EFFECT OF STEROLS AND SPHINGOLIPIDS DEPRIVATION ON POLLEN TUBE GROWTH IN NICOTIANA TABACUM (L.).

In Angiosperms, pollen tube can be considered a safe route to transport sperm cells in proximity of the female gamethophyte and its correct growth is necessary for a successful fertilization. Pollen tubes are highly specialized cells which follow an extreme form of polar growth, known as tip growth, based on transport of post-Golgi secretory vesicles (SVs) to the apical region where they fuse with a restricted area of the plasma membrane (PM) reversing outside cell wall components and supplying new surface of PM. This polarized secretion defines an apical growth PM domain and provides a specific protein/lipid composition. Recently, in tobacco pollen tube a polarized distribution of sterols and membrane lipid rafts have been reported and isolation and characterization of detergent insoluble membranes (DIMs) revealed that the physical basis for membrane raft formation is maintained in tobacco pollen tubes. In this work, using an experimental approach which considers the disorganization of lipid rafts by disturbing sterols and sphingolipids biosynthesis, we investigated the role of lipid rafts microdomains on tip growth in pollen tubes of the model specie Nicotiana tabacum (L.). The present results highligth a role for rafts microdomains in pollen tube growth and their involvement in cytoskeleton and organelles dynamics and consequently in the regulation of the polarized secretion. Biochemical and in vivo techniques show that changes in sterols/sphingolipids ratio in pollen tubes membranes alter actin association with microsomes and dynamics of the apical actin fringe. Alterations of morphology and dynamics of RabA4d compartments lead to postulate that the clear zone could represent a hub where different secretory pathways converge and functions as a sorting compartments. Also, perturbation of lipid rafts seems to mostly affect trans Golgi network (TGN), its maturation and the development of compartments which originate from it, involved both in secretion and in degradative pathway. Altogether these results support evidences of a role for sterols and sphingolipids, not only as fundamental structural components of membranes, but also in regulating intracellular membrane morphodynamics and polar secretion in pollen tubes.