Chemoattraction of neutrophils by substance P and transforming growth factor-beta 1 is inadequately explained by current models of lipid remodeling

"Classical" chemoattractants, such as FMLP, C5a, or leukotriene B4, not only elicit directed motility but also activate neutrophils (degranulation, release of active oxygen species). Signal transduction after ligation of receptors for these classical chemoattractants is mediated by pertussis toxin (PT)-sensitive, heterotrimeric G proteins and the early production of lipid messengers via phospholipases. In contrast, we have previously shown that substance P (SP) and transforming growth factor-beta 1 (TGF-beta 1) are "pure" chemoattractants in that they elicit chemotaxis without activating neutrophils. Paradoxically, pure chemoattractants also activate G proteins (plasmalemmal GTPase activity) without eliciting increments in cytosolic calcium ([Ca]i) and thus inositol trisphosphate. We therefore determined lipid remodeling and signal transduction in response to pure chemoattractants. Increments in plasmalemmal GTPase activated by SP (0.1 microM) and TGF-beta 1 (40 fM), like that after FMLP, were PT-sensitive (SP = 6.6 +/- 2 pm/mg/min vs SP + PT = 1.1 +/- 0.9 over basal activity; TGF-beta 1 = 4.3 +/- 1.6 vs TGF-beta 1 + PT = 2.3 +/- 0.9). In parallel, treatment of PMN with PT (1 microgram/ml, 30 min) inhibited chemotaxis (under agarose) after FMLP (2175 +/- 176 (SEM) microns vs 726 +/- 267) and SP (411 +/- 99 microns vs 103 +/- 62 microns) and TGF-beta 1 (40 fM, 375 +/- 53 microns vs 83 +/- 47). However, G proteins coupled to receptors for SP and TGF-beta 1, unlike FMLP, did not appear to be linked to phospholipases in that neither increments in diacylglycerol were detected after receptor ligation (FMLP = 152 +/- 22% resting levels; SP = 101 +/- 5%; TGF-beta 1 = 105 +/- 4%) nor was alkylacylglycerol increased by exposure to SP or TGF-beta 1 (SP = 92 +/- 4%; TGF-beta 1 = 101 +/- 8%; FMLP = 226 +/- 40%). Moreover, polymorphonuclear leukocytes failed to generate phosphatidates (PA) of either species after SP (DA-PA = 79 +/- 9% resting at 60 s; EA-PA = 103 +/- 4%) or TGF-beta 1 (DA-PA = 101 +/- 5%; EA-PA = 98 +/- 9%) in contrast to FMLP (DA-PA = 155 +/- 22%; EA-PA = 149 +/- 16%). The data clearly contravene the current dogma that all chemoattractants use inositol trisphosphate and diglycerides as intracellular signals and suggest the presence of a unique subset of PT-sensitive G proteins, not coupled to "classical" phospholipases, transduce chemoattraction.