X chromosome inactivation: Activation of Silencing

X chromosome inactivation is a process that ensures equal expression of the X chromosomes between males, which have one X and one Y chromosome, and females, which have two X chromosomes, in mammals. Females initiate inactivation of one of their two X chromosomes early during embryogenesis, by expressing an untranslated RNA from the X-encoded Xist gene. Xist RNA coats and silences the X chromosome in cis, after which the silenced state of the inactive X chromosome is propagated through many cell divisions. Three major research questions dominate the field of XCI. How is a cell able to count the number of X chromosomes present? How is it possible that only one of the two X chromosomes in female diploid cells is inactivated? And how is the X chromosome silenced in cis? Although many researchers have attempted to explain the initiation and regulation of the XCI process with a wide array of models, none of the proposed models is able to explain all observations made with deletion and overexpression studies in mouse ES cell lines and embryos. Therefore, the aim of this thesis is to obtain additional data, and to postulate a comprehensive model for the initiation of XCI in the mouse. This model will encompass how the cell is able to detect the number of X chromosomes and how it can inactivate only one of them without inactivating all X chromosomes present in a cell. The discovery of an important activator of XCI initiation strengthens this model. Furthermore, the mechanism of spreading of Xist RNA, and thus the silencing of the X chromosome in cis, will be addressed. A hypothesis regarding the spreading of Xist RNA over the inactive X, and how Xist RNA is restricted to the Xi, will be formulated. Overall, the most important conundrums in XCI will be addressed in this thesis, and we present a more logical and complete model explaining the regulation of XCI.