The labelling reagent 2-[¹⁸F]fluoroethylazide was used in a traceless Staudinger ligation. This reaction was employed to obtain the GABA_A receptor binding 6-benzyl-4-oxo-1,4-dihydro-quinoline-3-carboxylic acid (2-[¹⁸F]fluoroethyl) amide. The radiotracer was prepared with a non-decay corrected radiochemical yield of 7%, a radiochemical purity >95% and a specific radioactivity of 0.9 GBq/μmol. The compound showed low brain penetration in normal rats. A series of fluoroalkyl 4-quinolone analogues with nanomolar to sub-nanomolar affinity for the GABA_A receptor has been prepared as well.     

Early evidence of lymphoproliferative disorder: post-transplant monitoring of Epstein-Barr infection in adult and pediatric patients

Transplant patients are at high risk of post-transplant lymphoproliferative disorder (PTLD). A strong correlation between Epstein-Barr virus (EBV) and PTLD is observed in pediatric patients with primary infection after transplant. Because many patients have responded to reversal of immunosuppressive therapy, an early identification of EBV is essential for the reduction of immunosuppression and/or introduction of antiviral therapy to prevent PTLD. Polymerase chain reaction (PCR) is a specific and sensitive method to identify EBV DNA in blood. The aim of our study was to establish a protocol for monitoring EBV infection in transplanted patients for early identification those at high risk of PTLD. Viral presence in peripheral blood leukocytes (PBL) and serum samples was revealed by Nested PCR; positive specimens were quantified with Real Time PCR (RT-PCR). DNA in PBL was observed in 12 cases and 6 showed EBV in sera. Quantitative analysis showed a wide range of EBV DNA copies in leukocytes that were higher than in sera. Two patients displayed high viral load values in both PBL and sera associated with clinical evidence of PTLD. Our data suggest that the study of the EBV load represents an essential approach in the diagnosis of PTLD and the analysis of serum samples could provide useful information in the post-transplant monitoring of high-risk patients.     

Toxicity of rhizomes of the invasive Hedychium coronarium (Zingiberaceae) on aquatic species

The production and release of chemical compounds by invasive plants can affect competitors and native species overall, destabilizing ecological interactions and harming ecosystem functioning. Hedychium coronarium is an invasive macrophyte common on Brazilian riparian areas that produces a wide variety of allelochemicals, but little is known about their effect on aquatic species. Here, we identified the major chemical compounds of the aqueous extract of H. coronarium rhizomes and assessed its toxicity, evaluating the growth inhibition of one alga (Raphidocelis subcapitata) and one macrophyte (Lemna minor), and the lethality of cladoceran (Ceriodaphnia silvestrii and Daphnia similis) and Chironomidae larvae (Chironomus sancticaroli). The majoritarian compounds of H. coronarium rhizomes were Coronarin D and Coronarin D Ethyl Ether. The aqueous extract was toxic for all tested species. We observed growth inhibition in R. subcapitata, as well as reduction in biomass in L. minor. Chironomus sancticaroli and cladoceran were the most sensible species. The aqueous extract of H. coronarium rhizomes was toxic on tested conditions, suggesting that the rhizome compounds may interfere on aquatic organisms and in the dynamic of trophic webs of aquatic ecosystems on invaded areas.

     

A glucose biosensor operating under non-isothermal conditions: the dynamic response

The results obtained with a glucose biosensor operating under non-isothermal conditions are presented and discussed. Glucose oxidase, immobilized onto Nylon membranes, was used as biological element. An amperometric two electrodes system was employed to measure the anodic current produced by oxidation of hydrogen peroxide. Non-isothermal conditions were characterised in terms of the temperature difference, delta T = Tw - Tc, and of the average temperature of the system, Tav = (Tw + Tc)/2, Tw and Tc being the temperature in the warm and cold half-cells constituting the biosensor. Comparison between the functioning of the biosensor under isothermal and non-isothermal conditions was performed. It was found that, under non-isothermal conditions, the dynamic response and sensitivity increased, while the response times and the detection limit decreased, if comparison was done with the same parameters measured under isothermal conditions. The increase of the dynamic response was found to be proportional to the applied temperature gradient.     

3-O-methylfunicone, a secondary metabolite produced by Penicillium pinophilum, induces growth arrest and apoptosis in HeLa cells

3-O-Methylfunicone (OMF) is a secondary metabolite produced by the soil fungus Penicillium pinophilum which has cytostatic properties. The aim of this study was to investigate the mechanisms by which such properties are exerted, with special reference to any anti-proliferative and apoptotic potential, on HeLa cells. OMF treatment caused about 44% inhibition of cell growth after 24 h, and modifications in the tubulin fibre organization. In addition, a significant increase in p21 mRNA expression and a decrease in cyclin D1 and Cdk4 mRNA expression resulted at the same time. Apoptosis induction was demonstrated by the annexin V assay, cytofluorimetric analysis of the DNA content of the sub-G1 fraction and DNA laddering. Taken together, our data showed that the compound inhibits proliferation of HeLa cells by several mechanisms, such as disruption of tubulin fibres, cell cycle arrest and apoptosis induction. The capacity of the compound to affect the cell cycle and to modulate apoptosis is indicative of a potential for the development of a new agent for cancer chemotherapy.     

Glucose determination by means of a new reactor/sensor system operating under non-isothermal conditions

The dynamic and steady-state responses as well as the response times of a glucose biosensor have been studied under isothermal and non-isothermal conditions as a function of analyte concentration. The presence of a temperature gradient across the catalytic membrane system improved the biosensor characteristics, because the dynamic and steady-state responses increased and the response times decreased under non-isothermal conditions. For example, a macroscopic temperature difference of 20 degrees C applied across the catalytic membrane system increases the biosensor sensitivity of 70% and reduces of 50% its response time. The dependence of the observed effects on the magnitude of the temperature difference applied has been correlated with the substrate (and products) transport across the catalytic membrane system due to the process of thermodialysis.