Purine Catabolism in Wheat: Source of Nutrients and Protective Metabolites

Purine catabolism is known to have a dual function in recycling nitrogen (N) and carbon (C) atoms present in the heterocyclic purine ring and participating in stress signalling through the stimulation of ABA metabolism by allantoin, an intermediate in the pathway. However, little information was available of the functions in cereals and, in particular, bread wheat. The aims of this PhD thesis was to investigate the role of purine catabolism and allantoin in Australian bread wheat (Triticum aestivum) genotypes (RAC875 and Mace) grown under N deficiency and water deficit. Firstly, the purine catabolic genes, coding for seven enzymes in total, were identified and annotated in the hexaploid bread wheat genome of cv. Chinese Spring. The analysis revealed 24 loci associated with the enzyme genes. Interestingly, there was a duplication of the xanthine dehydrogenase gene, namely TaXDH1 and TaXDH2. Sequence analysis of the TaXDH2 homeologs located on chromosome group 6 appeared to be either non-functional (TaXDH2-6AS/6BS) or with an inactive xanthine substrate binding site (TaXDH2-6DS). Protein structure modelling and a unique expression pattern under stress indicated that, TaXDH2-6DS may have a novel function in wheat. Characterisation of allantoin and transcription of purine catabolic genes under N and water restrictions, revealed that allantoin levels were reduced (22-fold) when N was limiting, whilst it tended to accumulate in large amounts under drought (up to 30-fold compared to well-watered plants). The latter may suggest allantoin is used as a temporary N sink as the N assimilatory pathway (GS/GOGAT cycle) is likely to have a reduced capacity in plants growing in water deficit conditions. This would prevent the accumulation of ammonium that is toxic at high concentrations and reduce ammonia emissions from plants leaves. The reduction or accumulation of allantoin under drought and N stress appeared transcriptionally regulated by purine catabolic genes. In particular, transcription of TaALN, coding for the allantoin-degrading enzyme allantoinase, oppositely reflected the levels of allantoin in the tissue. Further growth studies showed that wheat seedlings, when re-supplied with xanthine or allantoin as their sole N source after short-term N starvation, had growth rates which were equivalent to plants grown with inorganic nitrogen. This suggests that the N recycled through the purine catabolic pathway can support the growth of wheat. The data also provided evidence that allantoin takes part in N remobilisation during natural senescence. Sequence analysis of TaALN homeologs in a large number of bread wheat accessions highlighted substantial genetic variability when compared to the reference genome of Chinese Spring. Candidate accessions with nucleotide polymorphisms in regulatory elements or in the coding sequence were identified and represent valuable material for future studies. The outcomes of this PhD project provide the ground work for future fundamental research in wheat focussing on the dual role of purine catabolism in N recycling and abiotic stress. In addition, the candidate accessions identified, besides representing useful experimental material, could ultimately be used for breeding purposes. Additional strategies will include genetic engineering of target genes in the pathway that may lead to improved wheat growth and yields in unfavourable environments.Thesis (Ph.D.) -- University of Adelaide, School of Agriculture, Food & Wine, 201

Attenuation of pattern recognition receptor signaling is mediated by a MAP kinase kinase kinase

Pattern recognition receptors (PRRs) play a key role in plant and animal innate immunity. PRR binding of their cognate ligand triggers a signaling network and activates an immune response. Activation of PRR signaling must be controlled prior to ligand binding to prevent spurious signaling and immune activation. Flagellin perception in Arabidopsis through FLAGELLIN‐SENSITIVE 2 (FLS2) induces the activation of mitogen‐activated protein kinases (MAPKs) and immunity. However, the precise molecular mechanism that connects activated FLS2 to downstream MAPK cascades remains unknown. Here, we report the identification of a differentially phosphorylated MAP kinase kinase kinase that also interacts with FLS2. Using targeted proteomics and functional analysis, we show that MKKK7 negatively regulates flagellin‐triggered signaling and basal immunity and this requires phosphorylation of MKKK7 on specific serine residues. MKKK7 attenuates MPK6 activity and defense gene expression. Moreover, MKKK7 suppresses the reactive oxygen species burst downstream of FLS2, suggesting that MKKK7‐mediated attenuation of FLS2 signaling occurs through direct modulation of the FLS2 complex

RNA catabolites contribute to the nitrogen pool and support growth recovery of wheat

