FUNCTIONAL COMPLEMENTATION OF MAJOR HISTOCOMPATIBILITY COMPLEX CLASS-II REGULATORY MUTANTS BY THE PURIFIED X-BOX-BINDING PROTEIN RFX

Major histocompatibility complex (MHC) class II deficiency, or bare ly mphocyte syndrome (BLS), is a disease of gene regulation. Patients wit h BLS have been classified into at least three complementation groups (A, B, and C) believed to correspond to three distinct MHC class II re gulatory genes. The elucidation of the molecular basis for this diseas e will thus clarify the mechanisms controlling the complex regulation of MHC class II genes. Complementation groups B and C are characterize d by a lack of binding of RFX, a nuclear protein that normally binds s pecifically to the X box cis-acting element present in the promoters o f all MHC class II genes. We have now purified RFX to near homogeneity by affinity chromatography. Using an in vitro transcription system ba sed on the HLA-DRA promoter, we show here that extracts from RFX-defic ient cells from patients with BLS (BLS cells) in groups B and C, which are transcriptionally inactive in this assay, can be complemented to full transcriptional activity by the purified RFX. As expected, purifi ed RFX also restores a completely normal pattern of X box-binding comp lexes in these mutant extracts. This provides the first direct functio nal evidence that RFX is an activator of MHC class II gene transcripti on and that its absence is indeed responsible for the regulatory defec t in MHC class II gene expression id patients with BLS.