GPI-anchored protein-mediated cell signaling system in B-precursor cells

我们先前报道了分化簇（CD）24的交联诱导伯基特淋巴瘤细胞以及B-前体细胞ALL细胞的凋亡。CD 24是一种GPI锚定蛋白，其凋亡诱导作用是由筏微结构域介导的细胞信号系统介导的。为了研究GPI锚定蛋白在正常B前体细胞中介导的细胞信号传导，我们建立了一种体外培养系统，将CD 34+细胞分化为前B细胞。利用该培养体系，我们发现CD 24的交联可诱导正常人pro-B细胞凋亡。B细胞连接蛋白（BLNK）是B细胞受体（BCR）信号通路的一个组成部分，筏微结构域在其中起着重要作用。我们研究了人前B ALL细胞系中的BLNK表达，发现检测的四种细胞系之一HPB-NULL细胞缺乏BLNK表达，我们使用这些表达和不表达BLNK的人前B ALL细胞系研究前BCR交联后的细胞内信号传导事件。当pre-BCR与抗μ重链抗体交联时，观察到细胞内分子（包括Syk、Shc、ERK MAP激酶和AKT）的显著磷酸化以及Ras的活化，而不考虑BLNK表达的缺陷，表明BLNK不是pre-BCR介导的MAP激酶和磷脂酰肌醇3（PI 3）激酶信号转导活化所必需的。相比之下，仅在表达BLNK的细胞中观察到磷脂酶C-γ 2（PLC-γ 2）磷酸化和由前BCR交联介导的细胞内Ca（2+）水平升高，表明BLNK对PLC-γ 2诱导的Ca（2+）内流至关重要。

We previously reported that the cross-linking of cluster of differentiation (CD)24 induces apoptosis in Burkitt's lymphoma cells as well as B-predursor ALL cells. CD24 is a GPI-anchored protein and this apoptosis induction is mediated by raft microdomain-mediated cell signaling system. To investigate the cell signaling mediated by GPI-anchored protein in normal B precursor cells, we developed an in vitro culture system to differentiate CD34+ cells into pro-B cells. Using this culture system, we found that the cross-linking of CD24 induced apoptosis in normal human pro-B cells.B-cell linker protein (BLNK) is a component of the B-cell receptor (BCR) signalling pathway, in which raft microdomain is deeply involved. We investigated BLNK expression in human pre-B ALL cell lines and found that one of the four cell lines tested, HPB-NULL cells, was found to lack BLNK expression, and we used these human pre-B ALL cell lines that express and do not express BLNK to investigate the intracellular signalling events following pre-BCR cross-linking. When pre-BCR was cross-linked with anti-μ heavy-chain antibodies, significant phosphorylation of intracellular molecules, including Syk, Shc, ERK MAP kinase, and AKT, and an activation of Ras were observed without regard to deficiency of BLNK expression, suggesting that BLNK is not required for pre-BCR-mediated activation of MAP kinase and phosphatidyl-inositol 3 (PI3) kinase signalling. By contrast, phospholipase C-gamma2 (PLC-gamma2) phosphorylation and an increase in intracellular Ca(2+) level mediated by pre-BCR cross-linking were observed only in the BLNK-expressing cells, indicating that BLNK is essential for PLC-gamma2-induced Ca(2+) influx.