Laboratory of Receptor Biology and Gene Expression, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America.
PLoS One. 2009 Sep 29;4(9):e7163. doi: 10.1371/journal.pone.0007163.
NF-kappaB is a prototypic stress-responsive transcription factor that acts within a complex regulatory network. The signaling dynamics of endogenous NF-kappaB in single cells remain poorly understood. To examine real time dynamics in living cells, we monitored NF-kappaB activities at multiple timescales using GFP-p65 knock-in mouse embryonic fibroblasts. Oscillations in NF-kappaB were sustained in most cells, with several cycles of transient nuclear translocation after TNF-alpha stimulation. Mathematical modeling suggests that NF-kappaB oscillations are selected over other non-oscillatory dynamics by fine-tuning the relative strengths of feedback loops like IkappaBalpha. The ability of NF-kappaB to scan and interact with the genome in vivo remained remarkably constant from early to late cycles, as observed by fluorescence recovery after photobleaching (FRAP). Perturbation of long-term NF-kappaB oscillations interfered with its short-term interaction with chromatin and balanced transcriptional output, as predicted by the mathematical model. We propose that negative feedback loops do not simply terminate signaling, but rather promote oscillations of NF-kappaB in the nucleus, and these oscillations are functionally advantageous.
NF-κB 是一种典型的应激反应转录因子，它在复杂的调控网络中发挥作用。内源性 NF-κB 在单细胞中的信号转导动力学仍知之甚少。为了在活细胞中实时检测动力学，我们使用 GFP-p65 基因敲入的鼠胚胎成纤维细胞在多个时间尺度上监测 NF-κB 活性。在 TNF-α刺激后，大多数细胞中 NF-κB 持续发生振荡，并且有几个短暂的核转位循环。数学模型表明，通过精细调节 IkappaBalpha 等反馈回路的相对强度，NF-κB 振荡被选择为其他非振荡动力学。通过光漂白后荧光恢复（FRAP）观察到，NF-κB 与基因组的体内扫描和相互作用的能力在早期到晚期循环中仍然保持惊人的恒定。长期 NF-κB 振荡的干扰与其与染色质的短期相互作用和平衡转录输出相平衡，正如数学模型所预测的那样。我们提出，负反馈回路并不简单地终止信号转导，而是促进核内 NF-κB 的振荡，并且这些振荡具有功能优势。
