Map4k4 negatively regulates peroxisome proliferator-activated receptor (PPAR) γ protein translation by suppressing the mammalian target of rapamycin (mTOR) signaling pathway in cultured adipocytes

Abstract

The receptor peroxisome proliferator-activated receptor γ (PPARγ) is considered a master regulator of adipocyte differentiation and promotes glucose and lipid metabolism in mature adipocytes. We recently identified the yeast Sterile 20 (Ste20) protein kinase ortholog, Map4k4, in an RNA interference-based screen as an inhibitor of PPARγ expression in cultured adipocytes. Here, we show that RNA interference-mediated silencing of Map4k4 elevates the levels of both PPARγ1 and PPARγ2 proteins in 3T3-L1 adipocytes without affecting PPARγmRNA levels, suggesting that Map4k4 regulates PPARγ at a post-transcriptional step. PPARγ degradation rates are remarkably rapid as measured in the presence of cycloheximide (t1/2 = 2 h), but silencing Map4k4 had no effect on PPARγ degradation. However, depletion of Map4k4 significantly enhances [ 35S]methionine/ cysteine incorporation into proteins, suggesting that Map4k4 signaling decreases protein translation. We show a function of Map4k4 is to inhibit rapamycin-sensitive mammalian target of rapamycin (mTOR) activity, decreasing 4E-BP1 phosphorylation. In addition, our results show mTOR and 4E-BP1 are required for the increased PPARγ protein expression upon Map4k4 knockdown. Consistent with this concept, adenovirus-mediated expression of Map4k4 decreased PPARγ protein levels and mTOR phosphorylation. These data show that Map4k4 negatively regulates PPARγ post-transcriptionally, by attenuating mTOR signaling and a 4E-BP1-dependent mechanism. © 2010 by The American Society for Biochemistry and Molecular Biology, Inc.