Changes in the expression of genes encoding xanthophyl acyltransferases during the postharvest ripening of avocado (Persea americana) fruit.

BACKGROUND: Avocado fruit is rich in xanthophylls, which have been related to positive effects on human health. Xanthophyl acetyltransferases (XATs) are enzymes catalyzing the esterification of carboxylic acids to the hydroxyl group of the xanthophyll molecule. This esterification is thought to increase the lipophilic nature of the xanthophyll and its stability in a lipophilic environment. Studies on XATs in fruits are very scarce, and no studies had been carried out in avocado fruit during postharvest. The objective of this work was to investigate the changes in the expression of genes encoding XAT, during avocado fruit ripening. RESULTS: Avocado fruits were obtained from a local market and stored at 15 °C for 8 days. The fruit respiration rate, ethylene production, and fruit peel's color space parameters (L*, a*, b*) were measured during storage. Fruit mesocarp samples were taken after 1, 3, 5, and 7 days of storage and frozen with liquid nitrogen. Total RNA was extracted from fruit mesocarp, and the quantification of the two genes designated as COGE_ID: 936743791 and COGE_ID: 936800185 encoding XATs was performed with real-time quantitative reverse transcription polymerase chain reaction using actin as a reference gene. The presence of a climacteric peak and large changes in color were recorded during postharvest. The two genes studied showed a large expression after 3 days of fruit storage. CONCLUSIONS: We conclude that during the last stages of ripening in avocado fruit there was an active esterification of xanthophylls with carboxylic acids, which suggests the presence of esterified xanthophylls in the fruit mesocarp. © 2024 Society of Chemical Industry.

Plant long non-coding RNAs: identification and analysis to unveil their physiological functions.

Eukaryotic genomes encode thousands of RNA molecules; however, only a minimal fraction is translated into proteins. Among the non-coding elements, long non-coding RNAs (lncRNAs) play important roles in diverse biological processes. LncRNAs are associated mainly with the regulation of the expression of the genome; nonetheless, their study has just scratched the surface. This is somewhat due to the lack of widespread conservation at the sequence level, in addition to their relatively low and highly tissue-specific expression patterns, which makes their exploration challenging, especially in plant genomes where only a few of these molecules have been described completely. Recently published high-quality genomes of crop plants, along with new computational tools, are considered promising resources for studying these molecules in plants. This review briefly summarizes the characteristics of plant lncRNAs, their presence and conservation, the different protocols to find these elements, and the limitations of these protocols. Likewise, it describes their roles in different plant physiological phenomena. We believe that the study of lncRNAs can help to design strategies to reduce the negative effect of biotic and abiotic stresses on the yield of crop plants and, in the future, help create fruits and vegetables with improved nutritional content, higher amounts of compounds with positive effects on human health, better organoleptic characteristics, and fruits with a longer postharvest shelf life.

Transcriptome assessment in 'Red Globe' grapevine zygotic embryos during the cooling and warming phase of the cryopreservation procedure.

Cryopreservation has the potential for long-term germplasm storage. The metabolic pathways and gene regulation involved in cryopreservation procedures are still not well documented. Hence, the genetic expression profile was evaluated using RNA-Seq in zygotic embryos of grapevines subjected to cryopreservation by vitrification. Sequencing was performed on the Illumina NextSeq 500. The average alignment of reads was 96% against the reference genome. The expression profiles showed 229 genes differentially expressed (186 repressed and 46 induced). The main biological processes showing upregulated enrichment were related to nucleosome assembly, while downregulated processes were related to organ growth. The most highly repressed processes were associated with the organization of the cell wall and membrane components. The unnamed protein product and 17.3 kDa class II heat shock protein (HSP) were highly induced, while ATPase subunit 1 and expansin-A1 were repressed. The response to the cooling and warming process during cryopreservation probably indicates that the changes occurring in transcription may be related to epigenetics. In addition, the cell exhibits an increase in the reserve of nutrients while seeking to survive modestly using available energy and pausing the plant's development. Additionally, energy containment occurred to cope with the stress caused by the treatment where deactivation of components of the cell membrane was observed, possibly due to changes in fluidity caused by alterations in temperature.

