Localization of Novel Adiponectin Receptor Constructs

Abstract

Adiponectin is one of the most abundant fat-derived hormones involved in a multitude of metabolism pathways. The receptors AdipoR1 and AdipoR2 of this metabolically active protein have been identified recently. AdipoR1 and AdipoR2 are most abundantly expressed in the skeletal muscle and in the liver, respectively. It has been postulated that although they both consist of seven transmembrane helices, they are distinct from other G protein-coupled receptors (GPCRs). We cloned both receptors as fusion proteins with enhanced yellow fluorescent protein (YFP) to determine their localization and orientation in the cell membrane. By confocal microscopy and immune staining we demonstrated that both receptor-YFP-fusion proteins are integral membrane proteins with the predicted topology—an intracellular N-terminus and an extracellular C-terminus. In parallel, comparative experiments were performed with the NPY Y₂–receptor, a classical rhodopsin-like GPCR.

Key Words: :

• Adiponectin receptors
• 7TM
• Yellow fluorescent fusion protein
• Western blotting