Early in an immune response, the predominant immunoglobulin heavy chain produced is mu, later in the immune response the predominant immunoglobulin heavy chain is gamma, alpha, or epsilon. This change in heavy chain usage is accomplished by a DNA rearrangement that moves the heavy chain variable region gene from the mu constant region to the alpha constant region, epsilon constant region, or to one of the four gamma gamma1, gamma2b, gamma2a, or gamma3 in mouse) constant regions. This DNA rearrangement involves segments of DNA (called switch segments) upstream of each constant region gene. Immunoglobulin producing cells have a choice of six different switch rearrangements. This choice is an important one; the seven different heavy chain constant regions mediate very different effector functions and so handle an infectious organism or its toxic products in very different ways. Previous results have demonstrated that this choice is not random in the immunoglobulin producing cell, but rather is regulated. Furthermore, it seems likely that the regulation is mediated by changes in the local structure of chromatin of specific switch regions. The overall goal of the proposed experiments is to understand the molecular mechanisms by which switch recombination is regulated in a gene specific way. We plan to identify nuclear proteins that might control heavy chain gene-specific rearrangement and to identify the DNA sequences to which they bind. The candidate regulatory DNA sequences will be tested for their ability to alter chromatin/DNA structure by measuring changes in transcription mediated by the DNA sequences. The DNA sequences will be tested by transient transfection of cells that produce and do not produce immunoglobulin, and by transient transfection of cells that will be treated in various ways to induce gene-specific recombination. We will test the ability of the same sequences to regulate transcription from DNA molecules inserted into transgenic mice. Finally, we will attempt to test the ability of the sequences to regulate switch recombination from DNA molecules introduced into transgenic mice. We will study two murine heavy chain genes, gamma1 and gamma2a, and two inducers of switch recombination, IL4 for gamma1 and interferon-gamma for gamma2a.
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