GAPs in Plant Reproduction: Uncovering the Role of Glycosylphosphatidylinositol-Anchoring of Proteins in Arabidopsis Gametophyte Function

Abstract

Glycosylphoshpatidylinositol (GPI) is a complex glycolipid molecule biosynthesized in eukaryotic cells that can be post-translationally attached to a broad range of proteins by the transamidase complex (GPI-T) to serve as a membrane anchor. Apart from tethering GPI-anchored proteins (GAPs) to the extracellular leaflet of plasma membranes, GPI confers many molecular attributes to the proteins it attaches to. While the synthesis and structure of GPI is well conserved within eukaryotes, the utility of GPI-anchoring as a mode of protein membrane attachment shows striking variation among the eukaryotic kingdoms. For example, in protozoan parasites, GAPs are the major form of cell surface proteins and can shield the cell from host defenses during infection. Fungi such as yeast produce GPI anchors that can covalently link to polysaccharides in the cell wall to regulate its architecture. Animal GAPs are essential for coordinated growth during embryonic development and can act as signaling molecules to mediate cell-cell communication. In land plants, GAPs play roles in the directional growth and expansion of sporophytic tissues and are critical for the fertility of the gametophyte generation. Despite GAPs constituting about 1% of the Arabidopsis thaliana genome, it is poorly understood how this family of proteins function in plant development. Therefore, in this thesis, I investigated the role of GPI in plant reproduction by reviewing all 19 A. thaliana GAPs described to date with mutant fertility defects and annotated the expression of all 250 predicted GAPs in the A. thaliana gametophytes. Additionally, I analyzed mutations in two components of the GPI-T complex to investigate the importance of GPI-anchoring in male and female gametophyte functions. I showed that GPI-anchoring in the female gametophyte is apparently dispensable but is essential in the male gametophyte for pollen tube germination and growth. Based on my work, I propose that key differences in the structure and development of the male and female gametophytes may account for the difference in necessity of GPI-anchoring within their compartments. Thus, any conclusion regarding the importance of GPI-anchoring of proteins in the female gametophyte must consider these unique attributes. Lastly, because at least two GAPs involved in the female gametophyte function associate with receptor-like kinases (RLKs) to initiate signal transduction, we established several tools that can be used to elucidate the binding site of a well-studied GAP-RLK pair, the LORELEI-FERONIA co-receptor complex. The conclusions and tools generated in this study will help close the gaps in our understanding of the role of GAPs in plant reproduction.