The ERK-dependent signalling is stage-specifically modulated by FSH, during primary Sertoli cell maturation Pascale Cre´ pieux* ,1 , Se´bastien Marion 1 , Nadine Martinat 1 , Ve´ronique Fafeur 3 , Yves Le Vern 2 , Dominique Kerboeuf 2 , Florian Guillou 1 and Eric Reiter 1 1 Laboratoire de Physiologie de la Reproduction et des Comportements, Institut National de la Recherche Agronomique/Centre National pour la Recherche Scientifique/Universite´ de Tours, UMR 6073, Nouzilly, France; 2 Laboratoire de Pathologie aviaire et Parasitologie, Centre de Recherches de Tours, 37380; 3 CNRS FRE 2353, Institut de Biologie de Lille, Institut Pasteur de Lille, 59021 Lille, France Primary cultures of Sertoli cells provide an interesting model to study how signalling pathways induced by a single hormone in a single cell type evolve, depending on the developmental stage. In vivo, follicle-stimulating hormone (FSH) induces proliferation of Sertoli cells in neonate and controls the subsequent dierentiation of the entire population. Molecular mechanisms underlying Sertoli cell pleiotropic responses to FSH have long been investigated. But to date, only cAMP-dependent kinase (PKA) activation has been reported to account for most FSH biological activities in male. Here, we demonstrate that FSH activates the ERK MAP kinase pathway following dual coupling of the FSH-R both to Gs and to Gi heterotrimeric proteins, in a PKA- and also Src- dependent manner. This activation is required for FSH- induced proliferation of Sertoli cells isolated 5 days after birth. Consistently, we show that the ERK-mediated FSH mitogenic eect triggers upregulation of cyclin D1. In sharp contrast, at 19 days after birth, as cells proceed through their dierentiation program, the ERK pathway is dramatically inhibited by FSH treatment. Taken together, these results show that FSH can exert opposite eects on the ERK signalling cascade during the maturation process of Sertoli cells. Thus, signalling modules triggered by the FSH-R evolve dynamically throughout development of FSH natural target cells. Oncogene (2001) 20, 4696 – 4709. Keywords: ERK MAP kinases; cAMP/PKA; FSH; cell proliferation; cell dierentiation Introduction Stimulation of cell-surface G protein-coupled receptors (GPCRs) elicits biological responses to a plethora of extracellular signals such as hormones and neurotrans- mitters. Ligand-bound GPCRs stimulate biochemical pathways initiated by the recruitment and activation of Ga and Gbg proteins, and subsequent release of intracellular second messengers such as cAMP. Over the last decade, it has become increasingly clear that GPCRs can also profoundly aect cell growth and dierentiation processes by interfering with the mito- gen-activated protein kinase (MAP kinase) cascades. MAP kinase-dependent pathways group a set of signalling modules (reviewed in van Biesen et al., 1996), including membrane recruitment by a small G protein of the Ras type of a Ser/Thr kinase of the Raf family (MAP kinase kinase kinase) which subsequently activates a Thr- and Tyr dual specificity kinase (MAP kinase kinase) such as MEK1 or MEK2 (reviewed in Tan and Kim, 1999). The outcomes of cAMP and PKA modulation on MAP kinase activity, which reflect tissue-specificity of the signalling modules recruited, are expected to ultimately elicit dierent biological responses. cAMP has long been viewed as an inhibitory signal towards mitogenic stimuli by antagonizing the MAP kinase pathway, as illustrated by Rat-1 fibroblast response to EGF (Wu et al., 1993) and to LPA (Cook and McCormick, 1993). Conversely, ERK activation by cAMP and/or PKA has mainly been evidenced by rare examples of transformed cell lines such as cAMP- treated Jurkat cells (Saxena et al., 1999), PC12 neuronal cells (Vossler et al., 1997), a-MSH-stimulated B16 melanoma cells (Busca et al., 2000), and isoproterenol-stimulated HEK293 cells (Daaka et al., 1997). These cell models have certainly broadened our current knowledge of signalling mechanisms. Notwith- standing, they are proven poorly helpful when a physiological issue is addressed. Physiological antagon- ism of MAP kinase and PKA signalling was first demonstrated in human arterial smooth muscle cells (Graves et al., 1993) in response to PDGF and in adipocytes stimulated by Insulin (Sevetson et al., 1993). Opposedly, EGF- or isoproterenol-induced prolifera- tion of rat parotid acinar cells via MAP kinase signalling is not impaired by a concomitant cAMP rise (Purushotham et al., 1994). Follicle-Stimulating Hormone (FSH) has long been considered as the pituitary key regulator orchestrating gametogenesis. Physiological functions of FSH are Oncogene (2001) 20, 4696 – 4709 ª 2001 Nature Publishing Group All rights reserved 0950 – 9232/01 $15.00 www.nature.com/onc *Correspondence: P Cre´pieux, E-mail: crepieux@tours.inra.fr Received 27 March 2001; revised 3 May 2001; accepted 10 May 2001