KDM1A, HDACs, and KAP1 cooperate to repress endogenous retroviral sequences in ES cells. (A,B) KDM1A complexes were affinity-purified from ES cells and subjected to LDS-PAGE and silver staining (A) or immunoblotting (B) with the indicated antibodies. (C) Wild-type ES cells were immunostained with KDM1A and KAP1 antibodies and subjected to confocal microscopy. (D) Bisulfite sequencing was performed on Kdm1a +/+ or GT/GT ES cells with primers amplifying the MERVL retrovirus. (E–G) Kdm1a +/+ or GT/GT ES cells were treated with TSA or 5Aza for 24 h. qRT–PCR was performed using primers specific for MERVL Pol (E), Tcstv3 (F), or Eif1a (Gm2022) (G) and normalized to Gapdh. The fold change was then plotted relative to untreated Kdm1a +/+ ES cells, with error bars representing SD. (H) Kdm1a +/+, +/GT, or GT/GT ES cells were treated with TSA for 24 h, and cells were subjected to immunoblotting with the indicated antibodies. (I) Kdm1a FL/FL Cre-ERT ES cells were treated with vehicle or 4OHT for 48 h, followed by treatment with TSA for 24 h. Immunoblotting was then performed with the indicated antibodies.