The GTP-binding protein G alpha s is present in dormant spores and expressed differentially during spore germination of the fungus Phycomyces blakesleeanus

G Martinez-Cadena, G Novoa-Martinez, A Gonzalez-Hernandez and J Garcia-Soto
Instituto de Investigacion en Biologia Experimental, Facultad de Quimica, Universidad de Guanajuato, Mexico.

A DNA sequence homologous to a G alpha s DNA probe, and the corresponding G alpha s protein (stimulatory alpha-subunit of GTP- binding protein) were detected in Phycomyces blakesleeanus. The protein was demonstrated in membrane fractions of dormant spores of this fungus using three different experimental approaches. Photoaffinity-labelling experiments with [alpha-32P]GTP of the membrane fraction revealed two bands, of 56 and 32 kDa. The 56 kDa GTP-binding protein was detected by this method in all the stages of early development and growth investigated. Also, a spore protein of 56 kDa was ADP-ribosylated by cholera toxin, and a 56 kDa protein was detected by Western blotting with a specific antibody against mammalian G alpha s. These results indicate that G alpha s (56 kDa) is present in dormant spores of P. blakesleeanus. Using the ADP-ribosylation and Western blotting assays, G alpha s was detected during all stages of spore germination before the hyphae became highly branched, but it was not detected in the branched hyphae that formed 18 h after the initiation of spore germination. Therefore, G alpha s is expressed differentially during Phycomyces development.