Identification of G proteins mediating fungal elicitor-induced dephosphorylation of host plasma membrane H+ -ATPase

Tomato cells (with the Cf-5 resistance gene) were treated withelicitor preparations containing the avr5 gene product fromtwo Cf-5 incompatible races of the fungal pathogen Cladosporiumfulvum (race 2.3 and race 4), or with elicitor preparationscontaining no avr5 gene product from two Cf-5 compatible races(race 5 and race 2.4.5.9 EC] .11). Elicitor preparations from race2.3 or race 4 caused dephosphorylation of host plasma membraneH⁺ -ATPase in isolated plasma membranes, while the preparationsfrom race 5 or race 2.4.5.9 EC] .11 did not. GTP()S, AlF₄^–and cholera toxin (CTX) each induced similar dephosphorylationin the absence of active elicitors. The elicitor-induced dephosphorylationof the H⁺ -ATPase was blocked by preincubation of membraneswith an antibody raised against a stimulatory G protein -subunit(anti-G_s This antibody cross-reacted with a 42 kDa polypeptidefrom tomato plasma membranes. A 42 kDa polypeptide was alsoADP-ribosylated by CTX. When plasma membranes were treated withelicitor preparations from race 4 and separated on non-dissociatingPAGE, two proteins were detected on Western blots with the antibodyraised against the -subunit, suggesting the dissociation ofthe trimeric complex. No dissociation of the complex was detectedwith antibodies raised against either the - or ß-subunitswhen the plasma membranes were treated with elicitor preparationsfrom race 5. The results provide evidence for the activationof a stimulatory subtype of trimeric G proteins in the stimulationof elicitor-induced host defences to fungal pathogens. Key words: G protein, dephosphorylation, H⁺ -ATPase, fungal elicitor, tomato