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https://hdl.handle.net/2440/9108
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Type: | Journal article |
Title: | Transformation of NIH3T3 fibroblasts by the c-kit receptor tyrosine kinase; effect of receptor density and ligand-requirement |
Author: | Caruana, G. Cambarer, A. Gonda, T. Ashman, L. |
Citation: | Oncogene, 1998; 16(2):179-190 |
Publisher: | STOCKTON PRESS |
Issue Date: | 1998 |
ISSN: | 0950-9232 1476-5594 |
Abstract: | Ectopic expression of the normal murine receptor tyrosine kinase, c-Kit, in NIH3T3 cells induced many phenotypic changes characteristic of transformation including anchorage-independent growth, focus formation and tumorigenicity in nude mice. Although transformation was largely dependent on the presence of recombinant murine Steel Factor (SLF), the ligand to the c-Kit receptor, anchorage independent growth did occur at a low frequency in the absence of added factor, and this could not be inhibited by neutralising antibodies or by SLF anti-sense mRNA. Clones from factor-independent colonies in semi-solid agar displayed a narrow range of c-Kit surface protein levels (4.3-6.4 x 10(4) receptors/cell) which was relatively high compared with the pool from which they were derived. Analysis of a larger series of random clones derived from adherent cultures expressing different levels of c-Kit demonstrated a positive correlation between SLF-dependent, anchorage-independent growth and c-Kit protein and mRNA expression levels (respectively, Rs = 0.58, P < 0.01; and Rs = 0.53, P < 0.01) with consistent colony formation observed with clones having > 2.5 x 10(4) receptors/cell. Interestingly, two of the three clones expressing the highest levels of c-Kit protein and mRNA produced few or no colonies in the presence or absence of SLF. Sequential overexpression of human c-KIT in NIH3T3 cells using a dihydrofolate reductase (DHFR)-encoding vector and gene co-amplification through methotrexate selection, which resulted in pools expressing up to 1.5 x 10(5) receptors/cell, confirmed that high receptor densities resulted in a decrease in colony numbers. Thus, analysis of clonal and selected populations has indicated that an optimal level of c-Kit is required for transformation of NIH3T3 cells in the presence of SLF, and that some ligand-independent transformation occurs. |
Keywords: | 3T3 Cells Animals Mice Cell Transformation, Neoplastic Methotrexate Ligands Cell Division Proto-Oncogene Proteins c-kit |
DOI: | 10.1038/sj.onc.1201494 |
Published version: | http://dx.doi.org/10.1038/sj.onc.1201494 |
Appears in Collections: | Aurora harvest 4 Medicine publications |
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