Up-modulation of the expression of functional keratinocyte growth factor receptors induced by high cell density in the human keratinocyte HaCaT cell line.
Keratinocyte growth factor (KGF) is involved in the control of proliferation and differentiation of human keratinocytes. It binds to, and activates, the tyrosine kinase KGF receptor (KGFR), a splicing transcript variant of the fibroblast growth factor receptor 2. We have previously shown (C. Marchese et al., Cell Growth Differ., 8: 989-997, 1997) that differentiation of primary cultured keratinocytes triggered by high Ca2+ concentrations in the growing medium induced up-regulation of KGFR expression, which suggested that KGFR may play a crucial role in the control of the proliferative/differentiative program during transition from basal to suprabasal cells. Here we analyzed the process of modulation of the expression of KGFRs in the human keratinocyte cell line HaCaT, widely used as a model to study keratinocyte differentiation. Western blot and double immunofluorescence for KGFR and the K1 differentiation marker showed that cell differentiation and stratification induced by confluence and high cell density correlated with an increase in KGFR expression. KGFRs, present on suprabasal differentiated cells, appeared to be efficiently tyrosine-phosphorylated by KGF, which indicated that the receptors up-regulated by differentiation can be functionally activated by ligand binding. Bromodeoxyuridine incorporation assay revealed that a significant portion of suprabasal differentiated cells expressing KGFR seemed to be still able to synthesize DNA and to proliferate in response to KGF, which suggested that increased KGFR expression may be required for retention of the proliferative activity.
Capone
A
A
Dipartimento di Medicina Sperimentale e Patologia, Università di Roma La Sapienza, Italy.
Visco
V
V
Belleudi
F
F
Marchese
C
C
Cardinali
G
G
Bellocci
M
M
Picardo
M
M
Frati
L
L
Torrisi
M R
MR
D015153:Blotting, Western; D001973:Bromodeoxyuridine / Q000378:metabolism; D002452:Cell Count; D002454:Cell Differentiation / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D051526:Fibroblast Growth Factor 10; D051523:Fibroblast Growth Factor 7; D005346:Fibroblast Growth Factors; D005455:Fluorescent Antibody Technique; D006133:Growth Substances / Q000494:pharmacology; D006801:Humans; D015603:Keratinocytes / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D008854:Microscopy, Electron; D010766:Phosphorylation / Q000187:drug effects; D019000:Phosphotyrosine / Q000378:metabolism; D051497:Receptor, Fibroblast Growth Factor, Type 2; D017468:Receptors, Fibroblast Growth Factor; D017978:Receptors, Growth Factor / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins; D015854:Up-Regulation / Q000187:drug effects
C496453:FGF7 protein, human; D051526:Fibroblast Growth Factor 10; D006133:Growth Substances; D017468:Receptors, Fibroblast Growth Factor; D017978:Receptors, Growth Factor; D011993:Recombinant Fusion Proteins; D051523:Fibroblast Growth Factor 7; D019000:Phosphotyrosine; D005346:Fibroblast Growth Factors; D051497:Receptor, Fibroblast Growth Factor, Type 2; C065104:keratinocyte growth factor receptor; D001973:Bromodeoxyuridine