Turn-over of RNA and catabolism of nucleotides releases one to four ammonia molecules; the released nutrients being reassimilated into primary metabolism. Preliminary evidence indicates that monocots store high levels of free nucleotides and nucleosides but their potential as a source of internal organic nitrogen for use and remobilization is uncharted. Early tillering wheat plants were therefore starved of N over a 5-day time-course with examination of nucleic acid yields in whole shoots, young and old leaves and roots. Nucleic acids constituted ∼4% of the total N pool of N starved wheat plants, which was comparable with the N available from nitrate (NO3 -) and greater than that available from the sum of 20 proteinogenic amino acids. Methods were optimized to detect nucleotide (purine and pyrimidine) metabolites, and wheat orthologs of RNA degradation (TaRNS), nucleoside transport (TaENT1, TaENT3) and salvage (TaADK) were identified. It was found that N starved wheat roots actively catabolised RNA and specific purines but accumulated pyrimidines. Reduced levels of RNA corresponded with induction of TaRNS2, TaENT1, TaENT3, and TaADK in the roots. Reduced levels of GMP, guanine, xanthine, allantoin, allantoate and glyoxylate in N starved roots correlated with accumulation of allantoate and glyoxylate in the oldest leaf, suggesting translocation of allantoin. Furthermore, N starved wheat plants exogenously supplied with N in the form of purine catabolites grew and photosynthesized as well as those plants re-supplied with NO3 -. These results support the hypothesis that the nitrogen and carbon recovered from purine metabolism can support wheat growth.Vanessa Jane Melino, Alberto Casartelli, Jessey George, Thusitha Rupasinghe, Ute Roessner, Mamoru Okamoto and Sigrid Heue

Additional file 3: Table S2. of Exploring traditional aus-type rice for metabolites conferring drought tolerance

Log2 ratios of Drought vs Well-Watered conditions per genotype and 2-way ANOVA. (XLSX 138 kb

Additional file 1: Table S1. of Exploring traditional aus-type rice for metabolites conferring drought tolerance

Experimental information and data on metabolite peaks. (XLSX 1006 kb

Allelic status of 1p and 19q in oligodendrogliomas and glioblastomas: multiplex ligation-dependent probe amplification versus loss of heterozygosity

Identification of the 1p/19q allelic status in gliomas, primarily those with a major oligodendroglial component, has become an excellent molecular complement to tumor histology in order to identify those cases sensitive to chemotherapy. In addition to loss of heterozygosity (LOH), fluorescence in situ hybridization (FISH), or comparative genomic hybridization (CGH), multiplex ligation-dependent probe amplification (MLPA) has been shown to be an alternative methodology to identify deletions of those chromosome arms. We used MLPA to explore the 1p and 19q glioblastomas, and a series of 76 gliomas: 41 tumors with a major oligodendroglial component, 34 glioblastomas, and one low-grade astrocytoma. We compared the MLPA findings of the oligodendroglial cases with those previously obtained using LOH in the same samples. Thirty-eight of 41 oligodendrogliomas displayed identical findings by both LOH and MLPA, and losses at either 1p and/or 19q were identified in 12 of 35 (34%) astrocytic tumors. These findings agree with data previously reported comparing MLPA versus FISH or CGH in gliomas and suggest that MLPA can be used in the identification 1p/19q allelic deletions on these brain neoplams. (c) 2009 Elsevier Inc. All rights reserved. reserved.Fondo de Investigaciones Sanitarias, Ministerio de Sanidad[PI-05-0829]Fundacion para la Investigacion Biomedica del Hospital Universitario La Pa

Un nuovo modello organizzativo ospedaliero pubblico: il week hospital sovradipartimentale

Proceeding with the description of innovative hospital organizational experiences, the article analyse preliminary data of the week hospital implementation model in the CTO - M. Adelaide hospital of Turin. The organizational change, driven by internal problems and by external benchmarking with others innovative models in orthopaedic hospitals, if planned and managed properly through coherent operational tools, proves its usefulness, even within the complex context of the operating theatre.Continuando a percorrere il filone delle esperienze organizzative innovative, in ambito sanitario, si presentano, in questo articolo, i primi dati derivanti dell'implementazione del modello organizzativo di week hospital, all'interno dell'A.O. CTO - M. Adelaide di Torino. Il cambiamento organizzativo, dettato da problematiche interne e da opportunità di confronti esterni con modelli innovativi sperimentati da altre strutture, se pianificato e gestito con leve operative coerenti e corrette, si dimostra utile e proficuo, anche nel complesso contesto delle attività di sala operatoria