Zuber J, Tchernitsa O I, Hinzmann B, Schmitz A C, Grips M, Hellriegel M, Sers C, Rosenthal A, Schäfer R
[1] Laboratory of Molecular Tumour Pathology, Institute of Pathology, Charité, Humboldt-University D-10117, Berlin, Germany.
Nat Genet. 2000 Feb;24(2):144-52. doi: 10.1038/72799.
An important aspect of multi-step tumorigenesis is the mutational activation of genes of the RAS family, particularly in sporadic cancers of the pancreas, colon, lung and myeloid system. RAS genes encode small GTP-binding proteins that affect gene expression in a global way by acting as major switches in signal transduction processes, coupling extracellular signals with transcription factors. Oncogenic forms of RAS are locked in their active state and transduce signals essential for transformation, angiogenesis, invasion and metastasis via downstream pathways involving the RAF/MEK/ERK cascade of cytoplasmic kinases, the small GTP-binding proteins RAC and RHO, phosphatidylinositol 3-kinase and others. We have used subtractive suppression hybridization (SSH), a PCR-based cDNA subtraction technique, to contrast differential gene expression profiles in immortalized, non-tumorigenic rat embryo fibroblasts and in HRAS- transformed cells. Sequence and expression analysis of more than 1,200 subtracted cDNA fragments revealed transcriptional stimulation or repression of 104 ESTs, 45 novel sequences and 244 known genes in HRAS- transformed cells compared with normal cells. Furthermore, we identified common and distinct targets in cells transformed by mutant HRAS, KRAS and NRAS, as well as 61 putative target genes controlled by the RAF/MEK/ERK pathway in reverted cells treated with the MEK-specific inhibitor PD 98059.
多步骤肿瘤发生的一个重要方面是RAS家族基因的突变激活，尤其是在胰腺、结肠、肺和髓系系统的散发性癌症中。RAS基因编码小GTP结合蛋白，这些蛋白通过作为信号转导过程中的主要开关，将细胞外信号与转录因子偶联，从而以全局方式影响基因表达。致癌形式的RAS被锁定在其活性状态，并通过涉及细胞质激酶RAF/MEK/ERK级联、小GTP结合蛋白RAC和RHO、磷脂酰肌醇3激酶等的下游途径转导转化、血管生成、侵袭和转移所必需的信号。我们使用了消减抑制杂交（SSH），一种基于PCR的cDNA消减技术，来对比永生化、无致瘤性的大鼠胚胎成纤维细胞和HRAS转化细胞中的差异基因表达谱。对1200多个消减cDNA片段的序列和表达分析显示，与正常细胞相比，HRAS转化细胞中有104个EST、45个新序列和244个已知基因的转录受到刺激或抑制。此外，我们在由突变型HRAS、KRAS和NRAS转化的细胞中鉴定出了共同和不同的靶点，以及在用MEK特异性抑制剂PD 98059处理的回复细胞中由RAF/MEK/ERK途径控制的61个推定靶点基因。
