Abscisic acid controls sugar accumulation essential to strawberry fruit ripening via the FaRIPK1-FaTCP7-FaSTP13/FaSPT module.

Strawberry is considered as a model plant for studying the ripening of abscisic acid (ABA)-regulated non-climacteric fruits, a process in which sugar plays a fundamental role, while how ABA regulates sugar accumulation remains unclear. This study provides a direct line of physiological, biochemical, and molecular evidence that ABA signaling regulates sugar accumulation via the FaRIPK1-FaTCP7-FaSTP13/FaSPT signaling pathway. Herein, FaRIPK1, a red-initial protein kinase 1 previously identified in strawberry fruit, not only interacted with the transcription factor FaTCP7 (TEOSINTE BRANCHEN 1, CYCLOIDEA, and PCF) but also phosphorylated the critical Ser89 and Thr93 sites of FaTCP7, which negatively regulated strawberry fruit ripening, as evidenced by the transient overexpression (OE) and virus-induced gene silencing transgenic system. Furthermore, the DAP-seq experiments revealed that FvTCP7 bound the motif "GTGGNNCCCNC" in the promoters of two sugar transporter genes, FaSTP13 (sugar transport protein 13) and FaSPT (sugar phosphate/phosphate translocator), inhibiting their transcription activities as determined by the electrophoretic mobility shift assay, yeast one-hybrid, and dual-luciferase reporter assays. The downregulated FaSTP13 and FaSPT transcripts in the FaTCP7-OE fruit resulted in a reduction in soluble sugar content. Consistently, the yeast absorption test revealed that the two transporters had hexose transport activity. Especially, the phosphorylation-inhibited binding of FaTCP7 to the promoters of FaSTP13 and FaSPT could result in the release of their transcriptional activities. In addition, the phosphomimetic form FaTCP7S89D or FaTCP7T93D could rescue the phenotype of FaTCP7-OE fruits. Importantly, exogenous ABA treatment enhanced the FaRIPK1-FaTCP7 interaction. Overall, we found direct evidence that ABA signaling controls sugar accumulation during strawberry fruit ripening via the "FaRIPK1-FaTCP7-FaSTP13/FaSPT" module.

Epigenomic and transcriptomic persistence of heat stress memory in strawberry (Fragaria vesca).

BACKGROUND: In plants, epigenetic stress memory has so far been found to be largely transient. Here, we wanted to assess the heritability of heat stress-induced epigenetic and transcriptomic changes following woodland strawberry (Fragaria vesca) reproduction. Strawberry is an ideal model to study epigenetic inheritance because it presents two modes of reproduction: sexual (self-pollinated plants) and asexual (clonally propagated plants named daughter plants). Taking advantage of this model, we investigated whether heat stress-induced DNA methylation changes can be transmitted via asexual reproduction. RESULTS: Our genome-wide study provides evidence for stress memory acquisition and maintenance in F. vesca. We found that specific DNA methylation marks or epimutations are stably transmitted over at least three asexual generations. Some of the epimutations were associated with transcriptional changes after heat stress. CONCLUSION: Our findings show that the strawberry methylome and transcriptome respond with a high level of flexibility to heat stress. Notably, independent plants acquired the same epimutations and those were inherited by their asexual progenies. Overall, the asexual progenies can retain some information in the genome of past stresses encountered by their progenitors. This molecular memory, also documented at the transcriptional level, might be involved in functional plasticity and stress adaptation. Finally, these findings may contribute to novel breeding approaches for climate-ready plants.

A histone deacetylase, FaSRT1-2, plays multiple roles in regulating fruit ripening, plant growth and stresses resistance of cultivated strawberry.

