Comparative transcriptomic and metabolomic analyses reveal differences in flavonoid biosynthesis between PCNA and PCA persimmon fruit

The fruit of the persimmon (Diospyros kaki.) has high economic and nutritional value and is rich in flavonoids. Flavonoids are essential secondary metabolisms in plants. The association between persimmon astringency and changes in the proanthocyanidins (a flavonoid subclass) content is well-known. However, information on the relationships between different astringency types and other flavonoid subclasses and biosynthetic genes is more limited. In this study, an initial correlation analysis between total flavonoids and fruit astringency type, and KEGG analysis of metabolites showed that flavonoid-related pathways were linked to differences between mature pollination-constant non-astringent (PCNA) varieties (‘Jiro’ and ‘Yohou’) and pollination-constant astringent (PCA) fruit varieties (‘Zhongshi5’ and ‘Huojing’). Based on these findings, variations in the expression of genes and metabolites associated with flavonoid biosynthesis were investigated between typical PCNA (‘Jiro’) and PCA (‘Huojing’) persimmons during fruit development. The flavonoid concentration in ‘Huojing’ fruit was significantly higher than that of ‘Jiro’ fruit, especially, in levels of proanthocyanin precursor epicatechin and anthocyanin cyanidin derivatives. Combined WGCNA and KEGG analyses showed that genes such as PAL, C4H, CHI, CHS, F3H, F3’5’H, FLS, DFR, ANR, ANS, and UF3GT in the phenylpropanoid and flavonoid biosynthesis pathways may be significant factors impacting the proanthocyanin precursor and anthocyanin contents. Moreover, interactions between the R2R3MYB (evm.TU.contig7272.598) and WD40 (evm.TU.contig3208.5) transcription factors were found to be associated with the above structural genes. These findings provide essential information on flavonoid biosynthesis and its regulation in the persimmon and lay a foundation for further investigation into how astringency types affect flavor components in PCNA and PCA persimmons

Genetic insights into the dissolution of dioecy in diploid persimmon Diospyros oleifera Cheng

BACKGROUND: Dioecy, a sexual system of single-sexual (gynoecious/androecious) individuals, is rare in flowering plants. This rarity may be a result of the frequent transition from dioecy into systems with co-sexual individuals. RESULTS: In this study, co-sexual expression (monoecy and hermaphroditic development), previously thought to be polyploid-specific in Diospyros species, was identified in the diploid D. oleifeara historically. We characterized potential genetic mechanisms that underlie the dissolution of dioecy to monoecy and andro(gyno)monoecy, based on multiscale genome-wide investigations of 150 accessions of Diospyros oleifera. We found all co-sexual plants, including monoecious and andro(gyno)monoecious individuals, possessed the male determinant gene OGI, implying the presence of genetic factors controlling gynoecia development in genetically male D. oleifera. Importantly, discrepancies in the OGI/MeGI module were found in diploid monoecious D. oleifera compared with polyploid monoecious D. kaki, including no Kali insertion on the promoter of OGI, no different abundance of smRNAs targeting MeGI (a counterpart of OGI), and no different expression of MeGI between female and male floral buds. On the contrary, in both single- and co-sexual plants, female function was expressed in the presence of a genome-wide decrease in methylation levels, along with sexually distinct regulatory networks of smRNAs and their targets. Furthermore, a genome-wide association study (GWAS) identified a genomic region and a DUF247 gene cluster strongly associated with the monoecious phenotype and several regions that may contribute to andromonoecy. CONCLUSIONS: Collectively, our findings demonstrate stable breakdown of the dioecious system in D. oleifera, presumably also a result of genomic features of the Y-linked region

Examples of MSAP profiles showing the inheritance and variation of DNA methylation sites in F1 hybrids relative to their parental lines.

