In vivo and in vitro effects of v-fos and EJ-Ha-ras oncogene expression in murine epidermal keratinocytes.
Primary neonatal Balb/c keratinocyte (NEK) cultures grown, using 3T3 feeder cell support in high calcium, serum supplemented medium, were transfected with EJ-Ha-ras or v-fos DNA sequences and pSV2 neo. Several neo resistant clones were isolated and several established cell lines expressing the transfected gene products derived. Two of these lines, Ras 8 and Fos 1, have been examined in detail with respect to their self renewal capacity and differentiation potential in vitro and in vivo. In vitro, both lines (when compared to normal NEK) have an extended probably immortal phenotype, enhanced colony forming efficiency (a measure of in vitro self renewal capacity) and a reduction in growth factor and serum dependence. When grafted onto syngeneic recipients neither cell line is tumourigenic. Instead a histologically abnormal epithelium with no stratum corneum and with features specific to the oncogene expressed is formed. The extent of the histological atypia correlates with the in vitro alterations in cytoskeletal peptides as revealed by 2D PAGE. However despite the gross histological abnormality there is no alteration in the in vivo self renewal capacity (measured as the number of grafted cells required for epidermal reformation) between normal NEK and the Ras 8 or Fos 1 lines; in each case a minimum of 10(5) cells/1.14 cm2 is required before a full thickness epithelium forms.
Appleby
M W
MW
Department of Pathology, Cambridge University, UK.
Greenfield
I M
IM
Crook
T
T
Parkinson
E K
EK
Stanley
M A
MA
D000818:Animals; D000831:Animals, Newborn; D002460:Cell Line; D002471:Cell Transformation, Neoplastic; D015681:Fusion Proteins, gag-onc / Q000235:genetics*; D015870:Gene Expression; D011905:Genes, ras; D015603:Keratinocytes / Q000378:metabolism*; D051379:Mice; D008807:Mice, Inbred BALB C; D009857:Oncogenes; D010957:Plasmids; D011505:Protein-Tyrosine Kinases / Q000235:genetics; D014162:Transfection
D015681:Fusion Proteins, gag-onc; D011505:Protein-Tyrosine Kinases