Although the existence of CpG islands has been known for over two decades, the mechanisms through which they contribute to genome function have remained poorly understood. Here we provide compelling evidence that CpG islands are actively recognized by KDM2A binding to nonmethylated CpG DNA through a ZF-CxxC domain (). Using ChIP-seq we show that KDM2A binds to 90% CpG island elements genome-wide and that this nucleation event is independent of the transcriptional status of the associated gene (). We demonstrate that KDM2A occupancy at CpG islands imposes a unique H3K36me2-depleted chromatin signature that differentiates these regulatory elements from bulk chromatin, and we provide functional data demonstrating that depletion of KDM2A results in H3K36me2 spreading into CpG island elements (). Together these observations provide insight into how CpG islands can impact chromatin architecture at regulatory elements by utilizing their underlying DNA sequence as a nucleation site for a chromatin-modifying enzyme (). Therefore, these data suggest a central function of CpG island DNA may be to impose a chromatin architecture that differentiates CpG island chromatin from bulk chromatin, highlighting these important regulatory regions within large and complex mammalian genomes. In support of this contention, CpG island chromatin is also enriched in H3K4me3 regardless of whether the associated gene is expressed (Bernstein et al., 2006; Guenther et al., 2007
). Intriguingly, Set1 and Mll H3K4 methyltransferase complexes that trigger this modification also have ZF-CxxC domains (Birke et al., 2002; Lee and Skalnik, 2005
), suggesting they may use a similar mechanism as KDM2A to target CpG islands (). This possibility is supported by evidence that the ZF-CxxC domain-containing protein, CFP1, which is a component of the Set1 H3K4 methyltransferase complex, is targeted to CpG island elements genome-wide, where it deposits H3K4me3 methylation in a manner that is independent of the transcriptional state of the associated gene (A. Bird, personal communication). These observations regarding CFP1 nucleation and H3K4me3 perfectly mirror our discovery that KDM2A binds CpG islands, resulting in enzymatic depletion of H3K36me2 independently of the transcriptional state of the associated gene. Together these data indicate that ZF-CxxC domain-containing proteins are important mediators of CpG island chromatin architecture and highlight the fact that CpG islands as a DNA-encoded genetic element are functioning to directly impact the epigenetic state of surrounding chromatin.
Unique Chromatin Architecture at CpG Islands
Interestingly, the processes that impose a unique chromatin environment at CpG islands are presumably even more complex. For example, from biochemical assays it is known that CpG island chromatin is specifically depleted of linker histone H1 (Tazi and Bird, 1990
) (). Given that CpG island chromatin architecture appears to form irrespective of transcriptional state, an interesting question is what impact this chromatin environment has on the function of associated genes. It is known that H3K36me2 and H1 can have an inhibitory effect on transcription initiation (Carrozza et al., 2005; Cheung et al., 2002; Levine et al., 1993; Li et al., 2009; Strahl et al., 2002; Youdell et al., 2008
), and H3K4me3 is generally associated with processes positively contributing to transcription (Kouzarides, 2007
). Therefore, one possibility is that unique CpG island chromatin architecture reinforced by several intersecting chromatin-regulating pathways may define regions of the genome that are more permissive to nucleation of the transcriptional machinery, effectively differentiating CpG islands from bulk chromatin and highlighting regulatory regions of the genome. The inherent complexity of the CpG chromatin signature makes this a difficult hypothesis to directly examine experimentally, but this concept is indirectly supported by the observation in genome-wide run-on transcription assays that CpG island promoters sustain nonproductive transcriptional initiation events in both sense and antisense directions even in the absence of activated directional transcription, whereas non-CpG island genes fail to show this property (Core et al., 2008; Seila et al., 2008
). An important implication of this hypothesis is that CpG island chromatin architecture would provide an environment that is permissive to transcription but not drive productive directional transcriptional output, a process that requires transcription factor binding and concerted gene activation mechanisms. This type of CpG island-specific transcriptional competence has recently been shown to contribute to the induction kinetics of CpG island-containing genes in activated macrophages (Ramirez-Carrozzi et al., 2009
In conclusion, we have provided a mechanistic link between CpG island elements and nucleation of a histone demethylase and demonstrated that CpG islands function to define cellular chromatin landscapes at these regulatory elements. A corollary of this observation from a genome evolution standpoint is that ZF-CxxC domain recognition of CpG island elements may also impose an additional selective pressure over evolutionary time to maintain the nonmethylated state of CpG island elements. Based on our understanding of CpG island function presented here, a future challenge will be to understand if chromatin modifications at CpG islands impact the transcriptional machinery and how non-CpG island promoters differ in the absence of these mechanisms.