JNK MAPK Pathway Regulates Constitutive Transcription of CCL5 by Human NK Cells through SP1

Abstract

N atural killer cells have features similar to CD8 ϩ T cells in that they recognize and lyse virally infected and neoplastic cells; express receptors such as CD28, CD43, and 2B4; and contain cytolytic granules (1-6). The activation of NK surface receptors triggers microtubule organization, granule polarization, and cytotoxicity signals through both ERK2 and JNK1 phosphorylation pathways (7,8).The chemokine CCL5 has been suggested to act as an important mediator of diverse inflammatory processes (9 -11). In CTL, the protein is associated with the development and expression of effector function (12)(13)(14). In CD8 ϩ T cells, CCL5 transcription occurs late (3-5 days) after the activation of naive cells and is coincident with the up-regulation of perforin, the granzymes, and granulysin (15)(16)(17). A complex of nuclear proteins recruited to the promoter by Krü ppel-like factor 13 (KLF13) 3 controls the transcription of CCL5 in CTL (18,19). KLF13 is translationally regulated and appears several days after T cell activation. The p38 and ERK1/2 MAPK pathways control KLF13 gene expression, whereas pre-mRNA processing factor 4 homolog B, a member of the MAPK family, regulates KLF13 phosphorylation in T cells and by extension CCL5 expression (20,21). In NK cells, the MAPKs and PI3K pathways are also involved in triggering the polarized release of cytolytic granules (7,8,(22)(23)(24); however, the role of these pathways in the control of CCL5 expression in NK cells has not been investigated.We report in this study that unlike in T cells, CCL5 mRNA is constitutively transcribed and translated in peripheral blood NK cells. The CCL5 protein is localized to secretory vesicles that are distinct from the cytolytic granules. The constitutive transcription of CCL5 mRNA in NK cells is mediated through binding of the SP1 transcription factor to a site just upstream from the TATA box of the CCL5 gene. The JNK/MAPK pathway in turn controls SP1 expression and, through this, constitutive CCL5 transcription. Materials and Methods Abs, inhibitors, and oligonucleotidesPhospho-specific Abs that recognize the activated forms of phospho-stressactivated protein kinase/JNK (Thr 183 /Tyr 185 ) and inactive forms of stressactivated protein kinase/JNK were purchased from Cell Signaling Technology (catalog 9251 and 9252). The Abs for NF-B, p50, p52, p65, SP1, and SP3 were purchased from Upstate Biotechnology (catalog 06-886, 06-413, 06-418, 07-645, and 07-107, respectively), and ␤ actin for proteinloading control and KLF13 was purchased from Abcam (catalog ab8227 and ab1127). Primary Abs were as follows: mouse IgG2b-anti-human CCL5 VL1 or VL2 hybridoma supernatant (25), mouse IgG1 anti-human granzyme B (Serotec; catalog MCA2120PE), mouse anti-human perforin (BD Biosciences; catalog 556434), goat anti-mouse IgG1 CD3 Pacific Blue (DakoCytomation; PB982), goat anti-mouse IgG1 CD56 allophycocyanin conjugated (BD Biosciences; catalog IM2474), and rabbit anti-human CD8 (Neomarkers; catalog RM-9116-R7 The costs of publication of this article were defray...

14-3-3ϵ and 14-3-3σ Inhibit Toll-like Receptor (TLR)-mediated Proinflammatory Cytokine Induction

“…Specifically, we have shown that although MyD88 inhibits poly(I:C)-mediated RANTES production, Mal does not (7,11). Therefore, our work correlates with other studies showing that CCL5 induction is a complex cell type-and ligand-dependant process in vivo (32,33).…”

“…In contrast, Pam 3 CSK 4 -, LPS-, R848-, and CpG-mediated 14-3-3 expression, facilitated by MyD88, is enhanced by 14-3-3. Because studies have shown that the CCL5 promoter contains numerous response elements for the transcriptional activators NF-B, AP-1, SP-1, IRF3, IRF-1-RE, and STAT1 (31)(32)(33), we propose that transcriptional induction of CCL5 mRNA may be differentially regulated by MyD88 and TRIF through the induction/suppression of various transcriptional activators. Herein, we preclude NF-B and IRF3 because they are comparably modulated following stimulation with the various PAMPs.…”

Sphingosine Kinase 1 Regulates Tumor Necrosis Factor-mediated RANTES Induction through p38 Mitogen-activated Protein Kinase but Independently of Nuclear Factor κB Activation

“…In the TNF response, many transcription factors have been implicated in RANTES induction including NF-B and interferon regulatory factor 1 (29,30), and activator protein 1 and NF-AT in airway epithelial and smooth muscle cells, respectively (31). Moreover, upstream of transcription factors, other signaling cascades have been implicated in RANTES regulation such as JNK (32), ERK (33), and p38 MAPK (34). More recently, our group identified sphingolipids as regulators of RANTES through the acid sphingomyelinase/ acid ceramidase pathway (16).…”

Angiotensin II inhibits chemokine CCL5 expression in vascular smooth muscle cells from spontaneously hypertensive rats

“…CCL5 transcription has been known to occur relatively late after the activation of naive cells and is coincident with the upregulation of cytolytic granules in cytotoxic T-cells. 29 However, CCL5 mRNA is constitutively transcribed and translated in peripheral blood natural killer cells 30 and the expression of CCL5 mRNA has been observed in naive WKY VSMCs. 25 Thus, constitutive expression of a chemokine may be dependent on cell type.…”