Autophagy is required for strawberry fruit ripening

Autophagy is a catabolic and recycling pathway that maintains cellular homeostasis under normal growth and stress conditions. Two major types of autophagy, microautophagy and macroautophagy, have been described in plants. During macroautophagy, cellular content is engulfed by a double-membrane vesicle called autophagosome. This vesicle fuses its outer membrane with the tonoplast and releases the content into the vacuole for degradation. During certain developmental processes, autophagy is enhanced by induction of several autophagy-related genes (ATG genes). Autophagy in crop development has been studied in relation to leaf senescence, seed and reproductive development, and vascular formation. However, its role in fruit ripening has only been partially addressed. Strawberry is an important berry crop, representative of non-climacteric fruit. We have analyzed the occurrence of autophagy in developing and ripening fruits of the cultivated strawberry. Our data show that most ATG genes are conserved in the genome of the cultivated strawberry Fragaria x ananassa and they are differentially expressed along the ripening of the fruit receptacle. ATG8-lipidation analysis proves the presence of two autophagic waves during ripening. In addition, we have confirmed the presence of autophagy at the cellular level by the identification of autophagy-related structures at different stages of the strawberry ripening. Finally, we show that blocking autophagy either biochemically or genetically dramatically affects strawberry growth and ripening. Our data support that autophagy is an active and essential process with different implications during strawberry fruit ripening.This work has received funding from the European Union’s Horizon 2020 research and innovation programme under the Marie Skłodowska-Curie to VS-V grant agreement 844365 project FrATGaria: Identification of the role of autophagy during fruit ripening and stress resistance in Strawberry: Autophagy manipulation as a way to improve Strawberry fitness, by the Ministerio de Economía y Competitividad, co-financed by the European Regional Development Fund (grant BIO2017-82609-R) to MB and by the Plan Propio of the University of Malaga

Generation and analysis of ESTs from strawberry (Fragaria xananassa) fruits and evaluation of their utility in genetic and molecular studies

<p>Abstract</p> <p>Background</p> <p>Cultivated strawberry is a hybrid octoploid species (<it>Fragaria xananassa </it>Duchesne ex. Rozier) whose fruit is highly appreciated due to its organoleptic properties and health benefits. Despite recent studies on the control of its growth and ripening processes, information about the role played by different hormones on these processes remains elusive. Further advancement of this knowledge is hampered by the limited sequence information on genes from this species, despite the abundant information available on genes from the wild diploid relative <it>Fragaria vesca</it>. However, the diploid species, or one ancestor, only partially contributes to the genome of the cultivated octoploid. We have produced a collection of expressed sequence tags (ESTs) from different cDNA libraries prepared from different fruit parts and developmental stages. The collection has been analysed and the sequence information used to explore the involvement of different hormones in fruit developmental processes, and for the comparison of transcripts in the receptacle of ripe fruits of diploid and octoploid species. The study is particularly important since the commercial fruit is indeed an enlarged flower receptacle with the true fruits, the achenes, on the surface and connected through a network of vascular vessels to the central pith.</p> <p>Results</p> <p>We have sequenced over 4,500 ESTs from <it>Fragaria xananassa</it>, thus doubling the number of ESTs available in the GenBank of this species. We then assembled this information together with that available from <it>F. xananassa </it>resulting a total of 7,096 unigenes. The identification of SSRs and SNPs in many of the ESTs allowed their conversion into functional molecular markers. The availability of libraries prepared from green growing fruits has allowed the cloning of cDNAs encoding for genes of auxin, ethylene and brassinosteroid signalling processes, followed by expression studies in selected fruit parts and developmental stages. In addition, the sequence information generated in the project, jointly with previous information on sequences from both <it>F. xananassa </it>and <it>F. vesca</it>, has allowed designing an oligo-based microarray that has been used to compare the transcriptome of the ripe receptacle of the diploid and octoploid species. Comparison of the transcriptomes, grouping the genes by biological processes, points to differences being quantitative rather than qualitative.</p> <p>Conclusions</p> <p>The present study generates essential knowledge and molecular tools that will be useful in improving investigations at the molecular level in cultivated strawberry (<it>F. xananassa</it>). This knowledge is likely to provide useful resources in the ongoing breeding programs. The sequence information has already allowed the development of molecular markers that have been applied to germplasm characterization and could be eventually used in QTL analysis. Massive transcription analysis can be of utility to target specific genes to be further studied, by their involvement in the different plant developmental processes.</p

