期刊名称：Proceedings of the National Academy of Sciences
印刷版ISSN：0027-8424
电子版ISSN：1091-6490
出版年度：2016
卷号：113
期号：47
页码：13516-13521
DOI：10.1073/pnas.1607984113
语种：English
出版社：The National Academy of Sciences of the United States of America
摘要：SignificanceCells sense changes in environmental conditions and translate them into physiological responses that are mediated by a plethora of molecular intermediaries that converge at critical nodes. This is particularly relevant to multiple levels of biological organization in which the circadian clock and cell division are interlocked. Unexpectedly, the circadian rhythms of two critical components of the interlocked system, Period 2 (Per2) and p53, were in antiphase. This finding was analyzed using a systematic modeling approach, and predictions of the model were validated by experimental studies that revealed a distinct and complex scenario. Accordingly, we show that the spatiotemporal organization of Per2:p53 interactions and the nature of their chemical modifications are critical for their time-dependent subcellular redistribution and potential biological functions. The circadian clock and cell cycle networks are interlocked on the molecular level, with the core clock loop exerting a multilevel regulatory role over cell cycle components. This is particularly relevant to the circadian factor Period 2 (Per2), which modulates the stability of the tumor suppressor p53 in unstressed cells and transcriptional activity in response to genotoxic stress. Per2 binding prevents Mdm2-mediated ubiquitination of p53 and, therefore, its degradation, and oscillations in the peaks of Per2 and p53 were expected to correspond. However, our findings showed that Per2 and p53 rhythms were significantly out-of-phase relative to each other in cell lysates and in purified cytoplasmic fractions. These seemingly conflicting experimental data motivated the use of a combined theoretical and experimental approach focusing on the role played by Per2 in dictating the phase of p53 oscillations. Systematic modeling of all possible regulatory scenarios predicted that the observed phase relationship between Per2 and p53 could be simulated if (i) p53 was more stable in the nucleus than in the cytoplasm, (ii) Per2 associates to various ubiquitinated forms of p53, and (iii) Per2 mediated p53 nuclear import. These predictions were supported by a sevenfold increase in p53s half-life in the nucleus and by in vitro binding of Per2 to the various ubiquitinated forms of p53. Last, p53s nuclear shuttling was significantly favored by ectopic expression of Per2 and reduced because of Per2 down-regulation. Our combined theoretical/mathematical approach reveals how clock regulatory nodes can be inferred from oscillating time course data.
