Unravelling the regulatory network putatively controlling flavonoid biosynthesis in grapevine

I flavonoidi sono una classe di metaboliti secondari fondamentali per la bacca d\u2019uva e per il vino. Nella vite, vengono sintetizzati in molti organi a diversi stadi di sviluppo. Studi recenti hanno dimostrato che la sintesi \ue8 controllata a livello trascrizionale da un complesso proteico WD40-bHLH-MYB, che in altre specie \ue8 responsabile della regolazione di altri processi metabolici anche attraverso l\u2019attivazione di altri fattori di trascrizione. In questo progetto di ricerca, \ue8 stata studiata la funzione di VvMYB5a e VvMYB5b, appartenenti al complesso regolatorio in vite (Deluc 2006, 2008). Inoltre, abbiamo identificato e studiato l\u2019ipotetico coinvolgimento di VvWRKY19 nel network regolatorio in vite, dimostrando che in vite il complesso proteico regola molteplici processi metabolici.Flavonoids are secondary metabolites important for grape and wine quality. The biosynthetic pathway is highly regulated in order that they are produced in different organs and at various stages of development. Recent studies demonstrated that a WD40-bHLH-MYB regulatory complex controls the flavonoid pathway in grapevine. In other species this complex is responsible for the regulation of other metabolic processes through the control of other intermediate regulators. In this PhD project, we studied the roles of VvMYB5a and VvMYB5b, two putative members of the complex in grapevine (Deluc 2006, 2008). Moreover, the involvement of VvWRKY19 as a putative intermediate regulator belonging to the regulatory network was investigated

A Grapevine TTG2-Like WRKY Transcription Factor Is Involved in Regulating Vacuolar Transport and Flavonoid Biosynthesis

open7noA small set of TTG2-like homolog proteins from different species belonging to the WRKY family of transcription factors were shown to share a similar mechanism of action and to control partially conserved biochemical/developmental processes in their native species. In particular, by activating P-ATPases residing on the tonoplast, PH3 from Petunia hybrida promotes vacuolar acidification in petal epidermal cells whereas TTG2 from Arabidopsis thaliana enables the accumulation of proanthocyanidins in the seed coat. In this work we functionally characterized VvWRKY26 identified as the closest grapevine homolog of PhPH3 and AtTTG2. When constitutively expressed in petunia ph3 mutant, VvWRKY26 can fulfill the PH3 function in the regulation of vacuolar pH and restores the wild type pigmentation phenotype. By a global correlation analysis of gene expression and by transient over-expression in Vitis vinifera, we showed transcriptomic relationships of VvWRKY26 with many genes related to vacuolar acidification and transport in grapevine. Moreover, our results indicate an involvement in flavonoid pathway possibly restricted to the control of proanthocyanidin biosynthesis that is consistent with its expression pattern in grape berry tissues. Overall, the results show that, in addition to regulative mechanisms and biological roles shared with TTG2-like orthologs, VvWRKY26 can play roles in fleshy fruit development that have not been previously reported in studies from dry fruit species. This study paves the way toward the comprehension of the regulatory network controlling vacuolar acidification and flavonoid accumulation mechanisms that contribute to the final berry quality traits in grapevine.openAmato, Alessandra; Cavallini, Erika; Zenoni, Sara; Finezzo, Laura; Begheldo, Maura; Ruperti, Benedetto; Tornielli, Giovanni BattistaAmato, Alessandra; Cavallini, Erika; Zenoni, Sara; Finezzo, Laura; Begheldo, Maura; Ruperti, Benedetto; Tornielli, Giovanni Battist

A molecular characterization of the invasive fig weevil Aclees taiwanensis in Italy

An economically important pest of Ficus carica L. is causing severe infestations in many fig nurseries and orchards in Italy. Belonging to the genus Aclees spp. (Coleoptera Curculionidae), this Asiatic species was accidentally introduced in Europe about 15 years ago, in a Tuscan nursery. Originally identified as Aclees cribratus Gyllenhal, it has been then reported as Aclees sp.cf. foveatus Voss and, more recently, identified as Aclees taiwanensis Kono. A serious damage to fig plants is caused mainly by the larvae, which drill tunnels into the wood and by adults that feed on buds, leaves and young fruits. The present survey applies molecular genetics techniques to reconstruct the genetic profile of the species. To this purpose, the partial sequence of the 18S rRNA gene and the hypervariable region ITS2 of the ribosomal cistron were used as molecular markers for specimens of A. taiwanensis collected in Italy and Aclees hirayamai Kono from Philippines. The analysis of the partial sequences of the 18S rRNA allowed the distinction of two haplotypes for each insect, except for a sample from Philippines, for which one haplotype does exist. The use of the ITS2 hypervariable region highlighted the existence of only one haplotype in the Italian accessions. Only in the sample collected in Lucca (2LU) two haplotypes were highlighted in ITS2. These results are discussed with the occurrence of A. taiwanensis in Italy

