THE INTRAMITOCHONDRIAL ARACHIDONIC ACID RELEASE IS NEEDED FOR CHOLESTEROL TRANSPORT

CASTILLA, ROCÍO; MALOBERTI, PAULA; PODESTÁ, ERNESTO J.

Pinamar, Buenos Aires, Argentina

Congreso; X Congress of Panamerican Association for Biochemistry and Molecular Biology; 2005

Sociedad Argentina de Investigación en Bioquímica y Biología Molecular

We have previously described that the Arachidonic Acid- preferring Acyl-CoA synthetase 4 and the mitochondrial Acyl-CoA thioesterase I (Acot2) regulate, in a concerted mode, the intracellular levels of AA and the steroids production. Steroidogenic cells express Acot2 but also a cytosolic isoform (Acot1) which is 92.5% homologous to the mitochondrial enzyme. The aim of this study was to determinate the role of Acot1 in the steroidogenesis. For this purpose, we overexpressed Acot1 in the MA-10 Leydig cell line. Cell transfection with a plasmid containing the full sense Acot1 cDNA produced a clear increase in the Acot1 expression level as analyzed by immunocitochemistry and western blot. The Acot1 overexpression did not affect the cell viability as assessed by Trypan Blue exclusion method. The 22-OH cholesterol–sustained steroid synthesis in the Acot1 over expressing cells was not affected. A little decrease of steroid production (% of inhibition: 21.99 ± 2.2) was evidenced when we studied the effect of Acot1 overexpression on cAMP-stimulated steroidogenesis although Acot2 overexpression produced a significant increase in the levels of steroid synthesis (% of stimulation: 66.3 ± 6.1) both compared with mock-transfected cells. From these results we conclude that only the isoform Acot2 participates in the steroid synthesis, hence, the intramitochondrial AA release is needed for cholesterol transport. Further studies have been made to determinate de role of Acot1 in these cells.