Figure 5

hSIN3B interacts with ETO in primary placenta cells. (A) Western blotting was used to detect hSIN3B and ETO homologues in TRIZOL extract of primary placental cells as described in Material and Methods. The following antibodies were used: α-hSIN3B (SIN), α-ETO (ETO), α-MTGR1 (R1), α-MTG16 (16) and α-MTG (MTG, reactive with all ETO homologues). Arrowhead shows the position of hSIN3B and ETO homologues. The blots were re-probed with pre-immune serum (PIS) in order to rule out non-specific binding of the antibodies. The blots were probed with α-actin to show equal loading. (B) IP-Western was used to examine the presence of complexes between hSIN3B and the ETO homologues in nuclear extracts. IP with α-MTG pulled down hSIN3B as detected on immunoblotting with α-SIN3B (SIN) (lane 2). In the reverse experiment, IP with α-SIN3B (SIN) pulled down ETO as detected on immunoblotting with α-ETO (ETO) (lane 4). However, IP with α-SIN3B (SIN) did not pull down MTGR1 or MTG16 as no signal was detected upon immunoblotting with α-MTGR1 or MTG16 (lanes 5–8). Lower panel shows input of MTGR1 and MTG16 in 2% of IP lysate. The positions of the molecular weight markers are indicated at the left.