Strategy-adaptation memory training: predictors of older adults' training gains

AbstractOver the past decades, memory training interventions have been developed in an attempt to stabilize or enhance memory functioning in aging. Only recently has attention been paid to individual differences in training gains and consequently to predictors of such gains. The aim of the present study was to identify which specific cognitive mechanisms/processes or components of the intervention were responsible for the desired change and which individuals were more responsive to memory strategic training. Eighty-one older adults (aged 55 to 82) were involved in a four-session strategy-adaptation training based on a learner-oriented approach that has previously been found to be effective in improving memory performance in practiced and untrained tasks. Results showed that baseline performance in memory tasks predicted the gains in the practiced task. Baseline performance in memory tasks and other cognitive variables, such as working memory, processing speed, and verbal knowledge predicted transfer effects. Interestingly, we found that the magnitude of training gain on the associative memory practiced task predicted the gains in the transfer tasks, suggesting those who best implemented the targeted strategies during training realized greater transfer to other tasks. Our study shows that older adults with larger cognitive resources will benefit more from interventions focused on the generalization via active processes

The photomorphogenic factors UV-B RECEPTOR 1, ELONGATED HYPOCOTYL 5, and HY5 HOMOLOGUE are part of the UV-B signalling pathway in grapevine and mediate flavonol accumulation in response to the environment

Grapevine (Vitis vinifera L.) is a species well known for its adaptation to radiation. However, photomorphogenic factors related to UV-B responses have not been molecularly characterized. We cloned and studied the role of UV-B RECEPTOR (UVR1), ELONGATED HYPOCOTYL 5 (HY5), and HY5 HOMOLOGUE (HYH) from V. vinifera We performed gene functional characterizations, generated co-expression networks, and tested them in different environmental conditions. These genes complemented the Arabidopsis uvr8 and hy5 mutants in morphological and secondary metabolic responses to radiation. We combined microarray and RNA sequencing (RNA-seq) data with promoter inspections to identify HY5 and HYH putative target genes and their DNA binding preferences. Despite sharing a large set of common co-expressed genes, we found different hierarchies for HY5 and HYH depending on the organ and stress condition, reflecting both co-operative and partially redundant roles. New candidate UV-B gene markers were supported by the presence of HY5-binding sites. These included a set of flavonol-related genes that were up-regulated in a HY5 transient expression assay. We irradiated in vitro plantlets and fruits from old potted vines with high and low UV-B exposures and followed the accumulation of flavonols and changes in gene expression in comparison with non-irradiated conditions. UVR1, HY5, and HYH expression varied with organ, developmental stage, and type of radiation. Surprisingly, UVR1 expression was modulated by shading and temperature in berries, but not by UV-B radiation. We propose that the UV-B response machinery favours berry flavonol accumulation through the activation of HY5 and HYH at different developmental stages at both high and low UV-B exposure

Inspection of the grapevine BURP superfamily highlights an expansion of RD22 genes with distinctive expression features in berry development and ABA-mediated stress responses

The RESPONSIVE TO DEHYDRATION 22 (RD22) gene is a molecular link between abscisic acid (ABA) signalling and abiotic stress responses. Its expression has been used as a reliable ABA early response marker. In Arabidopsis, the single copy RD22 gene possesses a BURP domain also located at the C-terminus of USP embryonic proteins and the beta subunit of polygalacturonases. In grapevine, a RD22 gene has been identified but putative paralogs are also found in the grape genome, possibly forming a large RD22 family in this species. In this work, we searched for annotations containing BURP domains in the Vitis vinifera genome. Nineteen proteins were defined by a comparative analysis between the two genome predictions and RNA-Seq data. These sequences were compared to other plant BURPs identified in previous genome surveys allowing us to reconceive group classifications based on phylogenetic relationships and protein motif occurrence. We observed a lineage-specific evolution of the RD22 family, with the biggest expansion in grapevine and poplar. In contrast, rice, sorghum and maize presented highly expanded monocot-specific groups. The Vitis RD22 group may have expanded from segmental duplications as most of its members are confined to a region in chromosome 4. The inspection of transcriptomic data revealed variable expression of BURP genes in vegetative and reproductive organs. Many genes were induced in specific tissues or by abiotic and biotic stresses. Three RD22 genes were further studied showing that they responded oppositely to ABA and to stress conditions. Our results show that the inclusion of RNA-Seq data is essential while describing gene families and improving gene annotations. Robust phylogenetic analyses including all BURP members from other sequenced species helped us redefine previous relationships that were erroneously established. This work provides additional evidence for RD22 genes serving as marker genes for different organs or stresses in grapevine

Genome and transcriptome analysis of the grapevine (Vitis vinifera L.) WRKY gene family

