Fig. 5.

CaMKII regulates K63-linked ubiquitination of GABA_B receptors. a Inhibition of CaMKII activity reduced K63-linked ubiquitination of GABA_B receptors. Neurons were incubated for 7.5 min in the absence (control) or presence of KN93 and analyzed for K63-linked ubiquitination of GABA_B receptor by in situ PLA using antibodies directed against GABA_B1- and K63-linked ubiquitin (white dots in representative images, scale bar 5 μm). Right, quantification of in situ PLA signals. The data represent the mean ± SD of 33 (control) and 30 (KN93) neurons from two independent experiments. **p < 0.01, two-tailed unpaired t test. b, c CaMKIIβ promotes Lys-63-linked ubiquitination of GABA_B receptors. Neurons were transfected with either CaMKIIα or CaMKIIβ or their functionally inactive mutants CaMKIIα(DN) or CaMKIIβ(DN). After 2 days, GABA_B receptors were tested for Lys-63-linked (b) or Lys-48-linked (c) ubiquitination by in situ PLA using antibodies directed against GABA_B1 and Lys-63 or Lys-48 ubiquitin. Left, representative images depicting PLA signals (white dots, scale bars 5 μm). Right, quantification of PLA signals. The data represent the mean ± SD of 17–21 neurons from two independent experiments. **p < 0.01, ****p < 0.0001, ^nsp > 0.05; two-tailed unpaired t test