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Figure 1 BRCA1 associates with SUMO1 in vitro and in vivo. (A) Schematic representation of BRCA1 constructs. The RING domain (RING), nuclear localization signal sequences (NLS), activation domain and two BRCT domains (BRCT) are indicated. Numbers above the BRCA1 constructs used in this study indicate the amino acid residues of the respective BRCA1 fragments. (B) Interaction of BRCA1 with SUMO1 in the yeast two hybrid system. Yeast cells transformed with two hybrid plasmids were grown under induced conditions for reporter gene activation. The streaks represent yeast cells cotransformed with either pLexA BRCA1 (1 324, 260 553, 502 802 or 758 1064) and pB42AD SUMO1. The ability of the transactivation domains of BRCA1 to interact with SUMO1 was measured following cotransformation of yeast cells with pB42AD BRCA1 (1005 1313 or 1314 1864) and pLexA SUMO1. (C) In vitro interaction of BRCA1 with SUMO1. The six GST BRCA1 (1 324, 260 553, 502 802, 758 1064, 1005 1313 and 1314 1864, indicated as #1 #6, respectively) and GST proteins were immobilized on GST Sepharose beads and incubated with His SUMO1 proteins. His SUMO1 proteins bound to immobilized GST BRCA1 proteins were analyzed by immunoblotting with an anti SUMO1 antibody (Upper). Twenty percent of the input SUMO1 proteins (input). SUMO1 did not bind to immobilized GST. An equivalent amount of GST BRCA1 protein was used for immobilization (Lower). (D) In vivo interaction of BRCA1 with SUMO1. Extracts of 293T cells were immunoprecipitated with anti BRCA1. Coimmunoprecipitated SUMO1 proteins were detected by immunoblots with anti SUMO1 antibody..
Image ReferenceSUMO1 negatively regulates BRCA1 mediated transcription, via modulation of promoter occupancy
Nucleic Acids Res. 2008 Jan; 36(1):263-283