Regulation of inositol phospholipid and inositol phosphate metabolism in chemoattractant‐activated human polymorphonuclear leukocytes

Abstract

Binding of chemoattractants to specific cell surface receptors on polymorphonuclear leukocytes (PMNs) initiates a series of biochemical responses leading to cellular activation. A critical early biochemical event in chemoattractant (CTX) receptor-mediated signal transduction is the phosphodiesteric cleavage of plasma membrane phosphatidylinositol 4,5-bisphosphate (PIP2), with concomitant production of the calcium mobilizing inositol-1,4,5-trisphosphate (IP3) isomer, and the protein kinase C activator, 1,2-diacylglycerol (DAG). The following lines of experimental evidence collectively suggest that CTX receptors are coupled to phospholipase C via a guanine nucleotide binding (G) protein. Receptor-mediated hydrolysis of PIP2 in PMN plasma membrane preparations requires both fMet-Leu-Phe and GTP, and incubation of intact PMNs with pertussis toxin (which ADP ribosylates and inactivates some G proteins) eliminates the ability of fMet-Leu-Phe plus GTP to promote PIP2 breakdown in isolated plasma membranes. Studies with both PMN particulate fractions and with partially purified fMet-Leu-Phe receptor preparations indicate that guanine nucleotides regulate CTX receptor affinity. Finally, fMet-Leu-Phe stimulates high-affinity binding of GTP gamma S to PMN membranes as well as GTPase activity. A G alpha subunit has been identified in phagocyte membranes which is different from other G alpha subunits on the basis of molecular weight and differential sensitivity to ribosylation by bacterial toxins. Thus, a novel G protein may be involved in coupling CTX receptors to phospholipase C. Studies in intact and sonicated PMNs demonstrate that metabolism of 1,4,5-IP3 proceeds via two distinct pathways: 1) sequential dephosphorylation to 1,4-IP2, 4-IP1 and inositol, or 2) ATP-dependent conversion to inositol 1,3,4,5-tetrakisphosphate (IP4) followed by sequential dephosphorylation to 1,3,4-IP3, 3,4-IP2, 3-IP1 and inositol. Receptor-mediated hydrolysis of PIP2 occurs at ambient intracellular Ca2+ levels; but metabolism of 1,4,5-IP3 via the IP4 pathway requires elevated cytosolic Ca2+ levels associated with cellular activation. Thus, the two pathways for 1,4,5-IP3 metabolism may serve different metabolic functions. Additionally, inositol phosphate production appears to be controlled by protein kinase C, as phorbol myristate acetate (PMA) abrogates PIP2 hydrolysis by interfering with the ability of the activated G protein to stimulate phospholipase C. This implies a physiologic mechanism for terminating biologic responses via protein kinase C mediated feedback inhibition of PIP2 hydrolysis.

Remodeling of neutrophil phospholipids with 15(S)-hydroxyeicosatetraenoic acid inhibits leukotriene B4-induced neutrophil migration across endothelium.

“…LTB4 provoked a rapid increase in lP3 formation in vehicle-treated PMNs (Fig. 6, top), as reported by other investigators (41). IP3 levels were maximal after 5 s of exposure and declined to basal levels within 1 min (P3 levels in picomoles per 106 PMNs: basal 1.16±0.36; 5 s after LTB4 stimulation 2.42±0.41, n = 7, P < 0.001 ).…”

“…Concurrent with the reduction in CD 11/CD18-mediated PMN adhesion and transmigration, 15(S)-HETE remodeling caused a striking decrease in the affinity of the high affinity LTB4 cell-surface receptors that subserve these responses (39). Furthermore, lP3 generation, a pivotal event in LTB4-triggered signal transduction events in PMN cell membranes (41 ), was inhibited in 15 (S)-HETE-remodeled cells. The mechanism by which 15 (S)-HETE esterification modulates the affinity of LTB4 receptors can now be addressed in future studies.…”

Human neutrophil cytosolic phospholipase C: partial characterization

“…Stimulation of inositol lipid hydrolysis by guanine nucleotides compatible with involvement of G proteins in the activity of neutrophil PLC has been described by many investigators [8][9][10][29][30][31][32]. In our hands, the activity of neutlophil cytosolic PLC assayed in lipid vesicles or in heat-inactivated membranes devoid of active G proteins was not affected by GTPyS (Table Ill).…”

1,2‐Diacylglycerol accumulation in human neutrophils does not correlate with respiratory burst activation

“…In spite of the fact that opsonized particles are strong activators of human neutrophils, little is known about the signal(s) mediating this response. Several authors have speculated on the importance of the activation of a phospholipase C acting on phosphatidylinositol bisphosphate (PIP2) during activation of neutrophils and the subsequent production of the second messengers inositol 1,4,5-trisphosphate (Ins (1,4,5)Pa) and 1,2-diacylglycerol (1,2-DG) [4,5,6]. IntraceUular 1,2-DG resulting from this hydrolysis might be, in analogy…”