Distinct functions of FASCILIN-LIKE ARABINOGALACTAN PROTEINS relate to domain structure.

The role of glycoproteins as key cell surface molecules during development and stress is well established, yet the relationship between their structural features and functional mechanisms are poorly defined. FASCICLIN-LIKE ARABINOGALACTAN PROTEINs (FLAs), which impact plant growth and development, are an excellent example of a glycoprotein family with a complex multidomain structure. FLAs combine globular fasciclin-like (FAS1) domains, disordered sequences containing glycomotifs for directing addition of O-linked arabinogalactan (AG) glycans, and additional post-translational modifications including N-linked glycans in the FAS1 domains, a cleaved signal peptide at the N-terminus, and often a glycosylphosphatidylinositol (GPI)-anchor signal sequence at the C-terminus. The roles of glycosylation, the GPI-anchor, and FAS1 domain functions in the polysaccharide-rich extracellular matrix of plants remain unclear, as do the relationships between them. In this study, we examined sequence-structure-function relationships of Arabidopsis (Arabidopsis thaliana) FLA11, demonstrated to have roles in secondary cell wall (SCW) development, by introducing domain mutations and functional specialisation through domain swaps with FLA3 and FLA12. We identified FAS1 domains as essential for FLA function, differentiating FLA11/FLA12, with roles in SCW development, from FLA3, specific to flowers and involved in pollen development. The GPI-anchor and AG glycosylation co-regulate the cell surface location and release of FLAs into cell walls. The AG glycomotif sequence closest to the GPI-anchor (AG2) is a major feature differentiating FLA11 from FLA12. Results of our study show that the multidomain structure of different FLAs influences their sub-cellular location and biological functions during plant development.