title: The v-ras oncogene inhibits the expression of differentiation markers and facilitates expression of cytokeratins 8 and 18 in mouse keratinocytes.
abstract: Cultured mouse keratinocytes can be initiated in vitro by the introduction of a v-rasHa gene by viral transduction. Previous studies indicated that v-rasHa-transduced keratinocytes have a high proliferation rate in medium with 0.05 mM Ca2+ and resist terminal differentiation in medium with greater than 0.1 mM Ca2+, a culture condition in which normal cells mature into squames. The current studies demonstrate that v-rasHa keratinocytes do not express transcripts or protein for epidermal early differentiation markers keratins 1 and 10 when cells are challenged with 0.12 mM Ca2+, which is a signal for expression of these genes in normal cells. Both transcript and protein for the late differentiation marker loricrin are also diminished in v-ras keratinocytes, but filaggrin, also a late differentiation-related gene product, is expressed in nearly normal amounts but at a different Ca2+ optimum. Modification of intracellular Ca2+ with ionomycin failed to restore the expression of any suprabasal keratinocyte markers. In contrast to the effects on normal products of keratinocyte differentiation, the introduction of the v-rasHa gene facilitated the expression of keratins 8 (K8) and 18 (K18). These keratins are characteristic of embryonic cells and cells of simple adult epithelia but not stratified squamous epithelia such as skin. Like normal differentiation markers, the expression of K8 and K18 was dependent both on the v-ras oncogene and the Ca2+ concentration of the culture medium, with greater than 0.1 mM Ca2+ being optimal. At the optimal Ca2+ level, the majority of v-ras keratinocytes expressed K8 and K18 after 96 h, and many cells had reduced amounts of the normal keratinocyte cytokeratin K14. These studies indicate that the v-ras gene causes substantial reprogramming of epidermal physiology, producing an unusual phenotype devoid of early suprabasal markers but at least partially permissive for late marker expression. Furthermore, the Ca2(+)-dependent expression of K8 and K18 suggests that a normal signalling pathway used in keratinocyte differentiation is diverted to an abnormal endpoint.
authors.affiliation: Laboratory of Cellular Carcinogenesis and Tumor Promotion, National Cancer Institute, Bethesda, Maryland 20892.
authors.forename: C
authors.initials: C
authors.lastname: Cheng
authors.forename: A E
authors.initials: AE
authors.lastname: Kilkenny
authors.forename: D
authors.initials: D
authors.lastname: Roop
authors.forename: S H
authors.initials: SH
authors.lastname: Yuspa
mesh_terms: D000818:Animals; D015152:Blotting, Northern; D015153:Blotting, Western; D002118:Calcium / Q000502:physiology; D002454:Cell Differentiation; D000091344:Filaggrin Proteins; D005455:Fluorescent Antibody Technique; D011905:Genes, ras; D066298:In Vitro Techniques; D007381:Intermediate Filament Proteins / Q000378:metabolism; D015603:Keratinocytes / Q000166:cytology / Q000378:metabolism*; D007633:Keratins / Q000235:genetics / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D013997:Time Factors
chemical_list: D000091344:Filaggrin Proteins; D007381:Intermediate Filament Proteins; D008565:Membrane Proteins; C064554:loricrin; D007633:Keratins; D002118:Calcium