Retinoblastoma-related protein pRb2/p130 and suppression of tumor growth in vivo.
BACKGROUND
The RB/p105 and p107 genes of the retinoblastoma family are tumor suppressor genes whose proteins are inactivated by interaction with T-antigen proteins encoded by polyomaviruses (e.g., simian virus 40 and human JC virus), which have been found to be highly tumorigenic in animals. A variety of indirect evidence suggests that another member of the retinoblastoma gene family, RB2/p130, is also a tumor suppressor gene. To investigate the putative tumor suppressor activity of RB2/p130 more directly, we utilized a tetracycline-regulated gene expression system to control expression of the encoded protein pRb2/p130 in JC virus-induced hamster brain tumor cells and to study the effects of pRb2/p130 on the growth of such tumor cells in nude mice. The ability of pRb2/p130 to interact with JC virus T antigen was also studied.


METHODS
Northern blot hybridization analyses were performed on samples of total cellular RNA to measure RB2/p130 and beta-actin messenger RNA levels. Immunoprecipitation and western blot analyses were used to determine T-antigen and pRb2/p130 protein levels and to assess the phosphorylation status of these proteins. Tumor cells were injected subcutaneously into nude mice, and tumor growth, with or without induced expression of pRb2/p130, was monitored.


RESULTS
Induction of pRb2/p130 expression brought about a 3.2-fold, or 69% (95% confidence interval = 64%-73%), reduction in final tumor mass in nude mice. We also demonstrated that JC virus T antigen binds hypophosphorylated pRb2/p130 and that stimulation of pRb2/p130 expression overcomes cellular transformation mediated by this antigen.


CONCLUSIONS
Our findings support the hypothesis that RB2/p130 is a tumor suppressor gene.
Department of Pathology, Anatomy, and Cell Biology, Jefferson Medical College, and Sbarro Institute for Cancer Research and Molecular Medicine, Philadelphia, PA 19102, USA.
C M
CM
Howard
P P
PP
Claudio
G L
GL
Gallia
J
J
Gordon
G G
GG
Giordano
W W
WW
Hauck
K
K
Khalili
A
A
Giordano
D000818:Animals; D000957:Antigens, Viral, Tumor / Q000096:biosynthesis / Q000378:metabolism*; D015152:Blotting, Northern; D015153:Blotting, Western; D001932:Brain Neoplasms / Q000235:genetics* / Q000821:virology*; D006224:Cricetinae; D004273:DNA, Neoplasm / Q000032:analysis; D004195:Disease Models, Animal; D005434:Flow Cytometry; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects*; D015967:Gene Expression Regulation, Viral / Q000187:drug effects; D016161:Genes, Retinoblastoma / Q000187:drug effects*; D016147:Genes, Tumor Suppressor / Q000187:drug effects*; D006801:Humans; D007577:JC Virus / Q000276:immunology; D051379:Mice; D008819:Mice, Nude; D010750:Phosphoproteins / Q000096:biosynthesis* / Q000494:pharmacology*; D011233:Precipitin Tests; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D011506:Proteins; D050717:Retinoblastoma-Like Protein p130; D013752:Tetracycline / Q000494:pharmacology; D014407:Tumor Cells, Cultured; D014410:Tumor Stem Cell Assay
D000957:Antigens, Viral, Tumor; D004273:DNA, Neoplasm; D010750:Phosphoproteins; D011500:Protein Synthesis Inhibitors; D011506:Proteins; C493837:RBL2 protein, human; C493838:Rbl2 protein, mouse; D050717:Retinoblastoma-Like Protein p130; D013752:Tetracycline