Uncovering a novel role of PLCß4 in selectively mediating TCR signaling in CD8+ but not CD4+ T cells.

Because of their common signaling molecules, the main T cell receptor (TCR) signaling cascades in CD4+ and CD8+ T cells are considered qualitatively identical. Herein, we show that TCR signaling in CD8+ T cells is qualitatively different from that in CD4+ T cells, since CD8α ignites another cardinal signaling cascade involving phospholipase C ß4 (PLCß4). TCR-mediated responses were severely impaired in PLCß4-deficient CD8+ T cells, whereas those in CD4+ T cells were intact. PLCß4-deficient CD8+ T cells showed perturbed activation of peripheral TCR signaling pathways downstream of IP3 generation. Binding of PLCß4 to the cytoplasmic tail of CD8α was important for CD8+ T cell activation. Furthermore, GNAQ interacted with PLCß4, mediated double phosphorylation on threonine 886 and serine 890 positions of PLCß4, and activated CD8+ T cells in a PLCß4-dependent fashion. PLCß4-deficient mice exhibited defective antiparasitic host defense and antitumor immune responses. Altogether, PLCß4 differentiates TCR signaling in CD4+ and CD8+ T cells and selectively promotes CD8+ T cell-dependent adaptive immunity.

Phospholipase C-ß in immune cells.

Great progress has recently been made in structural and functional research of phospholipase C (PLC)-ß. We now understand how PLC-ß isoforms (ß1-ß4) are activated by GTP-bound Gαq downstream of G protein-coupled receptors. Numerous studies indicate that PLC-ßs participate in the differentiation and activation of immune cells that control both the innate and adaptive immune systems. The PLC-ß3 isoform also interplays with tyrosine kinase-based signaling pathways, to inhibit Stat5 activation by recruiting the protein-tyrosine phosphatase SHP-1, with which PLC-ß3 and Stat5 form a multi-molecular signaling platform, named SPS complex. The SPS complex has important regulatory roles in tumorigenesis and immune cell activation.

Superantigen-induced steroid resistance depends on activation of phospholipase Cß2.

The glucocorticoid receptor is present in a TCR-associated complex, which includes the Src family tyrosine kinase Lck. Glucocorticoids rapidly dissociate this complex, resulting in the inhibition of canonical Lck-phospholipase C (PLC)γ-dependent TCR signaling. The relative importance of this nongenomic role for the glucocorticoid receptor compared with its direct transcriptional effects is not known. Superantigens induce a state of steroid resistance in activated T cells. It was reported that, in addition to canonical Lck-PLCγ signaling, superantigens can activate a noncanonical G protein-PLCß-dependent signaling pathway. In this study, we show that staphylococcal enterotoxin B activates a Gαq and PLCß2-dependent pathway in human T cells. We find that this pathway bypasses the need for canonical Lck-PLCγ signaling in T cell activation and renders superantigen-stimulated T cells insensitive to glucocorticoids in vitro. We show that the PLCß inhibitor U-73122 sensitizes staphylococcal enterotoxin B-treated mice to dexamethasone in vivo. In conclusion, we find that effects of glucocorticoids on TCR-induced T cell proliferation are mainly nongenomic and can be bypassed by the activation of an Lck-independent signaling pathway.

Immune regulation by phospholipase C-ß isoforms.

Rapid progress has recently been made regarding how phospholipase C (PLC)-ß functions downstream of G protein-coupled receptors and how PLC-ß functions in the nucleus. PLC-ß has also been shown to interplay with tyrosine kinase-based signaling pathways, specifically to inhibit Stat5 activation by recruiting the protein-tyrosine phosphatase SHP-1. In this regard, a new multimolecular signaling platform, named SPS complex, has been identified. The SPS complex has important regulatory roles in tumorigenesis and immune cell activation. Furthermore, a growing body of work suggests that PLC-ß also participates in the differentiation and activation of immune cells that control both the innate and adaptive immune systems.

Altered PLCß-1 expression in the gerbil hippocampal complex following spontaneous seizure.

