Exploring the Use of Fruit Callus Culture as a Model System to Study Color Development and Cell Wall Remodeling during Strawberry Fruit Ripening

Cell cultures derived from strawberry fruit at different developmental stages have been obtained to evaluate their potential use to study different aspects of strawberry ripening. Callus from leaf and cortical tissue of unripe-green, white, and mature-red strawberry fruits were induced in a medium supplemented with 11.3 µM 2,4-dichlorophenoxyacetic acid (2,4-D) under darkness. The transfer of the established callus from darkness to light induced the production of anthocyanin. The replacement of 2,4-D by abscisic acid (ABA) noticeably increased anthocyanin accumulation in green-fruit callus. Cell walls were isolated from the different fruit cell lines and from fruit receptacles at equivalent developmental stages and sequentially fractionated to obtain fractions enriched in soluble pectins, ester bound pectins, xyloglucans (XG), and matrix glycans tightly associated with cellulose microfibrils. These fractions were analyzed by cell wall carbohydrate microarrays. In fruit receptacle samples, pectins were abundant in all fractions, including those enriched in matrix glycans. The amount of pectin increased from green to white stage, and later these carbohydrates were solubilized in red fruit. Apparently, XG content was similar in white and red fruit, but the proportion of galactosylated XG increased in red fruit. Cell wall fractions from callus cultures were enriched in extensin and displayed a minor amount of pectins. Stronger signals of extensin Abs were detected in sodium carbonate fraction, suggesting that these proteins could be linked to pectins. Overall, the results obtained suggest that fruit cell lines could be used to analyze hormonal regulation of color development in strawberry but that the cell wall remodeling process associated with fruit softening might be masked by the high presence of extensin in callus cultures

Partial Activation of SA- and JA-Defensive Pathways in Strawberry upon Colletotrichum acutatum Interaction

[EN] Understanding the nature of pathogen host interaction may help improve strawberry (Fragaria x anahassa) cultivars. Plant resistance to pathogenic agents usually operates through a complex network of defense mechanisms mediated by a diverse array of signaling molecules. In strawberry, resistance to a variety of pathogens has been reported to be mostly polygenic and quantitatively inherited, making it difficult to associate molecular markers with disease resistance genes. Colletotrichum acutaturn spp. is a major strawberry pathogen, and completely resistant cultivars have not been reported. Moreover, strawberry defense network components and mechanisms remain largely unknown and poorly understood. Assessment of the strawberry response to C. acutatum included a global transcript analysis, and acidic hormones SA and JA measurements were analyzed after challenge with the pathogen. Induction of transcripts corresponding to the SA and JA signaling pathways and key genes controlling major steps within these defense pathways was detected. Accordingly, SA and JA accumulated in strawberry after infection. Contrastingly, induction of several important SA, JA, and oxidative stress-responsive defense genes, including FaPR1-1, FaLOX2, FaJAR1, FaPDF1, and FaGST1, was not detected, which suggests that specific branches in these defense pathways (those leading to FaPR1-2, FaPR2-1, FaPR2-2, FaAOS, FaPR5, and FaPR10) were activated. Our results reveal that specific aspects in SA and JA dependent signaling pathways are activated in strawberry upon interaction with C. acutatum. Certain described defense-associated transcripts related to these two known signaling pathways do not increase in abundance following infection. This finding suggests new insight into a specific putative molecular strategy for defense against this pathogen.Authors are grateful to Dr. JM Lopez-Aranda (IFAPA-Centro de Churriana) for providing micropropagated strawberry plants and to Nicolas Garcia-Caparros for technical assistance. Authors also want to thank Kevin M. Folta for his insightful comments on the paper. This work was supported by Junta de Andalucia, Spain [Proyectos de Excelencia P07-AGR-02482/P12-AGR-2174, and grants to Grupo-BIO278].Amil-Ruiz, F.; Garrido-Gala, J.; Gadea Vacas, J.; Blanco-Portales, R.; Munoz-Merida, A.; Trelles, O.; De Los Santos, B.... (2016). Partial Activation of SA- and JA-Defensive Pathways in Strawberry upon Colletotrichum acutatum Interaction. Frontiers in Plant Science. 7(1036). https://doi.org/10.3389/fpls.2016.01036S71036Acosta, I. F., & Farmer, E. E. (2010). Jasmonates. The Arabidopsis Book, 8, e0129. doi:10.1199/tab.0129Al-Shahrour, F., Diaz-Uriarte, R., & Dopazo, J. (2004). 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Evaluation of the role of the endo-beta-(1,4)-glucanase gene FaEG3 in strawberry fruit softening

In strawberry, Fragaria x ananassa Duch., fruit, two different endo-beta-1,4-glucanase genes, FaEG1 and FaEG3, also named Cel1 and Cel2, are expressed during the softening process that occurs during fruit ripening. It has also been suggested that FaEG3, which contains a putative cellulose-binding domain, could play a key role in fruit development, since previous attempts to down-regulate this gene through transgenesis have been unsuccessful. In this investigation, we obtained transgenic strawberry plants containing an antisense sequence of the FaEG3 gene under the control of the 35S promoter. Ripened fruit from transgenic lines (Acel lines) showed large variation in FaEG3 silencing, but fruit firmness was similar to control fruit in all the lines. Two Acel lines showing almost 95% reductions in FaEG3 mRNA levels were selected for further study. In these lines, FaEG3 down-regulation was high, from 78 to 95%, at all fruit developmental stages, whereas FaEG1 was only slightly suppressed. In spite of the high FaEG3 silencing achieved, EGase activity was not modified in ripe fruit. At the cell wall level, walls from transgenic ripe fruit contained a significantly higher amount of the 4 M KOH fraction, which is enriched in hemicellulosic polymers. The analysis of this fraction by size exclusion chromatography showed that transgenic cell walls contained a smaller amount of higher molecular mass polymers than controls. Altogether, these results indicate that FaEG3 does not play a key role either in fruit development or fruit softening. However, its silencing affects the amount and, in a minor way, the size of hemicellulosic polymers. (C) 2009 Elsevier B.V. All rights reserved

Transient silencing of the FaWRKY1 strawberry gene (Fragaria x ananassa) in fruit induces resistance to Colletotrichum acutatum infection

Trabajo presentado en el 15th Congress of the Mediterranean Phytopathological Union (Plant health sistaining Mediterranean ecosystems), celebrado en Córdoba (España) del 20 al 23 de junio de 2017.Anthracnose, caused by Colletotrichum acutatum, is responsible for significant yield losses in commercial strawberry production worldwide. For this reason, it is of interest to uncover the molecular basis underlying this strawberry/pathogen interaction. Previously, FaWRKY1 was identified as an important element mediating defence responses.This research was supported by the Project P12-AGR-2174 (Junta de Andalucía, Spain).Peer reviewe