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Color acquisition is one of the most distinctive features of fruit development and ripening processes. The color red is closely related to the accumulation of polyphenolic compounds, mainly anthocyanins, during sweet cherry fruit maturity. In non-climacteric fruit species like sweet cherry, the maturity process is mainly controlled by the phytohormone abscisic acid (ABA), though other hormones may also play a role. However, the coordinated stage-specific production of polyphenolic compounds and their relation with hormone content variations have not been studied in depth in sweet cherry fruits. To further understand the accumulation dynamics of these compounds (hormones and metabolites) during fruit development, two sweet cherry cultivars ("Lapins" and "Glenred") with contrasting maturity timing phenotypes were analyzed using targeted metabolic analysis. The ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) approach revealed that phenolic acids, flavonols, and flavan-3-ols accumulated mainly until the straw-yellow stage in the early-maturing cultivar, while accumulation was mainly at the green stage in the mid-maturing cultivar, suggesting a cultivar-dependent stage-specific production of secondary metabolites. In the mid-maturing cultivar, anthocyanins were detected only from the red stage onward, whereas detection began at the pink stage in the early-maturing cultivar. ABA negatively correlated (p-value < 0.05) with the flavonols and flavan-3-ols in both cultivars. ABA and anthocyanin content increased at the same time in the early-season cultivar. In contrast, anthocyanins accumulated and the pink color initiation started several days after the ABA increase in the mid-maturing cultivar. Differential accumulation patterns of GA4, a ripening antagonizing hormone, between the cultivars could explain this difference. These results showed that both red-colored cultivars presented different accumulation dynamics of phenolic compounds and plant hormones during fruit development, suggesting underlying differences in the sweet cherry fruit color evolution.

Ethylene plays an essential role in the ripening of peach, a climacteric stone fruit. However, several studies have shown that other phytohormones may alter fruit organoleptic properties, nutritional value, and overall quality. Although cytokinin levels are highest during fruit set and early stages of fruit development, exogenous cytokinin application on the late stages of peach fruit development (post-lignification stage) alters the peach fruit ripening indicators and delays ripening. To better understand the molecular mechanisms by which cytokinin alters fruit ripening and fruit quality traits, we identified a conserved set of 26 peach cytokinin-regulated orthologs of the Arabidopsis cytokinin-regulated genes. All but two of these genes show expression in ripe peach fruits. The peach cytokinin-regulated genes map throughout the Prunus persica genome, located on all eight chromosome-scale pseudomolecules. However, eight of these orthologs are located on pseudomolecule 1, while five orthologs are on pseudomolecule 2. Gene Ontology enrichment analyses revealed that many of these genes are associated with metabolic processes. The region upstream of the transcription start site contained Cytokinin Response Elements (CKREs) in 77% (20/26) of these putative cytokinin-regulated genes. The cytokinin responsiveness of eight of these genes (PpeBPH1, PpeCKX4, PpeCPK28, PpeCYP18-2, PpeHSP90-7, PpeNRP2, PpeNIP1-1, and PpePCRP1) was confirmed by RT-qPCR analyses of ripe peach fruits treated exogenously with trans-zeatin for one hour. Therefore, by using a comparative genomic analysis between peach and Arabidopsis, we have identified a conserved set of cytokinin-regulated genes expressed in peach fruits that respond to exogenous application of trans-zeatin and, based upon the conserved cis-regulatory regions, are most likely regulated by the cytokinin Type-B Response Regulators.

