RAFTK/Pyk2 mediates LPA-induced PC12 cell migration.
RAFTK/Pyk2 mediates LPA-induced PC12 cell migration.
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RAFTK/Pyk2 介导 LPA 诱导的 PC12 细胞迁移。
DOI：
10.1016/j.cellsig.2005.08.018
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发表时间：
2006
影响因子：
4.8
通讯作者：
Avraham,Shalom
中科院分区：
文献类型：
--
作者：
Park,Shin-Young;Schinkmann,KarinA;Avraham,Shalom
磷脂溶血磷脂酸（LPA）是血清的正常成分，其作为脂质生长因子和细胞内信号分子起作用。在这份报告中，我们研究了酪氨酸激酶RAFTK/Pyk 2在LPA诱导的细胞迁移中的信号转导机制和功能。在神经内分泌PC 12细胞中LPA刺激后的酪氨酸磷酸化分析显示6种主要的酪氨酸磷酸化蛋白，估计大小为180，120，115，68，44和42 kDa。这些蛋白质分别被鉴定为表皮生长因子受体（EGFR）、粘着斑激酶、RAFTK/Pyk 2、桩蛋白、Erk 1和Erk 2。使用特定的药理学抑制剂，我们发现，RAFTK/Pyk 2的酪氨酸磷酸化是细胞内Ca 2+依赖性的，但不是EGFR依赖性的，在LPA刺激这些细胞。此外，细胞骨架和信号支架蛋白，桩蛋白，与RAFTK/Pyk 2的钙依赖性的方式，并受其调节。LPA受体的鉴定表明，LPA 1（Edg 2）和LPA 2（Edg 4）是LPA的主要受体，而LPA 3受体（Edg 7）的表达是有限的。在使用LPA 1/LPA 3受体特异性拮抗剂VPC 32179后，我们观察到LPA 1/LPA 3受体的抑制对LPA诱导的RAFTK磷酸化没有影响，强烈地表明LPA 2受体是RAFTK磷酸化的关键介质。此外，LPA诱导PC 12细胞迁移，随后被显性负性形式的FAK，FRNK阻断。显性阴性形式的小GT3 Ras的表达也阻断了LPA诱导的细胞迁移和RAFTK磷酸化。总之，这些结果表明，RAFTK是一个关键的信号分子，介导的LPA诱导的PC 12细胞迁移的Ras依赖性方式。
The phospholipid lysophosphatidic acid (LPA) is a normal constituent of serum that functions as a lipid growth factor and intracellular signaling molecule. In this report, we have investigated the signaling mechanism and function of the tyrosine kinase RAFTK/Pyk2 in LPA-induced cell migration. Analysis of tyrosine phosphorylation upon LPA stimulation in neuroendocrine PC12 cells revealed 6 major tyrosine-phosphorylated proteins with estimated sizes of 180, 120, 115, 68, 44, and 42 kDa. These proteins were identified as epidermal growth factor receptor (EGFR), focal adhesion kinase, RAFTK/Pyk2, paxillin, Erk 1, and Erk 2, respectively. Using specific pharmacological inhibitors, we found that the tyrosine phosphorylation of RAFTK/Pyk2 was intracellular Ca2+-dependent, but not EGFR-dependent, during LPA stimulation of these cells. Moreover, the cytoskeletal and signal scaffolding protein, paxillin, associated with and was regulated by RAFTK/Pyk2 in a Ca2+-dependent manner. Characterization of LPA receptors showed that LPA1(Edg2) and LPA2(Edg4) are major receptors for LPA, while LPA3receptor (Edg7) expression was limited. Upon using the LPA1/LPA3receptor-specific antagonist VPC 32179, we observed that inhibition of the LPA1/LPA3receptors had no effect on the LPA-induced phosphorylation of RAFTK, strongly suggesting that the LPA2receptor is a key mediator of RAFTK phosphorylation. Furthermore, LPA induced PC12 cell migration, which was subsequently blocked by the dominant-negative form of FAK, FRNK. Expression of a dominant-negative form of the small GTPase Ras also blocked LPA-induced cell migration and RAFTK phosphorylation. Taken together, these results indicate that RAFTK is a key signaling molecule that mediates LPA-induced PC12 cell migration in a Ras-dependent manner.