Department of Chemistry and Molecular Biology, University of Gothenburg, Gothenburg, Sweden.
FEBS J. 2013 Aug;280(16):3854-67. doi: 10.1111/febs.12382. Epub 2013 Jul 5.
The control of activity and localization of transcription factors is critical for appropriate transcriptional responses. In eukaryotes, signal transduction components such as mitogen-activated protein kinase (MAPK) shuttle into the nucleus to activate transcription. It is not known in detail how different amounts of nuclear MAPK over time affect the transcriptional response. In the present study, we aimed to address this issue by studying the high osmolarity glycerol (HOG) system in Saccharomyces cerevisiae. We employed a conditional osmotic system, which changes the period of the MAPK Hog1 signal independent of the initial stress level. We determined the dynamics of the Hog1 nuclear localization and cell volume by single-cell analysis in well-controlled microfluidics systems and compared the responses with the global transcriptional output of cell populations. We discovered that the onset of the initial transcriptional response correlates with the potential of cells for rapid adaptation; cells that are capable of recovering quickly initiate the transcriptional responses immediately, whereas cells that require longer time to adapt also respond later. This is reflected by Hog1 nuclear localization, Hog1 promoter association and the transcriptional response, but not Hog1 phosphorylation, suggesting that a presently uncharacterized rapid adaptive mechanism precedes the Hog1 nuclear response. Furthermore, we found that the period of Hog1 nuclear residence affects the amplitude of the transcriptional response rather than the spectrum of responsive genes.
转录因子的活性和定位控制对于适当的转录反应至关重要。在真核生物中，信号转导成分，如丝裂原活化蛋白激酶（MAPK），穿梭进入细胞核以激活转录。目前还不清楚不同时间内核 MAPK 的不同数量如何影响转录反应。在本研究中，我们旨在通过研究酿酒酵母中的高渗甘油（HOG）系统来解决这个问题。我们采用了一种条件渗透压系统，该系统独立于初始应激水平改变 MAPK Hog1 信号的周期。我们通过单细胞分析在微流控系统中精确控制的条件下，确定了 Hog1 核定位和细胞体积的动力学，并将其与细胞群体的全局转录输出进行了比较。我们发现，初始转录反应的开始与细胞快速适应的潜力相关；能够快速恢复的细胞立即启动转录反应，而需要更长时间适应的细胞则反应较晚。这反映在 Hog1 的核定位、Hog1 启动子结合和转录反应上，但不反映在 Hog1 的磷酸化上，这表明在 Hog1 核反应之前存在一种目前尚未表征的快速适应机制。此外，我们发现 Hog1 核居留时间会影响转录反应的幅度，而不是响应基因的谱。
