RESUMEN
Toll-like receptor (TLR) signals that initiate innate immune responses to pathogens must be tightly regulated to prevent excessive inflammatory damage to the host. The adaptor protein Mal is specifically involved in signaling via TLR2 and TLR4. We demonstrate here that after TLR2 and TLR4 stimulation Mal becomes phosphorylated by Bruton's tyrosine kinase (Btk) and then interacts with SOCS-1, which results in Mal polyubiquitination and subsequent degradation. Removal of SOCS-1 regulation potentiates Mal-dependent p65 phosphorylation and transactivation of NF-kappaB, leading to amplified inflammatory responses. These data identify a target of SOCS-1 that regulates TLR signaling via a mechanism distinct from an autocrine cytokine response. The transient activation of Mal and subsequent SOCS-1-mediated degradation is a rapid and selective means of limiting primary innate immune response.
Asunto(s)
Proteínas Portadoras/metabolismo , Proteínas de Transporte de Membrana/metabolismo , Proteínas de la Mielina/metabolismo , Proteolípidos/metabolismo , Proteínas Represoras/metabolismo , Proteínas Supresoras de la Señalización de Citocinas/metabolismo , Receptores Toll-Like/metabolismo , Animales , Proteínas Portadoras/genética , Proteínas Portadoras/inmunología , Línea Celular , Humanos , Inmunidad Innata , Proteínas de Transporte de Membrana/inmunología , Ratones , Ratones Endogámicos CBA , Ratones Noqueados , Proteínas de la Mielina/inmunología , Proteínas Proteolipídicas Asociadas a Mielina y Linfocito , Fosforilación , Proteolípidos/inmunología , Proteínas Represoras/genética , Proteínas Represoras/inmunología , Transducción de Señal , Proteína 1 Supresora de la Señalización de Citocinas , Proteínas Supresoras de la Señalización de Citocinas/deficiencia , Proteínas Supresoras de la Señalización de Citocinas/genética , Proteínas Supresoras de la Señalización de Citocinas/inmunología , Receptor Toll-Like 2/metabolismo , Receptor Toll-Like 4/metabolismo , Factor de Transcripción ReIA/genética , Factor de Transcripción ReIA/metabolismo , Activación Transcripcional , Tirosina/metabolismo , Ubiquitina/metabolismoRESUMEN
UNLABELLED: PAR-2 is expressed by osteoblasts and activated by proteases present during inflammation. PAR-2 activation inhibited osteoclast differentiation induced by hormones and cytokines in mouse bone marrow cultures and may protect bone from uncontrolled resorption. INTRODUCTION: Protease-activated receptor-2 (PAR-2), which is expressed by osteoblasts, is activated specifically by a small number of proteases, including mast cell tryptase and factor Xa. PAR-2 is also activated by a peptide (RAP) that corresponds to the "tethered ligand" created by cleavage of the receptor's extracellular domain. The effect of activating PAR-2 on osteoclast differentiation was investigated. MATERIALS AND METHODS: Mouse bone marrow cultures have been used to investigate the effect of PAR-2 activation on osteoclast differentiation induced by parathyroid hormone (PTH), 1,25 dihydroxyvitamin D3 [1,25(OH)2D3], and interleukin-11 (IL-11). Expression of PAR-2 by mouse bone marrow, mouse bone marrow stromal cell-enriched cultures, and the RAW264.7 osteoclastogenic cell line was demonstrated by RT-PCR. RESULTS: RAP was shown to inhibit osteoclast differentiation induced by PTH, 1,25(OH)2D3, or IL-11. Semiquantitative RT-PCR was used to investigate expression of mediators of osteoclast differentiation induced by PTH, 1,25(OH)2D3, or IL-11 in mouse bone marrow cultures and primary calvarial osteoblast cultures treated simultaneously with RAP. In bone marrow and osteoblast cultures treated with PTH, 1,25(OH)2D3, or IL-11, RAP inhibited expression of RANKL and significantly suppressed the ratio of RANKL:osteoprotegerin expression. Activation of PAR-2 led to reduced expression of prostaglandin G/H synthase-2 in bone marrow cultures treated with PTH, 1,25(OH)2D3, or IL-11. RAP inhibited PTH- or 1,25(OH)2D3-induced expression of IL-6 in bone marrow cultures. RAP had no effect on osteoclast differentiation in RANKL-treated RAW264.7 cells. CONCLUSION: These observations indicate that PAR-2 activation inhibits osteoclast differentiation by acting on cells of the osteoblast lineage to modulate multiple mediators of the effects of PTH, 1,25(OH)2D3, and IL-11. Therefore, the role of PAR-2 in bone may be to protect it from uncontrolled resorption by limiting levels of osteoclast differentiation.