Sirtuins (SRTs) are a group of nicotinamide adenine dinucleotide (NAD+)-dependent deacetylase that target both histone and nonhistone proteins. The biological function of SRT in horticultural plants has been rarely studied. In this study, FaSRT1-2 was identified as a key member of the 8 FaSRTs encoded in cultivated strawberry genome. Transient overexpression of FaSRT1-2 in strawberry fruit accelerated ripening, increased the content of anthocyanins and sugars, enhanced ripening-related gene expression. Moreover, stable transformation of FaSRT1-2 in strawberry plants resulted in enhanced vegetative growth, increased sensitivity to heat stress and increased susceptibility to Botrytis cinerea infection. Interestingly, knocking out the homologous gene in woodland strawberry had the opposite effects. Additionally, we found the content of stress-related hormone abscisic acid (ABA) was decreased, while the growth-related gibberellin (GA) concentration was increased in FaSRT1-2 overexpression lines. Gene expression analysis revealed induction of heat shock proteins, transcription factors, stress-related and antioxidant genes in the FaSRT1-2-overexpressed plants while knocked-out of the gene had the opposite impact. In conclusion, our findings demonstrated that FaSRT1-2 could positively promote strawberry plant vegetative growth and fruit ripening by affecting ABA and GA pathways. However, it negatively regulates the resistance to heat stress and B. cinerea infection by influencing the related gene expression.

Strawberry phenotypic plasticity in flowering time is driven by the interaction between genetic loci and temperature.

Flowering time (FT), which determines when fruits or seeds can be harvested, is subject to phenotypic plasticity, that is, the ability of a genotype to display different phenotypes in response to environmental variation. Here, we investigated how the environment affects the genetic architecture of FT in cultivated strawberry (Fragaria × ananassa) and modifies its quantitative trait locus (QTL) effects. To this end, we used a bi-parental segregating population grown for 2 years at widely divergent latitudes (five European countries) and combined climatic variables with genomic data (Affymetrix SNP array). Examination, using different phenological models, of the response of FT to photoperiod, temperature, and global radiation indicated that temperature is the main driver of FT in strawberry. We next characterized in the segregating population the phenotypic plasticity of FT by using three statistical approaches that generated plasticity parameters including reaction norm parameters. We detected 25 FT QTLs summarized as 10 unique QTLs. Mean values and plasticity parameter QTLs were co-localized in three of them, including the major 6D_M QTL whose effect is strongly modulated by temperature. The design and validation of a genetic marker for the 6D_M QTL offers great potential for breeding programs, for example selecting early-flowering strawberry varieties well adapted to different environmental conditions.

Cold tolerance of woodland strawberry (Fragaria vesca) is linked to Cold Box Factor 4 and the dehydrin Xero2.

Domesticated strawberry is susceptible to sudden frost episodes, limiting the productivity of this cash crop in regions where they are grown during early spring. In contrast, the ancestral woodland strawberry (Fragaria vesca) has successfully colonized many habitats of the Northern Hemisphere. Thus, this species seems to harbour genetic factors promoting cold tolerance. Screening a germplasm established in the frame of the German Gene Bank for Crop Wild Relatives, we identified, among 70 wild accessions, a pair with contrasting cold tolerance. By following the physiological, biochemical, molecular, and metabolic responses of this contrasting pair, we identified the transcription factor Cold Box Factor 4 and the dehydrin Xero2 as molecular markers associated with superior tolerance to cold stress. Overexpression of green fluorescent protein fusions with Xero2 in tobacco BY-2 cells conferred cold tolerance to these recipient cells. A detailed analysis of the metabolome for the two contrasting genotypes allows the definition of metabolic signatures correlated with cold tolerance versus cold stress. This work provides a proof-of-concept for the value of crop wild relatives as genetic resources to identify genetic factors suitable to increase the stress resilience of crop plants.

Woodland strawberry axillary bud fate is dictated by a crosstalk of environmental and endogenous factors.

