Gene trap analysis of germ cell signaling to Sertoli cells: NGF-TrkA mediated induction of Fra1 and Fos by post-meiotic germ cells

Abstract

Analysis of complex signalisation networks involving distinct cell types is required to understand most developmental processes. Differentiation of male germ cells in adult mammals involves such a cross-talk between Sertoli cells, the somatic component which supports and controls germinal differentiation, and germ cells at their successive maturation stages. We developed a gene trapping strategy to identify genes, which, in Sertoli cells, are either up- or down-regulated by signals emitted by the germinal component. A library of approximately 2,000 clones was constituted from colonies independently selected from the Sertoli line 15P-1 by growth in drug-containing medium after random integration of a promoter-less (beta)geo transgene (neo(r)-lacZ fusion), which will be expressed as a fusion transcript from a ‘trapped’ cellular promoter, different in each clone. A first screen conducted on 700 events identified six clones in which beta-galactosidase activity was increased and one in which it was repressed upon addition of germ cells. The targeted loci were identified by cloning and sequencing the genomic region 5′ of the insert. One of them was identified as the gene encoding Fra1, a component of the AP1 transcription regulatory complex. Accumulation of Fra1 mRNA was induced, both in 15P-1 and in freshly explanted Sertoli cells, by addition of either round spermatids or nerve growth factor (NGF). The effect of NGF was mediated by the TrkA receptor and the ERK1-ERK2 kinase kinase pathway. Fos and Fra1 transcription were induced within the first hour after addition of the neurotrophin, but, unlike what is observed after serum induction in the same cells, a second wave of transcription of Fra1, but not of Fos, started 16 hours later and peaked at higher levels at about 20 hours. These results suggest that AP1 activation may be an important relay in the Sertoli-germ cell cross-talk, and validate the gene trapping approach as a tool for the identification of target genes in cell culture systems.

A Synthetic Peptide Modeled on PDNF, Chagas' Disease Parasite Neurotrophic Factor, Promotes Survival and Differentiation of Neuronal Cells through TrkA Receptor

“…However, the mutation of the CRE site in the pFLuc promoter rendered cells transfected with pCREunresponsive to either cAMP stimulation (data not shown), PDNF, or Y 21 (Figure 8B). The response to NGF was significantly reduced but not completely abolished because NGF could also induce transcriptional activation via the AP1 response element (34), also present in the TH promoter region (inset of Figure 8B).…”

Testicular Steroidogenesis after Pinealectomy - the Role of BDNF Signaling System

“…Recently, an object of research has been the potential role of the BDNFsignaling system, consisting of the brain derived neurotrophic factor (BDNF) and its corresponding receptor TrkB, outside the development and proliferation of neurons, namely: in the autocrine and paracrine control of testicular steroidogenesis [11,12]. The expression of this signaling molecule and its corresponding receptor in testicular Leydig cells (LCs) supports the hypothesis that neurotrophins produced and/or acting on LCs are involved in the process of their differentiation and functional activity [3][4][5][6][7][8].…”