MPK4 phosphorylation dynamics and interacting proteins in plant immunity

The Arabidopsis MAP kinase 4 (MPK4) has been proposed as negative player in plant immunity, but it is activated by pathogen-associated molecular patterns (PAMPs), such as flg22. The molecular mechanisms by which MPK4 is activated and controls plant defense remain elusive. In this study, we investigated the Arabidopsis defense against a bacterial pathogen Pseudomonas syringae pv. tomato (Pst) DC3000 when the MPK4 is overexpressed. We showed an increase in pathogen resistance and suppression of jasmonic acid (JA) signaling in the MPK4 overexpressing (OE) plants. We also showed that the OE plants are very sensitive to flg22-triggered reactive oxygen species (ROS) burst in guard cells, which resulted in an enhanced stomatal closure, compared to wild-type (WT). During the flg22 activation, dynamic phosphorylation of MPK4 within and outside of the conserved TEY activation loop was observed. To elucidate how MPK4 functions during the defense response, we used immunoprecipitation coupled with mass spectrometry (IP-MS) to identify MPK4 interacting proteins in the absence and presence of flg22. Quantitative proteomic analysis revealed a shift in MPK4-associated protein network, providing insight into the molecular functions of MPK4 at a systems level.