McCarrey 94-23850 Abstract In the embryo proper of eutherians X-chromosome inactivation is random with respect to whether the maternally- or paternally- derived chromosome becomes inactivated, whereas in marsupials it is exclusively the paternal X chromosome. However, in eutherians X chromosome inactivation is non-random in the extra-embryonic membranes and occurs earlier than in the embryo proper. A second difference is that whereas X chromosome inactivation is stable and can remain so for over 100 years, reactivation of the previously inactivated X chromosome occurs routinely with age in a tissue- specific manner in marsupials. These differences predict that much could be learned of the mechanism of X inactivation as well as the mechanism of maintenance of inactivation if the gene(s) involved could be compared in structure as well as function. A candidate gene has been isolated and sequenced from humans and mice, the Xist gene. Neither seems to encode a protein and both have high degrees of homology in the non-codine regions. However, the homology consist of tandem direct repeats which are not particularly useful for isolating homologues from other species. An attempt will be made to isolate the Xist homologue from the marsupial, Monodelphis domestica using PCR and the small regions of homology. Although this is not a routine cloning exercise and the risks are high, the isolation of the Xist homologue from a marsupial would make possible approaches leading to an understanding of one of the fundamental mechanisms of eukaryotic genetics. %%% The mouse Xist gene will be used in an attempt to isolate the marsupial homologue. The successful cloning and characterization of this gene will allow a comparative approach to understanding the differences and similarities between X chromosome inactivation in placental animals (eutharians) and non-placental animals (metherians). ***
