Dynamics of ribosome composition and ribosomal protein phosphorylation in immune signaling in Arabidopsis thaliana.

In plants, the detection of microbe-associated molecular patterns (MAMPs) induces primary innate immunity by the activation of mitogen-activated protein kinases (MAPKs). We show here that the MAMP-activated MAPK MPK6 not only modulates defense through transcriptional regulation but also via the ribosomal protein translation machinery. To understand the effects of MPK6 on ribosomes and their constituent ribosomal proteins (RPs), polysomes, monosomes and the phosphorylation status of the RPs, MAMP-treated WT and mpk6 mutant plants were analysed. MAMP-activation induced rapid changes in RP composition of monosomes, polysomes and in the 60S ribosomal subunit in an MPK6-specific manner. Phosphoproteome analysis showed that MAMP-activation of MPK6 regulates the phosphorylation status of the P-stalk ribosomal proteins by phosphorylation of RPP0 and the concomitant dephosphorylation of RPP1 and RPP2. These events coincide with a significant decrease in the abundance of ribosome-bound RPP0s, RPP1s and RPP3s in polysomes. The P-stalk is essential in regulating protein translation by recruiting elongation factors. Accordingly, we found that RPP0C mutant plants are compromised in basal resistance to Pseudomonas syringae infection. These data suggest that MAMP-induced defense also involves MPK6-induced regulation of P-stalk proteins, highlighting a new role of ribosomal regulation in plant innate immunity.

Essential role of the CD docking motif of MPK4 in plant immunity, growth, and development.

MAPKs are universal eukaryotic signaling factors whose functioning is assumed to depend on the recognition of a common docking motif (CD) by its activators, substrates, and inactivators. We studied the role of the CD domain of Arabidopsis MPK4 by performing interaction studies and determining the ligand-bound MPK4 crystal structure. We revealed that the CD domain of MPK4 is essential for interaction and activation by its upstream MAPKKs MKK1, MKK2, and MKK6. Cys181 in the CD site of MPK4 was shown to become sulfenylated in response to reactive oxygen species in vitro. To test the function of C181 in vivo, we generated wild-type (WT) MPK4-C181, nonsulfenylatable MPK4-C181S, and potentially sulfenylation mimicking MPK4-C181D lines in the mpk4 knockout background. We analyzed the phenotypes in growth, development, and stress responses, revealing that MPK4-C181S has WT activity and complements the mpk4 phenotype. By contrast, MPK4-C181D cannot be activated by upstream MAPKK and cannot complement the phenotypes of mpk4. Our findings show that the CD motif is essential and is required for activation by upstream MAPKK for MPK4 function. Furthermore, growth, development, or immunity functions require upstream activation of the MPK4 protein kinase.

A Chimeric IDD4 Repressor Constitutively Induces Immunity in Arabidopsis via the Modulation of Salicylic Acid and Jasmonic Acid Homeostasis.

INDETERMINATE DOMAIN (IDD)/BIRD proteins belong to a highly conserved plant-specific group of transcription factors with dedicated functions in plant physiology and development. Here, we took advantage of the chimeric repressor gene-silencing technology (CRES-T, SRDX) to widen our view on the role of IDD4/IMPERIAL EAGLE and IDD family members in plant immunity. The hypomorphic idd4SRDX lines are compromised in growth and show a robust autoimmune phenotype. Hormonal measurements revealed the concomitant accumulation of salicylic acid and jasmonic acid suggesting that IDDs are involved in regulating the metabolism of these biotic stress hormones. The analysis of immunity-pathways showed enhanced activation of immune MAP kinase-signaling pathways, the accumulation of hydrogen peroxide and spontaneous programmed cell death. The transcriptome of nonelicited idd4SRDX lines can be aligned to approximately 40% of differentially expressed genes (DEGs) in flg22-treated wild-type plants. The pattern of DEGs implies IDDs as pivotal repressors of flg22-dependent gene induction. Infection experiments showed the increased resistance of idd4SRDX lines to Pseudomonas syringae and Botrytis cinerea implying a function of IDDs in defense adaptation to hemibiotrophs and necrotrophs. Genome-wide IDD4 DNA-binding studies (DAP-SEQ) combined with DEG analysis of idd4SRDX lines identified IDD4-regulated functional gene clusters that contribute to plant growth and development. In summary, we discovered that the expression of idd4SRDX activates a wide range of defense-related traits opening up the possibility to apply idd4SRDX as a powerful tool to stimulate innate immunity in engineered crops.