Proteome and transcriptome analysis of retinoic acid-induced differentiation of human acute promyelocytic leukemia cells, NB4.
Proteome and transcriptome analysis of retinoic acid-induced differentiation of human acute promyelocytic leukemia cells, NB4.
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DOI：
10.1021/pr049976r
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发表时间：
2004-04
影响因子：
4.4
通讯作者：
Daojing Wang;Ronald H. Jensen;G. Gendeh;Katherine E. Williams;M. Pallavicini
中科院分区：
文献类型：
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作者：
Daojing Wang;Ronald H. Jensen;G. Gendeh;Katherine E. Williams;M. Pallavicini
急性早幼粒细胞白血病 (APL) 的特征是 t(15:17) 易位，该易位将视黄酸受体基因与早幼粒细胞基因融合，从而阻止分化为正常粒细胞。 NB4 细胞源自人类急性早幼粒细胞白血病，显示出这种基因型和表型。全反式视黄酸 (ATRA) 在体外诱导 NB4 细胞培养物的分化，在体内诱导 APL 的分化，尽管经常出现对分化治疗的耐药性。为了鉴定参与分化的基因，我们使用微阵列分析和二维 (2D) 差异凝胶电泳 (DIGE) 以及 MALDI-TOF-TOF 质谱法在 mRNA 和蛋白质水平上比较基因表达。使用基于寡核苷酸的微阵列识别差异表达的转录本，其靶标代表近 14000 个基因。使用蛋白质组学和/或基因组方法对基因子集进行实时 PCR，其产物显示差异表达。我们的分析确定了 46 个在 NB4 +/- ATRA 中差异表达的基因； 22 种是使用 2D-DIGE 鉴定的，24 种是使用微阵列分析鉴定的。除四个基因外，所有这些基因在分化细胞中均以较高水平表达，并且几个控制细胞结构（内部和细胞骨架）或信号转导。我们观察到，DIGE 蛋白质组分析和银染二维凝胶电泳分析揭示了两种测量方法之间的显着差异。此外，我们的数据显示蛋白质和转录水平的基因表达之间存在显着不一致。
Acute promyelocytic leukemia (APL) is characterized by a translocation t(15:17) that fuses the retinoic acid receptor gene with the promyelocytic gene, which blocks differentiation to normal granulocytes. NB4 cells, derived from human acute promyelocytic leukemia, display this genotype and phenotype. All trans-retinoic acid (ATRA) induces differentiation of NB4 cell cultures in vitro and APL in vivo, although resistance to differentiation therapy frequently develops. To identify genes involved in differentiation, we compared gene expression at the mRNA and protein levels using microarray analyses and two-dimensional (2D) difference gel electrophoresis (DIGE), plus MALDI-TOF-TOF mass spectrometry. Differentially expressed transcripts were identified using oligonucleotide-based microarrays with targets representing almost 14 000 genes. Real time PCR was performed on a subset of genes whose products were shown to be differentially expressed using proteomic and/or genomic approaches. Our analyses identified 46 genes that were differentially expressed in NB4 +/- ATRA; 22 were identified using 2D-DIGE and 24 using microarray analysis. All but four of these genes were expressed at higher levels in differentiated cells, and several controlled cell structure (internal and cytoskelatal) or signal transduction. We observed that proteome analysis with DIGE and silver-stained 2D gel electrophoresis analyses revealed significant differences between the two measurement approaches. Furthermore, our data showed significant discordance between gene expression at the protein and transcript levels.