ABSTRACT
The rat pheochromocytoma cell line (PC-12) offers a powerful in vitro model to study the mechanism of growth factor-induced differentiation and proliferation. Within minutes of addition, agents such as nerve growth factor (NGF), epidermal growth factor (EGF), basic fibroblast growth factor (bFGF) and dibutyryl cyclic AMP (db cAMP) rapidly activate cellular immediate early genes such as c-fos, c-jun, jun-B, and egr-1. fra-1, a member of the immediate early gene family, follows a distinctly later time course of induction than c-fos, c-jun, jun-B, and egr-1, suggesting that fra-1 may attenuate the action of genes induced earlier. We demonstrate that constitutive expression of fra-1 in PC-12 cells results in pronounced inhibition of NGF-induced differentiation. Transcriptional activation of c-fos, c-jun, jun-B, and egr-1 by NGF, EGF, and db cAMP was down-regulated to a varying extent whereas NGF-induced ornithine decarboxylase (ODC) was not affected. Expression of jun-D was not affected in PC-12 fra-1 cells. Transfection of fos and egr-1 promoter-chloramphenicol acetyl transferase (CAT) plasmid into these stable fra-1-expressing PC-12 cells revealed that repression of fos and egr-1 was exerted at the promoter level. Thus deregulated fra-1 expression may inhibit PC-12 cell differentiation by altering the patterns of immediate early gene expression.
Subject(s)
Adrenal Gland Neoplasms/pathology , Cell Transformation, Neoplastic/pathology , Gene Expression Regulation, Neoplastic/drug effects , Immediate-Early Proteins , Pheochromocytoma/pathology , Promoter Regions, Genetic/genetics , Adrenal Gland Neoplasms/genetics , Adrenal Gland Neoplasms/metabolism , Animals , Bucladesine/pharmacology , Cell Differentiation , Cell Transformation, Neoplastic/drug effects , Cell Transformation, Neoplastic/genetics , Cells, Cultured , DNA-Binding Proteins/genetics , DNA-Binding Proteins/metabolism , Down-Regulation/drug effects , Down-Regulation/genetics , Early Growth Response Protein 1 , Epidermal Growth Factor/pharmacology , Nerve Growth Factors/pharmacology , Pheochromocytoma/genetics , Pheochromocytoma/metabolism , Promoter Regions, Genetic/physiology , Proto-Oncogene Proteins/genetics , Proto-Oncogene Proteins/metabolism , Proto-Oncogene Proteins c-fos , Proto-Oncogene Proteins c-jun , Rats , Signal Transduction/drug effects , Signal Transduction/physiology , Transcription Factors/genetics , Transcription Factors/metabolism , Transcription, Genetic/drug effects , Transfection , Tumor Cells, Cultured/drug effects , Tumor Cells, Cultured/pathologyABSTRACT
A spontaneously immortalized, nontumorigenic mouse mammary epithelial cell line (MMEC) was transfected with an activated myc construct by electroporation. Constitutive expression of myc in MMEC resulted in anchorage independence in soft agar and tumorigenicity in nude mice. The myc-expressing MMEC showed higher saturation density, faster growth rate, and partial abrogation of serum-derived growth factor(s) requirement compared with parent MMEC. Epidermal growth factor or transforming growth factor alpha stimulated the anchorage-independent growth, but not the anchorage-dependent growth, of MMEC-myc cells. Type 1 transforming growth factor beta, on the other hand, inhibited both the anchorage-independent and anchorage-dependent growth of MMEC-myc cells. These results demonstrate that deregulated expression of myc results in neoplastic transformation iin mammary epithelial cells. Accompanying the transformation is altered sensitivity to polypeptide growth factors.
