Identification of Candidate Expansin Genes Associated with Seed Weight in Pomegranate (Punica granatum L.).

Seed weight is an important target trait in pomegranate breeding and culture. Expansins act by loosening plant cell walls and cellulosic materials, permitting turgor-driven cell enlargement. However, the role of expansin genes (EXPs) in pomegranate seed weight remains elusive. A total of 29 PgrEXPs were identified in the 'Dabenzi' genome. These genes were classified into four subfamilies and 14 subgroups, including 22 PgrEXPAs, 5 PgrEXPBs, 1 PgrEXPLA, and 1 PgrEXPLB. Transcriptome analysis of PgrEXPs in different tissues (root, leaf, flower, peel, and seed testa) in 'Dabenzi', and the seed testa of the hard-seeded pomegranate cultivar 'Dabenzi' and soft-seeded cultivar 'Tunisia' at three development stages showed that three PgrEXPs (PgrEXPA11, PgrEXPA22, PgrEXPA6) were highly expressed throughout seed development, especially in the sarcotesta. SNP/Indel markers of these PgrEXPs were developed and used to genotype 101 pomegranate accessions. The association of polymorphic PgrEXPs with seed weight-related traits (100-seed weight, 100-kernel weight, 100-sarcotesta weight, and the percentage of 100-sarcotesta to 100-seed weight) were analyzed. PgrEXP22 was significantly associated with 100-seed weight and 100-sarcotesta weight and is a likely candidate for regulating seed weight and sarcotesta development in particular. This study provides an effective tool for the genetic improvement of seed weight in pomegranate breeding programs.

Engineering Yarrowia lipolytica for Sustainable Production of the Pomegranate Seed Oil-Derived Punicic Acid.

Punicic acid is a conjugated linolenic acid with various biological activities including antiobesity, antioxidant, anticancer, and anti-inflammatory effects. It is often used as a nutraceutical, dietary additive, and animal feed. Currently, punicic acid is primarily extracted from pomegranate seed oil, but it is restricted due to the extended growth cycle, climatic limitations, and low recovery level. There have also been reports on the chemical synthesis of punicic acid, but it resulted in a mixture of structurally similar isomers, requiring additional purification/separation steps. In this study, a comprehensive strategy for the production of punicic acid in Yarrowia lipolytica was implemented by pushing the supply of linoleic acid precursors in a high-oleic oil strain, expressing multiple copies of the fatty acid conjugase gene from Punica granatum, engineering the acyl-editing pathway to improve the phosphatidylcholine pool, and promoting the assembly of punicic acid in the form of triglycerides. The optimal strain with high oil production capacity and a significantly increased punicic acid ratio accumulated 3072.72 mg/L punicic acid, accounting for 6.19% of total fatty acids in fed-batch fermentation, providing a viable, sustainable, and green approach for punicic acid production to substitute plant extraction and chemical synthesis production.

Chromosome-level assemblies of cultivated water chestnut Trapa bicornis and its wild relative Trapa incisa.

Water chestnut (Trapa L.) is a floating-leaved aquatic plant with high edible and medicinal value. In this study, we presented chromosome-level genome assemblies of cultivated large-seed species Trapa bicornis and its wild small-seed relative Trapa incisa by using PacBio HiFi long reads and Hi-C technology. The T. bicornis and T. incisa assemblies consisted of 479.90 Mb and 463.97 Mb contigs with N50 values of 13.52 Mb and 13.77 Mb, respectively, and repeat contents of 62.88% and 62.49%, respectively. A total of 33,306 and 33,315 protein-coding genes were predicted in T. bicornis and T. incisa assemblies, respectively. There were 159,232 structural variants affecting more than 11 thousand genes detected between the two genomes. The phylogenetic analysis indicated that the lineage leading to Trapa was diverged from the lineage to Sonneratia approximately 23 million years ago. These two assemblies provide valuable resources for future evolutionary and functional genomic research and molecular breeding of water chestnut.

Ripening of Pomegranate Skin as Revealed by Developmental Transcriptomics.

