[Growth of stable clones of mouse fibroblast cell line C3H10T1/2 expressing the human retinoblastoma gene product].
Phosphorylation and dephosphorylation of the retinoblastoma gene product (pRB) are recognized as necessary events in the cell cycle progression. To study the role of pRB in regulation of cell proliferation, the stable cell lines with constitutive expression of the exogenous RB gene can be employed. In order to obtain such cell lines in this work C3H10T1/2 mouse fibroblasts were infected with defective retrovirus encompassing the RB and Neo gene conferring resistance to geniticine (G418). The pRB production and its phosphorylation pattern were analyzed by immunoblotting in cell lysates considering well known data on correlation between pRB phosphorylation pattern and its electrophoretic mobility. Cell lines subjected to G418 selection with the following cloning procedure were identical to the control cells expressing beta-galactosidase, when compared for pRB production and phosphorylation in the cell cycle stages characterized by hyperphosphorylated pRB. However, cells of the experimental cell lines hypophosphorylated pRB much faster and accumulated much more underphosphorylated protein compared to the control cell lines. The doubling time of the cells was not affected either by changes in the pRB phosphorylation pattern or by its overproduction during separate cell cycle stages. These results suggest that maintaining of the physiological level of pRB phosphorylation in cycling cells is strictly controlled and is considered to be a more important condition of the cell cycle progression than pRB dephosphorylation.
Popov
B V
BV
Institute of Cytology, Russian Academy of Sciences, St. Petersburg.
Kulakova
I A
IA
Popov
N B
NB
Bondar'
T O
TO
Frenkel'
Z M
ZM
Nikolaev
A V
AV
D016475:3T3 Cells; D000818:Animals; D002453:Cell Cycle / Q000502:physiology; D002455:Cell Division / Q000502:physiology; D002999:Clone Cells / Q000166:cytology / Q000378:metabolism*; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism; D015972:Gene Expression Regulation, Neoplastic / Q000502:physiology*; D016161:Genes, Retinoblastoma; D006801:Humans; D051379:Mice; D010766:Phosphorylation
Izuchenie rosta stabil'nykh klonov myshinykh fibroblastov linii C3H10T1/2, ékspressiruiushchikh produkt gena retinoblastomy cheloveka]