Differential equation methods for simulation of GFP kinetics in non-steady state experiments.
Differential equation methods for simulation of GFP kinetics in non-steady state experiments.
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用于模拟非稳态实验中 GFP 动力学的微分方程方法。
DOI：
10.1091/mbc.e17-06-0396
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发表时间：
2018
影响因子：
3.3
通讯作者：
Phair,RobertD
中科院分区：
文献类型：
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作者：
Phair,RobertD
基因编码的荧光蛋白与荧光显微镜相结合，广泛应用于细胞生物学中，以收集细胞内运输的动力学数据。从这些数据中提取定量信息的方法基于扩散和示踪动力学的数学。目前的方法虽然有用且强大，但依赖于所研究的细胞系统处于稳定状态的假设，即所有分子浓度和通量在实验期间保持恒定的假设。在这里，我们通过构建基础细胞生物过程的机械非线性微分方程模型并将其与控制荧光示踪剂动力学的一组单独的微分方程联系起来，得出了用于非稳态生物系统的新示踪剂动力学分析方法。连接两组方程是基于不可区分性基本示踪原理的新应用，并且与当前方法不同，支持示踪动力学对细胞动力学的正确依赖性。因此，这种方法为 GFP 荧光显微镜的应用（包括光漂白 [FRAP、FLIP] 和光活化到经常遇到的涉及生理或药理扰动（例如生长因子、神经递质、急性敲除、抑制剂、激素、细胞因子和代谢物）的实验方案提供了通用的数学框架，这些扰动启动了细胞内的机械信息 瞬变。当达到新的稳态时，这些方法会自动简化为经典的稳态示踪动力学分析。
Genetically encoded fluorescent proteins, combined with fluorescence microscopy, are widely used in cell biology to collect kinetic data on intracellular trafficking. Methods for extraction of quantitative information from these data are based on the mathematics of diffusion and tracer kinetics. Current methods, although useful and powerful, depend on the assumption that the cellular system being studied is in a steady state, that is, the assumption that all the molecular concentrations and fluxes are constant for the duration of the experiment. Here, we derive new tracer kinetic analytical methods for non–steady state biological systems by constructing mechanistic nonlinear differential equation models of the underlying cell biological processes and linking them to a separate set of differential equations governing the kinetics of the fluorescent tracer. Linking the two sets of equations is based on a new application of the fundamental tracer principle of indistinguishability and, unlike current methods, supports correct dependence of tracer kinetics on cellular dynamics. This approach thus provides a general mathematical framework for applications of GFP fluorescence microscopy (including photobleaching [FRAP, FLIP] and photoactivation to frequently encountered experimental protocols involving physiological or pharmacological perturbations (e.g., growth factors, neurotransmitters, acute knockouts, inhibitors, hormones, cytokines, and metabolites) that initiate mechanistically informative intracellular transients. When a new steady state is achieved, these methods automatically reduce to classical steady state tracer kinetic analysis.