Pink lemon is a spontaneous bud mutation of lemon (Citrus limon, L. Burm. f) characterized by the production of pink-fleshed fruits due to an unusual accumulation of lycopene. To elucidate the genetic determinism of the altered pigmentation, comparative carotenoid profiling and transcriptional analysis of both the genes involved in carotenoid precursors and metabolism, and the proteins related to carotenoid-sequestering structures were performed in pink-fleshed lemon and its wild-type. The carotenoid profile of pink lemon pulp is characterized by an increased accumulation of linear carotenoids, such as lycopene, phytoene and phytofluene, from the early stages of development, reaching their maximum in mature green fruits. The distinctive phenotype of pink lemon is associated with an up-regulation and down-regulation of the genes upstream and downstream the lycopene cyclase, respectively. In particular, 9-cis epoxycarotenoid dioxygenase genes were overexpressed in pink lemon compared with the wild-type, suggesting an altered regulation of abscisic acid biosynthesis. Similarly, during early development of the fruits, genes of the carotenoid-associated proteins heat shock protein 21, fibrillin 1 and 2 and orange gene were overexpressed in the pulp of the pink-fleshed lemon compared to the wild-type, indicating its increased capacity for sequestration, stabilization or accumulation of carotenes. Altogether, the results highlighted significant differences at the transcriptomic level between the pink-fleshed lemon and its wild-type, in terms of carotenoid metabolism and the capacity of stabilization in storage structures between the two accessions. Such changes may be either responsible for the altered carotenoid accumulation or in contrast, a metabolic consequence.

Rootstock choice has important effects on the horticultural and pathological traits of the citrus cultivars. Thus, the scion/rootstock combination can affect tree vigour, nutrition, and stress resistance; it can also have positive influences on the fruit quality traits. Although the study of rootstock effects has been a relevant research topic in citrus for many years, the main body of such study has been conducted at the biochemical level, while little effort has been directed to the determination of the rootstock influences at the molecular level. A comparative study of three combinations of scion and rootstock shows a positive correlation between the regulation of the fruit quality-related genes and the accumulations of bioactive compounds, as well as with acid degradation. Monitoring the anthocyanin accumulation during ripening shows the scion/rootstock combination can increase anthocyanin synthesis in the fruit, as well as vitamin C accumulation and acid degradation. Our results show that the rootstock genotype can exert important influences on citrus fruit quality by affecting gene expression in the scion. New insights into the molecular interactions between scion and rootstock may help unravel the systems through which rootstocks exert their influences on the regulatory networks in the scion, so as to influence relevant agronomic traits. This information should result in an improved rootstock breeding selection and definition of scion/rootstock combinations to enhance fruit quality traits.

Male gametophyte development is a critical phase of the plant life cycle due to its high sensitivity to environmental stresses. The rise in the average global temperature, often accompanied by extreme fluctuations, has an important impact on biological processes. Among those, male gametophytes are particularly sensitive to temperature stress during flower bud development and anthesis. Male gametophyte development was extensively studied in several plant species, but little information is available about the effects of temperature stress on male gametophyte development in the genus Citrus. We evaluated the effects of cold and hot temperatures during microsporogenesis and microgametogenesis on one of the most economically valuable citrus species, the "Comune" clementine (Citrus clementina Hort. ex. Tan.). The effect of constant temperature on the androecium was evaluated by a time course histological analysis performed on the anthers and by monitoring in vitro pollen germination. The results revealed how suboptimal hot and cold temperatures induce drastic alterations on the morphology of the tapetal cells, microspores and mature pollen grains. Shifting from the optimal temperature affected the timing of starch depletion in the anther walls, such as epidermis, endothecium and middle layer,influencing the pollen germination rate and pollen tube growth. To the best of our knowledge this is the first study attempting to assess how temperature stress affects male reproductive development in citrus. A better understanding of the mechanisms underlining male sterility will provide novel insights to elucidate the physiology of this agronomical important quality trait.

In Citrus, flower induction represents the transition from vegetative to reproductive growth. The regulation of flower induction is mainly triggered by exposure to low temperatures and water-deficit stress, which activates the signaling cascade leading to an increased expression of the citrus orthologs of the FLOWERING LOCUS T (CiFT). In this study, the relationship between rootstock and flower induction under Mediterranean field conditions was investigated by monitoring the expression levels of the floral promoter CiFT2 in leaves of the pigmented sweet orange “Tarocco Scirè” grafted onto “C35” citrange and “Swingle” citrumelo rootstocks. The latter two are known to confer, respectively, high and low yield efficiency to the scion. In both rootstock/scion combinations, CiFT2 showed a seasonal expression with a peak during the inductive period in January triggered by cold temperature. The “Tarocco Scirè”/”C35” citrange combination showed the highest expression levels for CiFT2; this increased expression was correlated with yield and a higher number of flowers in the following spring, suggesting a significant effect of rootstocks on flower induction mediated by the overexpression of the CiFT2 gene.