The appearance of pomegranate (Punica granatum L.) fruit is highly important for its marketing. The primary concerns are obtaining sufficient red pigment accumulation and minimal cracking of the fruit skin (the outer red layer of the peel). We analyzed the skin transcriptome of pomegranate cv. Wonderful at distinct time points of fruit development to characterize the processes that occur in the skin during fruit ripening and which may reflect on processes in the whole fruit, such as the non-climacteric nature of pomegranate. The data suggested a ripening mechanism in pomegranate skin that differs from that in strawberry-the model plant for non-climacteric fruit where abscisic acid is the growth regulator that drives ripening-involving ethylene, polyamine, and jasmonic acid pathways. The biosynthetic pathways of important metabolites in pomegranate-hydrolyzable tannins and anthocyanins-were co-upregulated at the ripening stage, in line with the visual enhancement of red coloration. Interestingly, cuticle- and cell-wall-related genes that showed differential expression between the developmental stages were mainly upregulated in the skin of early fruit, with lower expression at mid-growth and ripening stages. Nevertheless, lignification may be involved in skin hardening in the mature fruit.

Fifteen complete chloroplast genomes of Trapa species (Trapaceae): insight into genome structure, comparative analysis and phylogenetic relationships.

BACKGROUND: Trapa L. is a floating-leaved aquatic plant with important economic and ecological values. However, the species identification and phylogenetic relationship within Trapa are still controversial, which necessitates the need for plastid genome information of Trapa. In this study, complete chloroplast genomes of 13 Trapa species/taxa were sequenced and annotated. Combined with released sequences, comparative analyses of chloroplast genomes were performed on the 15 Trapa species/taxa for the first time. RESULTS: The Trapa chloroplast genomes exhibited typical quadripartite structures with lengths from 155,453 to 155,559 bp. The gene orders and contents within Trapa were conservative, but several changes were found in the microstructure. The intron loss of rpl2, also detected in Lythraceae, was found in all Trapa species/taxa, suggesting close genetic relationship between Lythraceae and Trapaceae. Notably, two small-seed species (T. incisa and T. maximowiczii) showed the smallest genome size with 155,453 and 155,477 bp, respectively. Each cp genome contained the same 130 genes consisting of 85 protein-coding genes, 37 tRNA genes and 8 rRNA genes. Trapa species/taxa showed 37 (T. incisa and T. maximowiczii) to 41 (T. sibirica) long repeats, including forward, palindromic, reversed and complementary repeats. There were 110 (T. quadrispinosa) to 123 (T. incisa and T. maximowiczii) SSR (simple sequence repeat) loci in Trapa chloroplast genomes. Comparative analyses revealed that two hotspot regions (atpA-atpF and rps2-rpoC2) in Trapa chloroplast genomes could be served as potential molecular markers. Three phylogenetic analyses (ML, MP and BI) consistently showed that there were two clusters within Trapa, including large- and small-seed species/taxa, respectively; for the large-seed Trapa, they clustered according to their geographical origin and tubercle morphology on the surface of seeds. CONCLUSION: In summary, we have acquired the sequences of 13 Trapa chloroplast genomes, and performed the comparative analyses within Trapa for the first time. The results have helped us better identify the Trapa species/taxa and deepen the understanding of genetic basis and phylogenetic relationship of Trapa, which will facilitate the effective management and utilization of the important genetic resources in the future.

Identification, Analysis and Gene Cloning of the SWEET Gene Family Provide Insights into Sugar Transport in Pomegranate (Punica granatum).

Members of the sugars will eventually be exported transporter (SWEET) family regulate the transport of different sugars through the cell membrane and control the distribution of sugars inside and outside the cell. The SWEET gene family also plays important roles in plant growth and development and physiological processes. So far, there are no reports on the SWEET family in pomegranate. Meanwhile, pomegranate is rich in sugar, and three published pomegranate genome sequences provide resources for the study of the SWEET gene family. 20 PgSWEETs from pomegranate and the known Arabidopsis and grape SWEETs were divided into four clades (Ⅰ, Ⅱ, Ⅲ and Ⅳ) according to the phylogenetic relationships. PgSWEETs of the same clade share similar gene structures, predicting their similar biological functions. RNA-Seq data suggested that PgSWEET genes have a tissue-specific expression pattern. Foliar application of tripotassium phosphate significantly increased the total soluble sugar content of pomegranate fruits and leaves and significantly affected the expression levels of PgSWEETs. The plant growth hormone regulator assay also significantly affected the PgSWEETs expression both in buds of bisexual and functional male flowers. Among them, we selected PgSWEET17a as a candidate gene that plays a role in fructose transport in leaves. The 798 bp CDS sequence of PgSWEET17a was cloned, which encodes 265 amino acids. The subcellular localization of PgSWEET17a showed that it was localized to the cell membrane, indicating its involvement in sugar transport. Transient expression results showed that tobacco fructose content was significantly increased with the up-regulation of PgSWEET17a, while both sucrose and glucose contents were significantly down-regulated. The integration of the PgSWEET phylogenetic tree, gene structure and RNA-Seq data provide a genome-wide trait and expression pattern. Our findings suggest that tripotassium phosphate and plant exogenous hormone treatments could alter PgSWEET expression patterns. These provide a reference for further functional verification and sugar metabolism pathway regulation of PgSWEETs.

