Fig. 3. LEF1 is a novel identified PRMT6 interaction partner.

Co-streptavidin-precipitation (CoSP). The biotin tagged bait was co-transfected with the prey expression vector into HEK293 cells. The biotinylated bait was pulled out from extracts with streptavidin beads (SP). Coprecipitated bait protein is shown in the upper lane, (CoSP). A CoSP of PRMT6 and LEF1. Coprecipitation of LEF1 was detected with an anti-LEF1 antibody. B CoSP of PRMT6 and RUNX1. Coprecipitation of RUNX1 was detected with an anti-RUNX1 antibody. C CoSP of RUNX1 and LEF1. Biotinylated RUNX1 protein was pulled out from extracts with streptavidin beads. Coprecipitation of LEF1 was detected with an anti-LEF1 antibody. D CoSP of PRMT6 and PRMT1. Coprecipitation of PRMT1 was detected with an anti-HA antibody. E GST-pulldown with GST-PRMT6 as bait and LEF1 protein expressed in HEK293 cells. GST-PRMT6 was incubated with cell extracts of from LEF1 over expressing cells. GST-pulldown with GST protein served as negative control. The GST proteins were pulled out with glutathione beads. Pulled out LEF1 was detected by western blot with anti-LEF1 antibody. F GST-pulldown with GST-PRMT6 and in vitro translated LEF1. Pulled out LEF1 was detected by western blot with anti-LEF1 antibody. G Schematic representation of the LEF1 protein. H GST-pulldown with GST-PRMT6 and in vitro translated ³⁵S labeled LEF1 deletion constructs. GST protein served as negative control and the GST proteins were pulled out with glutathione beads. Detection of pulled out LEF1 protein was done by radiography.