Department of Bioscience and Biotechnology, Sejong University, Seoul 143-747, Republic of Korea.
Exp Cell Res. 2011 Oct 15;317(17):2447-55. doi: 10.1016/j.yexcr.2011.07.003. Epub 2011 Jul 14.
B-cell activating factor (BAFF) plays a role in the generation and the maintenance of mature B cells. Lipopolysaccharide (LPS) increased BAFF expression through the activation of toll-like receptor 4 (TLR4)-dependent signal transduction. Here, we investigated the mechanism of action on mouse BAFF (mBAFF) expression by cAMP production in Raw264.7 mouse macrophages. mBAFF expression was increased by the treatment with a cAMP analogue, dibutyryl-cAMP which is the activator of protein kinase A (PKA), cAMP effector protein. PKA activation was measured by the phosphorylation of cAMP-response element binding protein (CREB) on serine 133 (S133). cAMP production and CREB (S133) phosphorylation were augmented by LPS-stimulation. While mBAFF promoter activity was enhanced by the co-transfection with pS6-RSV-CREB, it was reduced by siRNA-CREB. PKA inhibitor, H-89, reduced CREB (S133) phosphorylation and mBAFF expression in control and LPS-stimulated macrophages. Another principal cAMP effector protein is cAMP-responsive guanine nucleotide exchange factor (Epac), a Rap GDP exchange factor. Epac was activated by the treatment with 8-(4-chloro-phenylthio)-2'-O-methyladenosine-3',5'-cyclic monophosphate (CPT), Epac activator, as judged by the measurement of Rap1 activation. Basal level of mBAFF expression was increased by CPT treatment. LPS-stimulated mBAFF expression was also slightly enhanced by co-treatment with CPT. In addition, dibutyryl-cAMP and CPT enhanced mBAFF expression in bone marrow-derived macrophages (BMDM). With these data, it suggests that the activation of PKA and cAMP/Epac1/Rap1 pathways could be required for basal mBAFF expression, as well as being up-regulated in the TLR4-induced mBAFF expression.
B 细胞激活因子 (BAFF) 在成熟 B 细胞的生成和维持中发挥作用。脂多糖 (LPS) 通过激活 Toll 样受体 4 (TLR4) 依赖性信号转导来增加 BAFF 的表达。在这里，我们研究了 cAMP 产生对 Raw264.7 小鼠巨噬细胞中小鼠 BAFF (mBAFF) 表达的作用机制。cAMP 类似物二丁酰基-cAMP（蛋白激酶 A (PKA) 的激活剂，cAMP 效应蛋白）处理可增加 mBAFF 的表达。通过检测丝氨酸 133 (S133) 磷酸化的 cAMP 反应元件结合蛋白 (CREB) 来测量 PKA 的激活。LPS 刺激可增加 cAMP 产生和 CREB (S133) 磷酸化。虽然 mBAFF 启动子活性通过 pS6-RSV-CREB 的共转染而增强，但通过 siRNA-CREB 则降低。PKA 抑制剂 H-89 可降低对照和 LPS 刺激的巨噬细胞中 CREB (S133) 磷酸化和 mBAFF 的表达。另一种主要的 cAMP 效应蛋白是 cAMP 反应性鸟嘌呤核苷酸交换因子 (Epac)，一种 Rap GDP 交换因子。通过用 8-(4-氯-苯硫基)-2'-O-甲基腺苷-3'，5'-环单磷酸 (CPT)（Epac 激活剂）处理来判断 Rap1 的激活，来判断 Epac 的激活。CPT 处理可增加 mBAFF 基础表达水平。CPT 共处理也略微增强了 LPS 刺激的 mBAFF 表达。此外，二丁酰基-cAMP 和 CPT 增强了骨髓来源的巨噬细胞 (BMDM) 中的 mBAFF 表达。有了这些数据，表明 PKA 和 cAMP/Epac1/Rap1 途径的激活可能是基础 mBAFF 表达所必需的，并且在 TLR4 诱导的 mBAFF 表达中也被上调。
