Pertussis toxin‐insensitive effects of mastoparan, a wasp venom peptide, in PC 12 cells

Abstract

Recent studies have shown that mastoparan, an amphiphilic peptide derived from wasp venom, modifies the secretion of neurotransmitters and hormones from a variety of cell types. Mastoparan interacts with heterotrimeric guanine nucleotide-binding proteins (G proteins) such as Gi and G(o), which are ADP-ribosylated by pertussis toxin (PTX) and thereby uncoupled from receptors. Previously, some of the effects of mastoparan including secretion were reported to be modified selectively by PTX but not by cholera toxin (CTX). In the present study, we examined the influence of bacterial toxins on the effects of mastoparan in PC12 cells. Mastoparan stimulated [3H]noradrenaline (NA) release from prelabeled PC12 cells in the absence of CaCl2, although high K+ or ATP-stimulated the release in a Ca(2+)-dependent manner. Pretreatment with CTX, not PTX, for 24 h inhibited mastoparan-stimulated [3H]NA release. Mastoparan inhibited forskolin-stimulated cyclic AMP accumulation in a dose-dependent manner, although mastoparan had no effect by itself. Pretreatment with PTX completely abolished the inhibitory effect of carbachol via Gi on cyclic AMP accumulation and partially reduced the effect of mastoparan. However, the inhibitory effect of 20 microM mastoparan was not modified by pretreatment with PTX. Thus, we investigated the effect of mastoparan on CTX-catalyzed [32P]ADP-ribosylation of proteins in PC12 cells. A subunit of CTX (CTX-A) catalyzed [32P]ADP-ribosylation of many proteins in the cytosolic fraction of PC12 cells. One of these was a 20 kDa protein, named ADP-ribosylating factor (ARF). The addition of mastoparan to assay mixtures inhibited ADP-ribosylation of many proteins including ARF and CTX-A in the presence of the cytosolic fraction. In the absence of the cytosolic fraction, however, mastoparan slightly enhanced ADP-ribosylation of bovine serum albumin and auto-ADP-ribosylation by CTX-A. Mastoparan did not inhibit ADP-ribosylation of the alpha subunit of Gs in the membrane fraction. These findings suggest that 1) mastoparan interacts with PTX-insensitive and CTX-sensitive factor(s) to stimulate NA release, and 2) mastoparan interacts with ARF inhibiting its activity to enhance the ADP-ribosylation reaction by CTX. ARF may be an exocytosis-linked G protein.

Structural determinants for binding to CGRP receptors expressed by human SK‐N‐MC and Col 29 cells: studies with chimeric and other peptides

“…Ostenson et al , 1997). Indeed, mastoparan can inhibit forskolin‐induced cyclic AMP accumulation in PC12 cells (Murayama et al , 1996). As mastoparan itself failed to inhibit the response to CGRP and the mastoparan‐CGRP chimeras were able to displace specifically bound CGRP from SK‐N‐MC membranes, it is unlikely that they antagonize CGRP by any indirect mechanism.…”

Crystal structure of mastoparan from Polistes jadwagae at 1.2 Å resolution

“…They were originally discovered as the agents that induce histamine release from mast cells and later found to affect a variety of biological activities, including induction of secretion in different types of cells (Mizuno et al, 1998;Murayama et al, 1996;Ohara-Imaizumi et al, 2001;Straub et al, 1998;Wu et al, 1999;Yajima et al, 1997), hemolytic activity (Mendes et al, 2005), antimicrobial activity (Park et al, 1995), etc.…”

Crystallization and Preliminary X-Ray Diffraction Analysis of Three Mastoparans

“…They have a variety of biological activities. Mastoparans can induce secretion of neurotransmitters [1], hormones [2][3][4] and cytokines [5] in different types of cells. They can promote histamine release from mast cells [6], and some of these peptides also exhibit potent hemolytic activity [7,8] and antimicrobial activity [9].…”