Properties and hormonal regulation of two structurally related cAMP phosphodiesterases from the rat Sertoli cell

Upon exposure to follicle-stimulating hormone (FSH), the gonadotropin-responsive Sertoli cell expresses increased rolipram-sensitive cAMP-specific phosphodiesterase (cAMP-PDE) activity. To understand the mechanisms leading to this activation, the cAMP-PDEs present in the Sertoli cell were characterized and their regulation studied. Comparison of the conceptual translates of two groups of PDE cDNA clones isolated from a Sertoli cell cDNA library (ratPDE3 and ratPDE4) showed that the encoded proteins were structurally similar, containing a core region of 455 amino acids with a sequence identity of 87%. The amino and carboxyl termini were divergent. Expression of these cDNAs in Escherichia coli and monkey COS-7 cells demonstrated that the encoded cAMP-PDEs had similar affinities for the cAMP substrate and were equally sensitive to a number of PDE inhibitors (rolipram greater than Ro 20-1724 greater than cilostamide). FSH stimulation of the Sertoli cell produced an increased rate of transcription of the ratPDE3 gene and elevated mRNA levels for ratPDE3 and to a lesser extent of ratPDE4. The increase in mRNA levels was detected after 1 h of stimulation. Forskolin, cholera toxin, and N6, O2'-dibutyryl cAMP produced a similar increase in rate of transcription and elevated mRNA levels, indicating that this activation is mediated by an increase in intracellular cAMP. RatPDE4 mRNA levels were maximal upon exposure to 10 ng of FSH/ml, whereas ratPDE3 mRNA levels could be further elevated, with higher FSH concentrations. The intensity of an immunoreactive band with characteristics identical to a purified cAMP-PDE, correlated with the increased cAMP hydrolytic activity after FSH or dibutyryl cAMP treatment, demonstrating that changes in cAMP-PDE protein levels are involved in this regulation. These data provide evidence that multiple cAMP-PDE forms are expressed in the rat Sertoli cell. Although differences in the pattern of activation of these forms were observed, these data show, that in the rat Sertoli cell, the cAMP-PDE activity is regulated by hormones via a novel mechanism that involves a cAMP-dependent activation of transcription of a PDE gene.