Genetic Analysis of Adhesion Protein ELMO3 in Arabidopsis thaliana

The Extracellular Matrix (ECM) between plant cells is vital for structure, development, and intercellular adhesion. A pectin rich layer in between cells, the middle lamella, is largely responsible for regulating the adhesive properties of adjacent plant cells. Homogalacturonan (HG) pectin, the most common, is synthesized in the Golgi and secreted into the ECM where it undergoes calcium crosslinking, increasing its adhesive properties. Mutations in proteins essential for HG synthesis can reveal a severe adhesion defective phenotype, where the hypocotyls of dark grown Arabidopsis exhibit cell sloughing, curling, and general disorganization. A family of five ELMO proteins are suspected to act as scaffolds for pectin biosynthesis enzymes. ELMO1 and ELMO4 mutants exhibit an adhesion deficient phenotype, and a double mutant provides evidence of redundancy in function between ELMO1 and ELMO2. ELMO1-GFP co-immunoprecipitated with enzymes required for HG synthesis indicating its role as a scaffold protein. Double mutants of the other ELMO homologues were created to determine if they exhibit functional redundancy, and ELMO1 and ELMO3 appear partially redundant. A gene deletion of ELMO3 was also created using the CRSPR/Cas9 system, resulting in two distinct elmo3 deletion alleles, which were phenotypically identical to the original elmo3-/- mutant. All adhesion defective phenotypes can be partially suppressed by altering the osmoticum and hence turgor that provides pressure on adhesive cells. Lastly, ELMO3-GFP was localized to the Golgi, the site of pectin biosynthesis, further supporting a common role of the ELMOs in pectin biosynthesis.