Plant architecture is defined by fates and positions of meristematic tissues and has direct consequences on yield potential and environmental adaptation of the plant. In strawberries (Fragaria vesca L. and F. × ananassa Duch.), shoot apical meristems can remain vegetative or differentiate into a terminal inflorescence meristem. Strawberry axillary buds (AXBs) are located in leaf axils and can either remain dormant or follow one of the two possible developmental fates. AXBs can either develop into stolons needed for clonal reproduction or into branch crowns (BCs) that can bear their own terminal inflorescences under favorable conditions. Although AXB fate has direct consequences on yield potential and vegetative propagation of strawberries, the regulation of AXB fate has so far remained obscure. We subjected a number of woodland strawberry (F. vesca L.) natural accessions and transgenic genotypes to different environmental conditions and growth regulator treatments to demonstrate that strawberry AXB fate is regulated either by environmental or endogenous factors, depending on the AXB position on the plant. We confirm that the F. vesca GIBBERELLIN20-oxidase4 (FvGA20ox4) gene is indispensable for stolon development and under tight environmental regulation. Moreover, our data show that apical dominance inhibits the outgrowth of the youngest AXB as BCs, although the effect of apical dominance can be overrun by the activity of FvGA20ox4. Finally, we demonstrate that the FvGA20ox4 is photoperiodically regulated via FvSOC1 (F. vesca SUPPRESSOR OF OVEREXPRESSION OF CONSTANS1) at 18°C, but at higher temperature of 22°C an unidentified FvSOC1-independent pathway promotes stolon development.

Physiological and biochemical responses of strawberry crown and leaf tissues to freezing stress.

BACKGROUND: In northern Iran and other cold regions, winter freezing injury and resultant yield instability are major limitations to strawberry production. However, there is scarcity of information on the physiological and biochemical responses of strawberry cultivars to freezing stress. This study aimed to investigate the physiological and biochemical responses of strawberry cultivars (Tennessee Beauty, Blakemore, Kurdistan, Queen Elisa, Chandler, Krasnyy Bereg, and Yalova) to different freezing temperature treatments (- 5, - 10, - 15, - 20, and - 25 °C) under controlled conditions. RESULTS: All measured physiological and biochemical features were significantly affected by the interaction effect between low temperatures and cultivars. Tennessee Beauty showed the highest RWC at - 25 °C. The highest Fv/Fm was observed in Queen Elisa. Krasnyy Bereg had the least freezing injury (FI) in crown and leaf, while Yalova and Chandler showed the highest crown and leaf FI, respectively. At - 20 to - 25 °C, the highest carbohydrates contents of crown and leaf were noted in Blakemore and Krasnyy Bereg cultivars, respectively. The Yalova showed the highest protein content in both crown and leaf tissues at - 25 °C. The Tennessee Beauty and Blackmore cultivars showed the highest proline in crowns and leaves at - 15 °C, respectively. The highest ThioBarbituric Acid Reactive Substances (TBARS) contents in the crown and leaf were observed in Kurdistan and Queen Elisa, respectively. Queen Elisa and Krasnyy Bereg cultivars showed SOD and POD peaks in the crown at - 15 °C, respectively. CONCLUSION: Freezing stress was characterized by decreased Fv/Fm and RWC, and increased FI, TBARS, total carbohydrates, total proteins, proline content, and antioxidant enzyme activity. The extent of changes in above mentioned traits was cultivar dependent. FI and TBARS were the best traits among destructive parameters for evaluating freezing tolerance. Moreover, maximum quantum yield of PSII (Fv/Fm index), as non-destructive parameters, showed a significant efficiency in rapid assessment for screening of freezing tolerant strawberry cultivars. The cultivars Krasnyy Bereg, Queen Elisa, and Kurdistan were the most tolerant cultivars to freezing stress. These cultivars can be used as parents in breeding programs to develop new freezing tolerant cultivars.

Genome-wide identification of AP2/EREBP in Fragaria vesca and expression pattern analysis of the FvDREB subfamily under drought stress.

BACKGROUND: Drought is a common phenomenon worldwide. It is also one of the main abiotic factors that affect the growth and quality of strawberry. The dehydration-responsive element binding protein (DREB) members that belong to the APETALA2/ethylene-responsive element binding protein (AP2/EREBP) superfamily are unique transcription factors in plants that play important roles in the abiotic stress response. RESULTS: Here, a total of 119 AP2/EREBP genes were identified in Fragaria vesca, and the AP2/EREBP superfamily was divided into AP2, RAV, ERF, DREB, and soloist subfamilies, containing 18, 7, 61, 32, and one member(s), respectively. The DREB subfamily was further divided into six subgroups (A-1 to A-6) based on phylogenetic analysis. Gene structure, conserved motifs, chromosomal location, and synteny analysis were conducted to comprehensively investigate the characteristics of FvDREBs. Furthermore, transcriptome analysis revealed distinctive expression patterns among the FvDREB genes in strawberry plants exposed to drought stress. The expression of FvDREB6 of the A-2 subgroup was down-regulated in old leaves and up-regulated in young leaves in response to drought. Furthermore, qRT-PCR analysis found that FvDREB8 from the A-2 subgroup had the highest expression level under drought stress. Together, analyses with the expression pattern, phylogenetic relationship, motif, and promoter suggest that FvDREB18 may play a critical role in the regulation of FvDREB1 and FvDREB2 expression. CONCLUSIONS: Our findings provide new insights into the characteristics and potential functions of FvDREBs. These FvDREB genes should be further studied as they appear to be excellent candidates for drought tolerance improvement of strawberry.

