Abstract 8924: Heterochromatin Protein 1 Acts as an Intra-nuclear Redox Sensor
Background: Pro-atherogenic stimuli generates reactive oxygen species (ROS) in endothelial cells. To adapt to extrinsic ROS, cells possess the protective mechanisms using various sensor proteins including oxygen-reactive cysteine residues. However, little is known about the mechanism to sense intra-nuclear redox state. Heterochromatin protein 1 (HP1) is a chromatin-bound protein which forms a transcriptional repressor complex with other proteins and play diverse roles in gene regulation. Recently the specific role for the post-translational modification of HP1 in transcriptional regulation has been highlighted.
Hypothesis: HP1 acts as a sensor protein against intra-nuclear redox state in endothelial cells.
Methods and Results: During the purification of HP1, we found two fractions of HP1 one of which was speculated as a post-translationally modified form. Using mutation analysis and mass spectrometry, this separated fraction turned out to be the intermolecular disulfide homodimers of HP1, a novel oxidative modification. Among three isoforms of HP1 (α, β and γ), only HPγ readily and reversibly forms disulfide homodimers via cysteine 177 (C177) under oxidative conditions. To elucidate the physiological role for this novel modification of HP1γ, we screened the interacting protein of HP1γ under oxidative conditions using human umbilical vein endothelial cells (HUVECs). We found that HP1γ strongly interacts with TIF1β, a universal transcriptional repressor protein, only when oxidatively dimerized via C177. As a consequence, HP1γ holds TIF1β in chromatin component in the nucleus of HUVECs under oxidative conditions. Transcriptional reporter assay revealed that the oxidative modification of HP1γ has the reversing effect on the transcriptional repression activity of TIF1β. Finally, expression analysis using undimerizable HP1γ C177S mutant-replaced HUVECs clarified that HP1γ dimerization is involved in upregulating a subset of genes associated with transcriptional regulation and apoptosis.
Conclusion: In conclusion, our study suggests that in endothelial cells HP1γ acts as an intra-nuclear redox sensor and connects the intra-nuclear redox state with transcriptional regulation via interacting with TIF1β under oxidative conditions.
- © 2010 by American Heart Association, Inc.