The plant WRKY gene family represents an ancient and complex class of zinc-finger transcription factors (TFs) that are involved in the regulation of various physiological processes, such as development and senescence, and in plant response to many biotic and abiotic stresses. Despite the growing number of studies on the genomic organisation of WRKY gene family in different species, little information is available about this family in grapevine (Vitis vinifera L.). In the present study, a total number of 59 putative grapevine WRKY transcription factors (VvWRKYs) were identified based on the analysis of various genomic and proteomic grapevine databases. According to their structural and phylogentic features, the identified grapevine WRKY transcription factors were classified into three main groups. In order to shed light into their regulatory roles in growth and development as well as in response to biotic and abiotic stress in grapevine, the VvWRKYs expression profiles were examined in publicly available microarray data. Bioinformatics analysis of these data revealed distinct temporal and spatial expression patterns of VvWRKYs in various tissues, organs and developmental stages, as well as in response to biotic and abiotic stresses. To also extend our analysis to situations not covered by the arrays and to validate our results, the expression profiles of selected VvWRKYs in response to drought stress, Erysiphe necator (powdery mildew) infection, and hormone treatments (salicilic acid and ethylene), were investigated by quantitative real-time reverse transcription PCR (qRT-PCR). The present study provides a foundation for further comparative genomics and functional studies of this important class of transcriptional regulators in grapevine

Integration of B-cell receptor-induced ERK1/2 phosphorylation and mutations of SF3B1 gene refines prognosis in treatment-na\uefve chronic lymphocytic leukemia

ADP-ribosyl Cyclase 1; Disease Progression; Humans; Immunoglobulin Heavy Chains; Immunoglobulin Variable[no abstract available

The photomorphogenic factors UV-B RECEPTOR 1, ELONGATED HYPOCOTYL 5, and HY5 HOMOLOGUE are part of the UV-B signalling pathway in grapevine and mediate flavonol accumulation in response to the environment

Grapevine (Vitis vinifera L.) is a species well known for its adaptation to radiation. However, photomorphogenic factors related to UV-B responses have not been molecularly characterized. We cloned and studied the role of UV-B RECEPTOR (UVR1), ELONGATED HYPOCOTYL 5 (HY5), and HY5 HOMOLOGUE (HYH) from V. vinifera. We performed gene functional characterizations, generated co-expression networks, and tested them in different environmental conditions. These genes complemented the Arabidopsis uvr8 and hy5 mutants in morphological and secondary metabolic responses to radiation. We combined microarray and RNA sequencing (RNA-seq) data with promoter inspections to identify HY5 and HYH putative target genes and their DNA binding preferences. Despite sharing a large set of common co-expressed genes, we found different hierarchies for HY5 and HYH depending on the organ and stress condition, reflecting both co-operative and partially redundant roles. New candidate UV-B gene markers were supported by the presence of HY5-binding sites. These included a set of flavonol-related genes that were up-regulated in a HY5 transient expression assay. We irradiated in vitro plantlets and fruits from old potted vines with high and low UV-B exposures and followed the accumulation of flavonols and changes in gene expression in comparison with non-irradiated conditions. UVR1, HY5, and HYH expression varied with organ, developmental stage, and type of radiation. Surprisingly, UVR1 expression was modulated by shading and temperature in berries, but not by UV-B radiation. We propose that the UV-B response machinery favours berry flavonol accumulation through the activation of HY5 and HYH at different developmental stages at both high and low UV-B exposures. © The Author 2016. Published by Oxford University Press on behalf of the Society for Experimental Biology

Further Clarification of Pain Management Complexity in Radiotherapy: Insights from Modern Statistical Approaches

Background: The primary objective of this study was to assess the adequacy of analgesic care in radiotherapy (RT) patients, with a secondary objective to identify predictive variables associated with pain management adequacy using a modern statistical approach, integrating the Least Absolute Shrinkage and Selection Operator (LASSO) algorithm and the Classification and Regression Tree (CART) analysis. Methods: This observational, multicenter cohort study involved 1387 patients reporting pain or taking analgesic drugs from 13 RT departments in Italy. The Pain Management Index (PMI) served as the measure for pain control adequacy, with a PMI score < 0 indicating suboptimal management. Patient demographics, clinical status, and treatment-related factors were examined to discern the predictors of pain management adequacy. Results: Among the analyzed cohort, 46.1% reported inadequately managed pain. Non-cancer pain origin, breast cancer diagnosis, higher ECOG Performance Status scores, younger patient age, early assessment phase, and curative treatment intent emerged as significant determinants of negative PMI from the LASSO analysis. Notably, pain management was observed to improve as RT progressed, with a greater discrepancy between cancer (33.2% with PMI < 0) and non-cancer pain (73.1% with PMI < 0). Breast cancer patients under 70 years of age with non-cancer pain had the highest rate of negative PMI at 86.5%, highlighting a potential deficiency in managing benign pain in younger patients. Conclusions: The study underscores the dynamic nature of pain management during RT, suggesting improvements over the treatment course yet revealing specific challenges in non-cancer pain management, particularly among younger breast cancer patients. The use of advanced statistical techniques for analysis stresses the importance of a multifaceted approach to pain management, one that incorporates both cancer and non-cancer pain considerations to ensure a holistic and improved quality of oncological care