UC Davis

Regulation of the genome is a delicate, but dynamic process. It can be seen on many levels from chemical modifications on DNA and histones which modulate gene expression and regulate physical access to the genome, to interactions between macromolecules, and the formation of higher ordered structures. Subtle changes can be advantageous to the cell, as demonstrated by functionally divergent histone variants like histone H3.3. Though its sequence differs from canonical H3 by a handful of amino acids, it has been demonstrated to be essential in development and cell differentiation. Because of its distinct roles, we set out to examine the H3.3 interactome with the goal of identifying novel associations which may facilitate H3.3’s functionality, particularly in the context of human induced pluripotent stem cells (hiPSC). We employed a maltose-binding protein (MBP) assay to generate H3.1 and H3.3 MBP fusions to investigate the interactions via MBP pull down and liquid chromatography tandem mass spectrometry (LC-MS/MS). Our screen identified over 2,300 proteins, including a novel interaction between the H3 family and transcription factor ZHX2. Although our biochemical studies determined this interaction was not H3.3-specific, given the roles these proteins play in tumorigenesis (pediatric high grade gliomas, hepatocellular cancer, etc), we find this association to be noteworthy. We further identified modified H3 preferentially binds to ZHX2 over unmodified H3. The significance of this interaction remains to be characterized, but upon further examination, H3 and ZHX2 share mutual pathways and interacting partners potentially suggesting a functional link.