Endocrinology, Vol 136, 2889-2895, Copyright © 1995 by Endocrine Society

ARTICLES

Regulation of follistatin messenger ribonucleic acid in steroidogenic rat granulosa cell lines

L Shukovski, I Keren-Tal, A Dantes and A Amsterdam
Department of Hormone Research, Weizmann Institute of Science, Rehovot, Israel.

Follistatin is a 35-kilodalton monomer isolated from follicular fluid that acts on pituitary gonadotropes to suppress the production of FSH. Transfection of rat granulosa cells with specific oncogenes, such as simian virus 40 (SV40) DNA and Ha-ras oncogene, leads to their immortalization concomitant with preservation of their capacity for inducible steroidogenesis. Experiments were designed to investigate the regulation of follistatin messenger RNA (mRNA) accumulation upon stimulation with forskolin, 2-O-tetradecanol-phorbol-13-acetate (TPA), FSH, and hCG in four different granulosa cell lines. Granulosa cells were transfected with SV40 DNA alone (POGS5); with SV40 DNA and Ha-ras oncogene (POGRS1); with SV40 DNA, Ha-ras oncogene, and LH/CG receptor (GLHR15); or with FSH receptor (GFSHR17) expression plasmid. Cells were cultured to reach confluence and then stimulated for 6 or 24 h with ovine FSH (oFSH; 0.004-4 nM), hCG (9 nM), forskolin (50 microM), and TPA (50 nM), alone or in combination. In the POGS5 cell line, forskolin caused a 15-fold stimulation of follistatin mRNA after 24-h incubation. The POGRS1 cell line showed a time-dependent stimulation of follistatin gene expression induced by both forskolin (5.7-fold) and TPA (9.4- fold). In the GFSHR17 cells, forskolin, oFSH, and TPA induced an increase in follistatin mRNA. When oFSH (1.6 nM) was added to cells treated with forskolin (50 microM) or TPA (50 nM), no additional stimulation was observed. The GLHR15 cell line treated with hCG showed a 2.7-fold increase in follistatin mRNA accumulation within 6 h. Our data demonstrate that 1) follistatin mRNA is detectable and induced by forskolin and TPA in transformed granulosa cell lines that do not express the FSH or LH receptors; 2) in the GFSHR17 cell line, FSH, forskolin, and TPA caused a time- and dose-dependent regulation of the gene; and 3) follistatin gene expression is up-regulated by hCG in the GLHR15 cell line. We conclude that these transformed steroidogenic cell lines can serve as a useful model to study the regulation of follistatin gene expression, a peptide known to regulate pituitary and ovarian hormone secretion and differentiation of granulosa cells by its activin-binding action. Moreover, this gene can be regulated in immortalized granulosa cells by both the protein kinase A and protein kinase C pathways, although these cells express the large T antigen and the Ha-ras oncogenic proteins.

This article has been cited by other articles:

				R. SASSON, A. DANTES, K. TAJIMA, and A. AMSTERDAM Novel genes modulated by FSH in normal and immortalized FSH-responsive cells: new insights into the mechanism of FSH action FASEB J, July 1, 2003; 17(10): 1256 - 1266. [Abstract] [Full Text] [PDF]

				J. Grosse, A. Bulling, C. Brucker, U. Berg, A. Amsterdam, A. Mayerhofer, and M. Gratzl Synaptosome-Associated Protein of 25 Kilodaltons in Oocytes and Steroid-Producing Cells of Rat and Human Ovary: Molecular Analysis and Regulation by Gonadotropins Biol Reprod, August 1, 2000; 63(2): 643 - 650. [Abstract] [Full Text]

				S. J. Winters, A. C. Dalkin, and T. Tsujii Evidence That Pituitary Adenylate Cyclase Activating Polypeptide Suppresses Follicle-Stimulating Hormone-{beta} Messenger Ribonucleic Acid Levels by Stimulating Follistatin Gene Transcription Endocrinology, October 1, 1997; 138(10): 4324 - 4329. [Abstract] [Full Text] [PDF]

				A. Amsterdam and N. Selvaraj Control of Differentiation, Transformation, and Apoptosis in Granulosa Cells by Oncogenes, Oncoviruses, and Tumor Suppressor Genes Endocr. Rev., August 1, 1997; 18(4): 435 - 461. [Abstract] [Full Text] [PDF]