Insights into the Cytochrome P450 Monooxygenase Superfamily in Osmanthus fragrans and the Role of OfCYP142 in Linalool Synthesis.

Osmanthus fragrans flowers have long been used as raw materials in food, tea, beverage, and perfume industries due to their attractive and strong fragrance. The P450 superfamily proteins have been reported to widely participate in the synthesis of plant floral volatile organic compounds (VOCs). To investigate the potential functions of P450 superfamily proteins in the fragrance synthesis of O. fragrans, we investigated the P450 superfamily genome wide. A total of 276 P450 genes were identified belonging to 40 families. The RNA-seq data suggested that many OfCYP genes were preferentially expressed in the flower or other organs, and some were also induced by multiple abiotic stresses. The expression patterns of seven flower-preferentially expressed OfCYPs during the five different flower aroma content stages were further explored using quantitative real-time PCR, showing that the CYP94C subfamily member OfCYP142 had the highest positive correlation with linalool synthesis gene OfTPS2. The transient expression of OfCYP142 in O. fragrans petals suggested that OfCYP142 can increase the content of linalool, an important VOC of the O. fragrans floral aroma, and a similar result was also obtained in flowers of OfCYP142 transgenic tobacco. Combined with RNA-seq data of the transiently transformed O. fragrans petals, we found that the biosynthesis pathway of secondary metabolites was significantly enriched, and many 2-C-methyl-D-erythritol 4-phosphate (MEP) pathway genes were also upregulated. This evidence indicated that the OfCYP proteins may play critical roles in the flower development and abiotic response of O. fragrans, and that OfCYP142 can participate in linalool synthesis. This study provides valuable information about the functions of P450 genes and a valuable guide for studying further functions of OfCYPs in promoting fragrance biosynthesis of ornamental plants.

Characterization of OfWRKY3, a transcription factor that positively regulates the carotenoid cleavage dioxygenase gene OfCCD4 in Osmanthus fragrans.

The sweet osmanthus carotenoid cleavage dioxygenase 4 (OfCCD4) cleaves carotenoids such as ß-carotene and zeaxanthin to yield ß-ionone. OfCCD4 is a member of the CCD gene family, and its promoter contains a W-box palindrome with two reversely oriented TGAC repeats, which are the proposed binding sites of WRKY transcription factors. We isolated three WRKY cDNAs from the petal of Osmanthus fragrans. One of them, OfWRKY3, encodes a protein containing two WRKY domains and two zinc finger motifs. OfWRKY3 and OfCCD4 had nearly identical expression profile in petals of 'Dangui' and 'Yingui' at different flowering stages and showed similar expression patterns in petals treated by salicylic acid, jasmonic acid and abscisic acid. Activation of OfCCD4pro:GUS by OfWRKY3 was detected in coinfiltrated tobacco leaves and very weak GUS activity was detected in control tissues, indicating that OfWRKY3 can interact with the OfCCD4 promoter. Yeast one-hybrid and electrophoretic mobility shift assay showed that OfWRKY3 was able to bind to the W-box palindrome motif present in the OfCCD4 promoter. These results suggest that OfWRKY3 is a positive regulator of the OfCCD4 gene, and might partly account for the biosynthesis of ß-ionone in sweet osmanthus.

A one-locus model of androdioecy with two homomorphic self-incompatibility groups: expected vs. observed male frequencies.

Androdioecy, the occurrence of males and hermaphrodites in a single population, is a rare breeding system because the conditions for maintenance of males are restrictive. In the androdioecious shrub Phillyrea angustifolia, high male frequencies are observed in some populations. The species has a sporophytic self-incompatibility (SI) system with two self-incompatibility groups, which ensures that two groups of hermaphrodites can each mate only with the other group, whereas males can fertilize hermaphrodites of both groups. Here, we analyse a population genetic model to investigate the dynamics of such an androdioecious species, assuming that self-incompatibility and sex phenotypes are determined by a single locus. Our model confirms a previous prediction that a slight reproductive advantage of males relative to hermaphrodites allows the maintenance of males at high equilibrium frequencies. The model predicts different equilibria between hermaphrodites of the two SI groups and males, depending on the male advantage, the initial composition of the population and the population size, whose effect is studied through stochastic simulations. Although the model can generate high male frequencies, observed frequencies are considerably higher than the model predicts. We finally discuss how this model may help explain the large male frequency variation observed in other androdioecious species of Oleaceae: some species show only androdioecious populations, as P. angustifolia, whereas others show populations either completely hermaphrodite or androdioecious.

The role of staminate flowers in the breeding system of Olea europaea (Oleaceae): an andromonoecious, wind-pollinated taxon.

BACKGROUND AND AIMS: Andromonoecy, as a breeding system, has generated a considerable body of theory in terms of sexual selection, but extended records comparing the performance of pollen grains from staminate versus hermaphrodite flowers are still sparse. The objective in this study was to elucidate the role of staminate flowers in the andromonoecious breeding system of olive (Olea europaea). METHODS: To determine the meaning of staminate flowers, an evaluation was made of resource allocation to, and phenology of, staminate and hermaphrodite flowers in the cultivar 'Mission', and a comparison was made of the male function between both kinds of flowers. KEY RESULTS: Dry weight of hermaphrodite flowers was 19 % greater than dry weight of staminate flowers arising in comparable positions of the panicle. This difference was mainly due to pistil and petal weight; there were no significant differences in stamen weight. There were no significant differences between staminate and hermaphrodite flowers in either amount of pollen per anther, or pollen quality as determined by pollen viability, germinability or ability to fertilize other flowers. There was no significant link between gender and time of anthesis. However, position of the flower within the panicle correlated with time of anthesis and gender. Flowers at the apex and at primary pedicels tended to be hermaphrodite and open earlier, whereas flowers arising in secondary pedicels were mainly staminate and were commonly the last to reach anthesis. CONCLUSIONS: It is proposed that the main advantage provided by production of staminate flowers in olive is to enhance male fitness by increasing pollen output at the whole plant level, although a relict function of attracting pollinators cannot be completely discarded.