Thinning is an agronomic practice that consists in the removal of a certain number of fruits, thereby altering the source–sink equilibrium in the tree and favoring carbon allocation to the remaining fruits. If thinning is performed at the appropriate fruit developmental stage and at an adequate intensity, the remaining fruits will have a larger final diameter and higher soluble solids content. However, the amount of carbon translocated to a fruit depends on the photosynthetic rate in the source leaves, the production of “translocation sugars”, as well as the fruit sink strength, which is directly related to the phloem unloading. It is not clear which of these steps is altered by thinning. In this work, we evaluated the effect of thinning in an early season peach variety. Photosynthetic parameters, quantification of soluble sugars in leaves and fruits, and the expression of sucrose and sorbitol transporters were evaluated in leaves and fruits during whole fruit development. We showed that thinning was effective in improving final fruit size and sugar accumulation without inhibiting photosynthesis. We also observed a 10-day delay in stone lignification when the trees were not thinned. The stone lignification phase was very short in the thinned treatment, while it seems to last for longer under unthinning. Furthermore, it was shown that thinning affects the expression of translocation sugar transporters genes (PpeSUT1 and PpeSUT4) in both leaves and fruits, which could be increasing the fruit sink strength and subsequently improving translocation sugars transport to the fruit when trees are thinned.

In sweet cherry, as in most non-climacteric species, abscisic acid (ABA) plays a major role in the control of fruit ripening and color development. Although the ABA treatment of sweet cherry fruits has been reported to upregulate anthocyanin pathway-related genes or ABA pathway-related genes, the temporality of molecular and physiological events occurring during color development and the ABA control of these events during the color initiation are lacking in this species. In this work, we analyzed variations in the Index of Absorbance Difference (IAD), a maturity index, and total anthocyanins along with changes in transcript abundance of ABA and anthocyanin pathway-related genes, from light green to red fruit stages. PavNCED1 and ABA signaling pathway-related genes upregulated when fruits transitioned from light green to pink stage, whereas anthocyanin pathway-related transcripts increased from pink to the red stage, together with increases in the anthocyanin content and IAD, suggesting sequentiality in molecular and physiological events during color development. Additionally, ABA applied at color initiation in planta advanced IAD, increased anthocyanin content, and yielded darker fruits at harvest. These changes were accompanied by changes in the transcript accumulation of ABA and anthocyanin pathway-related genes. This in planta treatment of sweet cherry fruits with ABA confirms that ABA is a central player in the control of color initiation in sweet cherries, associated with the transcript accumulation of genes involved in ABA homeostasis and signaling, which is followed by the up-regulation of anthocyanin pathway-related genes and color development.

Gibberellin (GA) negatively affects color evolution and other ripening-related processes in non-climacteric fruits. The bioactive GA, gibberellic acid (GA3), is commonly applied at the light green-to-straw yellow transition to increase firmness and delay ripening in sweet cherry (Prunus avium L.), though causing different effects depending on the variety. Recently, we reported that GA3 delayed the IAD parameter (a ripening index) in a mid-season variety, whereas GA3 did not delay IAD but reduced it at ripeness in an early-season variety. To further explore this contrasting behavior between varieties, we analyzed the transcriptomic responses to GA3 applied on two sweet cherry varieties with different maturity time phenotypes. At harvest, GA3 produced fruits with less color in both varieties. Similar to our previous report, GA3 delayed fruit color initiation and IAD only in the mid-season variety and reduced IAD at harvest only in the early-season variety. RNA-seq analysis of control- and GA3-treated fruits revealed that ripening-related categories were overrepresented in the early-season variety, including ‘photosynthesis’ and ‘auxin response’. GA3 also changed the expression of carotenoid and abscisic acid (ABA) biosynthetic genes in this variety. In contrast, overrepresented categories in the mid-season variety were mainly related to metabolic processes. In this variety, some PP2Cs putative genes were positively regulated by GA3, which are negative regulators of ABA responses, and MYB44-like genes (putative repressors of PP2Cs expression) were downregulated. These results show that GA3 differentially modulates the transcriptome at the onset of ripening in a variety-dependent manner and suggest that GA3 impairs ripening through the modification of ripening associated gene expression only in the early-season variety; whereas in the mid-season variety, control of the ripening timing may occur through the PP2C gene expression regulation. This work contributes to the understanding of the role of GA in non-climacteric fruit ripening.

