Regulation of mitochondrial dynamics proteins in MA-10 Leydig cells: the role of Mitofusin 2 on Acyl-CoA synthetase expression, a key enzyme in steroidogenesis

ANA Z. FIORE; KATIA E. HELFENBERGER; LUCÍA HERRERA; GIULIANA ARGENTINO; ANA FERNANDA CASTILLO; CECILIA PODEROSO

Congreso; LXIV Reunión Anual de la Sociedad Argentina de Investigación Clínica (SAIC); 2019

Mitochondrial fusion is driven by GTPases like Mitofusins 1 and 2 (Mfn1/2) and Optic Atrophy 1 (OPA1) whereas mitochondrial fission mainly involves Dynamin-related protein 1 (Drp1). Mitochondrial dynamics (fusion/fission events) participate in many physiological processes and in different human pathologies. Steroidogenesis depends on several mitochondrial proteins as PKA, StAR, ERK/MEK, SHP2 (tyrosine phosphatase) and Acyl-CoA synthetase 4 (Acsl4). We have previously demonstrated that Mfn2 participates in steroids production and in StAR expression and mitochondrial localization, in MA-10 murine Leydig cells. However, the regulation of mitochondrial dynamics proteins under hormonal stimulation has been little explored. The aim of this study was to evaluate the hormone-regulation of Mfn2, OPA1 and Drp1 and a possible role of Mfn2 in Acsl4 expression and mitochondrial localization, in MA-10 Leydig cells. We observed that Mfn2 levels significantly increase after 8Br-cAMP (cAMP analogue) stimulation, in a time-dependent manner up to 24h (control vs. cAMP 4h,***p0.001). We detected by immunoblot two main OPA1 isoforms that are regulated by 8Br-cAMP. Mitochondrial Drp1 is significantly phosphorylated by human chorionic gonadotropin (hCG) and 8Br-cAMP stimulation in serine 637 (control vs 1h, ***p0.001), a PKA-phosphorylation site associated with a decrease in Drp1 fission activity. By decreasing Mfn2 expression with a shRNA, we observed that the presence of Mfn2 is necessary for the expression and mitochondrial localization of Acsl4, detected by real time PCR and immunoblot (***p0.001 mock vs shRNA); whereas mitochondrial localization of PKA did not depend on Mfn2. Phospho-Drp1 levels were not significantly affected by Mfn2 down-regulation, in agreement with the presence of mitochondrial PKA. Our results indicate that mitochondrial dynamics proteins are hormone-regulated and Mfn2 plays a role in the expression and mitochondrial localization of Acsl4 in MA-10 cells.