The mobility of phytochrome within protonemal tip cells of the moss Ceratodon purpureus, monitored by fluorescence correlation spectroscopy
The mobility of phytochrome within protonemal tip cells of the moss Ceratodon purpureus, monitored by fluorescence correlation spectroscopy
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DOI：
10.1529/biophysj.103.038521
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发表时间：
2004-09-01
影响因子：
3.4
通讯作者：
Lamparter, T
中科院分区：
文献类型：
--
作者：
Böse, G;Schwille, P;Lamparter, T
荧光相关光谱（FCS）是一种研究荧光分子在细胞中迁移的通用工具。在这篇文章中，我们展示了在活细胞中参与信号转导的生物分子的自由扩散和膜结合形式之间的区别是可能的。采用荧光相关光谱法测定了光敏色素的迁移率，光敏色素在紫角藓（Ceratodon purpureus）原毛尖细胞的向光性和向极性中起作用。光敏色素装载植红蛋白，仅在光敏色素结合状态下才荧光。采用激光共聚焦扫描显微镜成像，选择尖端细胞顶区xy测量位置。在细胞的古z位置测量荧光相关性。通过非线性最小二乘拟合对扩散系数进行分析，发现光敏色素亚细胞组分在细胞外围的扩散系数比核心组分高6倍。这种光敏色素显然与膜结合，可能控制着趋光性和偏光性反应。
Fluorescence correlation spectroscopy (FCS) is a versatile tool for investigating the mobilities of fluorescent molecules in cells. In this article, we show that it is possible to distinguish between freely diffusing and membrane-bound forms of biomolecules involved in signal transduction in living cells. Fluorescence correlation spectroscopy was used to measure the mobility of phytochrome, which plays a role in phototropism and polarotropism in protonemal tip cells of the moss Ceratodon purpureus. The phytochrome was loaded with phycoerythrobilin, which is fluorescent only in the phytochrome-bound state. Confocal laser scanning microscopy was used for imaging and selecting the xy measuring position in the apical zone of the tip cell. Fluorescence correlation was measured at ancient z-positions in the cell. Analysis of the diffusion coefficients by nonlinear least-square fits showed a subcellular fraction of phytochrome at the cell periphery with a sixfold higher diffusion coefficient than in the core fraction. This phytochrome is apparently bound to the membrane and probably controls the phototropic and polarotropic response.