Binding of transcription factors of the CREB/ATF family to the CRE-like sequence in the Fuc-T VII promoter.

Electrophoretic mobility shift assays were conducted with Jurkat T cell nuclear extracts, as described in “Materials and methods.” (A) A 27-bp oligonucleotide containing the CRE-like sequence was used as a probe in the absence and presence of competitor oligonucleotides. Complexes were resolved on a 4% nondenaturing gel. Sequence-specific binding species obtained were competed with a 20- to 50-fold molar excess of the cold competitor oligonucleotide. DNA–protein complexes are numbered. (B) The probe was incubated with specific antibodies against ATF1, ATF3, ATF4 (CREB-2), or CREB-1. Anti-Sp1 antibody was used as a negative control. Specific DNA–protein complexes are indicated by roman numerals. NS indicates nonspecific complex; SS, supershifted band; and CRE-con, CRE-consensus.