X chromosome dosage compensation and the regulation of the feto-maternal interface

PROJECT SUMMARY The goal of this proposal is to investigate how gene expression dosage on the X chromosomes is regulated during embryonic and extraembryonic development and how its deregulation impairs the feto-maternal interface within the placenta. X-chromosome dosage compensation in female placental mammals occurs through two

epigenetic processes: X-chromosome inactivation (XCI), which induces the inactive X chromosome (Xi), and X- chromosome upregulation (XCU), which increases the expression of the single active X chromosome (Xa) to match the gene dosage of autosomes. XCI happens twice in mouse development. First, an imprinted mechanism

silences the paternal X (imprinted XCI, iXCI) in pre-implantation embryos. iXCI is reversed in epiblast cells of the blastocyst and induced again on either the maternal or paternal X in embryonic cells upon implantation (random XCI, rXCI); in contrast, extraembryonic cells, which form the placenta, maintain iXCI. XCU accompanies both

iXCI and rXCI. XCI mechanisms are well-understood, but those governing XCU remain largely elusive. The long non-coding RNA Xist is the master regulator of both forms of XCI. Conserved repeats of Xist, termed A-F, are critical for Xist’s function and recruit specific proteins to block transcription and induce a heterochromatin state.

For instance, the B-repeat recruits repressive proteins including polycomb complexes and is required for heterochromatin formation and Xi compaction. We showed that without the B-repeat, silencing of most genes on the Xi is impaired in vitro. To investigate the functional consequences of increased gene expression from the Xi,

we created mice with a B-repeat deletion (ΔB-Xist). Our preliminary data show that the ΔB-Xi disrupts the development of the placenta of female embryos. Female ∆B-Xi placentas have severe depletion of the junctional zone (JZ) and limited trophoblast invasion of blood vessels, causing female-specific embryo mortality and growth

retardation. In the ΔB-Xist mouse model, developmental defects only manifest in the placenta, even though gene expression on the ΔB-Xi is increased in placental and embryonic tissues. Our data suggest that Xa expression is typically decreased in cells with the ΔB-Xi and that this does not happen in the JZ, suggesting that XCU

regulation varies in different cell types and providing a possible explanation for the JZ-specific sensitivity to X- chromosome dosage. Here, we will follow up on these exciting findings by further defining the role of the Xist B- repeat in the placenta (Aim1); exploring the chromatin state on the wildtype and ΔB-Xi and the crosstalk between

the Xi and Xa in embryonic and extraembryonic tissues (Aim 2); and uncovering mechanisms underlying JZ dysplasia in placentas of mouse interspecies hybrids (Aim 3). Overall, our work will elucidate the role of XCI and XCU in the regulation of the feto-maternal interface and explain the high X-dosage sensitivity of the JZ.