An interaction of Bcl-2 with SERCA had been documented in vitro using the SERCA1a isoform isolated from rat skeletal muscle [Dremina， E. S.， Sharov， V. S.， Kumar， K.， Azidi， A.， Michaelis， E. K.， Schöneich， C. (2004) Biochem. J. 383 (361-370)]. Here， we demonstrate the interaction of Bcl-2 with the SERCA3b isoform both in vitro and in cell culture. In vitro， the interaction of Bcl-2 with SERCA3b was studied using Bcl-2∆21， a truncated form of human Bcl-2， and microsomes isolated from SERCA3b-overexpressing HEK-293 cells. For these experiments， SERCA3b was quantified by a combination of amino acid analysis and Western blotting. We observed that Bcl-2∆21 both inactivates SERCA... More
An interaction of Bcl-2 with SERCA had been documented in vitro using the SERCA1a isoform isolated from rat skeletal muscle [Dremina， E. S.， Sharov， V. S.， Kumar， K.， Azidi， A.， Michaelis， E. K.， Schöneich， C. (2004) Biochem. J. 383 (361-370)]. Here， we demonstrate the interaction of Bcl-2 with the SERCA3b isoform both in vitro and in cell culture. In vitro， the interaction of Bcl-2 with SERCA3b was studied using Bcl-2∆21， a truncated form of human Bcl-2， and microsomes isolated from SERCA3b-overexpressing HEK-293 cells. For these experiments， SERCA3b was quantified by a combination of amino acid analysis and Western blotting. We observed that Bcl-2∆21 both inactivates SERCA3b and co-immunoprecipitates with SERCA3b. The incubation with Bcl-2∆21 changes the distribution of SERCA3b during sucrose density gradient centrifugation， likely as the result of Bcl-2∆21-induced conformational change of SERCA3b. When SERCA3b-overexpressing HEK-293 cells were co-transfected with Bcl-2， Bcl-2-dependent SERCA3b inactivation was observed. In these cells， Bcl-2 interaction with SERCA3b was demonstrated by co-immunoprecipitation. Furthermore， overexpression of Bcl-2 reduced fluorescein isothiocyanate (FITC) labeling of SERCA3b. Together， our data provide evidence for the interaction of Bcl-2 with SERCA3b in vitro and in cell culture， and for Bcl-2-dependent conformational and functional changes of SERCA3b.