Although the phospholipase C (PLC)ß-1 isoform is associated with spontaneous seizure and distinctively expressed in the telencephalon, the distribution of PLCß-1 expression in the epileptic gerbil hippocampus remains controversial. Therefore, we determined whether PLCß-1 is associated with spontaneous seizure in an animal model of genetic epilepsy. In the present study, PLCß-1 immunoreactivity was down-regulated in seizure-sensitive (SS) gerbils more than in seizure-resistant (SR) gerbils. The expression of PLCß-1 within calretinin (CR)- positive neurons was rarely detected within the dentate hilar region of SS gerbils. PLCß-1 immunoreactivity in the hippocampus was significantly elevated as compared to that in pre-seizure SS gerbil 3 h post-ictal. These findings suggest that alterations in PLCß-1 immunoreactivity in the SS gerbil hippocampus may be closely related to the epileptic state of the gerbil brain and transiently elevated PLCß-1 protein levels following seizure episodes. Such alterations may be compensatory responses in the SS gerbil hippocampus.

Phospholipase C-ß3 regulates FcɛRI-mediated mast cell activation by recruiting the protein phosphatase SHP-1.

Mast cells are major effectors in high-affinity IgE receptor (FcɛRI)-dependent allergic reactions. Here we show that phospholipase C (PLC)-ß3 is crucial for FcɛRI-mediated mast cell activation. Plcb3(-/-) mice showed blunted FcɛRI-dependent late-phase, but not acute, anaphylactic responses and airway inflammation. Accordingly, FcɛRI stimulation of Plcb3(-/-) mast cells exhibited reduced cytokine production but normal degranulation. Reduced cytokine production in Plcb3(-/-) cells could be accounted for by increased activity of the negative regulatory Src family kinase Lyn and reduced activities of the positive regulatory protein kinases MAPKs. Mechanistically, PLC-ß3 constitutively interacts with FcɛRI, Lyn, and SHP-1 (protein phosphatase). SHP-1 probably recognizes its substrates Lyn and MAPKs via the recently described kinase tyrosine-based inhibitory motif, KTIM. Consistent with PLC-ß3- and SHP-1-mediated repression of Lyn activity by dephosphorylation at Tyr396, FcɛRI-mediated phenotypes were similar in Plcb3(-/-) and SHP-1 mutant mast cells. Thus, we have defined a PLC-ß3- and SHP-1-mediated signaling pathway for FcɛRI-mediated cytokine production.

Suppression of PLCbeta2 by endotoxin plays a role in the adenosine A(2A) receptor-mediated switch of macrophages from an inflammatory to an angiogenic phenotype.

Toll-like receptor (TLR) 2, 4, 7, and 9 agonists, together with adenosine A(2A) receptor (A(2A)R) agonists, switch macrophages from an inflammatory (M1) to an angiogenic (M2-like) phenotype. This switch involves induction of A(2A)Rs by TLR agonists, down-regulation of tumor necrosis factor alpha (TNFalpha) and interleukin-12, and up-regulation of vascular endothelial growth factor (VEGF) and interleukin-10 expression. We show here that the TLR4 agonist lipopolysaccharide (LPS) induces rapid and specific post-transcriptional down-regulation of phospholipase C(PLC)beta1 and beta2 expression in macrophages by de-stabilizing their mRNAs. The PLCbeta inhibitor U73122 down-regulates TNFalpha expression by macrophages, and in the presence of A(2A)R agonists, up-regulates VEGF, mimicking the synergistic action of LPS with A(2A)R agonists. Selective down-regulation of PLCbeta2, but not PLCbeta1, using small-interfering RNA resulted in increased VEGF expression in response to A(2A)R agonists, but did not suppress TNFalpha expression. Macrophages from PLCbeta2(-/-) mice also expressed increased VEGF in response to A(2A)R agonists. LPS-mediated suppression of PLCbeta1 and beta2 is MyD88-dependent. In a model of endotoxic shock, LPS (35 microg/mouse, i.p.) suppressed PLCbeta1 and beta2 expression in spleen, liver, and lung of wild-type but not MyD88(-/-) mice. These studies indicate that LPS suppresses PLCbeta1 and beta2 expression in macrophages in vitro and in several tissues in vivo. These results suggest that suppression of PLCbeta2 plays an important role in switching M1 macrophages into an M2-like state.