Genome sequencing and transcriptome analyses provide insights into the origin and domestication of water caltrop (Trapa spp., Lythraceae).

Humans have domesticated diverse species from across the plant kingdom; however, our current understanding of plant domestication is largely founded on major cereal crops. Here, we examine the evolutionary processes and genetic basis underlying the domestication of water caltrop (Trapa spp., Lythraceae), a traditional, yet presently underutilized non-cereal crop that sustained early Chinese agriculturalists. We generated a chromosome-level genome assembly of tetraploid T. natans, and then divided the allotetraploid genome into two subgenomes. Based on resequencing data from 57 accessions, representing cultivated diploid T. natans, wild T. natans (2x and 4x) and diploid T. incisa, we showed that water caltrop was likely first domesticated in the Yangtze River Valley as early as 6300 yr BP, and experienced a second improvement c. 800 years ago. We also provided strong support for an allotetraploid origin of T. natans within the past 230 000-310 000 years. By integrating selective sweep and transcriptome profiling analyses, we identified a number of genes potentially selected and/or differentially expressed during domestication, some of which likely contributed not only to larger fruit sizes but also to a more vigorous root system, facilitating nutrient uptake, environmental stress response and underwater photosynthesis. Our results shed light on the evolutionary and domestication history of water caltrop, one of the earliest domesticated crops in China. This study has implications for genomic-assisted breeding of this presently underutilized aquatic plant, and improves our general understanding of plant domestication.

Chloroplast phylogenomics and divergence times of Lagerstroemia (Lythraceae).

BACKGROUND: Crape myrtles, belonging to the genus Lagerstroemia L., have beautiful paniculate inflorescences and are cultivated as important ornamental tree species for landscaping and gardening. However, the phylogenetic relationships within Lagerstroemia have remained unresolved likely caused by limited sampling and the insufficient number of informative sites used in previous studies. RESULTS: In this study, we sequenced 20 Lagerstroemia chloroplast genomes and combined with 15 existing chloroplast genomes from the genus to investigate the phylogenetic relationships and divergence times within Lagerstroemia. The phylogenetic results indicated that this genus is a monophyletic group containing four clades. Our dating analysis suggested that Lagerstroemia originated in the late Paleocene (~ 60 Ma) and started to diversify in the middle Miocene. The diversification of most species occurred during the Pleistocene. Four variable loci, trnD-trnY-trnE, rrn16-trnI, ndhF-rpl32-trnL and ycf1, were discovered in the Lagerstroemia chloroplast genomes. CONCLUSIONS: The chloroplast genome information was successfully utilized for molecular characterization of diverse crape myrtle samples. Our results are valuable for the global genetic diversity assessment, conservation and utilization of Lagerstroemia.

Punicic acid production in Brassica napus.

Punicic acid (PuA; 18:3Δ9cis,11trans,13cis), a conjugated linolenic acid isomer bearing three conjugated double bonds, is associated with various health benefits and has potential for industrial use. The major nature source of this unusual fatty acid is pomegranate (Punica granatum) seed oil, which contains up to 80% (w/w) of its fatty acids as PuA. Pomegranate seed oil, however, is low yielding with unstable production and thus limits the supply of PuA. Metabolic engineering of established temperate oil crops for PuA production, therefore, has the potential to be a feasible strategy to overcome the limitations associated with sourcing PuA from pomegranate. In this study, the cDNAs encoding a pomegranate fatty acid conjugase and a pomegranate oleate desaturase were co-expressed in canola-type Brassica napus. Transgenic B. napus lines accumulated up to 11% (w/w) of the total fatty acids as PuA in the seed oil, which is the highest level of PuA reported in metabolically engineered oilseed crops so far. Levels of seed oil PuA were stable over two generations and had no negative effects on seed germination. The transgenic B. napus lines with the highest PuA levels contained multiple transgene insertions and the PuA content of B. napus seed oil was correlated with efficiency of oleic acid desaturation and linoleic acid conjugation. In addition, PuA accumulated at lower levels in polar lipids (5.0-6.9%) than triacylglycerol (7.5-10.6%), and more than 60% of triacylglycerol-associated PuA was present at the sn-2 position. This study provides the basis for the commercial production of PuA in transgenic oilseed crops and thus would open new prospects for the application of this unusual fatty acid in health and industry.

