Islet-activating protein toxin

Figure 11. Effects of EGTA or treatment with islet activating protein (lAP or pertussis toxin) on the 6-HCH-induced Ca response detected in Indo-l-labeled cells. Cells were treated for 2 hours at 37 C with (lAP) or without (Control) 10 Jg/mL lAP, then labeled with Indo-1. Cells were washed and resuspended at 2 x 10 cells/mL buffer and stimulated with 6-HCH (solid trace). In some cases (dashed traces), stimulation was preceded by the addition of 5-mAf EGTA 10 s before stimulation. Other experimental conditions are as in Figure 9.

Fig. 1. Mechanisms of activation and inactivation of adenylate cyclase. Abbreviations not given in the text or figure are cholera T, Vibrio cholerae toxin IAP, islet activating protein, a Bordetetla pertussis toxin guanine N, guanine nucleotide 0- and a2-agonists, 0- and a2-adrenergic agonists 0. and a, a subunits of Gs and Gj, respectively.

G-protein identification has been aided considerably by the ability of their a subunits to be specifically ADP-ribosylated by either cholera or pertussis toxins. Cholera toxin ribosylates both transducin and Gs, causing dissociation of the holomeric form of these G-proteins to release a permanently activated and ADP-ribosylated a subunit. In contrast, pertussis toxin (islet activating protein, IAP) ribosylates the a subunit of the holomeric forms of Gi G0, Gh and transducin preventing their dissociation and, in the case of Gj, blocking the action of inhibitory hormones on adenylate cyclase activity. In the case of Gj, G0 and transducin, it has been clearly demonstrated that pertussis toxin can only ribosylate the holomeric form of these G-proteins. Cholera toxin, however, can ADP-ribosylate the free a subunits of Gs and transducin equally well, if not better, than when they form part of the holomeric complex. 

Katada T, Oinuma M, Ui M (1986) Two guanine nucleotide-binding proteins in rat brain serving as the specific substrate of islet-activating protein, pertussis toxin. Interaction of the a-subunits with 3y-subunits in development of their biological activities. In J. Biol. Chem. 261 8182-8191. 

Moss J, Stanley SJ, Burns DL, etal. (1983) Activation by thiol of the latent NAD gly-cohydrolase and ADP-ribosyltransferase activities of Bordefella pertussis toxin (Islet activating protein). In J. Biol. Chem. 258 11879-11882. 

Tamura M, Nogimori K, Murai S, et a/. (1982) Subunit structure of islet-activating protein, pertussis toxin, in conformity with the A-B model. In Biochemistry 21 5516-5522. 

Ui, M (1984) Islet-activating protein, pertussis toxin a probe for functions of the inhibitory guanine nucleotide regulatory component of adenylate cyclase. In Trends Pharmacol Sci 5 277—279... 

Okajima, F. and Ui, M. (1984). ADP-ribosylation of the specific membrane protein by islet-activating protein, pertussis toxin, associated with inhibition of a chemotactic peptide-induced arachidonate release in neutrophils A possible role of the toxin substrate in Ca " -mobilizing biosignalling. ]. Biol Chem., 259, 13863-13871... 

Katada T, Tamura M, Ui M. The A protomer of islet-activating protein, pertussis toxin, as an active peptide catalyzing ADP-ribosylation of a membrane protein. Arch Biochem Biophys 1983 224(1 ) 290-298. 

ADP-Ribosylation of a Membrane Protein Catalyzed by Islet-Activating Protein, Pertussis Toxin... 

Murayama T, Katada T, Ui M (1983) Guanine nucleotide activation and inhibition of adenylate cyclase as modified by islet-activating protein, pertussis toxin, in mouse 3T3 fibroblasts. Arch Biochem Biophys 221 381-390... 

Kurose H, Ui M (1983) Functional uncoupling of muscarinic receptors from adenylate cyclase in rat cardiac membranes by the active component of islet-activating protein, pertussis toxin. J Cycl Nucleotide Protein Phosphoryl Res 9 305-318... 

Ui M, Katada T, Murayama T, Kurose H, Yajima M, Tamura M, Nakamura T, Nogimori K (1984) Islet-activating protein, pertussis toxin A specific uncoupler of receptor-mediated inhibition of adenylate cyclase. In Greengard P et al. (eds) Advances in cyclic nucleotide and protein phosphorylation research, vol 17. Raven Press, New York, pp 145-151... 

Nogimori K, Tamura M, Yajima M, Ito K, Nakamura T, Kajikawa N, Maruyama Y, Ui M (1984) Dual mechanisms involved in development of diverse biological activities of islet-activating protein, pertussis toxin, as revealed by chemical modification of lysine residues in the toxin molecule. Biochim Biophys Acta 801 232-243... 

Nakamura T, Ui M (1983) Suppression of passive cutaneous anaphylaxis by pertussis toxin, an islet-activating protein, as a result of inhibition of histamine release from mast cells. Biochem Pharmacol 32 3435-3441... 

Nakamura T, Ui M (1985) Simultaneous inhibitions of inositol phospholipid breakdown, arachidonic acid release and histamine secretion in mast cells by islet-activating protein, pertussis toxin. A possible involvement of the toxin-specific substrate in the Ca -mobilizing receptor-mediated biosignaling system. J Biol Chem 260 3584-3593... 

Kurose H, Ui M (1985) Dual pathways of receptor-mediated cyclic GMP generation of NG108-15 cells as differentiated by susceptibility to islet-activating protein, pertussis toxin. Arch Biochem Biophys 238 424-434... 

Thus, lAP is, like diphtheria and cholera toxins, a protein toxin with an A-B structure. The holotoxin is bound to particular sites on the cell surface via its B-oligomer moiety as the first step of its interaction with mammalian cells. The A-protomer (or holotoxin itself) is then inserted to the plasma membrane traversing the lipid bilayer gradually. This slow process of the entrance of the toxin molecule is reflected in a definite lag time invariably preceding the onset of the action of lAP on intact cells as analyzed with rat pancreatic islets by kinetic and immunological approaches [25]. The A-protomer is finally activated by certain processing enzyme(s) inside the membrane to catalyze ADP-ribosylation of the target membrane protein with intracellular NAD as substrate. 

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