Heterologous expression of Arabidopsis thaliana rty gene in strawberry (Fragaria × ananassa Duch.) improves drought tolerance.

BACKGROUND: Strawberry (Fragaria × ananassa Duch.) is an important fruit crop worldwide. It was particularly sensitive to drought stress because of their fibrous and shallow root systems. Mutant rty of Arabidopsis thaliana ROOTY (RTY) results in increased endogenous auxin levels, more roots, and shoot growth. It is still unclear whether the rty gene improves stress tolerance in strawberry. RESULTS: rty gene was isolated from Arabidopsis thaliana and placed under the control of the cauliflower mosaic virus (CaMV) 35S promoter in the pBI121-rty binary vector carrying the selectable marker of neomycin phosphotransferase II (NPT II). Seven transgenic lines were confirmed by PCR and western blot analysis. Accumulations of IAA and ABA were significantly increased in the transgenic plants. The endogenous IAA contents were 46.5 ng g- 1 and 66.0 ng g- 1in control and transgenic plants respectively. The endogenous ABA contents in the control plant were 236.3 ng g- 1 and in transgenic plants were 543.8 ng g- 1. The production of adventitious roots and trichomes were enhanced in the transgenic plants. Furthermore, transcript levels of the genes including IAA and ABA biosynthetic, and stress-responsive genes, were higher in the transgenic plants than in the control plants under drought conditions. Water use efficiency and a reduced water loss rate were enhanced in the transgenic strawberry plants. Additionally, peroxidase and catalase activities were significantly higher in the transgenic plants than in the control plants. The experiment results revealed a novel function for rty related to ABA and drought responses. CONCLUSIONS: The rty gene improved hormone-mediated drought tolerance in transgenic strawberry. The heterologous expression of rty in strawberry improved drought tolerance by promoting auxin and ABA accumulation. These phytohormones together brought about various physiological changes that improved drought tolerance via increased root production, trichome density, and stomatal closure. Our results suggested that a transgenic approach can be used to overcome the inherent trade-off between plant growth and drought tolerance by enhancing water use efficiency and reducing water loss rate under water shortage conditions.

Salicylic acid-primed defence response in octoploid strawberry 'Benihoppe' leaves induces resistance against Podosphaera aphanis through enhanced accumulation of proanthocyanidins and upregulation of pathogenesis-related genes.

BACKGROUND: Podosphaera aphanis, a predominately biotrophic fungal pathogen, causes significant yield losses of strawberry. China is the largest strawberry producer in the world, and selecting for powdery mildew-resistant cultivars is desirable. However, the resistance mechanism against P. aphanis in the octoploid strawberry remains unclear. RESULTS: To understand possible mechanisms of disease resistance, we inoculated strawberry leaves with P. aphanis, and examined the expression profiles of candidate genes and the biochemical phenotypes in strawberry leaves of two groups. The unigenes obtained from ddH2O- and SA-pretreated leaves resulted in a total of 48,020 and 45,896 genes, respectively. KEGG enrichment showed that phenylpropanoid biosynthesis and plant hormone signal transduction pathways were enriched to a noticeable extent. DEG analysis showed that key TFs genes associated with the SA signaling pathway could play important role in the strawberry-P. aphanis interaction. In particular, FaWRKY70, FaJAZ1 and FaMYC2-like, involved in regulating the antagonistic effect of SA and JA signaling pathway, leading to increased expression of SA-responsive genes (in particular PR1, PR2, PR3, and PR5) compared to a decline in expression of JA-responsive genes (FaJAR1, FaAOS, and FaLOX2). Furthermore, SA pretreatment induced accumulation of PAs by activating the MBW complex and inhibit powdery mildew growth. CONCLUSIONS: This study describes the role of the proanthocyanidins (PAs), pathogenesis-related (PR) genes, SA, and transcription factors in regulatory model against P. aphanis, which coincided with an early activation of defense, leading to the accumulation of PAs and the PR proteins.

