Experiments are planned for a detailed analysis of the regulation of the rearrangement of immunoglobulin genes. Previous work with transgenic mice has shown that after rearrangement of a functional heavy and light chain gene a feedback mechanism shuts off the further production of the Ig gene specific recombinase. 1) It will now be determined whether the same mechanism operates in all types of B cells, or whether two different B cell lineages exist with respect to control mechanisms at the pre-B cell stage. 2) The role of gamma 2b in feedback inhibition has been controversial, apparently because the timing and/or quantities and/or membrane versus secreted forms of transgenic gamma 2b varied in different transgenic lineages. The different transgenic gamma 2b lines will now be exploited to shed more light on these parameters of the feedback control. 3) The isotypic exclusion of kappa and lambda genes will be further studied by determining the location of the lambda gene enhancer(s) and then producing transgenic mice with a gene under control of its own enhancer. 4) In order to define the molecular composition of the signal for feedback inhibition, recombinase positive pre-cell lines will be transfected with modified immunoglobulin genes and their effect on the recombinase will be determined. 5) Individual pre-cells from Ig gene transgenic mice will be analyzed for the timing of the expression of transgenic and endogenous Ig genes using the polymerase chain reaction. This may help unravel the kinetics of the feedback response. 6) By targeting the lambda 5 or C mu gene in embryonic stem cells with mutated gene constructs, mice will be produced which lack lambda 5 or mu production in order to study the role of these genes in early B cell development. It is hoped that these experiments may help in defining the control of immunoglobulin gene rearrangement and thus solve a well-known example of a developmental control, as well as be of clinical importance for immunological disorders with a B cell component.
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