Identification and cloning of genes displaying elevated expression as a consequence of metastatic progression in human melanoma cells by rapid subtraction hybridization
Identification and cloning of genes displaying elevated expression as a consequence of metastatic progression in human melanoma cells by rapid subtraction hybridization
复制标题
DOI：
10.1016/j.gene.2004.09.002
复制
发表时间：
2004-12-08
期刊：
影响因子：
3.5
通讯作者：
Fisher, PB
中科院分区：
文献类型：
--
作者：
Boukerche, H;Su, ZZ;Fisher, PB
虽然广泛的研究，完整的基因库相关和转移的原因仍然在很大程度上是未知的。我们开发了一种有效的方法，用于识别差异表达的基因，涉及快速消减杂交（RaSH）的cDNA克隆制备从两个细胞群，司机和测试。这种RaSH方法先前已经证明了在鉴定作为人黑素瘤细胞中诱导终末分化的函数而差异表达的基因方面的高灵敏度和有效性，人T细胞对人免疫缺陷病毒-1（HIV-1）感染的抗性或敏感性，以及感染HIV-1或用HIV-1治疗的正常人胎儿星形胶质细胞中基因表达的扰动，1 gp 120病毒包膜糖蛋白或肿瘤坏死因子-α（TNF-α）。在本研究中，RaSH已应用于转移性黑素瘤模型，其模拟人类中的早期转移事件，包括弱转移性与免疫抑制的新生大鼠选择的高转移性变体。这现在已经导致鉴定出在高转移性变体中相对于正常永生黑素细胞或弱转移性亲本克隆显示升高表达的八个基因。这些基因包括67-kDa层粘连蛋白受体（671-R）、内皮素受体B（ENDR B）、Na+/K+-ATP酶、Ku抗原、白细胞介素受体相关激酶-1（IRAK-1）和核糖体蛋白RPLA，它们可能参与黑色素瘤转移的复杂过程。此外，还鉴定了两种可能影响转移表型的未知基因（在当前数据库中未报道）。这些研究提供了使用RaSH方法的额外支持，在本申请中，在具有不同转移潜能的密切相关的变异细胞系的背景下，用于有效的差异基因鉴定，并阐明了8个先前未被识别的基因，其在黑色素瘤进展至转移能力中的作用现在可以被仔细检查。(C)2004 Elsevier B. V.保留所有权利。
Although extensively investigated, the complete repertoire of genes associated with and causative of metastasis remain largely unknown. We developed an efficient approach for identifying differentially expressed genes that involves rapid subtraction hybridization (RaSH) of cDNA clones prepared from two cell populations, a driver and a tester. This RaSH approach has previously documented high sensitivity and effectiveness in identifying genes that are differentially expressed as a function of induction of terminal differentiation in human melanoma cells, resistance or sensitivity to human immunodeficiency virus-1 (HIV-1) infection of human T cells and perturbation in gene expression in normal human fetal astrocytes infected with HIV-1 or treated with HIV-1 gp120 viral envelope glycoprotein or tumor necrosis factor-alpha (TNF-alpha). In the present study, RaSH has been applied to a metastatic melanoma model, which mimics the early events of metastasis in humans, comprising weakly metastatic vs. immunosuppressed newborn rat-selected highly metastatic variants. This has now resulted in the identification of eight genes displaying elevated expression in the high metastatic variants vs. normal immortal melanocytes or weakly metastatic parental clones. These include six known genes, 67-kDa laminin receptor (671-R), endothelin receptor B (ENDRB), Na+/K+-ATPase, Ku antigen, interleukin-receptor-associated kinase-1 (IRAK-1) and ribosomal protein RPLA, which may contribute to the complex process of melanoma metastasis. Additionally, two unknown genes (not reported in current databases) that may also impact on the metastatic phenotype have also been identified. These studies provide additional support of the use of the RaSH approach, in this application in the context of closely related variant cell lines with different metastatic potential, for effective differential gene identification and elucidate eight previously unrecognized genes whose role in melanoma progression to metastatic competence can now be scrutinized. (C) 2004 Elsevier B.V. All rights reserved.