Inhibitors of PI 3-kinase and MEK kinase differentially affect mediator secretion from immunologically activated human basophils

Previous studies have indicated a redundancy in the effects of the cytokines, IL-3, IL-5, and nerve growth factor (NGF) on acute priming of human basophils. In the current study, we have examined the effects of these three cytokines on 18-h priming for leukotriene C4 generation, their ability to induce FcεRIβ mRNA expression, or their ability to sustain basophil viability in culture. We also examine a variety of the signaling steps that accompany activation with these cytokines. In contrast with the ability of IL-3 to alter secretagogue-mediated cytosolic calcium responses following 18-h cultures, 18-h treatment with IL-5 or NGF did not affect C5a-induced leukotriene C4 generation or alter C5a-induced intracellular Ca2+ concentration elevations. IL-3 and IL-5, but not NGF, induced FcεRIβ mRNA expression and all three improved basophil viability in culture with a ranking of IL-3 > IL-5 ≥ NGF. All three cytokines acutely activated the extracellular signal-regulated kinase pathway and the signaling elements that preceded extracellular signal-regulated kinase and cytosolic phospholipase A2 phosphorylation, consistent with their redundant ability to acutely prime basophils. However, only IL-3 and IL-5 induced Janus kinase 2 and STAT5 phosphorylation. This pattern of signal element activation among the three cytokines most closely matched their ability to induce expression of FcεRIβ mRNA. Induction of the sustained calcium signaling that follows overnight priming with IL-3 appeared to be related to the strength of the early signals activated by these cytokines but the relevant pathway required was not identified. None of the signaling patterns matched the ability of the cytokines to promote basophil survival.

Section: Signaling Following Il-3supporting

confidence: 73%

Differences in Functional Consequences and Signal Transduction Induced by IL-3, IL-5, and Nerve Growth Factor in Human Basophils

Miura¹,

Saini²,

Gauvreau³

et al. 2001

“…3). Examples of PI 3-kinase-dependent activation of ERK have been cited previously in the literature (18,19). In contrast to the ERKs, we did not detect ULBPinduced activation of JNK or p38 in NK cells (data not shown).…”

Section: Ulbps Induce Activation Of the Erk Map Kinase Pathwaysupporting

confidence: 57%

UL16-Binding Proteins, Novel MHC Class I-Related Proteins, Bind to NKG2D and Activate Multiple Signaling Pathways in Primary NK Cells

Sutherland

Chalupny

Schooley

et al. 2002

234

186

The UL16-binding proteins (ULBPs) are a novel family of MHC class I-related molecules that were identified as targets of the human CMV glycoprotein, UL16. We have previously shown that ULBP expression renders a relatively resistant target cell sensitive to NK cytotoxicity, presumably by engaging NKG2D, an activating receptor expressed by NK and other immune effector cells. In this study we show that NKG2D is the ULBP counterstructure on primary NK cells and that its expression is up-regulated by IL-15 stimulation. Soluble forms of ULBPs induce marked protein tyrosine phosphorylation, and activation of the Janus kinase 2, STAT5, extracellular signal-regulated kinase, mitogen-activated protein kinase, and phosphatidylinositol 3-kinase (PI 3-kinase)/Akt signal transduction pathways. ULBP-induced activation of Akt and extracellular signal-regulated kinase and ULBP-induced IFN-γ production are blocked by inhibitors of PI 3-kinase, consistent with the known binding of PI 3-kinase to DAP10, the membrane-bound signal-transducing subunit of the NKG2D receptor. While all three ULBPs activate the same signaling pathways, ULBP3 was found to bind weakly and to induce the weakest signal. In summary, we have shown that NKG2D is the ULBP counterstructure on primary NK cells and for the first time have identified signaling pathways that are activated by NKG2D ligands. These results increase our understanding of the mechanisms by which NKG2D activates immune effector cells and may have implications for immune surveillance against pathogens and tumors.

“…These elements are also shared with IL-3 (unpublished studies) and probably C5a (although we have not yet examined p21 ras activation following C5a). The transient activity of the ERK pathway probably accounts for the cessation of LTC 4 following either stimulus (12), but as we have previously shown, the activity of ERK itself does not influence histamine or IL-4 secretion (12,14,37). It remains possible that signaling upstream of MEK-1, e.g., p21…”

Section: Discussionmentioning

confidence: 93%

“…We have recently demonstrated that extracellular signal-related kinase 1/2 (ERK1/2) and cytosolic phospholipase A 2 (cPLA 2 ) are phosphorylated during stimulation of basophils with anti-IgE Ab (12), FMLP, C5a, and IL-3 (13,14). For all stimuli, inhibition of mitogen-activated protein/ERK kinase (MEK) activity with PD98059 inhibits ERK1/2 and cPLA 2 phosphorylation, without effects on some other putative upstream signaling elements.…”

mentioning

confidence: 99%

Localizing a Control Region in the Pathway to Leukotriene C4 Secretion Following Stimulation of Human Basophils with Anti-IgE Antibody

Miura¹,

Lavens-Phillips²,

MacGlashan³

2001

Mediator release from human basophils is a self-limited process, but down-regulation of the signaling cascades leading to secretion of leukotriene C4 (LTC4) is controlled independently of the pathway leading to IL-4 secretion. In the current studies, we have explored the regulation of upstream signaling events leading to activation of extracellular signal-related kinases (ERKs; previously shown to be required for LTC4 generation) in human basophils. IgE-, but not FMLP-mediated activation, induced sustained tyrosine phosphorylation of syk, of shc, and an association of shc to the Grb2/son of sevenless 2 complex. In contrast, IgE-mediated activation resulted in transient activation of p21ras and mitogen-activated protein/ERK kinase 1, which were kinetically associated with phosphorylation of ERKs. The canonical Shc/Grb2/son of sevenless pathway to activation of p21ras is therefore sustained, while p21ras activity is not. We have previously shown that phosphatidylinositol 3 kinase activity is required for p21ras activity and, in the current studies, we show that of the p85-sensitive forms of p110 possible, basophils express only p110 δ and that there are no changes in association between p21ras and p110 δ in stimulated basophils. We used the generation of phospho-Akt as a marker of the presence of phosphatidylinositol-3,4,5-trisphosphate and found that phospho-Akt is transient on a time scale consistent with p21ras activity. On the basis of information obtained in these and other studies, we localize down-regulation of IgE-mediated LTC4 secretion to a region of the signaling cascade antecedent to p21ras activation, downstream of phosphatidylinositol 3 kinase activity and probably involving regulation of phosphatidylinositol-3,4,5-trisphosphate levels.