Shimadzu Shunji, Furuya Tomoyuki, Ozawa Yasuko, Fukuda Hiroo, Kondo Yuki
Graduate School of Science, The University of Tokyo, Tokyo, Japan.
Graduate School of Science, Kobe University, Kobe, Japan.
Quant Plant Biol. 2022 Jul 18;3:e15. doi: 10.1017/qpb.2022.12. eCollection 2022.
Stem cell fates are spatio-temporally regulated during plant development. Time-lapse imaging of fluorescence reporters is the most widely used method for spatio-temporal analysis of biological processes. However, excitation light for imaging fluorescence reporters causes autofluorescence and photobleaching. Unlike fluorescence reporters, luminescence proteins do not require excitation light, and therefore offer an alternative reporter for long-term and quantitative spatio-temporal analysis. We established an imaging system for luciferase, which enabled monitoring cell fate marker dynamics during vascular development in a vascular cell induction system called VISUAL. Single cells expressing the cambium marker, , had sharp luminescence peaks at different time points. Furthermore, dual-color luminescence imaging revealed spatio-temporal relationships between cells that differentiated into xylem or phloem, and cells that transitioned from procambium to cambium. This imaging system enables not only the detection of temporal gene expression, but also facilitates monitoring of spatio-temporal dynamics of cell identity transitions at the single cell level.
在植物发育过程中，干细胞命运受到时空调控。荧光报告基因的延时成像技术是生物过程时空分析中应用最广泛的方法。然而，用于成像荧光报告基因的激发光会导致自发荧光和光漂白现象。与荧光报告基因不同，发光蛋白不需要激发光，因此为长期和定量的时空分析提供了一种替代报告基因。我们建立了一种用于荧光素酶的成像系统，该系统能够在一个名为VISUAL的血管细胞诱导系统中监测血管发育过程中细胞命运标记物的动态变化。表达形成层标记物的单个细胞在不同时间点有明显的发光峰值。此外，双色发光成像揭示了分化为木质部或韧皮部的细胞与从原形成层过渡到形成层的细胞之间的时空关系。这种成像系统不仅能够检测基因的瞬时表达，还便于在单细胞水平上监测细胞身份转变的时空动态。