Beyond rest and quiescence (endodormancy and ecodormancy): A novel model for quantifying plant-environment interaction in bud dormancy release.

Bud dormancy of plants has traditionally been explained either by physiological growth arresting conditions in the bud or by unfavourable environmental conditions, such as non-growth-promoting low air temperatures. This conceptual dichotomy has provided the framework also for developing process-based plant phenology models. Here, we propose a novel model that in addition to covering the classical dichotomy as a special case also allows the quantification of an interaction of physiological and environmental factors. According to this plant-environment interaction suggested conceptually decades ago, rather than being unambiguous, the concept of "non-growth-promoting low air temperature" depends on the dormancy status of the plant. We parameterized the model with experimental results of growth onset for seven boreal plant species and found that based on the strength of the interaction, the species can be classified into three dormancy types, only one of which represents the traditional dichotomy. We also tested the model with four species in an independent experiment. Our study suggests that interaction of environmental and physiological factors may be involved in many such phenomena that have until now been considered simply as plant traits without any considerations of effects of the environmental factors.

Exogenous 5-aminolevulinic acid improves strawberry tolerance to osmotic stress and its possible mechanisms.

Cultivated strawberry, one of the major fruit crops worldwide, is an evergreen plant with shallow root system, and thus sensitive to environmental changes, including drought stress. To investigate the effect of 5-aminolevulinic acid (ALA), a new environment-friendly plant growth regulator, on strawberry drought tolerance and its possible mechanisms, we treated strawberry (Fragaria × annanasa Duch. cv. 'Benihoppe') with 15% polyethylene glycol 6000 to simulate osmotic stress with or without 10 mg l-1 ALA. We found that ALA significantly alleviated PEG-inhibited plant growth and improved water absorption and xylem sap flux, indicating ALA mitigates the adverse effect of osmotic stress on strawberry plants. Gas exchange and chlorophyll fluorescence analysis showed that ALA mitigated PEG-induced decreases of Pn , Gs , Tr , Pn /Ci , photosystem I and II reaction center activities, electron transport activity, and photosynthetic performance indexes. Equally important, ALA promoted PEG-increased antioxidant enzyme activities and repressed PEG-increased malondialdehyde and superoxide anion in both leaves and roots. Specially, ALA repressed H2 O2 increase in leaves, but stimulated it in roots. Furthermore, ALA repressed abscisic acid (ABA) biosynthesis and signaling gene expressions in leaves, but promoted those in roots. In addition, ALA blocked PEG-downregulated expressions of plasmalemma and tonoplast aquaporin genes PIP and TIP in both leaves and roots. Taken together, ALA effectively enhances strawberry drought tolerance and the mechanism is related to the improvement of water absorption and conductivity. The tissue-specific responses of ABA biosynthesis, ABA signaling, and H2 O2 accumulation to ALA in leaves and roots play key roles in ALA-improved strawberry tolerance to osmotic stress.

Compatibility of pesticides used in strawberry crops with predatory mites Stratiolaelaps scimitus (Womersley) and Cosmolaelaps brevistilis (Karg).