<p>H genomic DNA was digested with <i>Hpa</i>II and <i>Eco</i>RI; M genomic DNA was digested with <i>Msp</i>I and <i>Eco</i>RI; Primer combinations are E₀₀+GC/ HM₀₀+CT (a), E₀₀+CC/HM₀₀+TC (b), E₀₀+GG/ HM₀₀+TG (c). (A) Monomorphic inheritance; (B) Additivity inheritance; (C) Demethylated; (D) Hyper- methylated; (E) Unidentified types.</p

Inheritance and Variation of Cytosine Methylation in Three <i>Populus</i> Allotriploid Populations with Different Heterozygosity

<div><p>DNA methylation is an epigenetic mechanism with the potential to regulate gene expression and affect plant phenotypes. Both hybridization and genome doubling may affect the DNA methylation status of newly formed allopolyploid plants. Previous studies demonstrated that changes in cytosine methylation levels and patterns were different among individual hybrid plant, therefore, studies investigating the characteristics of variation in cytosine methylation status must be conducted at the population level to avoid sampling error. In the present study, an F1 hybrid diploid population and three allotriploid populations with different heterozygosity [originating from first-division restitution (FDR), second-division restitution (SDR), and post-meiotic restitution (PMR) 2n eggs of the same female parent] were used to investigate cytosine methylation inheritance and variation relative to their common parents using methylation-sensitive amplification polymorphism (MSAP). The variation in cytosine methylation in individuals in each population exhibited substantial differences, confirming the necessity of population epigenetics. The total methylation levels of the diploid population were significantly higher than in the parents, but those of the three allotriploid populations were significantly lower than in the parents, indicating that both hybridization and polyploidization contributed to cytosine methylation variation. The vast majority of methylated status could be inherited from the parents, and the average percentages of non-additive variation were 6.29, 3.27, 5.49 and 5.07% in the diploid, FDR, SDR and PMR progeny populations, respectively. This study lays a foundation for further research on population epigenetics in allopolyploids.</p></div

Inheritance and variation of cytosine methylation patterns in the four hybrid progeny populations.

<p>Note: a, the number of corresponding sites; b, the frequency of corresponding sites (%).</p><p>Inheritance and variation of cytosine methylation patterns in the four hybrid progeny populations.</p

Result of ploidy level detection.

<p>(A) Flow cytometric analysis of nuclei mixtures from young leaves of the diploid control and triploid hybrid. (B) Somatic chromosome number in the diploid control (2n = 2x = 38), scale bar = 5 μm.</p

Genome-Wide Identification and Expression Analysis of the PME and PMEI Gene Families in <i>Diospyros kaki</i>: A Bioinformatics Study

Pectins are major components of cell walls in plants. Pectin methylesterases (PMEs) and pectin methylesterase inhibitors (PMEIs) play crucial roles in pectin synthesis and metabolism. Overall, 28 putative DkPMEs and 29 putative DkPMEIs were identified from the D. kaki genome. According to phylogenetic analysis, DkPME/DkPMEI proteins can be classified into four and five clades, respectively. Motif and gene structure analysis showed that DkPME/DkPMEI are highly conserved in the same clades, which indicates that the function of these DkPME/DkPMEI were similar. Besides, DkPME/DkPMEI genes were distributed unevenly on their corresponding chromosomes. Synteny analysis showed that PME or PMEI gene usually matched with more than one DkPME/DkPMEI in D. oleifera, D. lotus, and A. thaliana, implying that the function of these genes in D. kaki may be diverse. Expression analysis showed that DkPME/DkPMEI from the same clade exhibited diverse expression patterns, indicating that these genes might have diverse functions. Functional protein–protein interaction network analysis showed that DkPMEI21 and DkPMEI15 were core nodes and were, respectively, positive and negative regulators for carbohydrate metabolism, stress responses, and sugar signaling. This study provides a theoretical basis for the functional characteristics, evolutionary relationship, and role of these gene families in developing persimmon fruit

Genome-Wide Identification and Expression Analysis of the PME and PMEI Gene Families in Diospyros kaki: A Bioinformatics Study

Pectins are major components of cell walls in plants. Pectin methylesterases (PMEs) and pectin methylesterase inhibitors (PMEIs) play crucial roles in pectin synthesis and metabolism. Overall, 28 putative DkPMEs and 29 putative DkPMEIs were identified from the D. kaki genome. According to phylogenetic analysis, DkPME/DkPMEI proteins can be classified into four and five clades, respectively. Motif and gene structure analysis showed that DkPME/DkPMEI are highly conserved in the same clades, which indicates that the function of these DkPME/DkPMEI were similar. Besides, DkPME/DkPMEI genes were distributed unevenly on their corresponding chromosomes. Synteny analysis showed that PME or PMEI gene usually matched with more than one DkPME/DkPMEI in D. oleifera, D. lotus, and A. thaliana, implying that the function of these genes in D. kaki may be diverse. Expression analysis showed that DkPME/DkPMEI from the same clade exhibited diverse expression patterns, indicating that these genes might have diverse functions. Functional protein&ndash;protein interaction network analysis showed that DkPMEI21 and DkPMEI15 were core nodes and were, respectively, positive and negative regulators for carbohydrate metabolism, stress responses, and sugar signaling. This study provides a theoretical basis for the functional characteristics, evolutionary relationship, and role of these gene families in developing persimmon fruit

