RNA-Seq analysis of murine splenic B cells with deletion of YY1

X chromosome inactivation (XCI) in female lymphocytes is uniquely regulated, as the inactive X (Xi) chromosome lacks localized Xist RNA and heterochromatin modifications. Epigenetic profiling reveals that Xist RNA is lost from the Xi at the pro-B cell stage and that additional heterochromatic modifications are gradually lost during B cell development. Activation of mature B cells restores Xist RNA and heterochromatin to the Xi in a dynamic two-step process that differs in timing and pattern, depending on the method of B cell stimulation. Finally, we find that DNA binding by YY1 maintains XCI in activated B cells, as ex-vivo YY1 deletion results in loss of Xi heterochromatin marks and up-regulation of X-linked genes. Ectopic expression of the YY1 zinc finger domain is sufficient for Xist RNA localization during B cell activation. Together, our results indicate that Xist RNA localization is critical for maintaining XCI in female lymphocytes, and that chromatin changes on the Xi during B cell development and the dynamic nature of YY1-dependent XCI maintenance in mature B cells predisposes X- linked immunity genes to reactivation. Overall design: Follicular B cells were isolated from the spleens of six (three males and three females) C57Bl/6 yy1 fl/fl mice. For each spleen, 1/3 of the cells were not treated with any reagent (naive cells), 1/3 the cells received mock treatment (mock cells) and 1/3 received TATCRE (tatcre cells). The mock and tatcre samples were then grown in RPMI medium along with anti-IgM and 20% FBS for 48 hours. All cells were lysed and RNA was isolated for library preparation. Expression differences between Mock and TATCRE treated cells were determined to understand the role of yy1 in B cell activation.