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To haplotype the X chromosome of two additional samples (MB59 and GBM103), we performed ChIP-seq for histone marks H3K36me3 and macroH2A1. Histone variant macroH2A1 is known to show an ?1.5-fold uniform Ribonucleotide reductase enrichment along the inactive X chromosome (Mietton et?al., 2009). In contrast, H3K36me3 is enriched in actively transcribed regions and therefore on Xa. Thus, by sequencing both histone marks to sufficiently high depth to infer allele frequencies of mutations, a mutation on Xi should have a high allele frequency in the macroH2A1 reads and a low allele frequency in H3K36me3, which we clearly observed for the germline mutations (data not shown). Integrating matching RNA-seq data with mutations showing particularly high/low macroH2A1/H3K36me3 allele frequencies (Extended Experimental Procedures), 31 SNVs were haplotyped in glioblastoma GBM103, with only 3 SNVs locating to Xa (p?CH5424802 female patients in which the inactive X?chromosome was present, and haplotyping of four individual samples clearly indicates that X hypermutation is confined to the inactive X chromosome. To assess whether X chromosome hypermutation is a general feature of multiple tumor types, we analyzed the somatic mutation rate of the X chromosome in an additional ?300 whole-cancer genomes, including our own published and unpublished data from five different cancer entities (pilocytic VX-770 astrocytoma, glioblastoma, ependymoma, B cell lymphoma, and prostate carcinoma) complemented by published mutation call sets of six different cancer types: breast cancer (Nik-Zainal et?al., 2012), neuroblastoma (Molenaar et?al., 2012), chronic lymphocytic leukemia (Puente et?al., 2011), acute myeloid leukemia (Welch et?al., 2012), colorectal carcinoma (Bass et?al., 2011), and retinoblastoma (Zhang et?al., 2012). We observed X chromosome hypermutation in a significant fraction of cancer genomes from female samples (56/191, 29%) comprising nine different cancer entities across a diverse set of childhood tumors as well as adult solid and hematopoietic malignancies (Tables 1 and S1 and Figure?4).
