Platelets play a major role in preventing bleeding. They are produced by giant and polyploid cells, megakaryocytes, present in the bone marrow, following a process involving cytoskeletal proteins and more particularly microtubules (MT).
MT is one of the three major components of the cell cytoskeleton with microfilaments and intermediate filaments. MTs result from the assembly of alpha and beta tubulins and form a dynamic network that plays a central role in multiple biological functions (cell division, intracellular trafficking and cellular morphogenesis). In cells, microtubules can be longitudinal or curved suggesting the existence of a local regulation of their mechanical properties, in connection with the functions in which they are engaged. In platelets, microtubules bend by adopting a characteristic ring structure consisting of a 10 to 15 microtubule coil called the marginal band. This original structure, specific to mammalian platelets, supports the discoid form of platelets and contributes to the maintenance of vascular homeostasis. In some patients with thrombopathies, there is a lack of formation of the marginal band leading to the impairment of platelet functions resulting in hemostasis defects.
The various functions related to MTs seem to depend on particular combinations of tubulin alpha / beta isoforms and post-translational modifications (MPTs) (acetylation, glycylation, glutamylation, tyrosination) thus allowing the recruitment of effectors (MAPs) dedicated to different functions ..
The thesis project we propose aims to study the particular functions of platelet MTs by a complete characterization of MPTs present on alpha and beta tubulins during platelet biogenesis.
A first study in platelets demonstrates an important role of acetylation in the platelet shape change in response to an agonist. Complete characterization of TMPs will be by immunoblotting and confocal microscopy using specific antibodies and identifying the enzymes involved in TMPs by real-time quantitative PCR. The role of these MPTs will be evaluated on megakaryocytes differentiated in culture i) from human hematopoietic progenitors (CD34 +) transduced by shRNAs specific for the identified enzymes and ii) from murine progenitors (Lin) from inactivated mice for enzymes interest.
This work will clarify the type and role of MPTs involved in megakaryocyte maturation and platelet formation. It is part of a broader team project that aims to better understand and characterize the role of microtubules during platelet biogenesis.

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