Molecular Biology, Composition and Physiological Functions of Cuticle Lipids in Fleshy Fruits.

Fleshy fruits represent a valuable resource of economic and nutritional relevance for humanity. The plant cuticle is the external lipid layer covering the nonwoody aerial organs of land plants, and it is the first contact between fruits and the environment. It has been hypothesized that the cuticle plays a role in the development, ripening, quality, resistance to pathogen attack and postharvest shelf life of fleshy fruits. The cuticle's structure and composition change in response to the fruit's developmental stage, fruit physiology and different postharvest treatments. This review summarizes current information on the physiology and molecular mechanism of cuticle biosynthesis and composition changes during the development, ripening and postharvest stages of fleshy fruits. A discussion and analysis of studies regarding the relationship between cuticle composition, water loss reduction and maintaining fleshy fruits' postharvest quality are presented. An overview of the molecular mechanism of cuticle biosynthesis and efforts to elucidate it in fleshy fruits is included. Enhancing our knowledge about cuticle biosynthesis mechanisms and identifying specific transcripts, proteins and lipids related to quality traits in fleshy fruits could contribute to the design of biotechnological strategies to improve the quality and postharvest shelf life of these important fruit crops.

Toward Understanding the Molecular Recognition of Fungal Chitin and Activation of the Plant Defense Mechanism in Horticultural Crops.

Large volumes of fruit and vegetable production are lost during postharvest handling due to attacks by necrotrophic fungi. One of the promising alternatives proposed for the control of postharvest diseases is the induction of natural defense responses, which can be activated by recognizing molecules present in pathogens, such as chitin. Chitin is one of the most important components of the fungal cell wall and is recognized through plant membrane receptors. These receptors belong to the receptor-like kinase (RLK) family, which possesses a transmembrane domain and/or receptor-like protein (RLP) that requires binding to another RLK receptor to recognize chitin. In addition, these receptors have extracellular LysM motifs that participate in the perception of chitin oligosaccharides. These receptors have been widely studied in Arabidopsis thaliana (A. thaliana) and Oryza sativa (O. sativa); however, it is not clear how the molecular recognition and plant defense mechanisms of chitin oligosaccharides occur in other plant species or fruits. This review includes recent findings on the molecular recognition of chitin oligosaccharides and how they activate defense mechanisms in plants. In addition, we highlight some of the current advances in chitin perception in horticultural crops.

Abiotic stress hormesis: An approach to maintain quality, extend storability, and enhance phytochemicals on fresh produce during postharvest.

Postharvest losses of whole and fresh-cut fruits and vegetables cause significant reductions in food availability and an increase in economic losses/damages. Additionally, regulatory agencies are increasingly restricting the postharvest use of synthetic chemicals. This has strengthened the need to develop environmentally friendly approaches to postharvest management, such as utilization of natural compounds, antagonist microorganisms, and treatments with abiotic stresses, among others. The current review focuses on the potential of low doses of abiotic stresses to extend the shelf life, increase the amount of health beneficial phytochemicals, and reduce postharvest losses of fresh produce. The positive effects of the responses to low doses of abiotic stresses are based on a biological phenomenon termed hormesis. Research to develop new technologies to improve postharvest handling of fresh fruit and vegetables as well as minimally processed products is critical. The phenomenon of abiotic stress hormesis in fresh fruit and vegetables shows the potential not only to enhance defense compounds that could reduce diseases during postharvest storage and extend shelf life but also to elevate the content of health-promoting substances. The beneficial effects of UV-C hormesis have been extensively investigated in numerous types of fresh produce. However, our knowledge on hormesis exhibited by other abiotic stresses is still limited. Hence, the objective of this review is to discuss the relevance of hormesis for postharvest research by examining whether all abiotic stresses exhibit the phenomenon, its biological significance, the potential application in various commodities, and how it may direct the future of postharvest research.

