Activation of cGMP-dependent Protein Kinase Iβ Inhibits Interleukin 2 Release and Proliferation of T Cell Receptor-stimulated Human Peripheral T Cells

Abstract

Several major functions of type I cGMP-dependent protein kinase (cGK I) have been established in smooth muscle cells, platelets, endothelial cells, and cardiac myocytes. Here we demonstrate that cGK I␤ is endogenously expressed in freshly purified human peripheral blood T lymphocytes and inhibits their proliferation and interleukin 2 release. Incubation of human T cells with the NO donor, sodium nitroprusside, or the membrane-permeant cGMP analogs PET-cGMP and 8-pCPTcGMP, activated cGK I and produced (i) a distinct pattern of phosphorylation of vasodilator-stimulated phosphoprotein, (ii) stimulation of the mitogen-activated protein kinases ERK1/2 and p38 kinase, and, upon anti-CD3 stimulation, (iii) inhibition of interleukin 2 release and (iv) inhibition of cell proliferation. cGK I was lost during in vitro culturing of primary T cells and was not detectable in transformed T cell lines. The proliferation of these cGK I-deficient cells was not inhibited by even high cGMP concentrations indicating that cGK I, but not cGMP-regulated phosphodiesterases or channels, cAMP-dependent protein kinase, or other potential cGMP mediators, was responsible for inhibition of T cell proliferation. Consistent with this, overexpression of cGK I␤, but not an inactive cGK I␤ mutant, restored cGMP-dependent inhibition of cell proliferation of Jurkat cells. Thus, the NO/cGMP/cGK signaling system is a negative regulator of T cell activation and proliferation and of potential significance for counteracting inflammatory or lymphoproliferative processes.Effective T cell activation can be elicited by a combination of (i) engagement of the T cell antigen receptor (TCR) 1 ⅐CD3complex with foreign antigen presented on major histocompatibility complex molecules and (ii) ligation of the CD28 T cell antigen by B7 molecules (CD80 and CD86) on the surface of antigen-presenting cells, resulting in transcription factor activation, interleukin 2 (IL-2) production, and cell proliferation (1-4). Activation and subsequent proliferation of resting T lymphocytes is an essential process in the T cell-mediated immune response. In vitro, T cells respond to stimulation of the TCR⅐CD3 complex (without costimulation) with IL-2-dependent induction of cell proliferation (2, 5). A costimulatory signal from the CD28 pathway synergizes with the signal from the TCR⅐CD3 complex to further increase cell proliferation and IL-2 production, and T cell proliferation becomes largely independent of IL-2 (6). Intracellular signaling pathways, which may mediate T cell activation, include phosphorylation cascades that stimulate MAP kinases including ERK (3, 7, 8) and p38 kinase (9, 10), as well as phosphatidylinositol 3-kinase (11, 12) and its target Akt/ protein kinase B (13). Reports of cyclic nucleotide effects on T cell activation indicate that cAMP is inhibitory, whereas effects of cGMP have not been conclusively elucidated. In fact, lowering of cAMP levels by induction of phosphodiesterase-7 was reported to be required for T cell activation (14). Nitric oxide, which activ...

Loss of Sp1 function via inhibitory phosphorylation in antifolate-resistant human leukemia cells with down-regulation of the reduced folate carrier

“…44 However, we find here that PKGI and PKGII transcripts were completely absent from both parental CCRF-CEM and their antifolate-resistant cells. The finding that PKG is not expressed in the T-cell line CCRF-CEM used in this study is consistent with previous reports that upon in vitro culturing of primary T-cells, PKGI expression is lost, 45 whereas PKGII is not expressed at all in blood cells. 38 Hence, since cGMP exerts its biologic effect by modulating various proteins, including cGMP-dependent PKGI and PKGII, 38 PDEs, 43 and cyclic nucleotide-regulated ion channels, 46 it is clear that the putative cGMPdependent activation of PP2A is independent of PKG.…”

Quantitative analysis of the cardiac fibroblast transcriptome—implications for NO/cGMP signaling

“…Both isoforms of cGK I α and β were demonstrated to be present in cardiac fibroblasts and could potentially participate in antiproliferative action. Antiproliferative effects of cGK I have been described for the β isoform of cGK I in smooth muscle [36], BHK cells [37], mesangial cells [38] and T cells [39]. Recently cGMP and cAMP effects on PDGF-BB-induced proliferation were studied in rat cardiac fibroblasts [40].…”

“…For proliferation assays, mouse cardiac fibroblasts (passage 15-20 as well as passage [27][28][29][30][31][32][33][34][35][36][37][38][39] were seeded onto 12 well plates (Greiner, Frickenhausen, Germany) at a density of 500 cells/cm 2 . Subconfluent cells were starved in medium containing 0.1 % FCS for 2 days, then cell proliferation was stimulated for 48 h by the addition of fresh medium containing 5 % FCS in the presence or absence of 8-pCPT-cGMP (Biolog, Bremen, Germany).…”

Bacterial enterotoxins are associated with resistance to colon cancer

“…Thus, in human vascular smooth muscle cells, cGMP delays the G 1 ͞S transition by down-regulation of cyclin D1 and cyclin-dependent kinase 4 activities after platelet-derived growth factor stimulation (25). Also, cGMP suppresses human T cell proliferation by inhibiting IL-2 release (26). In addition, proliferation of glomerular mesangial cells by phorbol 12,13-dibutyrate-mediated activation of mitogen-activated protein kinase is antagonized by cGMPinduced expression of the specific phosphatase MKP-1 (27).…”