Rowat Amy C, Bird James C, Agresti Jeremy J, Rando Oliver J, Weitz David A
Department of Physics/School of Engineering and Applied Sciences, Harvard University, Cambridge, MA 02138, USA.
Proc Natl Acad Sci U S A. 2009 Oct 27;106(43):18149-54. doi: 10.1073/pnas.0903163106. Epub 2009 Oct 13.
Cells within a genetically identical population exhibit phenotypic variation that in some cases can persist across multiple generations. However, information about the temporal variation and familial dependence of protein levels remains hidden when studying the population as an ensemble. To correlate phenotypes with the age and genealogy of single cells over time, we developed a microfluidic device that enables us to track multiple lineages in parallel by trapping single cells and constraining them to grow in lines for as many as 8 divisions. To illustrate the utility of this method, we investigate lineages of cells expressing one of 3 naturally regulated proteins, each with a different representative expression behavior. Within lineages deriving from single cells, we observe genealogically related clusters of cells with similar phenotype; cluster sizes vary markedly among the 3 proteins, suggesting that the time scale of phenotypic persistence is protein-specific. Growing lines of cells also allows us to dynamically track temporal fluctuations in protein levels at the same time as pedigree relationships among the cells as they divide in the chambers. We observe bursts in expression levels of the heat shock protein Hsp12-GFP that occur simultaneously in mother and daughter cells. In contrast, the ribosomal protein Rps8b-GFP shows relatively constant levels of expression over time. This method is an essential step toward understanding the time scales of phenotypic variation and correlations in phenotype among single cells within a population.
在基因相同的群体中，细胞表现出表型变异，在某些情况下这种变异可以持续多代。然而，当将群体作为一个整体进行研究时，蛋白质水平的时间变异和家族依赖性信息仍然隐藏着。为了将表型与单个细胞随时间的年龄和谱系相关联，我们开发了一种微流控装置，该装置通过捕获单个细胞并限制它们成排生长多达8代，使我们能够并行追踪多个谱系。为了说明这种方法的实用性，我们研究了表达3种自然调节蛋白之一的细胞谱系，每种蛋白都有不同的代表性表达行为。在源自单个细胞的谱系中，我们观察到具有相似表型的细胞的谱系相关簇；这3种蛋白质的簇大小差异显著，表明表型持续的时间尺度是蛋白质特异性的。生长的细胞系还使我们能够在细胞在小室中分裂时，动态追踪蛋白质水平的时间波动以及细胞之间的谱系关系。我们观察到热休克蛋白Hsp12-GFP的表达水平在母细胞和子细胞中同时出现爆发。相比之下，核糖体蛋白Rps8b-GFP的表达水平随时间相对恒定。这种方法是理解群体中单个细胞表型变异的时间尺度以及表型相关性的重要一步。
