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. Author manuscript; available in PMC: 2013 Jan 1.
Published in final edited form as: J Neurochem. 2011 Nov 24;120(1):56–69. doi: 10.1111/j.1471-4159.2011.07559.x

Fig. 2. Effect of D2R or MOR co-expression of on TX100 solubility of RGS9-2 and effect of D2R co-expression on TX100 solubility of either the green fluorescent protein (GFP) or the transferrin receptors (TFR).

Fig. 2

A. HEK293T cells were transfected with RGS9-2-V5-tagged cDNA (RGS9) alone or with RGS9 and D2R cDNA (RGS9+D2R) or RGS9 and MOR cDNA (RGS9+MOR) as indicated. CTRL refers to a control group of cells transfected with empty plasmid vector. Cells were solubilized in TX100 detergent-containing buffer. Soluble and insoluble proteins were separated by centrifugation, resolved by SDS PAGE, and the RGS9-2 protein in TX100 soluble (S) and insoluble (P, pellet) cell fractions were visualized by immunoblotting using an anti-V5 epitope tag antibody, as described in the Materials and Methods section. B. Levels of RGS9-2 protein in TX100-soluble (closed black bars) and insoluble (open white bars) cellular fractions visualized in A and expressed as percentage of the total cellular RGS9-2 protein (mean ± SEM, n=4-5, *represents p<0.01 compared to RGS9-2 cDNA only transfected group). C. HEK293T cells were transfected with GFP cDNA alone or with GFP and D2R cDNA as indicated. CTRL refers to a control group of cells transfected with empty plasmid vector. Cells were solubilized in TX100 detergent-containing buffer. Soluble and insoluble proteins were separated by centrifugation, resolved by SDS PAGE, and the GFP protein in TX100 soluble (S) and insoluble (P, pellet) cell fractions were visualized by immuno-blotting using an anti-GFP antibody, as described in the Materials and Methods section. D. Levels of GFP protein in TX100-soluble (closed black bars) and insoluble (open bars) cellular fractions visualized in C and expressed as percentage of the total cellular GFP protein (mean ± SD, n=2). E. HEK293T cells were transfected with either empty vector control (CTRL) or D2R cDNA as indicated. Soluble and insoluble proteins were separated and resolved by SDS PAGE as described above and the endogenously expressed TFR in TX100 soluble (S) and insoluble (P, pellet) cell fractions were visualized by immuno-blotting using an anti-TFR antibody, as described in the Materials and Methods. F. Levels of TFR protein in TX100-soluble (closed black bars) and insoluble (open bars) cellular fractions visualized in E and expressed as percentage of the total cellular TFR protein (mean ± SEM, n=4).