Abscisic acid (ABA) plays a major role in promoting ripening in sweet cherry, a non-climateric fruit. Exogenous application of ABA has been performed to study fruit ripening and cracking, but this growth regulator is not used for commercial production. To determine the potential of this growth regulator to improve sweet cherry fruit quality, ABA canopy spraying was assayed in four cultivars. Canopy spraying of S-ABA significantly: 1) enhanced sweet cherry fruit color in ‘Glenred’, ‘Lapins’ and 'Bing' cultivars, but not in ‘Royal Rainier’ (a bi-colored cultivar), and 2) decreased fruit size and firmness in ‘Lapins’, ‘Bing’ and ‘Royal Rainier’. Seasonally reproducible effects were seen in ‘Lapins’ (mid/late-maturing) but not in ‘Glenred’ (early-maturing). Canopy spraying of nordihydroguaiaretic acid (NDGA) decreased color and increased fruit size in ‘Lapins’, but not in ‘Glenred’. Direct application of ABA on fruits attached to the tree, without application to the foliage, increased Lapins' fruit color without reducing size. These results suggest a localized fruit response to exogenous ABA application on fruit color development, but that a decrease in fruit size may be due to the effects of exogenous ABA on the tree canopy foliage.

Sweet cherry is a valuable non-climacteric fruit with elevated phytonutrients, whose fruit quality attributes are prone to rapid deterioration after harvest, especially peel damage and water loss of stem. Here the metabolic and transcriptional response of exogenous melatonin was assessed in two commercial cultivars of sweet cherry (Santina and Royal Rainier) during cold storage. Gene expression profiling revealed that cuticle composition and water movement may underlie the effect of melatonin in delaying weight loss. An effect of melatonin on total soluble solids and lower respiration rate was observed in both cultivars. Melatonin induces overexpression of genes related to anthocyanin biosynthesis, which correlates with increased anthocyanin levels and changes in skin color (Chroma). Our results indicate that along with modulating antioxidant metabolism, melatonin improves fruit quality traits by triggering a range of metabolic and gene expression changes, which ultimately contribute to extend sweet cherry postharvest storability.

Several phytohormones modulate ripening in non-climacteric fruits, which is triggered by abscisic acid (ABA). Gibberellins (GAs) are present during the onset of ripening in sweet cherry fruits, and exogenous gibberellic acid (GA3) application delays ripening, though this effect is variety-dependent. Although an ABA accumulation delay has been reported following GA3 treatment, the mechanism by which GA modulates this process has not been investigated at the molecular level in sweet cherry. Therefore, the aim of this work is to analyze the effect of GA3 on the fruit ripening process and the transcript levels of ABA pathway orthologs in two varieties having different maturity time phenotypes. The early-season variety had a rapid transition from yellow to pink fruit color, whereas pink color initiation took longer in the mid-season variety. GA3 increased the proportion of lighter colored fruits at ripeness in both varieties, but it produced a delay in IAD&mdash, a ripening index&mdash, only in the mid-season variety. This delay was accompanied by an increased transcript abundance of PavPP2Cs, which are putative negative regulators of the ABA pathway. On the other hand, the early-season variety had increased expression of PavCYP707A2&mdash, a putative ABA catabolic gene&ndash, and reduced transcript levels of PavPP2Cs and SnRK2s after the GA3 treatment. Together these results show that GA modulates fruit ripening, exerting its action in part by interacting with the ABA pathway in sweet cherry.