Stratiolaelaps scimitus (Womersley) and Cosmolaelaps brevistilis (Karg) (Acari: Laelapidae) are predatory mites of soil-inhabiting pests, mainly small insects. Fungus gnats fly species are found in greenhouse strawberry production and may be controlled with predatory mites, being important to know their compatibility with the pesticides used in strawberry crops. In this study, the compatibility of seven commercial pesticides used in strawberry cultivation with the predatory mites S. scimitus and C. brevistilis was assessed in laboratory conditions. Survival and oviposition rates were evaluated between 0.5 and 120 h after treatment (HAT). The results demonstrate that lambda-cyhalothrin treatment resulted in the lowest survival rate for both mites in the first evaluations, being moderately harmful, while spinetoran was slightly harmful to C. brevistilis. On the other hand, abamectin, azadirachtin, azoxystrobin + difenoconazole, iprodione and thiamethoxam were harmless for both mites and, oviposition rate was significantly different only at 72 and 120 HAT for S. scimitus and C. brevistilis respectively. These results may be used to develop guidelines for the adoption of selective pesticides in integrated pest management programs that conserves predatory mites.

Classification of Dried Strawberry by the Analysis of the Acoustic Sound with Artificial Neural Networks.

In this paper, the authors used an acoustic wave acting as a disturbance (acoustic vibration), which travelled in all directions on the whole surface of a dried strawberry fruit in its specified area. The area of space in which the acoustic wave occurs is defined as the acoustic field. When the vibrating surface-for example, the surface of the belt-becomes the source, then one can observe the travelling of surface waves. For any shape of the surface of the dried strawberry fruit, the signal of travelling waves takes the form that is imposed by this irregular surface. The aim of this work was to research the effectiveness of recognizing the two trials in the process of convection drying on the basis of the acoustic signal backed up by neural networks. The input variables determined descriptors such as frequency (Hz) and the level of luminosity (dB). During the research, the degree of crispiness relative to the degree of maturity was compared. The results showed that the optimal neural model in respect of the lowest value of the root mean square turned out to be the Multi-Layer Perceptron network with the technique of dropping single fruits into water (data included in the learning data set Z2). The results confirm that the choice of method can have an influence on the effectives of recognizing dried strawberry fruits, and also this can be a basis for creating an effective and fast analysis tool which is capable of analyzing the degree of ripeness of fruits including their crispness in the industrial process of drying fruits.

Phenotypic analysis combined with tandem mass tags (TMT) labeling reveal the heterogeneity of strawberry stolon buds.

BACKGROUND: Ramet propagation in strawberry (Fragaria × ananassa) is the most effective way in production. However, the lack of systematically phenotypic observations and high-throughput methods limits our ability to analyze the key factors regulating the heterogeneity in strawberry stolon buds. RESULTS: From observation, we found that the axillary bud located in the first node quickly stepped into dormancy (DSB), after several bract and leaf buds were differentiated. The stolon apical meristem (SAM) degenerated as the new ramet leaf buds (RLB), and the new active axillary stolon buds (ASB) differentiated continually after the differentiation of the first leaf. Using the tandem mass tags (TMT) labeling method, a total of 7271 strawberry proteins were identified. Between ASB and DSB, the spliceosome DEPs, such as Ser/Arg-rich (SR) and heterogeneous nuclear ribonucleoprotein particle (hnRNP), showed the highest enrichment and high PPI connectivity. This indicated that the differences in DEPs (e.g., SF-3A and PK) at the transcriptional level may be causing the differences between the physiological statuses of ASB and DSB. As expected, the photosynthetic pre-form RLB mainly differentiated from ASB and DSB judging by the DEP enrichment of photosynthesis. However, there are still other specialized features of DEPs between RLB and DSB and between ASB and DSB. The DEPs relative to DNA duplication [e.g., minichromosome maintenance protein (MCM 2, 3, 4, 7)], provide a strong hint of functional gene duplication leading the bud heterogeneity between RLB and DSB. In addition, the top fold change DEP of LSH 10-like might be involved in the degeneration of SAM into RLBs, based on its significant function in modulating the plant shoot initiation. As for RLB/ASB, the phenylpropanoid biosynthesis pathway probably regulates the ramet axillary bud specialization, and further promotes the differentiation of xylem when ASB develops into a new stolon [e.g., cinnamyl alcohol dehydrogenase 1 (CAD1) and phenylalanine ammonia-lyase 1 (PAL1)]. CONCLUSIONS: By using phenotypic observation combined with proteomic networks with different types of strawberry stolon buds, the definite dormancy phase of DSB was identified, and the biological pathways and gene networks that might be responsible for heterogeneity among different stolon buds in strawberry were also revealed.

