Tomato wall-associated kinase SlWak1 acts in an Fls2- and Fls3-dependent manner to promote apoplastic immune responses to Pseudomonas syringae

Wall-associated kinases (Waks) are known to be important components of plant immunity against various pathogens including pv. tomato () although their molecular mechanisms are largely unknown. In tomato, has been implicated in immunity because its transcript abundance increases significantly in leaves after treatment with the flagellin-derived peptides flg22 and flgII-28, which activate the receptors Fls2 and Fls3, respectively. We generated two tomato mutants (Δwak1) using CRISPR/Cas9 and investigated the role of in tomato- interactions. PTI activated in the apoplast by flg22 or flgII-28 was compromised in Δwak1 plants but PTI at the leaf surface was unaffected. The Δwak1 plants developed fewer callose deposits than wild-type plants but retained the ability to generate reactive oxygen species and activate MAPKs in response to flg22 and flgII-28. The induction of gene expression by flg22 and flgII-28 was greatly reduced in a tomato mutant lacking Fls2 and Fls3 but induction of gene expression by flgII-28 was unaffected in Δwak1 plants. After inoculation, Δwak1 plants developed disease symptoms more slowly than Δfls2.1/fls2.2/fls3 mutant plants, although both plants ultimately were similarly susceptible. SlWak1 co-immunoprecipitated with both Fls2 and Fls3 independently of flg22/flgII-28 or Bak1. These observations suggest that SlWak1 acts in a complex with Fls2/Fls3 and plays an important role at later stages of the PTI in the apoplast.