Background Through the diversity of cytokinin regulated processes, this phytohormone has a profound impact on plant growth and development. Cytokinin signaling is involved in the control of apical and lateral meristem activity, branching pattern of the shoot, and leaf senescence. These processes influence several traits, including the stem diameter, shoot architecture, and perennial life cycle, which define the development of woody plants. To facilitate research about the role of cytokinin in regulation of woody plant development, we have identified genes associated with cytokinin signaling and homeostasis pathways from two hardwood tree species. Results Taking advantage of the sequenced black cottonwood (Populus trichocarpa) and peach (Prunus persica) genomes, we have compiled a comprehensive list of genes involved in these pathways. We identified genes belonging to the six families of cytokinin oxidases (CKXs), isopentenyl transferases (IPTs), LONELY GUY genes (LOGs), two-component receptors, histidine containing phosphotransmitters (HPts), and response regulators (RRs). All together 85 Populus and 45 Prunus genes were identified, and compared to their Arabidopsis orthologs through phylogenetic analyses. Conclusions In general, when compared to Arabidopsis, differences in gene family structure were often seen in only one of the two tree species. However, one class of genes associated with cytokinin signal transduction, the CKI1-like family of two-component histidine kinases, was larger in both Populus and Prunus than in Arabidopsis.

Thapsigargin, a specific inhibitor of most animal intracellular SERCA-type Ca 2+ pumps present in thesarcoplasmic/endoplasmic reticulum, was originally isolated from the roots of the Mediterranean plant Thapsia gargancia L . Here, we demonstrate that this root-derived compound is capable of altering rootgravitropism in Arabidopsis thaliana . Thapsigargin concentrations as low as 0.1 µM alter root gravitropismwhereas under similar conditions cyclopiazonic acid does not. Furthermore, a fluorescently conjugatedthapsigargin (BODIPY FL thapsigargin) suggests that target sites for thapsigargin are located inintracellular organelles in the root distal elongation zone and the root cap, regions known to regulate rootgravitropism. Key terms: Arabidopsis thaliana , Ca 2+ -ATPases, calcium, gravitropism, thapsigargin, root. Corresponding author: Lee Meisel, Plant Biotechnology Center, Faculty of Ecology and Natural Resources, UniversidadAndres Bello, Av. Republica 217, Santiago, Chile, Tel: (56-2) 661-8449, Fax: (56-2) 661-5832, E-mail: lmeisel@unab.clReceived: October 3, 2005. In revised form: January 2. 2006. Accepted: January 9, 2006.INTRODUCTION

The synthesis of noncellulosic polysaccharides and glycoproteins in the plant cell Golgi apparatus requires UDP-galactose as a substrate. We have cloned and characterized a nucleotide sugar transporter from Arabidopsis thaliana (L.) Heynh. named AtUTr2. Expression in tobacco and Saccharomyces cerevisiae and subsequent biochemical characterization indicate that AtUTr2 transports UDP-galactose, but not UDP-glucose, UDP-N-acetyl glucosamine, UDP-xylose, UDP-glucuronic acid, GDP-fucose or GDP-mannose. Experiments expressing an AtUTr2-GFP fusion protein in onion epidermal cells suggest that AtUTr2 is located in the Golgi apparatus. Finally, northern analysis indicates that the AtUTr2 transcript was more abundant in roots and calli although it was also present in other Arabidopsis organs but at lower levels. Therefore, AtUTr2 is a nucleotide sugar transporter capable of transporting UDP-galactose that may play an important role in the synthesis of galactose-containing glycoconjugates in Arabidopsis.

http://www.scielo.cl/scielo.php?script=sci_arttext&pid=S0716-97602005000100010&lng=es&nrm=iso Prunus persica has been proposed as a genomic model for deciduous trees and the Rosaceae family. Optimized protocols for RNA isolation are necessary to further advance studies in this model species such that functional genomics analyses may be performed. Here we present an optimized protocol to rapidly and efficiently purify high quality total RNA from peach fruits (Prunus persica). Isolating high-quality RNA from fruit tissue is often difficult due to large quantities of polysaccharides and polyphenolic compounds that accumulate in this tissue and co-purify with the RNA. Here we demonstrate that a modified version of the method used to isolate RNA from pine trees and the woody plant Cinnamomun tenuipilum is ideal for isolating high quality RNA from the fruits of Prunus persica. This RNA may be used for many functional genomic based experiments such as RT-PCR and the construction of large-insert cDNA libraries.