Translational regulation of lipoprotein lipase in adipocytes: depletion of cellular protein kinase Cα activates binding of the C subunit of protein kinase A to the 3′-untranslated region of the lipoprotein lipase mRNA

Adipose LPL (lipoprotein lipase) plays an important role in regulating plasma triacylglycerols and lipid metabolism. We have previously demonstrated that PKCα (protein kinase Cα) depletion inhibits LPL translation in 3T3-F442A adipocytes. Using in vitro translation experiments, the minimum essential region on the 3′UTR (3′-untranslated region) of LPL mRNA required for the inhibition of translation was identified as the proximal 39 nt. These results were confirmed by RNase protection analysis using cytoplasmic proteins isolated from the adipocytes treated with PKCα antisense oligomers and the LPL 3′UTR transcript (LPL 3′UTR nt: 1512–1640). The protein components involved in this RNA-binding interaction from PKCα depletion were passed through an affinity column containing a sequence of the LPL 3′UTR and, after Western blotting, the RNA-binding proteins were identified as the catalytic and the regulatory subunits of PKA (protein kinase A), Cα and RIIβ, and AKAP (Akinase-anchoring protein) 121. This RNA inhibitory complex consisted of the same RNA-binding proteins that have been identified previously as mediators of LPL translational inhibition by PKA activation, suggesting that PKCα depletion inhibits LPL translation through PKA activation. In additional experiments, PKC depletion by prolonged PMA treatment or PKCα antisense oligomers resulted in an increase in PKA activity in 3T3-F442A adipocytes, comparable with PKA activation with adrenaline (epinephrine) treatment. These results demonstrate that LPL translational inhibition occurs through an RNA-binding complex involving PKA subunits and AKAP121, and this complex can be activated either through traditional PKA activation methods or through the depletion of PKCα.