Use of inductors in the control of Colletotrichum gloeosporioides and Rhizopus stolonifer isolated from soursop fruits: in vitro tests.

Soursop (Annona muricata) is a tropical fruit that can be infected by Colletotrichum gloeosporioides and Rhizopus stolonifer. Traditional methods used for postharvest disease control include the application of fungicides, however due to their excessive use, as well as their persistence in the environment, the development of new strategies that control pathogens are required. The application of chitosan (Chi), salicylic acid (SA) and methyl jasmonate (MJ) is an environmentally-friendly alternative with antimicrobial properties and also induces defense mechanisms in plant tissues. In this study, Colletotrichum was reactivated and Rhizopus was identified using morphological features and molecular tools. In vitro, the application of 0.5 and 1.0% of Chi alone or in combination with SA and MJ decreased mycelial growth and sporulation, a complete inhibition of spore germination was obtained. Thus, the application of Chi in combination with SA and MJ could be a smart strategy to inhibit the development of pathogens that attack soursop fruit.

Functional analysis of tomato rhamnogalacturonan lyase gene Solyc11g011300 during fruit development and ripening.

Rhamnogalacturonan I (RG-I) is a domain of plant cell wall pectin. The rhamnogalacturonan lyase (RGL) enzyme (EC 4.2.2.23) degrades RG-I by cleaving the α-1,4 glycosidic bonds located between the l-rhamnose and d-galacturonic residues of the main chain. While RGL's biochemical mode of action is well known, its effects on plant physiology remain unclear. To investigate the role of the RGL enzyme in plants, we have expressed the Solyc11g011300 gene under a constitutive promoter (CaMV35S) in tomato cv. 'Ohio 8245' and evaluated the expression of this and other RGL genes, enzymatic activity and alterations in vegetative tissue, and tomato physiology in transformed lines compared to the positive control (plants harboring the pCAMBIA2301 vector) and the isogenic line. The highest expression levels of the Solyc11g011300, Solyc04g076630, and Solyc04g076660 genes were observed in leaves and roots and at 10 and 20 days after anthesis (DAA). Transgenic lines exhibited lower RGL activity in leaves and roots and during fruit ripening, whereas higher activity was observed at 10, 20, and 30 DAA than in the isogenic line and positive control. Both transgenic lines showed a lower number of seeds and fruits, higher root length, and less pollen germination percentage and viability. In red ripe tomatoes, transgenic fruits showed greater firmness, longer shelf life, and reduced shriveling than did the isogenic line. Additionally, a delay of one week in fruit ripening in transgenic fruits was also recorded. Altogether, our data demonstrate that the Solyc11g011300 gene participates in pollen tube germination, fruit firmness, and the fruit senescence phenomena that impact postharvest shelf life.

Functional analysis of a tomato (Solanum lycopersicum L.) rhamnogalacturonan lyase promoter.

The enzyme rhamnogalacturonan lyase (RGL) cleaves α-1,4 glycosidic bonds located between rhamnose and galacturonic acid residues in the main chain of rhamnogalacturonan-I (RG-I), a component of the plant cell wall polymer pectin. Although the mode of action of RGL is well known, its physiological functions associated with fruit biology are less understood. Here, we generated transgenic tomato plants expressing the ß-glucuronidase (GUS) reporter gene under the control of a -504 bp or a -776 bp fragment of the promoter of a tomato RGL gene, Solyc11g011300. GUS enzymatic activity and the expression levels of GUS and Solyc11g011300 were measured in a range of organs and fruit developmental stages. GUS staining was undetectable in leaves and roots, but high GUS enzymatic activity was detected in flowers and red ripe (RR) fruits. Maximal expression levels of Solyc11g011300 were detected at the RR developmental stage. GUS activity was 5-fold higher in flowers expressing GUS driven by the -504 bp RGL promoter fragment (RGFL3::GUS) than in the isogenic line, and 1.7-fold higher when GUS gene was driven by the -776 bp RGL promoter fragment (RGLF2::GUS) or the constitutive CaMV35S promoter. Quantitative real-time polymerase chain reaction analysis showed that the highest expression of GUS was in fruits at 40 days after anthesis, for both promoter fragments. The promoter of Solyc11g011300 is predicted to contain cis-acting elements, and to be active in pollen grains, pollen tubes, flowers and during tomato fruit ripening, suggesting that the Solyc11g011300 promoter is transcriptionally active and organ-specific.

