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Confocal Laser Scanning Microscopy of Oncogene Localization in Rainbow Trout Cell Lines Derived from Normal and Tumor Tissue*¹

Laboratory of Experimental Pathology, P.O. Box 12233, National Institute of Environmental Health Sciences, Research Triangle Park, North Carolina 27709

William W. Ellington

Duke University School of the Environment, Marine Laboratory, Beaufort, North Carolina 28516

Rebecca J. Van Beneden

Duke University School of the Environment, Marine Laboratory, Beaufort, North Carolina 28516, Duke University Medical Center, Cell Biology Department, Durham North Carolina 27710

We examined the localization and expression of the nuclear oncoprotein c-myc and the cytoplasmic membrane-associated oncoprotein c-ras in rainbow trout cell lines derived from both normal and tumor tissue in order to question whether c-myc and ras oncoprotein immunostaining was increased in cells derived from tumors compared to cells derived from normal tissue. Cell lines examined were derived from normal rainbow trout gonadal cells (RTG-2), a rainbow trout hepatoma (RTH-149), and a rainbow trout mesothelioma (RTM). Protein products of c-ras and c-myc were visualized in these 3 cell lines by employing fluorescein-labeled anti-mouse pan-ras and c-myc antibodies. The RTG-2 cells were used in this study as normal, control cells, and they exhibited little pan-ras and c-myc staining. The RTH-149 cell line (a tumorigenic cell line) exhibited positive pan-ras staining in regions of the membrane and cell cytoplasm. Localization of c-myc staining to perinuclear regions was punctate in RTH-149 cells. RTM cells (also a tumorigenic cell line) displayed a ras staining localization similar to the pattern seen in RTH-149 cells. RTM cells exhibit a diffuse perinuclear staining and, thus, display a more ubiquitous localization of c-myc than RTH-149 cells. Northern blot analysis indicated that c-myc expression was highest in RTM cells, whereas RTG-2 cells and RTH-149 cells expressed similar lower levels of c-myc expression. We were unable to detect significant ras expression in any of the cell lines by Northern blot analysis. In summary, the cell line derived from normal tissue, the RTG-2 cells, displayed little ras and c-myc immunostaining, whereas the cell lines derived from tumorigenic tissue, RTH and RTM cells, displayed increased immunostaining for c-myc and ras proteins.

Key Words: Fish cell lines • oncogene expression • c-myc antibody • pan-ras antibody

Toxicologic Pathology, Vol. 24, No. 3, 339-345 (1996)
DOI: 10.1177/019262339602400310


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