Folding and unfolding cellular phenotype

描述（由申请人提供）：维持稳定的表型需要全基因组范围的转录振荡，转录物簇在稳态周围保持平衡-这是表型的动态结构。我们试图证明蛋白质水平和mRNA表达的变化是通过转录物圈所描述的表面折叠或展开发生的，并表明从这40分钟的振荡到细胞周期和昼夜节律的路径是通过一系列周期加倍分叉发生的。在连续培养的出芽酵母酿酒酵母中，细胞间的信号传导导致相互携带或同步，表现为氧化还原状态的振荡和转录的全基因组振荡。反过来，这种转录氧化还原吸引子周期（TRAC）计时，或限制DNA复制和其他细胞周期事件。使用荧光标记的蛋白质，我们期望利用我们系统中所有转录本的表达模式的详细知识来有效地选择被GFP标记的蛋白质。有了一个很好表征的蛋白质子集，我们将把这些蛋白质映射到假设的吸引子表面，并探索转录物与其蛋白质之间的相关系。利用该系统，可以理清细胞内噪声与振荡信号之间的关系。使用带有青色、绿色和tdTomato-FP标记蛋白的菌株，可以通过观察这些选定蛋白在非同步培养和早期对数培养中的相位关系和振幅，来测试振荡普遍存在的假设。早期，广泛使用的S288C菌株将被投入连续培养系统，期望使用该菌株可以高效地检测大量的GFP标记蛋白。最重要的目标是绘制细胞在周期加倍过程中所采取的路径，以了解哪些基因进入哪些吸引力盆地。这一努力是值得的，因为理解吸引子表面的折叠和展开可以对干细胞发育、癌细胞异质性和肿瘤进展进行易于测试的预测，从而带来概念上的好处。目标是利用这些信息设计干细胞模型系统中的实验。

DESCRIPTION (provided by applicant): The maintenance of a stable phenotype requires a genome wide transcriptional oscillation with clusters of transcripts poised around the steady-state - this is the dynamic architecture of phenotype. We propose to demonstrate that changes in protein levels and mRNA expression occur through a folding or unfolding of the surface described by this circle of transcripts and suggest that the path from this 40-minute oscillation to the cell cycle and circadian rhythms takes place through a series of period doubling bifurcations. Cell to cell signaling in continuous cultures of the budding yeast S. cerevisiae leads to mutual entrainment or synchronization that is manifested as an oscillation in redox state and a genome -wide oscillation in transcription. In turn, this transcriptional redox attractor cycle (TRAC) times, or gates DNA replication and other cell cycle events. Using fluorescently tagged proteins we expect to take advantage of the detailed knowledge of the patterns of expression of all of the transcripts in our system to efficiently choose proteins to be GFP tagged. Having a well characterized subset of proteins we will map these proteins into the hypothesized attractor surface and explore the phase relationships between a transcript and its protein. Using this system it will be possible to sort out the relationship between noise and oscillatory signal in cells. Using strains with Cyan, Green and tdTomato-FP tagged proteins will make it possible to test the hypothesis that oscillations are ubiquitous by looking at the phase relationships and amplitudes of these selected proteins in unsynchronized cultures and cultures in early log phase. Early on, the widely used S288C strain will be put in the continuous culture system, with the expectation that use of this strain will make it possible to efficiently examine large numbers of GFP tagged proteins. The most important aim is to map the path taken by cells during the period doubling process to see which genes settle into which basins of attraction. This effort is worthwhile because of the conceptual benefits that arise from an understanding of how the folding and unfolding of an attractor surface can make readily testable predictions regarding stem cell development, cancer cell heterogeneity and tumor progression The goal would be to use this information to design experiments in a stem cell model system.