Cell Signaling Technology, Inc., Danvers, Massachusetts, United States of America.
PLoS One. 2011 Apr 28;6(4):e19169. doi: 10.1371/journal.pone.0019169.
Activating mutations of FMS-like tyrosine kinase-3 (FLT3) are found in approximately 30% of patients with acute myeloid leukemia (AML). FLT3 is therefore an attractive drug target. However, the molecular mechanisms by which FLT3 mutations lead to cell transformation in AML remain unclear. To develop a better understanding of FLT3 signaling as well as its downstream effectors, we performed detailed phosphoproteomic analysis of FLT3 signaling in human leukemia cells. We identified over 1000 tyrosine phosphorylation sites from about 750 proteins in both AML (wild type and mutant FLT3) and B cell acute lymphoblastic leukemia (normal and amplification of FLT3) cell lines. Furthermore, using stable isotope labeling by amino acids in cell culture (SILAC), we were able to quantified over 400 phosphorylation sites (pTyr, pSer, and pThr) that were responsive to FLT3 inhibition in FLT3 driven human leukemia cell lines. We also extended this phosphoproteomic analysis on bone marrow from primary AML patient samples, and identify over 200 tyrosine and 800 serine/threonine phosphorylation sites in vivo. This study showed that oncogenic FLT3 regulates proteins involving diverse cellular processes and affects multiple signaling pathways in human leukemia that we previously appreciated, such as Fc epsilon RI-mediated signaling, BCR, and CD40 signaling pathways. It provides a valuable resource for investigation of oncogenic FLT3 signaling in human leukemia.
FMS 样酪氨酸激酶-3（FLT3）的激活突变存在于大约 30%的急性髓细胞白血病（AML）患者中。因此，FLT3 是一个有吸引力的药物靶点。然而，FLT3 突变导致 AML 细胞转化的分子机制尚不清楚。为了更好地了解 FLT3 信号及其下游效应物，我们对人类白血病细胞中的 FLT3 信号进行了详细的磷酸蛋白质组学分析。我们从 AML（野生型和突变型 FLT3）和 B 细胞急性淋巴细胞白血病（FLT3 正常和扩增）细胞系中的约 750 种蛋白质中鉴定出超过 1000 个酪氨酸磷酸化位点。此外，我们使用稳定同位素标记的氨基酸在细胞培养中（SILAC），能够定量测定 400 多个磷酸化位点（pTyr、pSer 和 pThr），这些磷酸化位点对 FLT3 驱动的人类白血病细胞系中的 FLT3 抑制有反应。我们还将这种磷酸蛋白质组学分析扩展到原发性 AML 患者样本的骨髓中，在体内鉴定出超过 200 个酪氨酸和 800 个丝氨酸/苏氨酸磷酸化位点。这项研究表明，致癌性 FLT3 调节涉及多种细胞过程的蛋白质，并影响我们以前认为的人类白血病中的多种信号通路，如 Fc epsilon RI 介导的信号、BCR 和 CD40 信号通路。它为研究人类白血病中的致癌性 FLT3 信号提供了有价值的资源。
