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Although efforts have been made to identify circadian-controlled genes regulating cell cycle progression and cell death, little is known about the metabolic signals modulating circadian regulation of gene expression. We have identified heme, an iron-containing prosthetic group, as a regulatory ligand controlling human Period-2 (hPer2) stability. Furthermore, we defined a novel heme-regulatory motif within the C terminus of hPer2 (SC(841)PA) as necessary for heme binding and protein destabilization. Spectroscopy revealed that whereas the PAS domain binds to both the ferric and ferrous forms of heme, SC(841)PA bound exclusively to ferric heme, thus acting as a redox sensor. Consequently, binding prevented hPer2 from interacting with its stabilizing counterpart cryptochrome. In vivo, hPer2 downregulation is suppressed by inhibitors of heme synthesis or proteasome activity, while SA(841)PA is sufficient to stabilize hPer2 in transfected cells. Moreover, heme binding to the SC(841)PA motif directly impacted circadian gene expression, resulting in altered period length. Overall, the data support a model where heme-mediated oxidation triggers hPer2 degradation, thus controlling heterodimerization and ultimately gene transcription. This project is in collaboration with Dr. Carla Finkielstein (Virginia Tech). |