Analysis of a suppressive subtractive hybridization library of Alternaria alternata resistant to 2-propenyl isothiocyanate

Background Isothiocyanates (ITCs) are natural products obtained from plants of the Brassicas family. They represent an environmentally friendly alternative for the control of phytopathogenic fungi. However, as it has been observed with synthetic fungicides, the possibility of inducing ITC-resistant strains is a major concern. It is, therefore, essential to understanding the molecular mechanisms of fungal resistance to ITCs. We analyzed a subtractive library containing 180 clones of an Alternaria alternata strain resistant to 2-propenyl ITC (2-pITC). After their sequencing, 141 expressed sequence tags (ESTs) were identified using the BlastX algorithm. The sequence assembly was carried out using CAP3 software; the functional annotation and metabolic pathways identification were performed using the Blast2GO program. Results The bioinformatics analysis revealed 124 reads with similarities to proteins involved in transcriptional control, defense and stress pathways, cell wall integrity maintenance, detoxification, organization and cytoskeleton destabilization; exocytosis, transport, DNA damage control, ribosome maintenance, and RNA processing. In addition, transcripts corresponding to enzymes as oxidoreductases, transferases, hydrolases, lyases, and ligases, were detected. Degradation pathways for styrene, aminobenzoate, and toluene were induced, as well as the biosynthesis of phenylpropanoid and several types of N-glycan. Conclusions The fungal response showed that natural compounds could induce tolerance/resistance mechanisms in organisms in the same manner as synthetic chemical products. The response of A. alternata to the toxicity of 2-pITC is a sophisticated phenomenon including the induction of signaling cascades targeting a broad set of cellular processes. Whole-transcriptome approaches are needed to elucidate completely the fungal response to 2-pITC.

Utilization of physicochemical variables developed from changes in sensory attributes and consumer acceptability to predict the shelf life of fresh-cut mango fruit.

Sensory evaluation is the ideal tool for shelf-life determination. With the objective to develop an easy shelf-life indicator, color (L*, a*, b*, chroma and hue angle), total soluble solids (TSS), firmness (F), pH, acidity, and the sensory attributes of appearance, brightness, browning, odor, flavor, texture, color, acidity and sweetness were evaluated in fresh cut mangoes (FCM) stored at 5, 10, 15 and 20 °C. Overall acceptability was evaluated by consumers. Correlation analysis between sensory attributes and physicochemical variables was carried out. Physicochemical cut-off points based on sensory attributes and consumer acceptability was obtained by regression analysis and utilized to estimate FCM shelf-life by kinetic models fitted to each variable. The validation of the model was done by comparing the shelf life estimated by kinetic models and consumers. It was recorded large correlations between appearance, brightness, and color with L*; appearance and color with chroma and hue angle; sweetness and flavor with TSS, and between F and texture. The shelf life estimated based on consumer using a 9 point hedonic scale was in the range of 10-12, 2.3-2.6, 1.3-1.5 and 1.0-1.1 days for 5, 10, 15 and 20 °C. It was recorded large correlation coefficients between the shelf life estimated by consumer acceptability scores and physicochemical variables. Kinetic models based on physicochemical variables showed a tendency to overestimate the shelf life as compared with the models bases on the sensory attributes. It was concluded that physicochemical variables can be used as a tool to estimate the FCM shelf life.

Data on antioxidant activity in grapevine (Vitis vinifera L.) following cryopreservation by vitrification.

