Identificación y caracterización de la superfamilia génica ALDH en garbanzo (Cicer arietinum) mediante herramientas bioinformáticas de acceso libre

Premio extraordinario de Trabajo Fin de Máster curso 2017/2018. Máster en Producción, Protección y Mejora VegetalLas aldehído deshidrogenasas (ALDHs) son una superfamilia de proteínas con una función importante en la detoxificación de los aldehídos producidos en respuesta a estreses bióticos/abióticos. La disponibilidad del genoma de referencia de garbanzo (Cicer arietinum) da la oportunidad de identificar y caracterizar a los miembros de esta familia en una leguminosa de importancia agronómica. En este estudio, se han identificado 37 ALDHs en el genoma de garbanzo y se ha realizado una caracterización completa de las mismas. Los análisis filogenéticos comparativos con Medicago sugieren una gran conservación de la familia entre las dos especies y los análisis de duplicaciones indican una leve incidencia de eventos de duplicación con posterioridad a la especiación. El análisis de expresión in silico apoya el papel de la mayoría de miembros de la familia CaALDH en la tolerancia al estrés abiótico, con una mayor representación de las secuencias de la familia 18 en librerías EST de tolerancia a sequía. Todos los scripts escritos en este trabajo y la secuencia de ejecución para el análisis de las bases de datos están disponibles públicamente en un repositorio online de acceso libre. En resumen, este trabajo proporciona una visión general de la superfamilia génica ALDH en garbanzo. Es la primera vez que la familia se estudia en este cultivo. Nuestros resultados respaldan que las proteínas ALDHs están implicadas en una amplia gama de rutas metabólicas y que participan en la respuesta al estrés. Esto proporciona nuevos conocimientos sobre la la presencia y función de la familia en esta especie, lo que puede ser útil para desarrollar estrategias de mejora genética de respuesta a diferentes estreses. Este trabajo también proporciona una base para análisis genómicos comparativos posteriores en el estudio de la evolución de los genes ALDH dentro de la familia de las leguminosas.Aldehyde dehydrogenases (ALDHs) constitute a protein superfamily with an important function in the detoxification of the aldehydes produced in response to biotic/abiotic stresses. The availability of the chickpea (Cicer arietinum) reference genome provides an opportunity to identify and characterize the members of this family in a legume of agronomic importance. In this study, 37 ALDHs have been identified in the chickpea genome and a complete characterization of them has been carried out. The comparative phylogenetic analysis with Medicago suggests a high conservation of the family between both species and the duplication analysis indicates a slight duplication incidence after the speciation. In silico expression analysis supports the abiotic stress tolerance role of most CaALDH family members, showing the family 18 sequences overrepresented in drought tolerance EST libraries. All the code and scripts written for this work are publicly available in an online open access repository. In summary, this work provides the first general overview of the ALDH gene superfamily in this crop. Our results support that ALDH proteins are involved in a wide range of metabolic pathways and they participate in the stress response. This provides new knowledge of the family presence and function in this species, what may be useful to develop chickpea breeding strategies to improve development or stress responses. This work also supplies a basis for further comparative genomic analysis and a framework to study the ALDH genes evolution within the legume family

Aldehyde Dehydrogenase 3 Is an Expanded Gene Family with Potential Adaptive Roles in Chickpea

Legumes play an important role in ensuring food security, improving nutrition and enhancing ecosystem resilience. Chickpea is a globally important grain legume adapted to semi-arid regions under rain-fed conditions. A growing body of research shows that aldehyde dehydrogenases (ALDHs) represent a gene class with promising potential for plant adaptation improvement. Aldehyde dehydrogenases constitute a superfamily of proteins with important functions as ‘aldehyde scavengers’ by detoxifying aldehydes molecules, and thus play important roles in stress responses. We performed a comprehensive study of the ALDH superfamily in the chickpea genome and identified 27 unique ALDH loci. Most chickpea ALDHs originated from duplication events and the ALDH3 gene family was noticeably expanded. Based on the physical locations of genes and sequence similarities, our results suggest that segmental duplication is a major driving force in the expansion of the ALDH family. Supported by expression data, the findings of this study offer new potential target genes for improving stress tolerance in chickpea that will be useful for breeding programs

Multiclass cyanotoxin analysis in reservoir waters: tandem solid-phase extraction followed by zwitterionic hydrophilic interaction liquid chromatography-mass spectrometry

The presence of cyanobacteria and cyanotoxins in all water bodies, including ocean water and fresh water sources, represents a risk for human health as eutrophication and climate change are enhancing their level of proliferation. For risk assessment and studies on occurrence, the development of reliable and sensitive analytical approaches able to cover a wide range of cyanotoxins is essential. This work describes the development of an HILIC-MS/MS multiclass method for the simultaneous analysis of eight cyanotoxins in reservoir water samples belonging to three different classes according to their chemical structure: cyclic peptides (microcystin-LR, microcystin-RR and nodularin), alkaloids (cylindrospermopsin, anatoxin-a) and three non-protein amino acids isomers such as β-methylamino-L-alanine, 2,4-diaminobutyric acid and N-(2-aminoethyl)glycine). A SeQuant ZIC-HILIC column was employed to achieve the chromatographic separation in less than 12 min. Previously, a novel sample treatment based on a tandem solid-phase extraction (SPE) system using mixed cation exchange (MCX) and Strata-X cartridges was investigated with the aim of extracting and preconcentrating this chemically diverse group of cyanotoxins. The Strata-X cartridge, which was configured first in the line of sample flow, retained the low polar compounds and the MCX cartridge, which was at the bottom of the dual system, retained mainly the non-protein amino acids. The optimization procedure highlighted the importance of sample ion content for the recoveries of some analytes such as the isomers β-Nmethylamino-L-alanine and 2-4-diaminobutyric acid. Method validation was carried out in terms of linearity, limit of detection (LOD) and quantification (LOQ), recoveries, matrix effect and precision in terms of repeatability and intermediate precision. This work represents the first analytical method for the simultaneous analysis of these multiclass cyanotoxins in reservoir water samples, achieving LOQs in the very low range of 7·10-3 – 0.1 µg·L-1. Despite high recoveries obtained at the LOQ concentration levels (101.0-70.9%), relative standard deviations lower than 17.5% were achieved.RTI2018-097043-B-I00 financed by MCIN/ AEI /10.13039/501100011033/ FEDER “Una manera de hacer Europa”Spanish Network of Excellence in Sample preparation (RED2018-102522-T)FPU17/0381