Physiological Characteristics and Transcriptional Differences of Growth Traits of Persimmon with Different Ploidy

Ploidy breeding is one of the important approaches for persimmon (Diospyros kaki Thunb.) genetic improvement, and vegetative growth of seedlings is the basis for subsequent fruit development. Therefore, the physiological characteristics and transcriptional differences of seedling growth traits in different ploidy persimmon germplasm were studied in this study, which provided a theoretical basis for fruit evaluation and breeding of new polyploid persimmon varieties. Nonuploid and its full-sib hexaploid seedlings obtained from endosperm culture were used as materials. By observing plant phenotype, leaf tissue section, endogenous hormone content, and transcriptome sequencing, the phenotype and physiological characteristics of different ploidy Persimmon seedlings were compared, as well as the differences in transcription levels. (1) Compared with hexaploid seedlings, the nonuploid were more robust and compact, and the leaves were obviously thicker. The cell size of leaf veins and parenchyma were significantly different between the different ploidy plants. (2) The contents of Salicylic Acid (SA), Jasmonic Acid (JA), gibberellin A1 (GA1), gibberellin A3 (GA3) and Indole-3-acetic acid (IAA) in nonuploid leaves were significantly higher than those in hexaploid leaves, while the contents of cytokinin trans-zeanoside (Tzt) and dihydrozeanoside (DZR), N6-isopentenyladenine (iP) and Jasmonoyl-L-isoleucine (JA-ILE) in nonuploid leaves were significantly lower than those in hexaploid leaves. (3) A total of 5796 differentially expressed genes were identified in nonuploid and hexaploid leaves. These differentially expressed genes were mainly related to photosynthesis, plant-pathogen interaction, etc. Among them, YUCCA genes, GA3ox genes, and IPT genes related to hormone synthesis were significantly differentially expressed in the nonuploid and hexaploid leaves. It is speculated that it may be the key regulatory gene that leads to the difference in IAA, gibberellin (GA), and indolepropionic acid (IPA) levels in the nonuploid and the hexaploid. The growth traits of the new Persimmon germplasms with different ploidy were significantly different. The nonuploid plants were shorter and more compact, and the leaves were larger and thicker. These traits were closely related to the content of endogenous hormones, and the balance of endogenous hormones was affected by gene expression. In addition, based on the biological processes involved in hormones and differentially expressed genes, it is speculated that the nonuploid may be superior to the hexaploid in terms of resistance

Study of Sexual-Linked Genes (<i>OGI</i> and <i>MeGI</i>) on the Performance of Androecious Persimmons (<i>Diospyros kaki</i> Thunb.)

It is reported that the production of floral sexual phenotype in hexaploid monoecious persimmon (Diospyros kaki) is closely related to a pseudogene called OGI, and a short interspersed nuclear element (SINE)-like insertion (named Kali) in the OGI promoter leads to the gene silence. As a result, DNA methylation level of MeGI promoter determines the development of male or female flowers. However, the molecular mechanism in androecious D. kaki, which only bear male flowers, remains elusive. Here, real-time quantitative polymerase chain reaction (RT-qPCR), molecular cloning, and bisulfite PCR sequencing technique were carried out using 87 materials, including 56 androecious resources, 15 monoecious, and 16 gynoecious cultivars, to investigate the performance of OGI and MeGI on the specific androecious type of D. kaki in China. In conclusion, the Kali insertion was exactly located in the OGI promoter region, and the OGI gene and the Kali sequence were existing and conserved in androecious D. kaki. Meanwhile, we also demonstrated that the MeGI gene was widespread in our investigated samples. Ultimately, our result convincingly provided evidence that the low expression of OGI is probably ascribed to the presence of Kali displaying strong methylation in the OGI promoter, and low expression of MeGI, as well as high DNA methylation level, in the promoter was closely connected with the production of male flowers; this result was consistent with the monoecious persimmon model. Our findings provide predominant genetic aspects for investigation into androecious D. kaki, and future perfecting the sex-determining mechanisms in persimmon