The major goal of this grant and its predecessor is to study normal B lymphocyte development in the context of activation and expression of lymphocyte-specific genes. In the first grant the major focus was the rearrangement of immunoglobulin heavy and light chain genes during normal B cell development. This study included the isolation and sequencing of the light chain VK24 variable region gene family to determine their evolutionary relationship. In this proposal we expand from a study of immunoglobulin gene expression in normal lymphocytes to include the analysis of other B cell expressed genes. Using the techniques lineage tracing the growth and differentiation of pre-B cells shall be defined. At each stage of antigen-independent and antigen-dependent B cell differentiation the molecular genetic events relating to the heavy chain class switch shall be monitored on the expressed and on the nonexpressed heavy chain chromosome. This will require an analysis of these heavy chain arrangements in 1) three different stages of pre-B cell development; 2) in virgin BMu and mature B MuDelta cells; 3) in antigen-activated germinal center B cells expressing IgG or IgA receptor immunoglobulins of either one parental allotype or the other; and 4) in plasma cells. The lymphocyte differentiation antigens which are expressed in pre-B cell development (TdT, B220), in subsets of resting mature B cells (Ly-1, B220), and in antigen activated germinal center cells (transferrin receptor) are the primary objects of a new cDNA cloning strategy developed in this laboratory. The eventual objective of this cloning strategy is to characterize each of these genes in order to examine at the DNA sequence level whether particular shared sequences might be involved in their selective expression in B cells. Finally, a complete study of VK24 (antiphosphorycholine and antistreptococcyl A carbohydrate) light chain variable region genes will be carried out to understand their mechanism of evolution and their current status in mouse and man. The studies on normal pre-B cell and B cell differentiation should highlight those potential defects which may lead to B cell immunodeficiency syndromes. A study of the role of terminal transferase in providing immunoglobulin gene diversification during B cell differentiation will use as a model a mouse genetic defect (motheaten) whose hallmarks are immunodeficiency and autoimmunity. The studies on the immunoglobulin class switch mechanism should reveal important information for the eventual study of Ataxia Telangiectasia and selective immunoglobulin deficiency diseases. The analysis of the V kappa 24 family might reveal a basis for inordinate susceptibility to bacterial infection in certain humans.
