β-Arrestins were identified as scaffold-proteins that have the capacity to desensitize G protein-coupled receptors. However, it has been found that β-arrestins activate signaling pathways independent of G protein activation. The diversity of these signaling pathways has also been recognized for receptor tyrosine kinase. The aim of the present study was to validate the β-arrestin-dependent signaling mechanism(s) responsible for regulation of adipogenesis. Two signal models were selected, ghrelin and insulin, based on its β-arrestin-associated Akt activity. Herein, we found that β-arrestin 1 and 2 were essential molecules for adipocyte differentiation. More specifically, the role of these scaffolding proteins was demonstrated by depletion of β-arrestin 1 and 2 during ghrelin-induced adipogenesis in 3T3-L1 cells, which decreased the adipocyte differentiation and the expression levels of master regulators of early, the CCAAT/enhancer-binding protein β (C/EBPβ) and the CCAAT/enhancer-binding protein δ (C/EBPδ), and terminal, the peroxisome proliferator-activated receptor (PPARγ) and the CCAAT/enhancer-binding protein α (C/EBPα), adipogenesis. Accordingly ghrelin-induced Akt activity and its downstream targets, the mammalian target of rapamycin complex 1 (mTORC1) and the ribosomal protein S6 kinase beta-1 (S6K1), were inhibited by β-arrestin 1 and 2 siRNAs. By contrast, assays performed during insulin-activated adipogenesis showed an intensifying effect on the adipocyte differentiation as well as on the expression of C/EBPβ, C/EBPδ, PPARγ and C/EBPα. The increase in insulin-induced adipogenesis by β-arrestin knock-down was concomitant to a decrease in the insulin receptor susbtrate-1 (IRS-1) serine phosphorylation, proving the loss of the negative feedback loop on IRS-1/phosphoinositide 3-kinase (PI3K)/Akt. Therefore, β-arrestins control the extent and intensity of the lipogenic and adipogenic factors associated to Akt signaling, although the mechanistic and functional principles that underlie the connection between signaling and β-arrestins are specifically associated to each receptor type.
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