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Theses

Cellular and molecular characterization of O. cf. ovata cell cycle

David Velasquez 1 
1 mitose et contrôle du fuseau - LBDV_MITOSE_ET_CONTROLE_DU_FUSEAU
LBDV - Laboratoire de Biologie du Développement de Villefranche sur mer
Abstract : Dinoflagellates are protists with unique features among eukaryotes. The particular nucleus with chromosomes that remain condensed through the cell cycle and their unique kind of mitosis, called dinomitosis, set them apart from canonical eukaryotes and rise questions of how their cell cycle is controlled. Moreover, dinoflagellates are interesting from an ecological perspective, as they are the cause of massive and sometimes toxic proliferation events, called Harmful Algae Blooms (HABs). HABs are seasonal phenomena, which result mainly from an increase in cell division and aggregation through cell migration (Burkholder, et al., 2006). Ostreopsis cf. ovata is a cosmopolitan toxic dinoflagellate that produces seasonal blooms. Many ecological studies have addressed the parameters involved in the dynamic of Ostreopsis proliferation (Cohu, et al., 2011). However, it is currently unknown how this organism divides or how its cell cycle is controlled. The aim of this thesis was to provide a cellular and molecular characterization of the mitotic cell cycle, which underlies cell proliferation during vegetative growth, for the dinoflagellate O. cf. ovata. Morphological classification of cells collected at different phases of the bloom, allowed to identify different phases of the O. cf. ovata vegetative cycle, including cells undergoing division. Using this classification, I showed that during blooming, O. cf. ovata divides exclusively at night. Using immuno-fluorescence and confocal microscopy, I then characterized the changes in cytoskeletal organization associated with the O. cf. ovata cell cycle. During interphase, a ventral microtubule bundle protrudes from the ventral area, where the basal bodies are located, towards the nucleus and moves it to the center of the cell. Cells with a central nucleus are considered as pre-dividing. Once the nucleus is in the cell center, the mitotic spindle is organized perpendicularly to the long axis of the cell and segregates chromosomes to opposite cell ends. A cytokinetic plate made of microtubules starts to form in the dorsal area, opposite the basal bodies, already in anaphase. The cytokinetic plate grows unidirectionally towards the ventral side, dividing the cell along the longitudinal axis. To characterize O. cf. ovata cell cycle at the molecular level, I searched for O. cf. ovata homologues of conserved cell cycle regulators identified in model organisms, using a de novo transcriptome that I generated from cultured cells. This analysis revealed the presence of major components of the cell cycle such as cyclins and CDKs, proteins that are known to regulate mitotic entry, mitotic checkpoint components, kinetochore components and components of the budding yeast mitotic exit network. Cytokinetic and basal body components were also identified in the analysis. A differential expression analysis using meta-trascriptomic datasets corresponding to bloom samples enriched in interphase cells, pre-dividing cells and dividing, which I generated during the 2019 bloom, showed that more than 5000 transcripts were differentially expressed when comparing samples enriched in interphase cells and samples enriched in either pre-dividing or dividing cells. Among these transcripts, cyclin B, kinetochore and SAC components were up-regulated during the period of division. Instead, only 241 differentially expressed transcripts were present between pre-dividing or dividing cells. None of these transcripts was related with cell division, suggesting that once the cell is committed to mitosis (pre-dividing stage), the major regulations occur at the post-transcriptional level. By integrating the results obtained from the transcriptomic analysis with the cytological characterization I propose a first model of O. cf. ovata cell cycle, in which cyclins and CDK activities dictate the transition from interphase to mitosis.
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https://hal.archives-ouvertes.fr/tel-03355199
Contributor : Stefania Castagnetti Connect in order to contact the contributor
Submitted on : Monday, September 27, 2021 - 11:14:50 AM
Last modification on : Sunday, June 26, 2022 - 3:13:56 AM

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  • HAL Id : tel-03355199, version 1

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David Velasquez. Cellular and molecular characterization of O. cf. ovata cell cycle. Life Sciences [q-bio]. Sorbonne Université, 2021. English. ⟨tel-03355199⟩

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