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Analyzing the Biochemical and Functional Interactions of the RALF1-FERONIA-LLG1 (a peptide ligand-receptor kinase-GPI-anchored protein complex) Signaling Pathways in Arabidopsis thaliana

Signal transduction pathways play a critical role in plant growth and reproduction by perceiving extracellular signals, leading to a cellular response. FERONIA (FER) is a transmembrane receptor kinase found on the plasma membrane in the model plant Arabidopsis thaliana and plays critical roles in growth, development, and fertilization. FER works upstream of master molecular switch RAC/ROP GTPase to regulate signaling into the cytoplasm. LORELEI-Like Glycosylphosphatidylinositol (GPI)-Anchored Protein 1(LLG1) is a GPI-anchored protein and co-receptor of FER on the plasma membrane. LLG1 is responsible for chaperoning FER from the endoplasmic reticulum (ER) to its functional location on the plasma membrane. Rapid Alkalinization Factor 1 (RALF1) is a small, secreted growth-regulatory peptide that interacts with FER, regulating signaling activity. This interaction, among other, regulates the activity of a downstream plasma membrane proton ATPase (AHA2) which impacts cell growth. Additionally, published pulldown data indicates LLG1, FER, and RALF1 complex together. My data suggests that LLG1, in addition to localizing and chaperoning FER, binds directly to RALF1. My results show that this RALF1-LLG1 interaction is required for proper RALF1 mediated signaling through FER. Data also indicates that FER and LLG1 regulate RALF1 location on the plasma membrane. Additionally, RALF1 binds the MALA domain of FER. Another aspect of my thesis focuses on LURE1. LURE1 is a secreted cysteine-rich, defensin like protein which guides incoming pollen tubes to the ovule in a process called pollen tube guidance. LURE1 guides pollen tubes by binding with pollen-specific receptor kinase 6 (PRK6), located on the plasma membrane of the incoming pollen tubes, to facilitate proper fertilization. My data also shows that the ovule derived signaling molecule nitric oxide (NO), also regulated by FER, negatively impacts the property of LURE1, causing it to fall out of solution and aggregate. Furthermore, the negative impact of NO on LURE1 disrupts the binding affinity of LURE1 to PRK6. Together with data from my lab showing pollen tube arrival at the ovule triggers NO production in a FER dependent manner, my findings provide a biochemical explanation for why pollen tubes do not target fertilized ovules.