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 presence 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 over a period of about 30 min. This is thought to be due to the loss of key proteins involved in the exocytotic mechanism. Using this effect as the basis of a biological assay, we have isolated a protein from bovine brain cytosol that retards the loss of responsiveness to stimulation by Ca2+ and GTP-gamma-S. Purification of this protein and peptide sequencing have enabled us to identify it as the small GTP-binding protein rac complexed to the guanine nucleotide exchange inhibitor rhoGDI. Both proteins are required to retard the loss of the secretory response, while purified rhoGDI applied alone accelerates the rundown.
O'Sullivan
A J
AJ
Department of Physiology, University College, London, United Kingdom.
Brown
A M
AM
Freeman
H N
HN
Gomperts
B D
BD
D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D015550:Cell Degranulation / Q000187:drug effects; D002463:Cell Membrane Permeability / Q000502:physiology*; D003600:Cytosol / Q000737:chemistry; D019204:GTP-Binding Proteins / Q000378:metabolism*; D020730:Guanine Nucleotide Dissociation Inhibitors; D008297:Male; D008407:Mast Cells / Q000166:cytology / Q000378:metabolism*; D008969:Molecular Sequence Data; D011506:Proteins / Q000302:isolation & purification / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D013301:Streptolysins / Q000494:pharmacology; D020744:rac GTP-Binding Proteins; D062947:rho-Specific Guanine Nucleotide Dissociation Inhibitors
D020730:Guanine Nucleotide Dissociation Inhibitors; D011506:Proteins; D013301:Streptolysins; D062947:rho-Specific Guanine Nucleotide Dissociation Inhibitors; D019204:GTP-Binding Proteins; D020744:rac GTP-Binding Proteins