title: The cell cycle regulator ecdysoneless cooperates with H-Ras to promote oncogenic transformation of human mammary epithelial cells.
abstract: 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.
authors.affiliation: a Departments of Genetics ; Cell Biology and Anatomy ; Nebraska Medical Center , Omaha , NE USA.
authors.forename: Aditya
authors.initials: A
authors.lastname: Bele
authors.forename: Sameer
authors.initials: S
authors.lastname: Mirza
authors.forename: Ying
authors.initials: Y
authors.lastname: Zhang
authors.forename: Riyaz
authors.initials: R
authors.lastname: Ahmad Mir
authors.forename: Simon
authors.initials: S
authors.lastname: Lin
authors.forename: Jun Hyun
authors.initials: JH
authors.lastname: Kim
authors.forename: Channabasavaiah Basavaraju
authors.initials: CB
authors.lastname: Gurumurthy
authors.forename: William
authors.initials: W
authors.lastname: West
authors.forename: Fang
authors.initials: F
authors.lastname: Qiu
authors.forename: Hamid
authors.initials: H
authors.lastname: Band
authors.forename: Vimla
authors.initials: V
authors.lastname: Band
mesh_terms: D000818:Animals; D001343:Autophagy; D001943:Breast Neoplasms / Q000378:metabolism* / Q000473:pathology; D002352:Carrier Proteins / Q000502:physiology*; D002465:Cell Movement; D049109:Cell Proliferation; D002471:Cell Transformation, Neoplastic; D002478:Cells, Cultured; D005260:Female; D006801:Humans; D042361:Mammary Glands, Human / Q000473:pathology*; D016688:Mice, Inbred NOD; D016513:Mice, SCID; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D009368:Neoplasm Transplantation; D016283:Proto-Oncogene Proteins p21(ras) / Q000502:physiology*
chemical_list: D002352:Carrier Proteins; C513410:ECD protein, human; C477012:MAP1LC3A protein, human; D008869:Microtubule-Associated Proteins; C501009:HRAS protein, human; D016283:Proto-Oncogene Proteins p21(ras)
keywords: Ecd, Ecdysoneless; KD, Knockdown; KO, Knockout; MEFs, Mouse Embryonic Fibroblasts; PCR, Polymerase Chain Reaction; Rb, Retinoblastoma; autophagy; breast cancer; cell cycle; ecdysoneless; hMEC, Human Mammary Epithelial Cells; ras