Disturbances in growth control and gene expression in a C3H/10T1/2 cell line that stably overproduces protein kinase C.
We have utilized a retroviral expression vector to construct a series of C3H/10T1/2 murine fibroblast cell lines that stably overexpress a full length cDNA encoding rat protein kinase C beta 1 (PKC). These cell lines contain 3-11 fold greater PKC enzyme activity than parental cells or control cells that carry an integrated vector lacking the cDNA insert. 10T1/2-PKC-4, a line with an 11-fold increase in PKC activity, is morphologically altered, grows to 4-fold higher saturation density, and has decreased adhesiveness when compared to control cells. These cells also show constitutive and inducible alterations in the levels of two PKC-regulated genes, phorbin and TPA-R1. However, 10T1/2-PKC-4 cells do not have a transformed morphology, are incapable of growth in soft agar, and are non-tumorigenic in nude mice. When 10T1/2-PKC-4 cells are cultured in 2.5% calf serum and 100 ng ml-1 TPA, numerous large, dense foci appear 2-3 weeks after the cells reach confluence. TPA is required for focus formation, and control cells do not form foci under the same conditions. However, when such foci are isolated and grown in the absence of TPA, the cells closely resemble later passage 10T1/2-PKC-4 cells. Therefore, the events leading to focus formation appear to be reversible. The 10T1/2-PKC-4 cell line should be valuable for more precisely defining molecular events relevant to tumor promotion and multistage carcinogenesis.
Comprehensive Cancer Center, College of Physicians and Surgeons, Columbia University, New York, New York 10032.
R S
RS
Krauss
G M
GM
Housey
M D
MD
Johnson
I B
IB
Weinstein
D000818:Animals; D015152:Blotting, Northern; D002455:Cell Division; D002460:Cell Line; D005786:Gene Expression Regulation / Q000187:drug effects; D066298:In Vitro Techniques; D051379:Mice; D008819:Mice, Nude; D009374:Neoplasms, Experimental / Q000473:pathology; D011493:Protein Kinase C / Q000502:physiology*; D012333:RNA, Messenger / Q000235:genetics; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014162:Transfection
D012333:RNA, Messenger; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate