EGR1 induced by EGF is sumoylated and ubiquitinated <i>in vivo.</i>

<p>(<b>A</b>) ECV304 cells were treated for 1 hour with 10 ng/ml EGF to induce Egr1 expression. Lysates were immunoprecipitated with polyclonal antibody to EGR1 and immunoblotted with anti EGR1 and, after stripping, with anti SUMO-1 antibodies. (<b>B</b>) ECV304 cells were treated 1 hour with 10 ng/ml EGF in the absence or in the presence of 100 µM NEM and lysates obtained were immunoprecipitated with a polyclonal antibodies to EGR1 and immunoblotted with antibodies anti EGR1. The arrows indicate bands corresponding to EGR1 and EGR1 plus SUMO-1. (<b>C</b>) ECV304 cells were treated for 15, 30 or 60 minutes with 10 ng/ml EGF. Lysates were immunoprecipitated with polyclonal antibody to EGR1 and immunoblotted with anti EGR1 and, after stripping, with anti K48 ubiquitin antibodies. (<b>D</b>) ECV304 cells were treated for different times with 10 ng/ml EGF in the absence or in the presence of 100 µM NEM and lysates obtained were resolved by SDS-PAGE and analyzed by Western blot with antibodies against EGR1 and tubulin. All experiments are representative of three independent.</p>