Previous work (6-10) has shown that allelic exclusion of Ig gene expression is controlled by functionally rearranged mu and kappa genes. This report deals with the comparison of membrane mu (micron) and secreted mu (microsecond) in promoting such feedback inhibition. Splenic B cell hybridomas were analyzed from transgenic mice harboring a rearranged kappa gene alone or in combination with either an intact rearranged mu gene or a truncated version of the mu gene. The intact mu gene is capable of producing both membrane and secreted forms of the protein, while the truncated version can only encode the secreted form. The role of the microsecond was also tested in pre-B cell lines. Analysis of the extent of endogenous Ig gene rearrangement revealed that (a) the production of micron together with kappa can terminate Ig gene rearrangement; (b) microsecond with kappa does not have this feedback effect; (c) microsecond may interfere with the effect of micron and kappa; and (d) the feedback shown here probably represents a complete shutoff of the specific recombinase by micron + kappa; the data do not address the question of mu alone affecting the accessibility of H genes for rearrangement.

Transgenic mice were produced that carried in their germlines rearranged kappa and/or mu genes with V kappa and VH regions from the myeloma MOPC-167 kappa and H genes, which encode anti-PC antibody. The mu genes contain either a complete gene, including the membrane terminus (mu genes), or genes in which this terminus is deleted and only the secreted terminus remains (mu delta mem genes). The mu gene without membrane terminus is expressed at as high a level as the mu gene with the complete 3' end, suggesting that this terminus is not required for chromatin activation of the mu locus or for stability of the mRNA. The transgenes are expressed only in lymphoid organs. In contrast to our previous studies with MOPC-21 kappa transgenic mice, the mu transgene is transcribed in T lymphocytes as well as B lymphocytes. Thymocytes from mu and kappa mu transgenic mice display elevated levels of M-167 mu RNA and do not show elevated levels of kappa RNA, even though higher than normal levels of M-167 kappa RNA are detected in the spleen of these mice. Approximately 60% of thymocytes of mu transgenic mice produce cytoplasmic mu protein. However, despite a large amount of mu RNA of the membrane form, mu protein cannot be detected on the surface of T cells, perhaps because it cannot associate with T cell receptor alpha or beta chains. Mice with the complete mu transgene produce not only the mu transgenic mRNA but also considerably increased amounts of kappa RNA encoded by endogenous MOPC-167 like kappa genes. This suggests that B cells are selected by antigen (PC) if they coexpress the mu transgene and appropriate anti-PC endogenous kappa genes. Mice with the mu delta mem gene, however, do not express detectable levels of the endogenous MOPC-167 kappa mRNA. Like the complete mu transgene, the M-167 kappa transgene also causes amplification of endogenous MOPC-167 related immunoglobulins; mice with the kappa transgene have increased amounts of endogenous MOPC-167-like mu or alpha or gamma in the spleen, all of the secreted form. Implications for the regulation of immunoglobulin gene expression and B cell triggering are discussed.

SummaryTransgenic mice with a y2b transgene were produced to investigate whether y2b can replace P in the development of B lymphocytes . Transgenic y2b is present on the surface of B cells. Young transgenic mice have a dramatic decrease in B cell numbers, however, older mice have almost normal B cell numbers . Strikingly, all y2b-expressing B cells in the spleen also express h . The same is true for mice with a hybrid transgene in which the A. transmembrane and intracytoplasmic sequences replace those of y2b (72b-,umem) . The B cell defect is not due to toxicity of y2b since crosses between y2b transgenic and A transgenic mice have normal numbers of B cells. Presence of the y2b transgene strongly enhances the feedback inhibition of endogenous heavy chain gene rearrangement. Light chain genes are expressed normally, and the early expression of transgenic light chains does not improve B cell maturation. When the endogenous A locus is inactivated, B cells do not develop at all in y2b transgenic mice. The data suggest that y2b cannot replace A in promoting the developmental maturation of B cells, but that it can cause feedback inhibition ofheavy chain gene rearrangement . Thus, the signals for heavy chain feedback and B cell maturation appear to be different .

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Immunoglobulin (Ig) genes are expressed sequentially (first H-, then L-chain genes) during the development of B lymphocytes. These studies, performed with transgenic mice and transfected cells, were aimed at the regulation of turning on and off the rearrangement of Ig genes. The specific recombinase is active in pre-B cells, but not in plasma cells. Production of membrane mu, but not secreted mu or gamma-2b, turns off rearrangement of H genes. Feedback inhibition of kappa-gene rearrangement requires kappa and membrane mu. Kappa alone or in combination with secreted mu does not stop recombination. Mouse lambda genes were mapped by deletion analysis and pulsed-field gel electrophoresis. The gene order is V2-C2,4-V1-C3,C1. The distance between V2 and C2 is 74 kb, but that between V1 and C3, 1 is only 20 kb. V2 and C3, 1 are over 190 kb apart. Lambda genes appear to be rearranged in a subset of B cells that do not respond to feedback inhibition at the pre-B cell stage. Lambda and kappa genes are both rearranged and potentially functional in these cells. Kappa genes may then be deleted by recombination of a sequence (described by Selsing and Siminovitch et al.) downstream of C-kappa with sequences upstream of C-kappa. Presumably the recombinase is eventually inactivated in kappa-lambda cells by a mechanism that is different from H-kappa feedback.