Molecular adaptation to salinity fluctuation in tropical intertidal environments of a mangrove tree Sonneratia alba.

BACKGROUND: Mangroves have adapted to intertidal zones - the interface between terrestrial and marine ecosystems. Various studies have shown adaptive evolution in mangroves at physiological, ecological, and genomic levels. However, these studies paid little attention to gene regulation of salt adaptation by transcriptome profiles. RESULTS: We sequenced the transcriptomes of Sonneratia alba under low (fresh water), medium (half the seawater salinity), and high salt (seawater salinity) conditions and investigated the underlying transcriptional regulation of salt adaptation. In leaf tissue, 64% potential salinity-related genes were not differentially expressed when salinity increased from freshwater to medium levels, but became up- or down-regulated when salt concentrations further increased to levels found in sea water, indicating that these genes are well adapted to the medium saline condition. We inferred that both maintenance and regulation of cellular environmental homeostasis are important adaptive processes in S. alba. i) The sulfur metabolism as well as flavone and flavonol biosynthesis KEGG pathways were significantly enriched among up-regulated genes in leaves. They are both involved in scavenging ROS or synthesis and accumulation of osmosis-related metabolites in plants. ii) There was a significantly increased percentage of transcription factor-encoding genes among up-regulated transcripts. High expressions of salt tolerance-related TF families were found under high salt conditions. iii) Some genes up-regulated in response to salt treatment showed signs of adaptive evolution at the amino acid level and might contribute to adaptation to fluctuating intertidal environments. CONCLUSIONS: This study first elucidates the mechanism of high-salt adaptation in mangroves at the whole-transcriptome level by salt gradient experimental treatments. It reveals that several candidate genes (including salt-related genes, TF-encoding genes, and PSGs) and major pathways are involved in adaptation to high-salt environments. Our study also provides a valuable resource for future investigation of adaptive evolution in extreme environments.

Molecular mechanisms involved in postharvest chilling tolerance of pomegranate fruit.

Cold storage of pomegranates is essential for prolonging postharvest storage and for the implementation of cold-quarantine insect disinfestation treatments required for international trading. However, pomegranates are chilling sensitive; they may develop chilling injuries upon exposure to unfavorable low temperatures. In this mini-review, we summarize molecular data obtained from three different RNA Seq transcriptome analyses of responses of pomegranate fruits to cold storage. These experiments included comparisons among the transcriptomic responses following a 2-week exposure to 1 °C in three different model systems: 1) unconditioned chilling-sensitive fruits versus relatively chilling-tolerant low-temperature-conditioned fruits; 2) chilling-sensitive early harvested fruits versus relatively chilling-tolerant late-harvested ones; and 3) chilling-sensitive 'Ganesh' variety versus the relatively chilling-tolerant 'Wonderful' variety. Comparisons among differentially expressed transcripts that were exclusively and significantly up-regulated in the relatively chilling-tolerant fruits in all three model systems enabled identification of 573 common chilling tolerance-associated genes in pomegranates. Functional categorization and classification of the differentially expressed transcripts revealed several regulatory, metabolic, and stress-adaptation pathways that were uniquely activated in response to cold storage in relatively chilling-tolerant fruits. More specifically, we identified common up-regulation of transcripts involved in activation of jasmonic acid and ethylene hormone biosynthesis and signaling, stress-related transcription factors, calcium and MAPK signaling, starch degradation and galactinol and raffinose biosynthesis, phenol biosynthesis, lipid metabolism, and heat-shock proteins. We hypothesized these pathways to be involved in imparting chilling tolerance to pomegranate fruits. © 2019 Society of Chemical Industry.

The Complete Chloroplast Genomes of Punica granatum and a Comparison with Other Species in Lythraceae.

