Comprehensive cell-specific protein analysis in early and late pollen development from diploid microsporocytes to pollen tube growth

Pollen development in angiosperms is one of the most important processes controlling plant productivity. At the same time pollen development is highly sensitive to environmental fluctuations including temperature, drought and nutrition. Therefore pollen biology is a major focus in applied studies and breeding approaches for improving plant productivity in a globally changing climate. The most accessible developmental stages of pollen are the mature pollen and the pollen tubes and are, thus, most frequently analyzed. To reveal a complete quantitative proteome map we specifically adressed very early stages of tobacco pollen development from diploid microsporocytes, meioses, tetrads, microspores, polarized microspores, bipolar pollen, desiccated pollen and finally pollen tubes. A protocol for the isolation of the early stages was established. Proteins were extracted and analysed by a new Gel-LC-MS fractionation protocol. In total, 3825 proteins were identified. Quantitative analysis was performed based on peptide count or ion intensity. Exceedingly stage-specific differential protein regulation was observed during the conversion from the sporophytic to the gametophytic proteome. A map of highly specialized functionality for the different stages can be revealed from metabolic activity, pronounced differentiation of proteasomal and ribosomal protein complex composition up to protective mechanisms such as high levels of heat shock proteins in the very early stages of development.