An atypical HLH transcriptional regulator plays a novel and important role in strawberry ripened receptacle.

BACKGROUND: In soft fruits, the differential expression of many genes during development and ripening is responsible for changing their organoleptic properties. In strawberry fruit, although some genes involved in the metabolic regulation of the ripening process have been functionally characterized, some of the most studied genes correspond to transcription factors. High throughput transcriptomics analyses performed in strawberry red receptacle (Fragaria x ananassa) allowed us to identify a ripening-related gene that codes an atypical HLH (FaPRE1) with high sequence homology with the PACLOBUTRAZOL RESISTANCE (PRE) genes. PRE genes are atypical bHLH proteins characterized by the lack of a DNA-binding domain and whose function has been linked to the regulation of cell elongation processes. RESULTS: FaPRE1 sequence analysis indicates that this gene belongs to the subfamily of atypical bHLHs that also includes ILI-1 from rice, SlPRE2 from tomato and AtPRE1 from Arabidopsis, which are involved in transcriptional regulatory processes as repressors, through the blockage by heterodimerization of bHLH transcription factors. FaPRE1 presented a transcriptional model characteristic of a ripening-related gene with receptacle-specific expression, being repressed by auxins and activated by abscisic acid (ABA). However, its expression was not affected by gibberellic acid (GA3). On the other hand, the transitory silencing of FaPRE1 transcription by agroinfiltration in receptacle produced the down-regulation of a group of genes related to the ripening process while inducing the transcription of genes involved in receptacle growth and development. CONCLUSIONS: In summary, this work presents for the first time experimental data that support an important novel function for the atypical HLH FaPRE1 during the strawberry fruit ripening. We hypothesize that FaPRE1 modulates antagonistically the transcription of genes related to both receptacle growth and ripening. Thus, FaPRE1 would repress the expression of receptacle growth promoting genes in the ripened receptacle, while it would activate the expression of those genes related to the receptacle ripening process.

Transcriptome and hormone analyses provide insights into hormonal regulation in strawberry ripening.

MAIN CONCLUSION: The possible molecular mechanisms regulating strawberry fruit ripening were revealed by plant hormone quantification, exogenous hormone application, and RNA-sequencing. Fruit ripening involves a complex interplay among plant hormones. Strawberry is a model for studies on non-climacteric fruit ripening. However, the knowledge on how plant hormones are involved in strawberry ripening is still limited. To understand hormonal actions in the ripening process, we performed genome-wide transcriptome and hormonal analysis for the five major hormones (abscisic acid and catabolites, auxins, cytokinins, gibberellins, and ethylene) in achenes and receptacles (flesh) at different ripening stages of the woodland strawberry Fragaria vesca. Our results demonstrate that the pre-turning stage (a stage with white flesh and red achenes defined in this study) is the transition stage from immature to ripe fruits. The combinatorial analyses of hormone content, transcriptome data, and exogenous hormone treatment indicate that auxin is synthesized predominantly in achenes, while abscisic acid (ABA), bioactive free base cytokinins, gibberellins, and ethylene are mainly produced in receptacles. Furthermore, gibberellin may delay ripening, while ethylene and cytokinin are likely involved at later stages of the ripening process. Our results also provide additional evidence that ABA promotes ripening, while auxin delays it. Although our hormone analysis demonstrates that the total auxin in receptacles remains relatively low and unchanged during ripening, our experimental evidence further indicates that ABA likely enhances expression of the endoplasmic reticulum-localized auxin efflux carrier PIN-LIKES, which may subsequently reduce the auxin level in nucleus. This study provides a global picture for hormonal regulation of non-climacteric strawberry fruit ripening and also evidence for a possible mechanism of ABA and auxin interaction in the ripening process.

Transcriptome analysis of strawberry (Fragaria × ananassa) fruits under osmotic stresses and identification of genes related to ascorbic acid pathway.

