Possible Role for Ha-ras Expression in Inducible Steroidogenesis in Immortalized Granulosa Cell Lines
Primary granulosa cells cotransfected with SV40 DNA and the Ha-ras oncogene can be induced to produce progestins (progesterone and 20α-dihydroprogesterone) when incubated with 8-Br-cyclic AMP and substances elevating intracellular cyclic AMP (cAMP) such as forskolin, choleratoxin and the Bordetella pertussis invasive adenylate cyclase (BPAC). In contrast, cells transfected with SV40 DNA alone show only traces of steroidogenic activity under similar stimulation. The steroidogenic capacity of the cotransfected lines was correlated with the epithelioid appearance of the cells and low expression of actin and actin binding proteins in these cells. Expression of isoforms 2 and 3 of tropomyosins which possess high affinity for actin filaments was extremely low in these cells compared to cells transfected with SV40 DNA alone. Expression of p21 in cotransfected individual lines was correlated to the steroidogenic capacity. Primary granulosa cells and luteinized cells also express modestly but significantly p21 precipitable by monoclonal antibodies against the proto/mutated oncogene product. The cotransfected cells were highly tumorigenic when injected to nude mice but pretreatment of the cells with BPAC, which resulted in prolonged intracellular accumulation of cAMP, prevented metastatic spread of the tumor cells. Therefore, high levels of intracellular cAMP may arrest proliferation of the transformed cells both in vivo and in vitro. It is suggested that the expression of the Ha-ras oncogene may be involved in inducible steroidogenesis in immortalized granulosa cell lines, while the product of the protooncogene may be implicated in this process in normal cells.
KeywordsGranulosa Cell Granulosa Cell Tumor Steroidogenic Enzyme Preantral Follicle Progesterone Production
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- 2.R. A. Weinberg, Oncogenes, antioncogenes, and the molecular basis of multistep carcinogenesis, Cancer Res. 49:3714(1989).Google Scholar
- 32.A. Amsterdam, S. Rotmensch, A. Furman, E. A. Venter, and I. Vlodavsky, Synergistic effect of human chorionic gonadotropin and extracellular matrix on in vitro differentiation of human granulosa cells: progesterone production and gap junction formation, Endocrinology 124:1956 (1989).PubMedCrossRefGoogle Scholar
- 40.D. Michalovitz, A. Amsterdam, and M. Oren, Interactions between SV40 and cellular oncogenes in the transformation of primary rat cells, in: “Current Topics in Microbiology and Immunology,” ed., Springer-Verlag, (1989).Google Scholar
- 42.I. Hanukoglu, Molecular biology of cytochrome P450 systems in steroidogenic tissues, in: “Follicular Development and the Ovulatory Response,” A. Tsafriri, and N. Dekel, ed., Ares-Serono Symposia Review, 23:233–252 (1989).Google Scholar
- 43.A. Amsterdam, and B. S. Suh, An inducible functional peripheral benzodiazepine receptor in mitochondria of steroidogenic granulosa cells, Endocrinology in press (1991).Google Scholar
- 44.H. Rennert, A. Amsterdam, J. T. Billheimer, and J. T. Strauss, Regulated expression of sterol carrier protein2 in the ovary: A key role for cyclic AMP., Submitted (1991).Google Scholar
- 47.B. S. Suh, R. Sprengel, P. H. Seeburg, and A. Amsterdam, Functional receptors to gonadotropins in oncogene-transformed steroidogenic granulosa cells, Proceedings of the 73rd Annual Meeting of the American Endocrine Society, 1991; Abstract 1851.Google Scholar
- 49.B. S. Suh, L. Eisenbach, and A. Amsterdam, Cyclic AMP suppresses metastatic spread in nude mice induced by steroidogenic rat granulosa cells transformed by SV40 and Ha-ras oncogene(s), Proceedings of the Annual Meeting of the Israeli Endocrine Society. (1991); Abstract 74.Google Scholar