Orientation and positional mapping of the subunits of the multicomponent transcription factors RFX and X2BP to the major histocompatibility complex class II transcriptional enhancer.

Major histocompatibility complex class II genes contain a common complex enhancer that allows for their coordinate regulation. The X box element of the enhancer cooperatively binds the multisubunit transcription factors RFX and X2BP. RFX is an essential class II transcription factor and contains three distinct proteins: RFX5, RFX-B/Ank and RFXAP. X2BP, a CREB/ATF family transcription factor, most likely binds as a homodimer. A site-specific protein-DNA photocrosslinking assay was used to investigate the interactions of the subunits of RFX and X2BP with X box DNA. Two of the RFX subunits, RFX5 and RFX-B/Ank, were found to bind defined sites within the X1 half of the X box. The third RFX subunit, RFXAP, made extensive X1 box contacts. The subunits of X2BP made contacts with the edges of the X2 half of the X box in a manner consistent with other bZIP transcription factor contact patterns. The resulting map provides specific base pair contacts and subunit orientation with respect to the DNA sequence of the RFX-X2BP-X box complex. Our results suggest possible stoichiometry of the RFX subunits and potential interaction between RFX-B/Ank and RFXAP with one of the subunits of X2BP.