Effect of heterologous expression of FT gene from Medicago truncatula in growth and flowering behavior of olive plants.

Olive (Olea europaea L. subsp. europaea) is one of the most important crops of the Mediterranean Basin and temperate areas worldwide. Obtaining new olive varieties adapted to climatic changing conditions and to modern agricultural practices, as well as other traits such as biotic and abiotic stress resistance and increased oil quality, is currently required; however, the long juvenile phase, as in most woody plants, is the bottleneck in olive breeding programs. Overexpression of genes encoding the 'florigen' Flowering Locus T (FT), can cause the loss of the juvenile phase in many perennials including olives. In this investigation, further characterization of three transgenic olive lines containing an FT encoding gene from Medicago truncatula, MtFTa1, under the 35S CaMV promoter, was carried out. While all three lines flowered under in vitro conditions, one of the lines stopped flowering after acclimatisation. In soil, all three lines exhibited a modified plant architecture; e.g., a continuous branching behaviour and a dwarfing growth habit. Gene expression and hormone content in shoot tips, containing the meristems from which this phenotype emerged, were examined. Higher levels of OeTFL1, a gene encoding the flowering repressor TERMINAL FLOWER 1, correlated with lack of flowering. The branching phenotype correlated with higher content of salicylic acid, indole-3-acetic acid and isopentenyl adenosine, and lower content of abscisic acid. The results obtained confirm that heterologous expression of MtFTa1 in olive induced continuous flowering independently of environmental factors, but also modified plant architecture. These phenotypical changes could be related to the altered hormonal content in transgenic plants.

Sexual and Apogamous Species of Woodferns Show Different Protein and Phytohormone Profiles.

The gametophyte of ferns reproduces either by sexual or asexual means. In the latter, apogamy represents a peculiar case of apomixis, in which an embryo is formed from somatic cells. A proteomic and physiological approach was applied to the apogamous fern Dryopteris affinis ssp. affinis and its sexual relative D. oreades. The proteomic analysis compared apogamous vs. female gametophytes, whereas the phytohormone study included, in addition to females, three apogamous stages (filamentous, spatulate, and cordate). The proteomic profiles revealed a total of 879 proteins and, after annotation, different regulation was found in 206 proteins of D. affinis and 166 of its sexual counterpart. The proteins upregulated in D. affinis are mostly associated to protein metabolism (including folding, transport, and proteolysis), ribosome biogenesis, gene expression and translation, while in the sexual counterpart, they account largely for starch and sucrose metabolism, generation of energy and photosynthesis. Likewise, ultra-performance liquid chromatography-tandem spectrometry (UHPLC-MS/MS) was used to assess the levels of indol-3-acetic acid (IAA); the cytokinins: 6-benzylaminopurine (BA), trans-Zeatine (Z), trans-Zeatin riboside (ZR), dyhidrozeatine (DHZ), dyhidrozeatin riboside (DHZR), isopentenyl adenine (iP), isopentenyl adenosine (iPR), abscisic acid (ABA), the gibberellins GA3 and GA4, salicylic acid (SA), and the brassinosteroids: brassinolide (BL) and castasterone (CS). IAA, the cytokinins Z, ZR, iPR, the gibberellin GA4, the brassinosteoids castasterone, and ABA accumulated more in the sexual gametophyte than in the apogamous one. When comparing the three apogamous stages, BA and SA peaked in filamentous, GA3 and BL in spatulate and DHRZ in cordate gametophytes. The results point to the existence of large metabolic differences between apogamous and sexual gametophytes, and invite to consider the fern gametophyte as a good experimental system to deepen our understanding of plant reproduction.

Hormonal and gene dynamics in de novo shoot meristem formation during adventitious caulogenesis in cotyledons of Pinus pinea.

KEY MESSAGE: Several members of WOX and KNOX gene families and several plant growth regulators, basically cytokinins and auxins, play a key role during adventitious caulogenesis in the conifer Pinus pinea. Similar to Arabidopsis thaliana, Pinus pinea shoot organogenesis is a multistep process. However, there are key differences between both species, which may alter the underlying physiological and genetic programs. It is unknown if the genic expression models during angiosperm development may be applicable to conifers. In this work, an analysis of the endogenous content of different plant growth regulators and the expression of genes putatively involved in adventitious caulogenesis in P. pinea cotyledons was conducted. A multivariate analysis of both datasets was also realized through partial least squares regression and principal component analysis to obtain an integral vision of the mechanisms involved in caulogenesis in P. pinea. Analyses show that cotyledons cultured in the presence of benzyladenine during long times (2-6 days) cluster separately from the rest of the samples, suggesting that the benzyladenine increase observed during the first hours of culture is sufficient to trigger the caulogenic response through the activation of specific developmental programs. In particular, the most relevant factors involved in this process are the cytokinins trans-zeatin, dihydrozeatin, trans-zeatin riboside and isopentenyl adenosine; the auxin indoleacetic acid; and the genes PpWUS, PpWOX5, PpKN2, PpKN3 and PipiRR1. WUS is functional in pines and has an important role in caulogenesis. Interestingly, WOX5 also seems to participate in the process, although its specific role has not been determined.

Interaction between environmental factors affects the accumulation of root proteins in hydroponically grown Eucalyptus globulus (Labill.).

Eucalyptus globulus (Labill.) is used for pulp and paper production worldwide. In this report we studied changes in protein expression in one osmotically stressed elite clone widely used in industrial plantations in Spain. High molecular weight polyethylene glycol (PEG) was used as an osmoticum in the growing medium. Roots of rooted cuttings were sampled after 3 and 36 h of treatment. Water potential and abscissic acid content were measured in shoot and root apices to characterize the physiological states of the plants. Total soluble proteins from roots were extracted and separated using two-dimensional gel electrophoresis (2-DE). Gels were stained with Coomassie brillant blue for quantitative analysis of protein accumulation. From a total of 406 reproducible spots, 34 were found to be differentially expressed depending on treatment (osmotic versus control condition) and/or stress duration (3 h versus 36 h), and were further characterized by tandem mass spectrometry. Several proteins were reliably identified including adenosine kinase, actin, stress-related proteins as well as proteins associated to cellular processes, among which some residents of the endoplasmic reticulum. This study constitutes the first investigation of the root proteome in this important forest tree genus.