Examining the role of GRP and LIK1 in Wall Associated Kinase (WAK) perception of pectin in the plant cell wall

Wall associated kinases (WAKs) are cell membrane bound receptor kinases that bind pectin and pectin fragments (OGs).The binding of WAKs to pectin sends a growth signal required for cell elongation and plant development. WAKs bind OGs with higher affinity than native pectin and instead activate a stress response. Glycine rich proteins (GRPs) are secreted cell wall proteins of unknown function. Seven GRPs with 65% sequence similarity are coded on a 90kb locus of Arabidopsis chromosome 2. GRP3 and WAK1 have been shown to bind in vitro, but single null mutations have no discernible phenotype, suggesting that the GRPs are redundant. Low recombination frequency has made multiple mutations difficult to achieve, but in this thesis, CRISPR/Cas9 technology was used to induce deletions of the GRP locus. The promoters pYAO and pICU2 drove Cas9 expression in transformed Arabidopsis plants. The presence of a deletion and Cas9 were detected by PCR. While somatic mutations were induced, there was no inheritance of the GRP deletion, indicating that pYAO and pICU2 do not drive Cas9 to induce deletions in progenitor cells. LIK1 is a CERK1 interacting kinase implicated in mediating response to various microbe associated molecular patterns (MAMP) such as chitin, flagellin, and peptidoglycans. LIK1 exhibits a drastic increase in phosphorylation in response to OG treatment, making it a candidate for a co-receptor to WAK. T-DNA insertions to the 5’UTR of LIK1 were used to examine the effect of a lik1 mutation on the OG induced stress response. lik1/lik1 mutant seedlings were grown in the presence and absence of OGs, and RNA was isolated. qPCR was used on cDNA to examine FADLOX expression, a reporter for the transcriptional response to OGs. The lik1/lik1 mutant caused a reduction in the OG induced transcriptional response. However, increased LIK1 expression was associated with the T-DNA insertion indicating that LIK1 inhibits the WAK stress response pathway. Understanding the roles of GRP and LIK1 in moderating WAK mediated pathogenic response in Arabidopsis will enable a better understanding of plant resistance to pathogen invasion in the greater plant kingdom.