Identification of class II transcriptional activator-induced genes by representational difference analysis:: Discoordinate regulation of the DNα/DOβ heterodimer

Class II transcriptional activator (CIITA) is a master regulator of MHC class II genes, including DR, DP, and DQ, and MHC class II-associated genes DM and invariant chain. To determine the repertoire of genes that is regulated by CIITA and to identify uncharacterized CIITA-inducible genes, we used representational difference analysis. Representational difference analysis screens for differentially expressed transcripts, All CIITA-induced genes were MHC class II related. We have identified the alpha subunit, DN alpha, of the class II processing factor DO as an additional CIITA-inducible gene. Northern analysis confirmed that DN alpha is induced by IFN-gamma in 2fTGH fibrosarcoma cells, and CIITA is necessary for high-level expression in B cells. The beta subunit, DO beta, is not inducible in fibrosarcoma cells by IFN-gamma or exogenous CIITA expression. Moreover, in contrast to other class II genes, DO beta expression remains high in the absence of CIITA in B cells. The promoters for DN alpha and DO beta contain the highly conserved WXY motifs, and, like other class II genes, expression of both DN alpha and DO beta requires RFX. These findings demonstrate that both DN alpha and DO beta are regulated by RFX, However, DN alpha is defined for the first time as a CIITA-inducible gene, and DO beta as a MHC class II gene whose expression is independent of CIITA.