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The Journal of Immunology, Vol 147, Issue 2 520-527, Copyright © 1991 by American Association of Immunologists
ARTICLES |
A Anisowicz, M Messineo, SW Lee and R Sager
Division of Cancer Genetics, Dana-Farber Cancer Institute, Boston, MA 02115.
Normal human foreskin fibroblasts were used to examine transcriptional induction by IL-1 and TNF-alpha of the novel cytokine gro (melanoma growth-stimulating activity). Gro mRNA was expressed at levels 100-fold above background within 45 min of exposure to either IL-1 or TNF-alpha, in growing or serum-starved cells and a similar response was shown by IL-6. In contrast, as shown previously, gro mRNA was elevated only 10- fold by serum in starved but not in growing cells, similar to fos. Thus gro expression appears to be regulated by at least two signal transduction systems: a cytokine pathway, and a growth-related pathway. Three closely related gro genes (alpha, beta, and gamma) have been described. Their proximal 5' regulatory sequences presented here show close similarity in the region to -136, which includes the NF-kappa B site at -66 to -76 in gro alpha and gro gamma, and -64 to -74 in gro beta, and sequence diversity further upstream. Transient transfection of HeLa cells with CAT constructs localized the cytokine response to a region between -84 and -65 in gro beta. Gel retardation studies with FS- 2 cells identified a cytokine-induced protein binding at the NF-kappa B site in all three gro genes as shown by competition studies with a pair of oligonucleotides representing wild-type and mutant sequences of the NF-kappa B binding site. Neither serum nor PMA induced a detectable gel shift at NF-kappa B or upstream to position -723. These results demonstrate conservation of the cytokine response element, NF-kappa B, in the three genes, consistent with the conservation of sequence in this region; and suggest that differential expression of the three gro genes may depend upon interactions with other sites located in the divergent upstream region.
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