Persistence of Ha-ras-induced metastatic potential of SP1 mouse mammary tumors despite loss of the Ha-ras shuttle vector.

Abstract

Previous studies have shown that the SP1 mouse mammary adenocarcinoma cell line, which is tumorigenic but nonmetastatic, acquires metastatic potential when transfected with the activated human Ha-ras gene. In addition, the process of calcium phosphate-mediated DNA transfection, as well as treatment with the calcium ionophore A23187 or with phorbol 12-myristate 13-acetate, can also result in heritable changes in the malignant behavior of SP1 cells. It was of interest, therefore, to determine whether the metastatic consequences of Ha-ras oncogene expression in SP1 cells are a primary effect of the transfected gene or whether heritable secondary changes are induced by Ha-ras oncogene expression. In the latter case, continued expression of the Ha-ras oncogene would not be required to maintain the metastatic phenotype. To test this hypothesis we introduced the Ha-ras oncogene into SP1 cells on a shuttle vector in which maintenance of the vector was dependent on selection for resistance to the antibiotic G418. Subclones which had lost the transfected Ha-ras gene were subsequently isolated following growth in nonselective medium. The Ha-ras-transfected clones and the revertant subclones were found to be equally metastatic, indicating that transfection with the Ha-ras gene does induce stable secondary changes in the metastatic phenotype of SP1 cells.Although there is a strong correlation between ras gene activation and oncogenesis (1, 2), it is not clear how this oncogene exerts its effect on the malignant phenotype. ras genes have been implicated in the initial stages of tumorigenesis as well as later stages of tumor progression, specifically metastasis. An activated Ha-ras gene can confer both tumorigenic and metastatic properties on NIH 3T3 cells (3)(4)(5)(6)(7)(8) or diploid rat embryo fibroblasts (9). Metastatic potential of many tumor cell lines is also enhanced by transfection of an activated Ha-rds oncogene (10-13). Expression of the rasencoded p21 protein is directly involved in normal cell proliferation (14), and morphological transformation by ras, and several other oncogenes, is dependent on ras gene expression (15). Many tumor cell lines, however, show no inhibition of growth when injected with anti-Ras antibodies, suggesting that additional genetic alterations render them independent of ras action (16). Evidence of "hit-and-run" tumorigenesis (17)(18)(19)(20) also suggests that secondary changes in the cellular genome are responsible for malignant transformation. Specific chromosome rearrangements are associated with Ha-ras oncogene-transformed CHEF (Chinese hamster embryo fibroblasts) cells as well as the "hit-andrun" foci recovered, following transfection with plasmid DNA alone (21). Oncogenes may, therefore, act by accelerating the frequency of genetic (or epigenetic) changes.Previous studies showed that the Ha-ras oncogene can induce metastasis of SP1 cells, which are normally tumorigenic but nonmetastatic (11), and that calcium phosphatemediated DNA transfection of SP1 cells results in the rec...

Loss of active MEK1-ERK1/2 restores epithelial phenotype and morphogenesis in transdifferentiated MDCK cells

“…To address this issue, we set out to isolate subpopulations of C7caMEK1 cells having lost constitutively active MEK1. For this purpose, starting at passage 11, we omitted hygromycin B from culture media, thereby relieving the selection pressure for caMEK1-expressing cells (22). After nine passages, small islands of epithelia-like cells were detected among transdifferentiated, fibroblastlike C7caMEK1 cells (Fig.…”

Expression of c‐myc and c‐ras oncogenes in the neoplastic and non‐neoplastic cells of Hodgkin's disease

“…It should be noted that normal cells have special checkpoint mechanisms that recognize DNA damage and, depending on the type of the damage, act to delay the cell cycle progress at either the G1 to S, or G2 to M transition, until the relevant damage is repaired. The tumor suppressor gene p53 is considered to be an important element in these checkpoints (45), whereas activated ras and some other oncogenes seem to abrogate these checkpoints and allow the progress of the cell cycle without repairing the DNA lesions (46). Therefore, continuous high expression of c-ras in the apparently normal cells of the Hodgkin's disease lymph nodes may lead to instability of their DNA by impairing their checkpoint mechanisms.…”

APOPTOSIS EN CÉLULAS MAMARIAS TRANSFECTADAS CON EL ONCOGEN ras

“…Estas células son estimula-363 REV CHIL OBSTET GINECOL 2007; 72 (6) das a proliferar mediante el factor de crecimiento epidérmico (2).…”