MCF-7 cells were maintained at 37 °C in Dulbecco’s modified Eagle’s medium supplemented with 10% fetal calf serum, 1% nonessential amino acids and 2% of penicillin/streptomycine. The cell line has been authenticated by Eurofins. Prior to treatment with ligands, cells were grown for 48 h in phenol red-free medium supplemented with 10% charcoal-stripped serum (Biowest), in order to remove steroid hormones or in serum-free medium for IGF-1 treatment. The cells were then treated for different times with E2 (Sigma) 10–8 M or IGF-1 (4×10–5 µg/µl) from Peprotech. When stated, cells were treated with the PRMT1 inhibitor MS023 (Tocris Bioscience).
For knockdown experiments, specific siRNAs or scramble siRNA (Eurogentec) (50 nM) were transfected into MCF-7 cells using the lipofectamine 2000 reagent (Invitrogen). The targeted sequences are given in Supplementary Table1 . After 72 h of transfection, proteins were analyzed.
For overexpression experiments, pSG5-Flag-tagged vectors were transfected into MCF-7 cells using Jetprime reagent (Ozyme) according to the manufacturer’s protocol. Thirty hours after transfection, cells were collected and analyzed.
For knockdown experiments, specific siRNAs or scramble siRNA (Eurogentec) (50 nM) were transfected into MCF-7 cells using the lipofectamine 2000 reagent (Invitrogen). The targeted sequences are given in Supplementary Table
For overexpression experiments, pSG5-Flag-tagged vectors were transfected into MCF-7 cells using Jetprime reagent (Ozyme) according to the manufacturer’s protocol. Thirty hours after transfection, cells were collected and analyzed.