Cryopreservation is used for the long-term conservation of plant genetic resources. This technique very often induces lethal injury or tissue damage. In this study, we measured indicators of viability and cell damage following cryopreservation and vitrification-cryopreservation in Vitis vinifera L. axillary buds cv. "Flame seedless" stored in liquid nitrogen (LN) for: three seconds, one hour, one day, one week and one month; after LN thawed at 38 °C for three minutes. The enzymatic activity of catalase (CAT) and superoxide dismutase (SOD), as well as the amount of malondialdehyde (MDA), total protein and viability were assayed.

Induction of the expression of defence genes in Carica papaya fruit by methyl jasmonate and low temperature treatments

The defence mechanisms that are activated by methyl jasmonate (MJ) in fruits are not well understood. In this work, we studied the expression of defence genes in papaya fruit that are induced by the exposure to MJ and/or low temperatures. The papaya fruits ‘Maradol’ were randomly divided into two groups: one group was the untreated control and the other was treated with 10-4 M of MJ. Half of the fruits from each of the two groups were stored after treatment for 5 days at 5ºC and 2 days at 20ºC. We studied the expression levels of the pdf1.1 and pdf1.2 genes by amplification from expression libraries created from the pulp and skin tissues of the papaya fruit. As a reference, the mRNA level of the 18s ribosomal gene was used. In the skin tissue, the expression levels of the pdf1.1 and pdf1.2 genes were higher immediately after MJ treatment compared to the control. Furthermore, the expression of pdf1.2 remained high after MJ treatment and subsequent storage compared to the control. It was therefore concluded that the activation of the pdf1.1 and pdf1.2 genes forms part of the molecular defence mechanism in fruits that is activated by exposure to MJ. To our knowledge, this is the first study that analyzes the gene expression in papaya fruit that is induced by the exogenous application of methyl jasmonate and cold treatment.

Differentially expressed cDNAs in Alternaria alternata treated with 2-propenyl isothiocyanate.

The molecular mechanism of the fungal tolerance phenotype to fungicides is not completely understood. This knowledge would allow for the development of environmentally friendly strategies for the control of fungal infection. With the goal of determining genes induced by 2p-ITC, a forward suppressive subtractive hybridization was performed using cDNAs from ITC-treated Alternaria alternata as a "tester" and from untreated fungus as a "driver." Using this approach, a library containing 102 ESTs was generated that resulted in 50 sequences after sequence assembly (17 contigs and 33 singletons). Blastx analysis revealed that 38% and 40% of the sequences showed significant similarity with known and hypothetical proteins, respectively, whereas 18% were not similar to known genes. These last sequences could represent novel genes that play an unknown role in the molecular responses of fungi during adaptation to 2p-ITC. Clones similar to opsins, ABC transporters, calmodulin, ATPases and SNOG proteins were identified. Using real-time RT-PCR analysis, significant inductions of an ABC transporter and a Ca(++) ATPase during 2p-ITC treatment were discovered. These results suggest that the fungal resistance phenotype to 2p-ITC involves calcium ions and 2p-ITC efflux via an ABC transporter.

Isolation and study of a ubiquitously expressed tomato pectin methylesterase regulatory region

Pectin methylesterase (PME) is an enzyme located in the plant cell wall of higher plants whose physiological role is largely unknown. We had isolated a PME gene from a tomato genomic library, including 2.59 kb of 5üL flanking region and the coding region. Both coding and promoter region were sequenced and computer analyzed. Tobacco transgenic plants were created harboring constructs in which 2.596 Kb, 1.306 Kb and 0.267 Kb sizes of the promoter were driving the expression of âÀ-Glucuronidase gene (GUS). GUS activity was studied by histochemical and fluorometric assays. Two introns of 106 and 1039 bp were found in the coding region and phylogenetic analysis placed this PME gene closer to genes from Citrus sinensis and Arabidopsis thaliana than tomato fruit-specific PME genes. In the promoter, it was found direct repeats, perfect inverted repeats and light responsive elements. GUS histochemical analysis showed activity in all plant tissues with the exception of pollen. The reduction in the promoter size induced a reduction in GUS activity in root, stem and leaf. Furthermore, root and leaf showed the highest and lowest activity, respectively. We had isolated a tomato PME gene with novel characteristics as compared with other known PME genes from tomato.