果蝇无生态型(Ecd)基因产物的哺乳动物直系同源调节Rb-E2F相互作用,是细胞周期进程所必需的。Ecd在乳腺癌中过表达,其过表达预测ErbB2阳性肿瘤患者的生存期较短。这里,我们证明Ecd敲低(KD)在人乳腺上皮细胞(hMECs)诱导生长停滞,与Ecd敲除(KO)对小鼠胚胎成纤维细胞的影响相似。此外,对照与对照的全基因组mRNA表达分析hMECs中的ECDKD表明,下调的前40个基因中的几个是E2F靶基因。为了探讨Ecd在乳腺肿瘤发生中的作用,我们在hTERT永生化hMEC中过表达Ecd和/或突变型H-Ras。细胞周期分析显示过表达Ecd+Ras的hMECs在生长因子剥夺后在G1期显示不完全停滞,在含生长因子的培养基中细胞周期进程更快。细胞迁移分析,入侵,3-DMatrigel中的腺泡结构和非锚定依赖性生长表明,与表达载体的细胞相比,EcdRas过表达的细胞表现出更明显的转化表型,Ras或Ecd。在营养匮乏的情况下,Ecd+Ras过表达hMECs表现出更好的存活率,自噬标志物LC3在mRNA和蛋白质水平均有显著上调。重要的是,而单独表达Ecd或突变型Ras的hMEC在NOD/SCID小鼠中不形成肿瘤,Ecd+Ras过表达hMECs形成肿瘤,清楚地表明Ecd和突变体Ras之间的致癌合作。总的来说,我们证明了Ecd通过促进细胞存活在乳腺肿瘤发生发展中的重要共癌作用。
The mammalian ortholog of Drosophila ecdysoneless (Ecd) gene product regulates Rb-E2F interaction and is required for cell cycle progression. Ecd is overexpressed in breast cancer and its overexpression predicts shorter survival in patients with ErbB2-positive tumors. Here, we demonstrate Ecd knock down (KD) in human mammary epithelial cells (hMECs) induces growth arrest, similar to the impact of Ecd Knock out (KO) in mouse embryonic fibroblasts. Furthermore, whole-genome mRNA expression analysis of control vs. Ecd KD in hMECs demonstrated that several of the top 40 genes that were down-regulated were E2F target genes. To address the role of Ecd in mammary oncogenesis, we overexpressed Ecd and/or mutant H-Ras in hTERT-immortalized hMECs. Cell cycle analyses revealed hMECs overexpressing Ecd+Ras showed incomplete arrest in G1 phase upon growth factor deprivation, and more rapid cell cycle progression in growth factor-containing medium. Analyses of cell migration, invasion, acinar structures in 3-D Matrigel and anchorage-independent growth demonstrated that Ecd+Ras-overexpressing cells exhibit substantially more dramatic transformed phenotype as compared to cells expressing vector, Ras or Ecd. Under conditions of nutrient deprivation, Ecd+Ras-overexpressing hMECs exhibited better survival, with substantial upregulation of the autophagy marker LC3 both at the mRNA and protein levels. Significantly, while hMECs expressing Ecd or mutant Ras alone did not form tumors in NOD/SCID mice, Ecd+Ras-overexpressing hMECs formed tumors, clearly demonstrating oncogenic cooperation between Ecd and mutant Ras. Collectively, we demonstrate an important co-oncogenic role of Ecd in the progression of mammary oncogenesis through promoting cell survival.