Percorrer por autor "Coesel, Sacha"
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- Evolutionary origins and functions of the carotenoid biosynthetic pathway in marine diatomsPublication . Coesel, Sacha; Obornik, Miroslav; Varela, J.; Falciatore, Angela; Bowler, ChrisCarotenoids are produced by all photosynthetic organisms, where they play essential roles in light harvesting and photoprotection. The carotenoid biosynthetic pathway of diatoms is largely unstudied, but is of particular interest because these organisms have a very different evolutionary history with respect to the Plantae and are thought to be derived from an ancient secondary endosymbiosis between heterotrophic and autotrophic eukaryotes. Furthermore, diatoms have an additional xanthophyll-based cycle for dissipating excess light energy with respect to green algae and higher plants. To explore the origins and functions of the carotenoid pathway in diatoms we searched for genes encoding pathway components in the recently completed genome sequences of two marine diatoms. Consistent with the supplemental xanthophyll cycle in diatoms, we found more copies of the genes encoding violaxanthin de-epoxidase (VDE) and zeaxanthin epoxidase (ZEP) enzymes compared with other photosynthetic eukaryotes. However, the similarity of these enzymes with those of higher plants indicates that they had very probably diversified before the secondary endosymbiosis had occurred, implying that VDE and ZEP represent early eukaryotic innovations in the Plantae. Consequently, the diatom chromist lineage likely obtained all paralogues of ZEP and VDE genes during the process of secondary endosymbiosis by gene transfer from the nucleus of the algal endosymbiont to the host nucleus. Furthermore, the presence of a ZEP gene in Tetrahymena thermophila provides the first evidence for a secondary plastid gene encoded in a heterotrophic ciliate, providing support for the chromalveolate hypothesis. Protein domain structures and expression analyses in the pennate diatom Phaeodactylum tricornutum indicate diverse roles for the different ZEP and VDE isoforms and demonstrate that they are differentially regulated by light. These studies therefore reveal the ancient origins of several components of the carotenoid biosynthesis pathway in photosynthetic eukaryotes and provide information about how they have diversified and acquired new functions in the diatoms.
- Isolation and characterization of a stress-inducible Dunaliella salina Lcy-β gene encoding a functional lycopene β-cyclasePublication . Ramos, Ana; Coesel, Sacha; Marques, Ana; Rodrigues, Marta; Baumgartner, Alexandra; Noronha, João; Rauter, Amélia; Brenig, Bertram; Varela, J.The halotolerant green alga Dunaliella salina accumulates large amounts of β-carotene when exposed to various stress conditions. Although several studies concerning accumulation and biotechnological production of β-carotene have been published, the molecular basis and regulation of the genes involved in carotenoid biosynthesis in D. salina are still poorly known. In this paper, we report the isolation and regulation of the lycopene β-cyclase (Lcy-β) gene by abiotic stress. The function of this gene was determined by heterologous genetic complementation in E. coli. Gene expression and physiological analyses revealed that D. salina Lcy-β steady-state transcript and carotenoid levels were up-regulated in response to all stress conditions tested (salt, light and nutrient depletion). The results presented here suggest that nutrient availability is a key factor influencing carotenogenesis as well as carotenoid biosynthesis-related gene expression in D. salina.
- Nutrient limitation is the main regulatory factor for carotenoid accumulation and for Psy and Pds steady state transcript levels in Dunaliella salina (Chlorophyta) exposed to high light and salt stressPublication . Coesel, Sacha; Baumgartner, Alexandra; Teles, Lícia; Ramos, Ana; Henriques, Nuno; Cancela, Leonor; Varela, J.Dunaliella salina (Dunal) Teodoresco (1905) is a green unicellular alga able to withstand severe salt, light, and nutrient stress, adaptations necessary to grow in harsh environments such as salt ponds. In response to such growth conditions, this microalga accumulates high amounts of β-carotene in its single chloroplast. In this study, we show that carotenoid accumulation is consistently inhibited in cells grown in nutrient-supplemented media and exposed either to high-light or medium-low-light conditions. Likewise, carotenogenesis in cells shifted to higher salinity (up to 27% NaCl) under medium-low-light conditions is inhibited by the presence of nutrients. The steady-state levels of transcripts encoding phytoene syn- thase and phytoene desaturase increased substantially in D. salina cells shifted to high light or high salt under nutrient- limiting conditions, whereas the presence of nutrients inhibited this response. The regulatory effect of nutrient availability on the accumulation of carotenoids and mes- senger RNA levels of the first two enzymes committed to carotenoid biosynthesis is discussed.
- Regulation of the carotenoid biosynthetic pathway in the green microalga Dunaliella salina and the diatom Phaeodactylum tricornutumPublication . Coesel, Sacha; Varela, João; Bowler, ChrisCarotenoids are produced by all photosynthetic organisms where they play indispensable roles in light-harvesting and photoprotection. This thesis has focused on carotenoid biosynthesis in Dunaliella salina and Phaeodactylum tricornutum, two phylogenetically diverse algae. Both algae are able to maintain high photosynthetic rates under fluctuating light intensities. We examined the effect of several environmental stress conditions on carotenoid biosynthesis in D. salina and found that nutrient level, light intensity and salinity have a differential effect on carotenogenesis. We also found that the steady-state transcript levels of two key-enzymes involved in the early steps of carotenoid biosynthesis are coordinately up-regulated in carotenoid-accumulating D. salina cells, indicating that carotenoid biosynthesis in this alga may be partly regulated at the transcriptional level. Analysis of the P. tricornutum genome, as well as that from another diatom, Thalassiosira pseudonana, revealed that the genes involved in xanthophyll biosynthesis and xanthophyll cycling in diatoms have diversified greatly with respect to green algae and higher plants. We showed that the steady-state mRNA levels of the P. tricornutum carotenoid biosynthesis-related genes are increased upon nutrient stress and blue light, and we were able to establish that light of different spectral quality has a differential effect on the mRNA levels of these genes. By using transgenic P. tricornutum cell lines containing elevated levels of a putative diatom blue light cryptochrome photoreceptor, we demonstrated that this protein is involved in the transcriptional regulation of blue light-responsive genes, ultimately resulting in an enhanced accumulation of xanthophyll pigments and a significantly altered chromatic adaptation to blue light. In conclusion, the work reported in this thesis will facilitate future work on both the regulatory and biotechnological aspects of the carotenoid biosynthetic pathway in unicellular algae.