Physical interaction between HIF-1α and Psen1. Expression vectors carrying no insert, Psen1 or HIF-1α were transfected into HEK 293 cells in the indicated combinations. In panel A, immunoprecipitations (IP) were performed with a polyclonal anti-Psen1 antibody which recognizes the Psen1 NTF or with rabbit IgG as control and Western blotting was performed for HIF-1α. Total extracts without IP (-) (5% of input) were loaded in the two far right lanes (input). Note the co-immunoprecipitation of HIF-1α in the cultures co-transfected with HIF-1α and Psen1. A similar experiment was performed in (B), except that immunoprecipitations were performed with the monoclonal antibody NT.1 which recognizes the Psen1 NTF or with an antibody against the Psen1 loop region. In panel C, immunoprecipitation was done with a monoclonal HIF-1α antibody followed by Western blotting with an antibody against the Psen1 NTF (NT.1) or CTF (33B10). Panels labeled "none" show blots of total extracts prior to immunoprecipitation (input). Bands marked with an asterisk (*) are likely IgGs based on the rate of migration. Panel D shows the co-immunoprecipitation of endogenous HIF-1α by endogenous Psen1. Wild type fibroblasts were treated for 4 hours with 100 μM CoCl2 and 10 μM MG132 to induce HIF-1α accumulation and co-immunoprecipitations were performed as in (A).