Protein kinase B (Akt) blockade inhibits LH/hCG-mediated 17,20-lyase, but not 17α-hydroxylase activity of Cyp17a1 in mouse Leydig cell steroidogenesis

• Published in: Cellular signalling 2023-11, Vol.111, p.110872-110872, Article 110872
• Main Authors: Paradiso, Elia, Lazzaretti, Clara, Sperduti, Samantha, Melli, Beatrice, Trenti, Tommaso, Tagliavini, Simonetta, Roli, Laura, D'Achille, Fabio, Beltrán-Frutos, Ester, Simoni, Manuela, Casarini, Livio
• Format: Article
• Language: English
• Subjects: Akt; Akt inhibitor; Cyp17a1; Leydig cells; Steroidogenesis
• ISSN: 0898-6568; 1873-3913
• DOI: 10.1016/j.cellsig.2023.110872

Androgens are produced by adrenal and gonadal cells thanks to the action of specific enzymes. We investigated the role of protein kinase B (Akt) in the modulation of Δ4 steroidogenic enzymes' activity, in the mouse Leydig tumor cell line mLTC1. Cells were treated for 0–24 h with the 3 × 50% effective concentration of human luteinizing hormone (LH) and choriogonadotropin (hCG), in the presence and in the absence of the specific Akt inhibitor 3CAI. Cell signaling analysis was performed by bioluminescence resonance energy transfer (BRET) and Western blotting, while the expression of key target genes was investigated by real-time PCR. The synthesis of progesterone, 17α-hydroxy (OH)-progesterone and testosterone was measured by immunoassay. Control experiments for cell viability and caspase 3 activation were performed as well. We found that both hormones activated cAMP and downstream effectors, such as extracellularly-regulated kinase 1/2 (Erk1/2) and cAMP response element-binding protein (Creb), as well as Akt, and the transcription of Stard1, Hsd3b1, Cyp17a1 and Hsd17b3 genes, boosting the Δ4 steroidogenic pathway. Interestingly, Akt blockade decreased selectively Cyp17a1 expression levels, inhibiting its 17,20-lyase, but not the 17-hydroxylase activity. This effect is consistent with lower Cyp17a1 affinity to 17α-OH-progesterone than progesterone. As a result, cell treatment with 3CAI resulted in 17α-OH-progesterone accumulation at 16–24 h and decreased testosterone levels after 24 h. In conclusion, in the mouse Leydig cell line mLTC1, we found substantial Akt dependence of the 17,20-lyase activity and testosterone synthesis. Our results indicate that different intracellular pathways modulate selectively the dual activity of Cyp17a1. •The role of protein kinase B (Akt) in Leydig cell steroidogenesis is unclear.•Akt blockade by 3CAI decreased selectively Cyp17a1 expression levels.•Low Cyp17a1 levels decreased 17,20-lyase, but not the 17-hydroxylase activity.•Low 17-hydroxylase activity is linked to decreased testosterone synthesis.