hBRAG, a novel B cell lineage cDNA encoding a type II transmembrane glycoprotein potentially involved in the regulation of recombination activating gene 1 (RAG1).
The different display reverse transcription-PCR (DD RT-PCR) technique was used to identify novel cDNA detecting mRNA transcripts co-expressed with human recombination activating gene-1 (RAG1). A 5.0-kb transcript detected by the differential display amplicon 3G1 was found to correlate strongly with RAG1 mRNA expression in various human cell lines. Subsequent screenings of a pre-B cDNA library with 3G1 led to the identification of a complete cDNA we have termed hBRAG (human B-cell RAG-Associated Gene). The hBRAG cDNA encodes a 503-amino acid (aa) protein with no known homology to any nucleotide or protein sequence. The predicted molecular mass of 55 kDa was confirmed by in vitro translation. Based on sequence analysis, the predicted open reading frame encodes for a type II transmembrane spanning glycoprotein with the N-terminal 81 -aa in the cytoplasm, a 17-aa transmembrane domain, and a C-terminal 405-aa extracellular domain with four potential N-glycosylation sites. Northern blot analysis indicated a close association of the 5.0-kb hBRAG mRNA transcript with RAG1 in numerous human pro-B, pre-B and mature B cell lines assessed, but not in human T cell lines. In human tissues, hBRAG is expressed at highest levels in B cell-enriched tissues, but is not expressed in fetal or adult thymus. Southern blotting analysis revealed that this gene is conserved across eukaryotes, is expressed as a single copy in the human genome, and is likely not a multigene family member. The hBRAG gene was localized to the long arm of chromosome 10 (10q26). Transfection of the full-length hBRAG cDNA increased levels of human RAG1 transcripts in the B cell line OCI LY8-C3P, but not in the non-lymphoid line K562, suggesting a B cell-specific role for the hBRAG product in regulating RAG expression.
Verkoczy, LK; Marsden, PA; Berinstein, NL
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