Received for publication November 28, 2007.
Revised February 13, 2008.
Accepted for publication February 13, 2008.

Translocation of Glycosylphosphatidylinositol-Anchored Proteins from Plasma Membrane Microdomains to Lipid Droplets in Rat Adipocytes Is Induced by Palmitate, H₂O₂ and the Sulfonylurea Drug, Glimepiride

Abstract

Inhibition of lipolysis by palmitate, H₂O₂ and the antidiabetic sulfonylurea drug, glimepiride, in rat adipocytes has previously been shown to rely on the concerted degradation of cAMP by the glycosylphosphatidylinositol (GPI)-anchored phosphodiesterase Gce1 and 5'-nucleotidase CD73, which both gain access to the lipid droplets (LD). The present report demonstrates the translocation of Gce1 and CD73, harbouring the intact GPI anchor, from detergent-insoluble glycolipid-enriched plasma membrane domains (DIGs) to the LD in response to palmitate, H₂O₂ and glimepiride by analysis of their steady state distribution using photoaffinity labeling and activity determination as well as of their redistribution after pulse or equilibrium metabolic labeling. Surprisingly, palmitate, H₂O₂ and glimepiride induced the activation of the GPI-specific phospholipase C (GPI-PLC) at DIGs of rat adipocytes leading to anchor-less Gce1 and CD73. Inhibition of the GPI-PLC or the presence of non-hydrolyzable substrate analogues of Gce1 and CD73 interfered with the palmitate-, H₂O₂- and glimepiride-induced (i) lipolytic cleavage of Gce1 and CD73, (ii) translocation of their GPI-anchored versions from DIGs to LD, (iii) upregulation of cAMP degradation, and (iv) inhibition of lipolysis. These data suggest a novel insulin-independent anti-lipolytic mechanism in rat adipocytes which relies on the palmitate-, H₂O₂- and glimepiride-induced and GPI-PLC-dependent translocation of (c)AMP-degrading GPI-anchored proteins from the adipocyte plasma membrane to LD. The findings may shed new light on the biogenesis and degradation of LD in response to physiological and pharmacological stimuli.

Key words: Insulin, cAMP, Phosphodiesterases, Protein Kinase A, Lipid rafts/microdomains, Signaling network analyses, Endocrine cells