Figure 7. Co-immunoprecipitation of the cleaved distal C-terminus with truncated CaV1.2 channels.
A, TsA-201 cells were cotransfected with CaV1.2▵1731 plus distal1900–2171. Cell lysates were immunoprecipitated (ip) with anti-CNC1 antibody (lane 3) or non-immune IgG (lane 2). Immunoblots were probed with anti-CNC1 to detect truncated (upper panel) or anti-CH1 (lower panel) to detect distal1900–2171. Positive control for immunoblotting was 20 μl of extract (lane 1). B, CaV1.2▵1701 plus distal1900–2171. C, CaV1.2▵1693 plus distal1900–2171. D, CaV1.2▵1701RRQQ plus distal1900–2171(EEDQQQ). In this experiment, a longer exposure time was used to detect the lower amount of coimmunprecipitated distal1900–2171(EEDQQQ), which resulted in more intense staining of the CaV1.2▵1693 band as well. Relative to the staining of CaV1.2▵1693, there is much less coimmunoprecipitated distal1900–2171(EEDQQQ), indicating weaker interaction with this mutant distal C-terminal peptide.