Functional analysis of the human RAG 2 promoter

Recombination activating genes RAG1 and RAG2 are essential components of V(D)J recombination, a process that generates the specific antigen receptors in lymphocytes. To understand the mechanisms underlying the lineage and developmental regulation of transcription of RAG2, we have characterized the human RAG2 exon 1A promoter. In this study, a series of deletion constructs were used to isolate the promoter while a linker scanning approach was taken to assess functionally relevant cis elements within the promoter. Two regulatory domains were identified. The -140 to -123 region is critical for promoter activity in all cell lines tested. Mutations to the putative Ets (-122 to -118) or to the C/EBP (-137 to -129) consensus core sequences did abrogate promoter activity, although specific DNA/protein interactions remained, as determined by EMSA. The -69 to -48 region demonstrates lineage specific promoter activity. Mutations to an overlapping, BSAP-myb-Ikaros-myb site (-65 to -39) resulted in differential promoter activity in human B and T cells. EMSA analysis of this region showed a B cell specific protein complex. Transfection of BSAP into cell lines trans-activates the human RAG2 promoter. We conclude that transcriptional regulation of the human RAG2 gene is complex, involving both tissue specific and ubiquitous factors, and both proximal and distal regulatory elements.