PURIFICATION AND IDENTIFICATION OF FOAD-II, A CYTOSOLIC PROTEIN THAT REGULATES SECRETION IN STREPTOLYSIN-O PERMEABILIZED MAST-CELLS, AS A RAC RHOGDI COMPLEX/

Mast cells permeabilized by treatment with streptolysin-O in the prese nce of Ca2+ and GTP-gamma-S can secrete almost 100% of their contained N-acetyl-beta-D-glucosaminidase. If these stimuli are provided to the permeabilized cells after a delay, the response is diminished and the ability of the cells to undergo secretion runs down progressively ove r a period of about 30 min. This is thought to be due to the loss of k ey proteins involved in the exocytotic mechanism. Using this effect as the basis of a biological assay, we have isolated a protein from bovi ne brain cytosol that retards the loss of responsiveness to stimulatio n by Ca2+ and GTP-gamma-S. Purification of tl-Lis protein and peptide sequencing have enabled us to identify it as the small GTP-binding pro tein rac complexed to the guanine nucleotide exchange inhibitor rhoGDI . Both proteins are required to retard the loss of the secretory respo nse, while purified rhoGDI applied alone accelerates the rundown.