Expression of antisense osteopontin RNA inhibits tumor promoter-induced neoplastic transformation of mouse JB6 epidermal cells.
Elevated expression of osteopontin (OPN), a secreted adhesive phosphoglycoprotein, is frequently associated with many transformed cell lines of epithelial and stromal origin. Moreover, several clonal lines of preneoplastic JB6 cells derived from Balb/c mouse epidermal cultures (Colburn et al., 1978, 1979), upon treatment with 12-O-tetradecanoyl phorbol-13-acetate (TPA), become irreversibly oncogenic and concomitantly synthesize OPN at elevated levels (Smith and Denhardt, 1989). In the present study we sought to determine whether OPN expression facilitates transformation of such preneoplastic (initiated) cells. We transfected TPA-promotable JB6 c141.5a cells with an expression vector containing mouse OPN cDNA in antisense orientation under transcriptional control of dexamethasone-inducible MMTV-LTR promoter. Four stably transfected clones, which expressed drastically reduced levels of OPN in the presence of both dexamethasone and TPA, were characterized. We found that (a) more than 20 copies of OPN antisense cDNA were stably incorporated into the genome of cells from two of these clones that were examined by Southern blot analysis; (b) dexamethasone-induced expression of antisense OPN RNA prevented augmented OPN expression at both mRNA and protein levels following TPA treatment; and (c) cells from all four clones failed to form colonies in soft agar medium containing both dexamethasone and TPA. Taken together, these data demonstrate that inhibition of elevated OPN expression blocks TPA-induced anchorage-independent growth of JB6 c141.5a cells, suggesting the possibility that OPN overproduction is causally related to transformation of preneoplastic cells.
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Department of Biology, McGill University, Montreal, Quebec, Canada.
Mukherjee
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Mukherjee
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D000818:Animals; D002273:Carcinogens / Q000633:toxicity*; D002461:Cell Line, Transformed; D002471:Cell Transformation, Neoplastic / Q000139:chemically induced / Q000235:genetics*; D018076:DNA, Complementary; D003907:Dexamethasone / Q000494:pharmacology; D008324:Mammary Tumor Virus, Mouse / Q000235:genetics; D051379:Mice; D008807:Mice, Inbred BALB C; D053495:Osteopontin; D010750:Phosphoproteins / Q000235:genetics*; D016372:RNA, Antisense / Q000235:genetics*; D012795:Sialoglycoproteins / Q000235:genetics*; D013755:Tetradecanoylphorbol Acetate / Q000633:toxicity*
D002273:Carcinogens; D018076:DNA, Complementary; D010750:Phosphoproteins; D016372:RNA, Antisense; D012795:Sialoglycoproteins; C506896:Spp1 protein, mouse; D053495:Osteopontin; D003907:Dexamethasone; D013755:Tetradecanoylphorbol Acetate