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.

Receptor kinases (RKs) are transmembrane proteins that have been shown to regulate an array of important processes in A. thaliana, including polar cell growth, plant reproduction, and many other plant growth processes. In this thesis, I examine RECEPTOR KINASE 7 (RK7) and FERONIA (FER), two closely related transmembrane RKs, and their effects on plant reproduction. The RK7 gene when knocked out (rk7) in conjunction with FER resulted in delayed plant growth, decreased seed yield, and a lower percentage of the seeds germinating as compared to the single FER knockout. Transgenic plants with GUS reporter driven by RK7 promoter and RK7 promoter expressed GFP-tagged RK7 (RK7-GFP) were generated to study, respectively, the expression property of the RK7 gene and characterize the location of the RK7 protein. RK7 expression increased in the papillary cells as a direct result of pollination. Transgenic plants with RK7-GFP showed that RK7 protein localizes to the plasma membrane of stigma cells and pollination induces prominent internalization of this protein. RK7 is also expressed during seedling growth. rk7 mutant seedlings had a much weaker physiological response to brassinosteroids than wild type plants, implicating an involvement of RK7 in brassinosteroid signaling. Taken together this data point to the importance of RK7 in plant growth and reproduction through its ability to receive and transduce signals.

Cell-to-cell communication is the driving force behind successful reproduction in flowering plants. Extensive extracellular communication events occur between the male and female gametophytes during pollen tube reception to facilitate successful fertilization. These signaling events culminate into a product of great importance for both animals and plants: the seed. In this study, the pathogen defense regulator PATTERN-TRIGGERED IMMUNITY COMPROMISED RECEPTOR-LIKE CYTOPLASMIC KINASE 1 (PCRK1) was identified to function in pollen tube reception from both the male and female gametophytes in the flowering plant <i>Arabidopsis thaliana</i> using a forward genetic screen. A knockout of <i>pcrk1</i> suppresses the pollen tube overgrowth phenotype leading to infertility in <i>nortia</i> mutants. In addition, <i>pcrk1</i> pollen affected the pollen tube overgrowth phenotypes of pollen tube reception mutants <i>feronia</i> and <i>turan</i>. Shared molecular components of pollen tube reception and pathogen invasion have been reported. This study reveals another link between pathogen defense and pollen tube reception. By studying the links between fertility and disease in plants, we may be able to uncover potential trade-offs with fertility when breeding for pathogen resistance.<br>

Fertilization, both in plants and animals, is at its core, a study of cell to cell communication. With respect to plants, the male gametophyte, the pollen tube, elongates within the female organ called the pistil, transporting in its cytoplasm two sperm cells. The pollen tube is attracted by signals secreted from the synergid cells that are located at the entrance to the female gametophyte that resides in the ovule. Secondary pollen tube visitors to the ovules are unwanted and repelled presumably by signals emitted by the fertilized female. The final communication between the pollen tube and female gametophyte is the induction of pollen tube rupture upon penetration of the synergid cell, an event that leads to the release of the two sperm cells, which go on to fertilize the central cell and egg cell within the female gametophyte, completing a double fertilization process that is unique to plants. My thesis research is centered on elucidating the mechanism behind the synergid cell-induced pollen tube rupture process. Studies in our laboratory have established that the synergid cell-expressed receptor like kinase, called FERONIA, mediates a highly oxidative environment in the female gematophyte that is necessary for the pollen tube rupture process. Using an in vitro pollen tube culture system, my research showed that reactive oxygen species (ROS) induces pollen tube rupture in a Ca2+-dependent manner. My results suggests a careful and truly fascinating, though still hypothetical, design of a two molecule, FERONIA and ROS, two step activation system that uses ROS to prime the pollen tube outside the synergid cell, then expose it to calcium within the synergid cell to ensure that pollen tube rupture happens in the synergid cell, enabling fertilization.