Cytoplasmic sequestering of a fungal stress-activated MAPK in response to a host plant phenolic acid.

Fungal pathogens employ conserved signaling pathways to survive in the host. The stress-activated MAP kinase Hog1 of the maize pathogen Cochliobolus heterostrophus undergoes dephosphorylation upon exposure to ferulic acid, a phenolic compound abundant in the host plant. Unlike its nuclear localization during osmotic stress, Hog1 forms cytoplasmic foci in response to FA, indicating its sequestering to a compartment or condensate. FA prevents several characteristic responses of the Hog1 pathway to osmotic stress: hyperphosphorylation of Hog1, nuclear localization, and expression of a monosaccharide transporter gene, MST1. Under FA stress, mRNA-containing foci are formed, as visualized by sm-FISH. Hog1 foci extensively colocalize with mRNA foci. Hog1 did not colocalize with nuclei or peroxisomes. Fragmented mitochondria, appearing upon FA exposure with a delayed time course compared to Hog1 sequestration, were mostly distinct from the Hog1 foci, with few instances of colocalization. With tagged Hog1 as an affinity purification bait, we isolated an FA-dependent sub-proteome from a subcellular fraction enriched with fluorescent foci. The identified proteins include RNA-binding proteins, translation initiation factors and mitochondrial proteins. The RRM and pumilio domain protein Puf2 was enriched, and live imaging confirmed the accumulation of a Puf2 fluorescent fusion and its colocalization with Hog1 foci following FA induction. Stress-induced sequestering of MAPK Hog1 to RNA-containing granules, together with dephosphorylation, has the potential to collectively promote survival on the plant host where stress might cause over-activation of Hog1. Conversely, FA as a host defense interferes with stress-induced Hog1 nuclear localization and downstream gene expression. The MAPK signaling mode defined by the response of Hog1 to FA is thus relevant to both host defense and pathogen survival.