Genomic Environment Predicts Expression Patterns on the Human Inactive X Chromosome

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Abstract
What genomic landmarks render most genes silent while leaving others expressed on the inactive X chromosome in mammalian females? To date, signals determining expression status of genes on the inactive X remain enigmatic despite the availability of complete genomic sequences. Long interspersed repeats (L1s), particularly abundant on the X, are hypothesized to spread the inactivation signal and are enriched in the vicinity of inactive genes. However, both L1s and inactive genes are also more prevalent in ancient evolutionary strata. Did L1s accumulate there because of their role in inactivation or simply because they spent more time on the rarely recombining X? Here we utilize an experimentally derived inactivation profile of the entire human X chromosome to uncover sequences important for its inactivation, and to predict expression status of individual genes. Focusing on Xp22, where both inactive and active genes reside within evolutionarily young strata, we compare neighborhoods of genes with different inactivation states to identify enriched oligomers. Occurrences of such oligomers are then used as features to train a linear discriminant analysis classifier. Remarkably, expression status is correctly predicted for 84% and 91% of active and inactive genes, respectively, on the entire X, suggesting that oligomers enriched in Xp22 capture most of the genomic signal determining inactivation. To our surprise, the majority of oligomers associated with inactivated genes fall within L1 elements, even though L1 frequency in Xp22 is low. Moreover, these oligomers are enriched in parts of L1 sequences that are usually underrepresented in the genome. Thus, our results strongly support the role of L1s in X inactivation, yet indicate that a chromatin microenvironment composed of multiple genomic sequence elements determines expression status of X chromosome genes. To match the amount of gene product produced in males (XY), most genes in mammalian females (XX) are active on one X chromosome and inactivated on the other. However, some genes “escape” inactivation and are expressed from both X chromosomes. This study investigates sequences that may control whether a gene undergoes or escapes X chromosome inactivation, including DNA sequences previously thought of as non-functional or “junk.” Earlier work suggested that one such sequence, L1 interspersed repeats, may be associated with inactivation, but the extent of such association, and whether it represented a consequence of the evolutionary history of X, remained unclear. This study utilized recently generated chromosome-wide data on sequence and gene expression for human X, with a particular focus on the Xp22 region, which is evolutionarily young and has had no time to accumulate many L1 elements. A rigorous statistical analysis identified with high accuracy a set of short sequences that discriminate between genes undergoing and those escaping X chromosome inactivation. Interestingly, the majority of such sequences enriched in the vicinity of inactivated genes were found within L1s. These results strengthen the case for an involvement of L1s in X chromosome inactivation and suggest other DNA elements that might also play a role.