Fungal Communication Requires the MAK-2 Pathway Elements STE-20 and RAS-2,the NRC-1 Adapter STE-50 and the MAP Kinase Scaffold HAM-5

Intercellular communication is critical for the survival of unicellular organisms as well as for the development and function of multicellular tissues. cell-to-cell signaling is also required to develop the interconnected mycelial network characteristic of filamentous fungi and is a prerequisite for symbiotic and pathogenic host colonization achieved by molds. somatic cell–cell communication and subsequent cell fusion is governed by the mak-2 mitogen activated protein kinase (mapk) cascade in the filamentous ascomycete model neurospora crassa,yet the composition and mode of regulation of the mak-2 pathway are currently unclear. in order to identify additional components involved in mak-2 signaling we performed affinity purification experiments coupled to mass spectrometry with strains expressing functional gfp-fusion proteins of the mapk cascade. this approach identified ste-50 as a regulatory subunit of the ste11p homolog nrc-1 and ham-5 as cell-communication-specific scaffold protein of the mapk cascade. moreover,we defined a network of proteins consisting of two ste20-related kinases,the small gtpase ras-2 and the adenylate cyclase capping protein cap-1 that function upstream of the mak-2 pathway and whose signals converge on the nrc-1/ste-50 map3k complex and the ham-5 scaffold. finally,our data suggest an involvement of the striatin interacting phosphatase and kinase (stripak) complex,the casein kinase 2 heterodimer,the phospholipid flippase modulators ypk-1 and nrc-2 and motor protein-dependent vesicle trafficking in the regulation of mak-2 pathway activity and function. taken together,these data will have significant implications for our mechanistic understanding of mapk signaling and for homotypic cell–cell communication in fungi and higher eukaryotes. © 2014 dettmann et al.