Several unique genetic recombination or mutation events occur during B cell differentiation. These include V-D-J joining, a site specific recombination event, immunoglobulin heavy chain (IgH) chain class switch and somatic mutation directed at variable (V) regions. The enzymology directing and controlling these events remain obscure. It is of considerable significance that we have detected a nuclease activity expressed at highly elevated levels in normal B lymphocytes. This nuclease activity is not detected in pre-B cells, thymocytes, mature T cells or HeLa cells. The spectrum of expression of this enzyme suggests that it is developmentally regulated. This nuclease, referred to here as B-cell associated nuclease (BCAN), has a 3'->5' exonuclease activity. Our studies demonstrate that BCAN is active on double stranded (ds) and single stranded (ss) DNA. Deoxynucleoside triphosphates, EDTA, poly dI-dC and glycerol are all inhibitory to enzymatic activity. BCAN requires divalent cations, Mg2+ or Mn2+ for activity. We have partially purified BCAN by column chromatography. Using this preparation, we have begun a rigorous analysis of its activity. Our objectives are to fully characterize the BCAN enzyme, purify it to homogeneity and raise a polyclonal antisera to it. Our long term goals are to clone the gene for BCAN, study its regulation in normal and malignant B cells and analyse its function in vivo. We hypothesize that BCAN is involved in the Ig switch recombination and/or somatic mutation leading to V gene diversity. Once the cDNA for BCAN is available we will begin to test these propositions by transfection experiments.
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