Pomegranates (Punica granatum L.) are one of the most popular fruit trees cultivated in arid and semi-arid tropics and subtropics. In this study, we determined and characterized three complete chloroplast (cp) genomes of P. granatum cultivars with different phenotypes using the genome skimming approach. The complete cp genomes of three pomegranate cultivars displayed the typical quadripartite structure of angiosperms, and their length ranged from 156,638 to 156,639 bp. They encoded 113 unique genes and 17 are duplicated in the inverted regions. We analyzed the sequence diversity of pomegranate cp genomes coupled with two previous reports. The results showed that the sequence diversity is extremely low and no informative sites were detected, which suggests that cp genome sequences may be not be suitable for investigating the genetic diversity of pomegranate genotypes. Further, we analyzed the codon usage pattern and identified the potential RNA editing sites. A comparative cp genome analysis with other species within Lythraceae revealed that the gene content and organization are highly conserved. Based on a site-specific model, 11 genes with positively selected sites were detected, and most of them were photosynthesis-related genes and genetic system-related genes. Together with previously released cp genomes of the order Myrtales, we determined the taxonomic position of P. granatum based on the complete chloroplast genomes. Phylogenetic analysis suggested that P. granatum form a single clade with other species from Lythraceae with a high support value. The complete cp genomes provides valuable information for understanding the phylogenetic position of P. gramatum in the order Myrtales.

Comparative analyses of chloroplast genomes from 22 Lythraceae species: inferences for phylogenetic relationships and genome evolution within Myrtales.

BACKGROUND: Lythraceae belongs to the order Myrtales, which is part of Archichlamydeae. The family has 31 genera containing approximately 620 species of herbs, shrubs and trees. Of these 31 genera, five large genera each possess 35 or more species. They are Lythrum, with 35; Rotala, with 45; Nesaea, with 50; Lagerstroemia, with 56; and Cuphea, with 275 species. RESULTS: We reported six newly sequenced chloroplast (cp) genomes (Duabanga grandiflora, Trapa natans, Lythrum salicaria, Lawsonia inermis, Woodfordia fruticosa and Rotala rotundifolia) and compared them with 16 other cp genomes of Lythraceae species. The cp genomes of the 22 Lythraceae species ranged in length from 152,049 bp to 160,769 bp. In each Lythraceae species, the cp genome contained 112 genes consisting of 78 protein coding genes, four ribosomal RNAs and 30 transfer RNAs. Furthermore, we detected 211-332 simple sequence repeats (SSRs) in six categories and 7-27 long repeats in four categories. We selected ten divergent hotspots (ndhF, matK, ycf1, rpl22, rpl32, trnK-rps16, trnR-atpA, rpl32-trnL, trnH-psbA and trnG-trnR) among the 22 Lythraceae species to be potential molecular markers. We constructed phylogenetic trees from 42 Myrtales plants with 8 Geraniales plants as out groups. The relationships among the Myrtales species were effectively distinguished by maximum likelihood (ML), maximum parsimony (MP) and Bayesian inference (BI) trees constructed using 66 protein coding genes. Generally, the 22 Lythraceae species gathered into one clade, which was resolved as sister to the three Onagraceae species. Compared with Melastomataceae and Myrtaceae, Lythraceae and Onagraceae differentiated later within Myrtales. CONCLUSIONS: The study provided ten potential molecular markers as candidate DNA barcodes and contributed cp genome resources within Myrtales for further study.

Characterization of a NAC transcription factor involved in the regulation of pomegranate seed hardness (Punica granatum L.).

The pomegranate, Punica granatum L., which has been cultivated since antiquity, is known to be a superfruit, possessing an array of functional anti-oxidants and various other health benefits. The hardness of pomegranate seeds is an important indicator of fruit quality, which in turn affects economic value and market demand. However, the molecular mechanism underlying pomegranate seed hardness remains to be fully understood. In this study, we found a positive correlation between seed hardness and lignin content in two pomegranate varieties: "Tunisia" and "Sanbai". Specifically, genes associated with lignin biosynthesis were differentially expressed in soft-seed and hard-seed pomegranate varieties. Among these differential genes, we cloned and characterized the NAC transcription factor PgSND1-like. Sequence alignment found a single base replacement at the 166-bp position of CDS in the PgSND1-like gene from "Tunisia" and "Sanbai". Both PgSND1-like (Sanbai) and PgSND1-like (Tunisia) proteins are localized in the cell nucleus and have a transcription activation domain in the C-terminus. Yeast two-hybrid analysis indicated that PgSND1-like protein interacts with itself to form a homodimer. Overexpression of PgSND1-like (Sanbai) in Arabidopsis showed a higher lignin content in inflorescence stem and mature seed compared with wild-type Arabidopsis. Accordingly, the expression levels of several lignin biosynthesis-associated genes were upregulated in stem cells and mature seeds of transgenic plants. However, PgSND1-like (Tunisia) transgenic Arabidopsis showed no phenotypic differences with wild-type Arabidopsis. Taken together, we suggest that PgSND1-like may regulate at least two different functions in two pomegranate varieties, promoting lignin biosynthesis and seed hardness of pomegranate.

