A novel role for p21-activated protein kinase 2 in T cell activation
A novel role for p21-activated protein kinase 2 in T cell activation
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DOI：
10.4049/jimmunol.172.12.7324
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发表时间：
2004-06-15
影响因子：
4.4
通讯作者：
Masuda, ES
中科院分区：
文献类型：
--
作者：
Chu, PC;Wu, J;Masuda, ES
为了鉴定TCR信号通路的新组分，使用CD 69表达作为T细胞活化的标志物进行大规模的基于逆转录病毒的功能筛选。除了已知的调节剂之外，在T细胞筛选中分离了两种截短形式的p21活化激酶2（PAK 2），PAK 2DeltaL（1-224）和PAK 2DeltaS（1-113），两者都缺乏激酶结构域。PAK 2截短物PAK 2DeltaL阻断Jurkat T细胞中Ag受体诱导的NFAT活化和TCR介导的钙流。然而，它对PMA/离子霉素诱导的Jurkat细胞中的CD 69上调、抗IgM介导的B细胞中的CD 69上调或静息T细胞对化学引诱物的迁移反应的影响极小。我们表明，PAK 2激酶活性增加，在响应TCR刺激。此外，全长激酶失活形式的PAK 2阻断了Jurkat细胞中TCR诱导的CD 69上调和NFAT活性，表明激酶活性是TCR下游PAK 2功能所需的。我们还产生了缺乏Cdc 42/Rac相互作用结合区结构域的GFP融合的PAK 2截短物GFP-PAK 2（83-149）。我们表明，这种结构直接结合到PAK 2的激酶结构域，并抑制抗TCR刺激的T细胞活化。最后，我们证明，在原代T细胞中，显性负性PAK 2阻止抗CD 3/CD 28诱导的IL-2产生，和TCR诱导的CD 40配体表达，这两个关键功能的活化T细胞。总之，这些结果表明PAK 2作为T细胞活化的正调节剂的新作用。
To identify novel components of the TCR signaling pathway, a large-scale retroviral-based functional screen was performed using CD69 expression as a marker for T cell activation. In addition to known regulators, two truncated forms of p21-activated kinase 2 (PAK2), PAK2DeltaL(1-224) and PAK2DeltaS(1-113), both lacking the kinase domain, were isolated in the T cell screen. The PAK2 truncation, PAK2DeltaL, blocked Ag receptor-induced NFAT activation and TCR-mediated calcium flux in Jurkat T cells. However, it had minimal effect on PMA/ionomycin-induced CD69 up-regulation in Jurkat cells, on anti-IgM-mediated CD69 up-regulation in B cells, or on the migratory responses of resting T cells to chemoattractants. We show that PAK2 kinase activity is increased in response to TCR stimulation. Furthermore, a full-length kinase-inactive form of PAK2 blocked both TCR-induced CD69 up-regulation and NFAT activity in Jurkat cells, demonstrating that kinase activity is required for PAK2 function downstream of the TCR. We also generated a GFP-fused PAK2 truncation lacking the Cdc42/Rac interactive binding region domain, GFP-PAK2(83-149). We show that this construct binds directly to the kinase domain of PAK2 and inhibits anti-TCR-stimulated T cell activation. Finally, we demonstrate that, in primary T cells, dominant-negative PAK2 prevented anti-CD3/CD28-induced IL-2 production, and TCR-induced CD40 ligand expression, both key functions of activated T cells. Taken together, these results suggest a novel role for PAK2 as a positive regulator of T cell activation.