Cell type-specific genome packaging defines the epigenome set during cellular differentiation. Epigenetic marks have been shown to specify the patterns of gene expression and a number of whole-genome epigenetic mapping studies of human cells have been done in the recent past to explore the functional states across genome (2
). None of these, however, have explored the epigenetic signatures on the Y chromosome (1
), presumably because of the presence of many repeats and the instability of the chromosome in various cultured cell lines. In this study, we have analyzed chromatin from WBCs and designed Y chromosome specific probes from both unique and duplicated regions to create an epigenetic map of the human Y chromosome. Unique probes were specific to unique regions of the Y chromosome, while probes for multi-copy loci were designed from sequences that are duplicated but are unique to Y chromosome and regions having homologues on X chromosome.
The evolutionary origin of the sequences that constitute the Y chromosome is known and it has been observed that genes distributed in sequences of the same origin have a similar expression pattern (13
). We observed a global conservation of the epigenetic pattern associated with sequences of the same origin, which changed sharply at sequences of different origin. This implies that similar regulatory mechanisms might operate across genes that share a common origin and epigenetic profile, resulting in their similar expression pattern (35
). This also insinuates that these sequences have not diverged enough to have different functions after they became part of the Y chromosome. Surprisingly, even regions of internal duplication within the Y chromosome showed non-conflicting epigenetic marks in our analysis, suggesting that after duplication, the associated loci have maintained similar regulatory environments. For instance, all copies of the TSPY
genes appear to be silenced in the WBCs by mechanisms that involve methylation of H3K9 around their promoters and methylation of DNA at associated CpG islands. This locus has been linked to oncogenesis and loss of methylation at both DNA and histone has been observed at the promoters of oncogenes in many cancers. It is possible that in the case of gonadoblastoma, the GBY locus may be hypomethylated, resulting in overexpression of TSPY
). Therefore, it would be interesting to test the epigenetic marks associated with this region in cancer tissues to explore the epigenetic regulation of GBY
loci in different male-specific cancers.
Among the internally duplicated regions (palindromes) on the long arm, only DAZ
genes at the AZF locus showed conflicting patterns of histone marks, suggesting that in these clusters, different copies of the genes may be regulated differently. It is possible that in the case of genes with non-redundant functions, mechanisms have evolved for silencing some of the copies to enable normal functioning of the cell, as an alternative to maintaining all the copies in a repressed or expressed state. The non-conflicting histone patterns observed for the rest of the duplicated genes suggest the ability of the cell to function normally with multiple copies of these genes being expressed after the duplication event. In the absence of meiotic recombination, palindromes may make the Y chromosome unstable, but our observation of very similar epigenetic marks on duplicated regions in contrast to expected results (53
) indicates a regulatory mechanism at play that maintains the integrity of the Y chromosome in the absence of meiotic recombination and keeps global epigenetic regulation unchanged during evolution (16
In this study, we have used profiles of histone modifications and CTCF binding to explore and correlate the transcriptional potential of most of the genes of the Y chromosome. The association of CTCF with different marks of histone and DNA methylation in the case of SRY, TSPY
and tandem repeats suggests how epigenetic changes may lead to chromatin-mediated regulation of different coding and non-coding components of the genome. Interestingly, non-overlapping enrichment of H3K9me3 and DNA methylation at CpG islands of SRY
and overlapping enrichment of the two at TSPY
locus suggests that same epigenetic tools can be used differently at different loci. This also suggests the differential regulation of H3K9me3 modification and CTCF binding at each locus by other locus-specific factors. As observed in genic regions, we found distinct patterns of histone modifications and CTCF binding associated with different tandem repeats as well. Our data indicates that tandem repeats are capable of recruiting different kinds of chromatin modifiers, irrespective of their position, by virtue of their association with specific histone marks and this may play a role in setting the functional state of the chromatin. This suggests a non-neutral role for tandem repeats in recruiting various epigenetic regulatory factors. Since tandem repeats are known to be associated with almost every component of the genome, their epigenetic regulation can govern a large number of genomic loci by influencing many important cell processes like genome organization, replication and transcription. As CTCF is a transcriptional repressor and also binds with chromatin insulators (44
), in these cases it may coordinate transcription of genes through higher order chromatin-mediated regulation.
This analysis also provides a way to study the epigenetic profile associated with multi-copy genes as a representation of the average pattern across all the copies. Analysis of ChIP data from probes representing repeated regions carries a risk of misinterpretation, however a cautious approach can yield new insights into the possible mechanisms of global epigenetic regulation. When duplicated loci carry the same epigenetic marks, the epigenetic pattern is observed to be non-conflicting with a clear association with specific marks, whereas in cases where the duplicated genes have diverged significantly and have distinct epigenetic marks, conflicting histone modifications can be expected. This strategy is, therefore, useful in identifying the diversion in epigenetic regulation at duplicated loci. As our understanding of histone marks and their functional meaning increases, it is becoming more and more apparent that the functional status of a locus is decided by a complex histone code based on a combination of multiple histone modifications (3
). Further mapping of other known histone marks, histone variants and chromatin modifiers would, therefore, be extremely useful in understanding various Y chromosome associated diseases.