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 with elicitor preparations containing the avr5 gene product from two Cf-5 incompatible races of the fungal pathogen Cladosporium fulvum (race 2.3 and race 4), or with elicitor preparations containing no avr5 gene product from two Cf-5 compatible races (race 5 and race 2.4.5.9.11). Elicitor preparations from race 2.3 or race 4 caused dephosphorylation of host plasma membrane H+-ATPase in isolated plasma membranes, while the preparations from race 5 or race 2.4.5.9.11 did not. GTP(gamma)S, AIF(4)(-), and cholera, toxin (CTX) each induced similar dephosphorylation in the absence of active elicitors. The elicitor-induced dephosphorylation of the H+-ATPase was blocked by preincubation of membranes with an antibody raised against a stimulatory G protein alpha-subunit (anti-G(s alpha)). This antibody cross-reacted with a 42 kDa polypeptide from tomato plasma membranes. A 42 kDa polypeptide was also ADP-ribosylated by CTX. When plasma membranes were treated with elicitor preparations from race 4 and separated on non-dissociating PAGE, two proteins were detected on Western blots with the antibody raised against the alpha-subunit, suggesting the dissociation of the trimeric complex. No dissociation of the complex was detected with antibodies raised against either the alpha- or beta-subunits when the plasma membranes were treated with elicitor preparations from race 5. The results provide evidence for the activation of a stimulatory subtype of trimeric G proteins in the stimulation of elicitor-induced host defences to fungal pathogens.