Clocking the assembly of double-barred galaxies with the MUSE TIMER project

The formation of two stellar bars within a galaxy has proved challenging for numerical studies. It is not yet clear whether the inner bar is born via a star formation process promoted by gas inflow along the outer bar or whether it is dynamically assembled from instabilities in a small-scale stellar disc. Observational constraints to these scenarios are scarce. We present a thorough study of the stellar content of two double-barred galaxies observed by the MUSE TIMER project, NGC 1291 and NGC 5850, combined with a two-dimensional multicomponent photometric decomposition performed on the 3.6 μμm images from S^4G. Our analysis confirms the presence of σ-hollows appearing in the stellar velocity dispersion distribution at the ends of the inner bars. Both galaxies host inner discs matching in size with the inner bars, suggestive of a dynamical formation for the inner bars from small-scale discs. The analysis of the star formation histories for the structural components shaping the galaxies provides constraints on the epoch of dynamical assembly of the inner bars, which took place >6.5 Gyr ago for NGC 1291 and >4.5 Gyr ago for NGC 5850. This implies that inner bars are long-lived structures

Identificación y caracterización de genes implicados en la maduración y la calidad de la fresa

Ripening is a critical step for the development of flavor quality in fruits. This character has significantly declined in many fleshy fruits over recent decades. This is particularly significant in strawberry (Fragaria × ananassa), where current cultivars are derived from a narrow germplasm collection. Improving fruit quality requires two important breakthroughs: 1) a precise understanding of the fruit ripening process that will allow the targeting of relevant genes, and 2) the identification of novel alleles responsible for fruit quality traits. In our project (TRANSFR-Q, Starting Grant-ERC), we aim at the identification and characterization of key transcription factors involved in fruit ripening regulation and their target genes, in order to infer the Gene Regulatory Network controlling this process. On the other hand, we are carrying out a Genome-Wide Association Study using a germplasm collection of the woodland strawberry (Fragaria vesca) in order to identify loci involved in important traits such as aroma, fruit size, and resistance to pathogens. Finally, we have implemented the use of the genome-editing tool CRISPR/Cas9 in the cultivated strawberry, which we expect to open opportunities for engineering this species to improve traits of economic importance.ERC Starting Grant ERC-2014-StG 63813

Transcriptional regulatory network controlling strawberry fruit ripening and quality

Ripening is a critical step for the development of flavor quality in fruits. This character has significantly declined in many fleshy fruits over recent decades. This is particularly significant in strawberry (Fragaria × ananassa), where current cultivars are derived from a narrow germplasm collection. Improving fruit quality requires two important breakthroughs: 1) a precise understanding of the fruit ripening process that will allow the targeting of relevant genes, and 2) the identification of novel alleles responsible for fruit quality traits. In our project, we aim at the identification and characterization of key transcription factors involved in fruit ripening regulation and their target genes, in order to infer the Gene Regulatory Network controlling this process. On the other hand, we are carrying out a Genome-Wide Association Study using a germplasm collection of the woodland strawberry (Fragaria vesca) in order to identify loci involved in important traits such as aroma, fruit size or resistance to pathogens. Finally, we have implemented the use of the genome-editing tool CRISPR/Cas9 in the cultivated strawberry, which we expect it might open opportunities for engineering this species to improve traits of economic importance.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Study of Transcriptional Regulatory Network Controlling Strawberry Fruit Ripening and Quality