Targeted metabolite profiling to gain chemometric insight into Indian pomegranate cultivars and elite germplasm.

BACKGROUND: Pomegranate fruit is an excellent source of bioactive polyphenolics, known to contribute significantly to human health. India is the largest producer of pomegranate in the world and produces the finest quality fruit with highly desirable consumer traits such as soft seeds, low acidity, and attractive fruit and aril color. Knowledge of the extent of variation in key metabolites (sugars, organic acids, phenolics, and anthocyanins) is key to selecting superior genotypes for germplasm improvement. Relevant information with respect to Indian genotypes is scarce. The present study therefore aims to evaluate quantitatively important metabolites in some cultivars and elite germplasm of pomegranate in India. RESULTS: Identification and quantification of primary and secondary metabolites such as sugars, organic acids, vitamin C, polyphenolics, and anthocyanins were conducted using a liquid chromatography - mass spectrometry (LC-MS) platform. Fructose and citric acid were the predominant sugar and organic acid, respectively. Wild genotypes had significantly higher concentrations of organic acids, antioxidant activity, and phenolics, namely punicalagin, ellagic acid, sinapic, and ferulic acid. CONCLUSION: Cyanidin and delphinidin derivatives of anthocyanins were more abundant in red aril commercial genotypes. Results suggest that wild-sour accessions represent a rich source of polyphenolics that can be utilized in future breeding programs to breed healthier varieties, food supplements, and pharmaceutical products. © 2019 Society of Chemical Industry.

Antioxidant activity and phenolic composition in pomegranate (Punica granatum L.) genotypes from south Italy by UHPLC-Orbitrap-MS approach.

BACKGROUND: Pomegranate fruits are a rich source of polyphenols with numerous health-promoting effects. Pomegranate juices of five genotypes ('Mollar', 'Kingdom', 'Dente di Cavallo', and two old populations 'Francofonte' and 'Santa Tecla') were evaluated regarding anthocyanin and non-anthocyanin phenolic contents using ultrahigh performance liquid chromatography (UHPLC)-Orbitrap-mass spectrometry (MS). Moreover, total antioxidant activity (TAA) was evaluated using a 2,2'-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS) assay. RESULTS: Twenty-three phenolic compounds were identified. Cyanidin-3,5-O-diglucoside and pelargonidin-3,5-O-diglucoside were the most representative anthocyanins in all genotypes; the Santa Tecla population had the highest content of these anthocyanins, 97.64 mg L-1 and 40.29 mg L-1 respectively. In the Francofonte population, ferulic acid hexoside was the most abundant compound (391.18 mg L-1 ). TAA values ranged between 221.5 and 36.73 µmol Trolox equivalents/100 mL of juice. A high TAA value was recorded for the Santa Tecla pomegranate population. CONCLUSION: The UHPLC-Orbitrap-MS approach was employed for the first time to identify the phenolic compound profiling in five pomegranate genotypes. TAA was analysed using an ABTS assay, and the results showed a significant variability in nutraceutical potential of the pomegranate genotypes studied. The inclusion of phenolic information in the linear discriminant analysis allowed very good discriminations among genotypes to be obtained. © 2018 Society of Chemical Industry.

Diversity among Pomegranate Varieties in Chilling Tolerance and Transcriptome Responses to Cold Storage.

We found great variability in chilling tolerance among 84 pomegranate varieties from the Newe Ya'ar collection; among them, 'Ganesh' was chilling-sensitive, whereas 'Wonderful' was relatively chilling-tolerant. To evaluate the different molecular responses of these varieties to cold storage, we analyzed the transcriptomic changes in the inner membrane tissues of 'Ganesh' and 'Wonderful' fruit after 2 weeks of cold storage at 1 °C. By functional categorization of the differentially expressed transcripts using MapMan, we found that many transcripts related to various pathways, such as jasmonic acid biosynthesis and signaling, galactinol, raffinose, phenol, and phenylpropanoid biosynthesis, calcium and mitogen-activated protein kinase signaling, lipid metabolism, and various transcription factors and heat-shock proteins, have been massively upregulated in 'Wonderful' but not in 'Ganesh' fruit. Thus, it is suggested that these pathways most likely participate in imparting chilling tolerance in pomegranate fruit.

