UCSF

The proper localization of chromatin modifications is critical to their function as signals for DNA compaction and transcription regulation. This localization is controlled by the activities of chromatin modifying enzymes, which “write” and “erase” chromatin modifications. While the chromatin field has identified the chromatin modifiers responsible for the deposition and removal of chromatin modifications, little is known about how the activities of chromatin modifiers are controlled to localize these modifications in the genome. In this thesis, we investigate the mechanisms that direct and control the activity of chromatin modifiers. We hypothesize that the identified mechanisms determine the localization of chromatin modifications and the biological roles of chromatin modifiers in the nucleus. To answer these questions we use a model family of chromatin modifiers, the human KDM4 histone demethylases, specifically KDM4C. Our approach is to reconstitute KDM4C and its substrates to probe the mechanisms which regulate demethylase activity. Moreover, we identify a transcriptional network reliant on KDM4C as a means to validate and discover novel regulatory mechanisms important to its role in controlling gene expression. In this thesis, we demonstrate that KDM4C can be regulated by chromatin modifications auxiliary to its substrates, methylated histone H3 lysine 9 and methylated histone H3 lysine 36. We show that auxiliary chromatin modifications can affect activity through directly disrupting interactions between the histone substrate and the catalytic domain. Alternatively, we show that recognition by KDM4C “reader” domains, the tandem tudor domain and the plant homeodomains, regulate the activity of the demethylase. For example, the recognition by the tandem tudor domain drastically stimulates the activity of KDM4C toward its substrate. Moreover, we have generated hypotheses about novel functions of the plant homeodomains that may regulate demethylase activity and localization. Thus, our work has identified many mechanisms fundamental to the regulation of KDM4C. We hypothesize that these modifications will inform future work on other members of the KDM4 demethylase family and broader classes of chromatin modifications. This work will also contribute to the broader understanding of how chromatin modifications are established and maintained in order to properly package the genome and regulate gene expression.