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Figure 6 | Journal of Molecular Signaling

Figure 6

From: Differential role of beta-arrestin ubiquitination in agonist-promoted down-regulation of M1 vs M2 muscarinic acetylcholine receptors

Figure 6

Expression of a chimeric ubiquitinated form of β-arrestin 2 enhances the agonist-promoted down-regulation of M 1 and M 2 mAChRs in MEF KO1/2. MEF KO1/2 cells were transfected with eGFP-M1 (A) or HA-M2 (B) mAChRs and either no β-arrestin, FLAG-β-arrestin 2 or YFP-β-arrestin 2-Ub. 24 hr following transfection, cells were treated with 1 mM carbachol for 12 hr. Down-regulation was determined using [3H]-QNB binding (fmol/mg protein) in crude membranes as described in methods. Expression of the constitutively ubiquitinated form of β-arrestin (β-arrestin 2-Ub) had a greater effect on down-regulation of the M2 mAChR compared to the M1 subtype. Data are expressed as percent of [3H]-QNB bound compared to the untreated control with no β-arrestin and are presented as mean ± standard deviation from two independent experiments with duplicate data points. Statistical analysis was performed using a repeated measures ANOVA with Bonferroni post test; * indicates p ≤ 0.05 and ** indicates p ≤ 0.001 (compared to untreated, no β-arrestin control), ns indicates not significant. Total mAChR expressed (fmol/mg) in the absence of β-arrestin was 1000 – 2000 for both receptor subtypes.

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