Tomato Pollen Wall Development, and the Role of FIN4 in the Formation of the Pollen Intine

The cell wall of a mature pollen grain is a highly specialized, multilayered structure. The outer, sporopollenin-based exine protects and supports the pollen grain, while the inner intine, composed primarily of cellulose, is essential for pollen germination. The process of pollen wall development is complex; multiple cell wall polymers are deposited, some transiently, then removed or remodeled in a controlled sequence of events. Here, I describe the structure of the tomato pollen wall and establish a developmental timeline of its formation. I also characterize a mutant with an intine phenotype, fin4, and describe its role in tomato pollen wall development. Using histology and immunostaining, I determined how key cell wall polymers were deposited with respect to overall pollen and anther development. Pollen development began in young flower buds when the pre-meiotic microspore mother cells (MMCs) began losing their cellulose primary cell wall. Following meiosis, the still conjoined microspores progressed to the tetrad stage characterized by a temporary, thick callose wall. Breakdown of the callose wall released the individual early microspores. Exine deposition began with the secretion of the sporopollenin foot layer. At the late microspore stage, exine deposition was completed, and the tapetum degenerated. The pollen underwent mitosis to produce bicellular pollen, at which point intine formation began, continuing through to pollen maturation. The entire cell wall development process was also punctuated by dynamic changes in pectin composition, particularly changes in methyl-esterified and de-methyl-esterified homogalacturonan. While the development of the exine layer is better understood due to the characterization of several mutants with exine phenotypes, isolation of homozygous mutants with intine phenotypes has presented significant challenges, limiting the understanding of the development and functioning of the intine layer. This work also characterizes the phenotype of the mutant of the hydroxyproline O-arabinosyltransferase FIN4 using the framework of the pollen wall development timeline. FIN4 was highly expressed during pollen development in the tapetum and developing microspores through mature pollen. In the absence of FIN4 activity, fin4 mutants had severely reduced male fertility and, while the released pollen was viable, it had poor germination and hydration ability. Closer examination uncovered that fin4 pollen had weaker cell walls. This was due to the failure to develop the intine layer of the pollen wall properly. This intine defect traces to reduced cellulose deposition, but an increased abundance of pectin in the bicellular pollen of fin4 compared to wild-type. This work helps elucidate not only the timeline of pollen wall development inside the anther, but it also furthers the understanding of pollen intine development process and the role of intine in the proper function of pollen grain and male fertility.PHDMolecular, Cellular, and Developmental BiologyUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/171404/1/srfj_1.pd