122 Locating the BPS Family Proteins Using Molecular Infusion Cloning

Faculty Mentor: Leslie Sieburth (School of Biological Sciences)

Bypass1 (bps1) mutants in Arabidopsis thaliana exhibit a severe growth arrest phenotype. This phenotype is caused by a root-derived, graft transmissible molecule known as dalekin. It was previously shown that the BPS2 allele in the Apost-1 natural variant accession of Arabidopsis suppresses the bps1 growth arrest, and the Columbia variant of BPS2 promotes growth arrest. We know that BPS2 acts in the same pathway as BPS1 and also contributes to regulation of dalekin synthesis, but the protein’s localization is still unknown. BPS2 (Columbia) and BPS2Apost-1 differ by only 6 base pair mutations, 2 of which are in the coding region of the gene. I want to determine whether these mutations change the localization of the BPS2 alleles from Col-0 and Apost-1. To answer this question, I generated constructs that fused the coding region of fluorescent proteins to the coding region of BPS2 from Col-0 and Apost-1. These will be transformed into Col-0 Arabidopsis, and in the future, they will be used to characterize the impact of Apost-1 mutations on BPS2 localization (tissue level and cellular level). To test whether BPS2 and BPS2Apost-1 have different protein environments, I established proximity labeled constructs by fusing the biotin ligase enzymes TurboID and miniTurbo to BPS2 and BPS2Apost-1 . Once transformed into plants, these enzymes will label proteins that are close to BPS2. The research I have done will be able to provide information regarding the BPS family proteins and their localizations; thus, leading to more information about dalekin’s pathway in Arabidopsis thaliana.