COORDINATED ACTION OF PKA AND ERK IN CHOLESTEROL TRANSPORT ACTIVATION.

CECILIA PODEROSO*, SOLEDAD GALLI¥, PAULA MALOBERTI*, JUAN JOSÉ PODEROSO¥ AND ERNESTO PODESTÁ

Pinamar, Buenos Aires

Congreso; XLI Reunion Anual de SAIB; 2005

Sociedad Argentina de Investigacion en Bioquimica y Biologia Molecular

We have shown a strict dependence of ERK1/2 activation for PKA-activated cholesterol transport in MA-10 Leydig cells. The goal of the present study was to analyze the interaction of PKA, ERK and StAR (Steroidogenic Acute Regulatory) protein on mitochondria isolated from MA-10 cells. The interaction of ERK1 with PKA and StAR was demonstrated by co-precipitation and Western blot of PKA and StAR from isolated mitochondria derived from stimulated cells and ERK1-GST bound to agarose. Mitochondria derived from cells transfected with either sense or antisense StAR cDNA were incubated with active or mutated inactive (Lys71 à Ala) ERK and catalytic PKA subunit. Mitochondrial StAR content increased after sense cDNA transfection and production of progesterone increased in the presence of PKA and wild-type ERK from 32±3 to 70±7 ng/ml (+112%, p<0.05). The effect was abolished by the use of mutated ERK or antisense StAR cDNA. The results indicate that an interaction between PKA, ERK and StAR is required to activate cholesterol transport. The presence of a consensus site for ERK phosphorylation within StAR protein suggests that, apart from PKA, ERK activity would also be required for the formation of a molten-globule StAR conformation leading to cholesterol interaction with hydrophobic domains.