Dynamics of Calcium during Microspore Embryogenesis and Microspore Development in and .

Calcium is widely known to have a role as a signaling molecule in many different processes, including stress response and activation of the embryogenic program. However, there are no direct clues about calcium levels during microspore embryogenesis, an experimental process that combines a developmental switch toward embryogenesis and the simultaneous application of different stressing factors. In this work, we used FluoForte, a calcium-specific fluorescent vital dye, to track by confocal microscopy the changes in levels and subcellular distribution of calcium in living rapeseed ( and eggplant microspores and pollen grains during development, as well as during the first stages of -induced microspore embryogenesis in rapeseed. During development, a clear peak of cytosolic Ca was observed in rapeseed vacuolate microspores and young pollen grains, the stages more suitable for embryogenesis induction. However, the Ca levels observed in eggplant were dramatically lower than in rapeseed. Just after induction, Ca levels increased specifically in rapeseed embryogenic microspores at levels dramatically higher than during development. The increase was observed in the cytosol, but predominantly in vacuoles. Non-embryogenic forms such as callus-like and pollen-like structures presented remarkably different calcium patterns. After the heat shock-based inductive treatment, Ca levels progressively decreased in all cases. Together, our results reveal unique calcium dynamics in rapeseed microspores, as well as in those reprogrammed to embryogenesis, establishing a link between changes in Ca level and subcellular distribution, and microspore embryogenesis.