Southwestern blotting to establish the composition of UGPB, a putative transcription factor that binds to the rabbit uteroglobin promotor

Abstract

Southwestern blotting identified two major proteins in heparin-agarose purified endometrial nuclear extract that bind to the -194/+9 region of the rabbit uteroglobin promoter, p105 and p70. These proteins were presented as candidate components of the uteroglobin promotor binding complex (UGPB), a protein complex proposed to regulate expression of the endometrial uteroglobin gene. Both p105 and p70 displayed similar binding characteristics as UGPB and p105 was established to co-purify with UGPB during affinity purification. Inclusion of competing DNA in Southwestern blotting suggested that p105 is a relatively non-specific DNA binding protein, whereas p70 is more specific. Both p105 and p70 were shown to bind to the UGPB DNase I footprint region of the uteroglobin promoter (-170/-85) and it was suggested that p105 may bind distally to the uteroglobin transcription start site, while p70 binds proximally. p105 and p70 were identified in unstimulated endometrial extracts as well as endometrial extracts from progesterone and oestrogen stimulated rabbits. pl05 may also be present in unstimulated and oestrogen-dominated lung extracts and in oestrogen-dominated ovary extracts. Several minor proteins were identified in heparin-agarose purified endometrium that may also play a role in the UGPB complex.