Thesis (Ph.D.)--University of Washington, 2013The plant epidermis is a critical interface between the atmosphere and internal plant tissues, which allows plants to succeed on land by restricting their exposure to the environment. Stomata, closable pores on the plant surface bounded by specialized guard cells, are an integral part of epidermal function. By controlling water loss and carbon dioxide uptake, stomata regulate global carbon and water cycles. Stomata undergo a complex course of development involving cell-cell signaling and sequential action of master regulatory transcription factors, making stomata a suitable system for studying widely applicable developmental processes necessary for tissue and organ development. This dissertation presents an examination of molecular characteristics, gene function, and protein dynamics in stomatal lineage cells, including preliminary results of manipulating cell fate during stomatal development. We performed molecular profiling of the stem-cell-like stomatal precursor, the meristemoid, by enriching cell types through synthetic mutations. This uncovered new genes involved in stomatal development as well as molecular commonalities with the main plant stem-cell niches at the shoot and root apices. A novel gene, POLAR, was found to localize asymmetrically in dividing stomatal-lineage cells. Also uncovered through meristemoid profiling was a transcription factor, HOMEODOMAIN GLABROUS 2 (HDG2), which is highly expressed in stomatal lineage cells and sufficient to convert internal leaf cells to stomata when ectopically expressed. Loss of function in HDG2 hinders stomatal development after initiation and causes aberrant stomata; further loss of its close relative AtML1 magnifies the effect. To further investigate the role of transcription factors in stomatal development, we used time lapse microscopy to observe protein dynamics of stomatal regulators in germinating cotyledons. Cell-cell signaling in the cotyledon was perturbed using laser ablation of stomatal-lineage cells, and preliminary results indicate that cell fate was thus affected. Cotyledon time lapse revealed an unexpected developmental sequence indicating possible epidermal prepatterning, so we employed an embryo time lapse technique to discover that both regulatory genes and signaling components were active in the embryo, indicating that stomatal development begins during embryonic development. This work demonstrates that stomatal development exemplifies crucial developmental processes and provides novel insight into how cell fate is dynamically specified by tissue-level regulation, cell-cell signaling, and cell-autonomous molecular mechanisms in plants
