MANIPULATING MHC GENE EXPRESSION IN ES CELL CHIMERAS

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The broad objective of this research is to analyse developmentally regulated expression of Class I products of the Major Histocompatability Complex in the mouse. In particular, the possibility that control of MHG gene expression may play a key role in maternal tolerance of the fetal allograft will be examined. We have designed a strategy that will allow us to study the consequences of ectopic expression of H-2D delta transplantation antigens in the developing embryo. A construct carrying the human Beta-actin gene promoter fused to the coding regions of the H-2D delta gene has been introduced into pluripotent embryonic stem (ES) cells. Although H-2D delta protein was not expressed at the cell surface in undifferentiated ES cells, significant amounts of H-2D delta membrane glycoproteins are detected on ES cells induced to differentiate in vitro. A major aim of the proposed investigation is to determine whether these transfected ES cell lines can contribute to all the somatic tissues when incorporated into normal embryos. Should our efforts to obtain live born chimeras be successful, the next goal will be to determine whether the introduced H-2D Delta gene can be transmitted in the germ line. Should transgenic chimeras fail to develop to term, we will describe the developmental failure and determine whether the abnormal phenotype (s) is attributable to defects in specific cell lineages. The spatial and temporal expression of Class I antigens in the developing embryos will be examined using immunohistochemical and in situ hybridization techniques. We will investigate the possibility that ectopic MHC gene expression in the extra- embryonic lineages may trigger a maternal immune response directed against the fetal allograft. A particularly important point will be to test whether maternal lymphocytes infiltrate and disrupt the abnormal embryos. These experiments will hopefully lead to a clearer understanding regulation of MHC gene expression during early mouse development and processes involved in maternal-fetal interactions.