Ripening is a critical step for the development of flavor quality in fruits. This character has significantly declined in many fleshy fruits over recent decades. This is particularly significant in strawberry (Fragaria × ananassa), where current cultivars are derived from a narrow germplasm collection. Improving fruit quality requires two important breakthroughs: 1) a precise understanding of the fruit ripening process that will allow the targeting of relevant genes, and 2) the identification of novel alleles responsible for fruit quality traits. In our project we aim at the identification and characterization of key transcription factors involved in fruit ripening regulation and their target genes, in order to infer the Gene Regulatory Network controlling this process. On the other hand, we are carrying out a Genome-Wide Association Study using a germplasm collection of the woodland strawberry (Fragaria vesca) in order to identify loci involved in important traits such as aroma, fruit size, and resistance to pathogens. Finally, we have implemented the use of the genome-editing tool CRISPR/Cas9 in the cultivated strawberry, which we expect to open opportunities for engineering this species to improve traits of economic importance.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tec

Characterizing the involvement of FaMADS9 in the regulation of strawberry fruit receptacle development

FaMADS9 is the strawberry (Fragaria x ananassa) gene that exhibits the highest homology to the tomato (Solanum lycopersicum) RIN gene. Transgenic lines were obtained in which FaMADS9 was silenced. The fruits of these lines did not show differences in basic parameters, such as fruit firmness or colour, but exhibited lower Brix values in three of the four independent lines. The gene ontology MapMan category that was most enriched among the differentially expressed genes in the receptacles at the white stage corresponded to the regulation of transcription, including a high percentage of transcription factors and regulatory proteins associated with auxin action. In contrast, the most enriched categories at the red stage were transport, lipid metabolism and cell wall. Metabolomic analysis of the receptacles of the transformed fruits identified significant changes in the content of maltose, galactonic acid-1,4-lactone, proanthocyanidins and flavonols at the green/white stage, while isomaltose, anthocyanins and cuticular wax metabolism were the most affected at the red stage. Among the regulatory genes that were differentially expressed in the transgenic receptacles were several genes previously linked to flavonoid metabolism, such as MYB10, DIV, ZFN1, ZFN2, GT2, and GT5, or associated with the action of hormones, such as abscisic acid, SHP, ASR, GTE7 and SnRK2.7. The inference of a gene regulatory network, based on a dynamic Bayesian approach, among the genes differentially expressed in the transgenic receptacles at the white and red stages, identified the genes KAN1, DIV, ZFN2 and GTE7 as putative targets of FaMADS9. A MADS9-specific CArG box was identified in the promoters of these genes

Strawberry GRN forever: insights into the transcriptional regulatory network controlling strawberry fruit ripening and quality

Ripening is a critical step for the development of flavor quality in fruits. This character has significantly declined in many fleshy fruits over recent decades. This is particularly significant in strawberry (Fragaria × ananassa), where current cultivars are derived from a narrow germplasm collection. Improving fruit quality requires two important breakthroughs: 1) a precise understanding of the fruit ripening process that will allow the targeting of relevant genes, and 2) the identification of novel alleles responsible for fruit quality traits. In our project, we aim at the identification and characterization of key transcription factors (TF) involved in fruit ripening regulation and their target genes, in order to infer the Gene Regulatory Network controlling this process. Among them, we have identified two TFs belonging to the NAC (FaRIF) and the BLH9 (FaRPL) family. Functional analyses establishing stable silencing and overexpression lines support that both TFs play a critical role in the regulation of fruit ripening and development. Furthermore, using a stage- and tissue-specific transcriptome analysis, we have identified TFs specifically expressed in the external layer of ripe receptacles of F. vesca fruits, which are involved in the regulation of wax and cuticle formation. Finally, we have implemented the use of the genome-editing tool CRISPR/Cas9 in the cultivated strawberry, which we expect to open opportunities for engineering this species to improve traits of economic importance