Strawberry (Fragaria ananassa Duch.) is an economically important fruit with a high demand owing to its good taste and medicinal properties. However, its cultivation is affected by various biotic and abiotic stresses. Plants exhibit several intrinsic mechanisms to deal with stresses. In the case of strawberry, the mechanisms highlighting the response against these stresses remain to be elucidated, which has hampered the efforts to develop and cultivate strawberry plants with high yield and quality. Although a virtual reference genome of F. ananassa has recently been published, there is still a lack of information on the expression of genes in response to various stresses. Therefore, to provide molecular information for further studies with strawberry plants, we present the reference transcriptome dataset of F. ananassa, assembled and annotated from deep RNA-Seq data of fruits cultivated under salinity and drought stresses. We also systematically arranged a series of transcripts differentially expressed during these stresses, with an emphasis on genes related to the accumulation of ascorbic acid (AsA). Ascorbic acid is the most potent antioxidant present in these fruits and highly considered during biofortification. A comparison of the expression profile of these genes by RT-qPCR with the content of AsA in the fruits verified a tight regulation and balance between the expression of genes, from biosynthesis, degradation and recycling pathways, resulting in the reduced content of AsA in fruits under these stresses. These results provide a useful repertoire of genes for metabolic engineering, thereby improving the tolerance to stresses.

Mixotrophic in vitro cultivations: the way to go astray in plant physiology.

Rate of photosynthesis and related plant carbohydrate status are crucial factors affecting plant vigor. Sugars providing carbon and energy sources serve also as important signaling molecules governing plant growth and development through a complex regulatory network. These facts are often neglected when mixotrophic cultivation of plants in vitro is used, where artificial exogenous sugar supply hinders studies of metabolism as well as sugar-driven developmental processes. We compared the growth, selected gas-exchange parameters and sugar metabolism characteristics in four model plants, potato (Solanum tuberosum 'Lada'), tobacco (Nicotiana tabacum 'Samsun'), rapeseed (Brassica napus 'Asgard') and strawberry (Fragaria vesca), under both photomixotrophic (PM) and photoautotrophic (PA) conditions. To ensure PA conditions, we used our improved sun caps that serve as gas and light permeable covers for cultivation vessels. We found bigger biomass accumulation, larger leaf areas, higher stomatal conductance and higher instantaneous water use efficiency and lower root sugar contents in PA plants compared to PM ones. However, for other characteristics (root biomass, root/shoot ratio, pigment contents, leaf sugar and starch levels and transpiration rates), a strong species-dependent reactions to the exogenous sugar supply was noted, which does not allow to create a general view on the overall impact of PM nutrition under in vitro conditions.

Mycorrhizal frequency, physiological parameters, and yield of strawberry plants inoculated with endomycorrhizal fungi and rhizosphere bacteria.

Due to the impoverishment of agricultural and horticultural soils and replant diseases, there is a need to use bioproducts and beneficial microorganisms in order to improve the quality of soils and growth substrates. For this reason, research was undertaken to assess the impact of arbuscular mycorrhizal fungi and rhizosphere bacteria on changes in soil microbiology, the degree of colonization of plant roots by mycorrhizal fungi, selected physiological parameters, and fruit quality and yield of the strawberry cultivar "Rumba." The plants were inoculated with the mycorrhizal preparation Mykoflor (Rhizophagus irregularis, Funneliformis mosseae, Claroideoglomus etunicatum), MYC 800 (Rhizophagus intraradices), and the bacterial preparation Rhizocell C (Bacillus amyloliquefaciens IT45). The applied preparations increased the total number of bacteria and fungi in the soil and mycorrhizal frequency in the roots of the strawberry plants. They increased the chlorophyll "a" and total chlorophyll concentrations in the leaves as well as the rate of transpiration and CO2 concentration in the intercellular spaces in the leaves. The plants treated with Rhizocell C and MYC 800 exhibited a higher CO2 assimilation rate than control plants. The biopreparations increased chlorophyll fluorescence parameters such as maximum fluorescence (FM) and the maximum potential photochemical reaction efficiency in PS II (FV/FM). The influence of the species of rhizosphere bacteria and mycorrhizal fungi used in the experiment on the physiological traits of strawberry plants contributed, especially in the second year of the study, to increase the yield and mean weight of strawberry fruit.