x chromosome inactivation and differentiation occur readily in es cells doubly-deficient for macroh2a1 and macroh2a2x染色体失活和分化容易发生在胚胎干细胞doubly-deficient macroh2a1和macroh2a2.pdfVIP

X Chromosome Inactivation and Differentiation Occur Readily in ES Cells Doubly-Deficient for MacroH2A1 and MacroH2A2 Borko Tanasijevic1,2, Theodore P. Rasmussen1,2,3* 1 Center for Regenerative Biology, University of Connecticut, Storrs, Connecticut, United States of America, 2 Department of Molecular and Cell Biology, University of Connecticut, Storrs, Connecticut, United States of America, 3 Department of Pharmaceutical Sciences, University of Connecticut, Storrs, Connecticut, United States of America Abstract Macrohistones (mH2As) are unusual histone variants found exclusively in vertebrate chromatin. In mice, the H2afy gene encodes two splice variants, mH2A1.1 and mH2A1.2 and a second gene, H2afy2, encodes an additional mH2A2 protein. Both mH2A isoforms have been found enriched on the inactive X chromosome (Xi) in differentiated mammalian female cells, and are incorporated into the chromatin of developmentally-regulated genes. To investigate the functional significance of mH2A isoforms for X chromosome inactivation (XCI), we produced male and female embryonic stem cell (ESC) lines with stably-integrated shRNA constructs that simultaneously target both mH2A1 and mH2A2. Surprisingly, we find that female ESCs deficient for both mH2A1 and mH2A2 readily execute and maintain XCI upon differentiation. Furthermore, male and female mH2A-deficient ESCs proliferate normally under pluripotency culture conditions, and respond to several standard differentiation procedures efficiently. Our results show that XCI can readily proceed with substantially reduced total mH2A content. Citation: Tanasijevic B, Rasmussen TP (2011) X Chromosome Inactivation and Differentiation Occur Readily in ES Cells Doubly-Deficient for MacroH2A1 and MacroH2A2. PLoS ONE 6(6): e21512. doi:10.1371/journal.pone.0021512 Editor: Brian P. Chadwick, Florida State Unive