Characterization of Novel Aperture Proteins in Arabidopsis thaliana

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The Ohio State University

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In many species, exine, the outer wall of pollen grains, is deposited non-uniformly on the pollen surface. The gaps left on the pollen surface by the absence of exine deposition are known as apertures. These apertures facilitate emergence of the pollen tubes, making them important for male fertility in many plants. The positioning of the apertures is highly conserved within species, yet it varies widely between species, making apertures an ideal model for studying how cells control formation of distinct extracellular domains. Very little is known about how the locations of apertures are specified and how apertures are formed. Dobritsa lab's current work involves identification and characterization of novel proteins in Arabidopsis required for aperture formation. My project focused on two of the mutants recently found during a mutagenesis screen, both of which completely lack apertures. The candidate genes responsible for these mutations, called STRUBBELIG-RECEPTOR FAMILY 2 (SRF2) and INAPERTURATE POLLEN 2 (INP2), were isolated, fused with YFP in some cases, and introduced into their respective mutants in Arabidopsis using Agrobacterium-mediated transformation. I created the constructs and confirmed the identity of these genes by phenotypic rescue of their mutants. The expression and localization of the SRF2 protein was characterized using fluorescence of the YFP tag. In addition, I also tested the effects of inactivation of ELMOD-A, a close paralog of a third gene involved in aperture formation, called MACARON, by mutating ELMOD-A with CRISPR and demonstrated that ELMOD-A is also involved in pollen aperture formation.