|M.Sc Student||Ipenberg Inbal|
|Subject||Interplay between Heat Shock Protein 90 (Hsp90)|
and CHIP E3 Ubiquitin Ligase Regulates the
Stability of KDM4B Histone Demethylase
|Department||Department of Biology||Supervisor||Professor Nabieh Ayoub|
|Full Thesis text|
Lysine methylation of histones is a common post-translational modification (PTM) that regulates gene transcription. The Lysine-specific demethylase 4 (KDM4) histone demethylase family (KDM4A-D), which selectively demethylates H3K9 and H3K36, is implicated in multiple cellular processes critical for carcinogenesis, including transcription, DNA damage response, cell-cycle regulation, cellular differentiation and senescence. Members of the KDM4 family are up-regulated in several types of human cancers and their depletion impairs tumor formation, suggesting that KDM4A-D have a causative role in carcinogenesis. How KDM4A-D over-activity promotes carcinogenesis remains unclear. Therefore, studying the mechanism regulating the stability of KDM4 proteins becomes critical. The main objective of my study is to identify novel KDM4A-D interacting proteins and to study the biological relevance of some of these interactions. Towards this, I have devised GFP-Trap methodology combined with mass spectrometry analysis to identify proteins that interact with EGFP-KDM4A, B and C, but not with the EGFP-NLS fusion. Results reveal over 150 novel interacting proteins including several molecular chaperones like heat shock protein 70 (Hsp70), heat shock protein 90 (Hsp90) and TCP-1 Ring Complex (TRiC). Given the established function of Hsp70 and Hsp90 proteins in regulating the stability of their client proteins, we sought to determine whether KDM4 proteins are new clients of these chaperones. Here I show that: (1) Inhibition of Hsp90 activity destabilizes KDM4B isoform, but not KDM4A, C or D. (2) Destabilization of KDM4B is accompanied by an increase in the methylation levels of H3K9. (3) Inhibition of Hsp90 activity gives rise to the polyubiquitinated form of the KDM4B protein and leads to its degradation through the proteasome. (4) The carboxyl terminus of the Hsc70-interacting protein (CHIP) E3 ubiquitin ligase interacts with KDM4B and its over-expression promotes KDM4B degradation. In agreement with this, CHIP depletion stabilizes KDM4B protein.
Collectively, these results reveal a new pathway by which Hsp90 activity impacts the methylation levels of H3K9 mark, via controlling CHIP-mediated degradation of the KDM4B protein. This novel pathway can be exploited to cure cancers that are driven by KDM4B upregulation.