Pomegranate chitinase III: Identification of a new allergen and analysis of sensitization patterns to chitinases.

Allergy to pomegranate is often associated with severe symptoms. Two allergens have previously been described: 9k-LTP Pun g 1 and pommaclein Pun g 7. This study describes the isolation of a chitinase III, identified by direct protein sequencing and mass spectrometry. It is a 29-kDa protein showing 69% sequence identity with the latex hevamine and IgE binding in dot blotting, immunoblotting and FABER®test. Chitinase-specific IgE were detected in 69 of 357 patients sensitized to one or more pomegranate allergenic preparations present on the FABER®test. Using this test, 19.2% of the patients sensitized to kiwifruit chitinase IV were also sensitized to pomegranate chitinase III, rather than to latex chitinase I (7.2%) with which it shares the N-terminal hevein-like domain. In conclusion, a new allergen has been identified, contributing to improving food allergy diagnosis. This study reveals the important role of chitinases III and IV as allergy sensitizers and prompts further investigations.

First reported chloroplast genome sequence of Punica granatum (cultivar Helow) from Jabal Al-Akhdar, Oman: phylogenetic comparative assortment with Lagerstroemia.

Pomegranate (Punica granatum L.) is one of the oldest known edible fruits. It has grown in popularity and is a profitable fruit crop due to its attractive features including a bright red appearance and its biological activities. Scientific exploration of the genetics and evolution of these beneficial traits has been hampered by limited genomic information. In this study, we sequenced the complete chloroplast (cp) genome of the native P. granatum (cultivar Helow) cultivated in the mountains of Jabal Al-Akhdar, Oman. The results revealed a P. granatum cp genome length of 158,630 bp, characterized by a relatively conserved structure containing 2 inverted repeat regions of 25,466 bp, an 18,686 bp small single copy regions, and an 89,015 bp large single copy region. The 86 protein-coding genes included 37 transfer RNA genes and 8 ribosomal RNA genes. Comparison of the P. granatum whole cp genome with seven Lagerstroemia species revealed an overall high degree of sequence similarity with divergence among intergenic spacers. The location, distribution, and divergence of repeat sequences and shared genes of the Punica and Lagerstroemia species were highly similar. Analyses of nucleotide substitution, insertion/deletions, and highly variable regions in these cp genomes identified potential plastid markers for taxonomic and phylogenetic studies in Myrtales. A phylogenetic study of the cp genomes and 76 shared coding regions generated similar cladograms. The complete cp genome of P. granatum will aid in taxonomical studies of the family Lythraceae.

Complementary iTRAQ-based proteomic and RNA sequencing-based transcriptomic analyses reveal a complex network regulating pomegranate (Punica granatum L.) fruit peel colour.

Peel colour is an important factor affecting the marketability of pomegranate fruits. Therefore, elucidating the genetic mechanism of fruit peel colour development may be useful for breeding pomegranate cultivars with enhanced fruit peel colours. In this study, we combined an iTRAQ-based proteome-level analysis with an RNA sequencing-based transcriptome-level analysis to detect the proteins and genes related to fruit peel colour development in pomegranate. We analysed the 'Tunisia' (red fruit) and 'White' (white fruit) pomegranate cultivars at two stages of fruit development. A total of 27 differentially abundant proteins (increased abundance) and 54 differentially expressed genes (16 up-regulated and 38 down-regulated) were identified from our proteomics and transcriptomics data. The identified proteins and genes contribute to pomegranate fruit peel colour by participating in the biosynthesis of anthocyanins, stilbenoids, diarylheptanoids, gingerols, flavonoids, and phenylpropanoids. Several candidate proteins and genes corresponded to enzymes related to general reactions (PAL, 4CL, DFR, LDOX/ANS, CHS, and F3'5'H) and glycosylation (GT1 and UGAT) of compounds and pigments related to the colour of pomegranate fruit peel. Complementary proteome- and transcriptome-level analyses revealed a complex molecular network controlling fruit peel colour. The candidate genes identified in this study may be useful for the marker-based breeding of new pomegranate cultivars.