Nuclear NF-kappaB p65 phosphorylation at serine 276 by protein kinase A contributes to the malignant phenotype of head and neck cancer.
Nuclear NF-kappaB p65 phosphorylation at serine 276 by protein kinase A contributes to the malignant phenotype of head and neck cancer.
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DOI：
10.1158/1078-0432.ccr-09-1352
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发表时间：
2009-10-01
期刊：
影响因子：
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通讯作者：
Van Waes C
中科院分区：
文献类型：
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作者：
Arun P;Brown MS;Ehsanian R;Chen Z;Van Waes C
抑制剂 Kappa B 激酶对经典 NF-κB 亚基 RELA/p65 在丝氨酸 536 处的异常核激活和磷酸化在头颈鳞状细胞癌 (HNSCC) 中普遍存在，但其他激酶在 NF-κB 激活中的作用尚未明确。在这里，我们研究了蛋白激酶 A (PKA) 磷酸化 p65-Ser276 在恶性表型、基因反式激活以及作为潜在治疗靶点中的普遍性和功能。在 HNSCC 组织阵列和细胞系中测定磷酸化和总 p65 蛋白的表达和定位。检查了 PKA 抑制剂 H-89 对细胞增殖和细胞周期的影响，以及 H-89 和 PKA 特异性 siRNA 敲低对 NF-κB 激活和下游基因表达的影响。核 NF-κB p65 Ser276 磷酸化在 HNSCC 和邻近的发育不良粘膜中普遍存在，但在正常粘膜中局限于细胞质。在 HNSCC 系中，TNF-α 显着增加，而 H-89 抑制组成型和 TNF-α 诱导的核 p65-Ser276 磷酸化，并显着抑制 NF-κB 和靶基因 IL-8 报告基因活性。通过 siRNA 敲除 PKA 可抑制 NF-κB、IL-8 和 BCL-XL 报告基因活性。 H-89抑制细胞增殖、诱导细胞死亡并阻断细胞周期于G1/S期。与其生物学效应一致，H-89 下调 NF-κB 相关基因 Cyclin D1、BCL2、BCL-XL、COX2、IL-8 和 VEGF 以及诱导细胞周期抑制剂 p21CIP1/WAF1 的表达，同时抑制增殖标记物 Ki67。 PKA 引起的 NF-κB RELA Ser276 磷酸化可促进恶性表型，并具有作为 HNSCC 治疗靶点的潜力。
Aberrant nuclear activation and phosphorylation of the canonical NF-κB subunit RELA/p65 at Serine-536 by Inhibitor Kappa B Kinase is prevalent in head and neck squamous cell carcinoma (HNSCC), but the role of other kinases in NF-κB activation has not been well defined. Here, we investigated the prevalence and function of p65-Ser276 phosphorylation by Protein Kinase A (PKA) in the malignant phenotype, gene transactivation, and as a potential target for therapy. Phospho and total p65 protein expression and localization was determined in HNSCC tissue array and in cell lines. The effects of PKA inhibitor H-89 on cell proliferation and cell cycle and of H-89 and PKA specific siRNA knockdown on NF-κB activation and downstream gene expression were examined. Nuclear NF-κB p65 phosphorylated at Ser276 was prevalent in HNSCC and adjacent dysplastic mucosa, but localized to the cytoplasm in normal mucosa. In HNSCC lines, TNF-α significantly increased while H-89 inhibited constitutive and TNF-α induced nuclear p65-Ser276 phosphorylation, and significantly suppressed NF-κB and target gene IL-8 reporter activity. Knock down of PKA by siRNA inhibited NF-κB, IL-8 and BCL-XL reporter gene activities. H-89 suppressed cell proliferation, induced cell death and blocked the cell cycle in G1/S phase. Consistent with its biological effects, H-89 down-modulated expression of NF-κB related genes Cyclin D1, BCL2, BCL-XL, COX2, IL-8, and VEGF, as well as induced cell cycle inhibitor p21CIP1/WAF1, while suppressing proliferative marker Ki67. NF-κB RELA Ser276 phosphorylation by PKA promotes the malignant phenotype and holds potential as a therapeutic target in HNSCC.