Analysis of the IL-6-mediated intracellular signaling molecules in myeloma cell growth

细胞因子白细胞介素-6（IL-6）是人骨髓瘤细胞的生长因子。在IL-6刺激后，信号转导和转录激活因子（STAT）3和细胞外信号调节激酶（ERK）1/2被激活。我们最近发现，在骨髓瘤细胞系中，STAT 3和ERK 1/2的激活对于IL-6诱导的增殖是不够的，这进一步需要src家族蛋白酪氨酸激酶（PTKs）的激活，例如与CD 45蛋白酪氨酸磷酸酶（PTP）相关的林恩。为了探索控制IL-6刺激如何诱导骨髓瘤细胞增殖的分子机制，例如林恩的下游分子，我们建立了过表达林恩的骨髓瘤细胞系ILKM 2和ILKM 8。我们已经发现Lyn-overexpressed ILKM 2和ILKM 8都具有比它们的模拟对应物更高的响应IL-6的增殖率。相反，在缺乏IL-6的情况下，这些Lyns过表达的细胞系显示出更高的susceptibility。 ...更多信息 对细胞死亡的敏感性。细胞表面IL-6受体α和CD 45的表达不受Lyn-R过表达的影响。我们还发现，在用IL-6过度表达的细胞中，磷脂酰肌醇3-激酶（PI 3-K）和Akt的活化增强，而另外两种主要的IL-6应答分子STAT 3和ERK 1/2的活化水平保持不变，表明这些是Lyn-PI 3 K-Akt的独立信号传导途径。在Lyn-overexpressed细胞中，林恩PTK似乎是组成型活性的，因为在催化结构域中的正调节酪氨酸残基高度磷酸化，而在COOH-末端的负调节酪氨酸残基没有磷酸化，推测是由于其通过CD 45 PTP去磷酸化。在免疫沉淀试验中，PI 3-K与林恩共沉淀，并且通过药理学试剂特异性抑制PI 3-K或Akt活性抑制IL-6诱导的Lyn过表达细胞的增殖，这表明PI 3 K-与活化的林恩相关的Akt通路确实与其伴随的骨髓瘤细胞生长增强有关，尽管仍需要下游信号分子需要解释实验也正在进行中，以澄清可能的机制，加快细胞死亡的林恩过度表达的骨髓瘤细胞作为一个结果的IL-6撤出，我们的工作将提供一个理解林恩的作用，在IL-6受体介导的信号。少

A cytokine, interleukin-6 (IL-6) is a growth factor for human myeloma cells. Upon IL-6 stimulation, signal transducer and activator of transcription (STAT) 3 and extracellular signal-regulated kinase (ERK) 1/2 are activated. We recently found that in myeloma cell lines the activation of both STAT3 and ERK1/2 is not sufficient for the IL-6-induced proliferation, which further requires the activation of the src family protein-tyrosine kinases (PTKs), such as Lyn, associated with the CD45 protein-tyrosine phosphatase (PTP). To explore the molecular mechanism that governs how IL-6 stimulation induces the proliferation of myeloma cells, such as downstream molecules of Lyn, we have established the myeloma cell lines, ILKM2 and ILKM8 overexpressing Lyn. We have found that both the Lyn-overexpressed ILKM2 and ILKM8 have the higher proliferative rate in response to IL-6 than that of their mock counterpart. In contrast, in the absence of IL-6 these Lyn-overexpressed cell lines showed higher susc … More eptibility to cell death. Cell surface expression of IL-6 receptor a and CD45 was not influenced by the Lyn-overexpression. We have also found that the enhanced activation of the phosphatidylinositol 3-kinase (PI3-K) and Akt in the Lyn-overexpressed cells with IL-6, whereas the activation levels of two other major IL-6-responsive molecules, STAT3 and ERK1/2 remained unchanged, indicating that these are the independent signaling pathways of the Lyn-PI3K-Akt. In the Lyn-overexpressed cells, the Lyn PTK seems to be constitutively active because the positive regulatory tyrosine residue in the catalytic domain was highly phosphorylated, while the negative regulatory tyrosine residue in the COOH-terminus was not phosphorylated, presumably due to its dephosphorylation by CD45 PTP. The PI3-K was co-precipitated with Lyn in the immunoprecipitation assay, and specific inhibition of PI3-K or Akt activities by the pharmacologic reagents suppressed the IL-6-induced proliferation of the Lyn-overexpressed cells, suggesting that the activation of the PI3K-Akt pathways associated with the activated Lyn is indeed related to their concomitant augmentation of myeloma cell growth although the downstream signaling molecules are still need to be explicated. Experiments are also underway to clarify the possible mechanisms of the accelerated cell death of the Lyn-overexpressed myeloma cells as a consequence of the IL-6 withdrawal, our work will provide an understanding of Lyn's role in the IL-6 receptor-mediated signaling. Less