通过全内反射荧光显微镜对肌动蛋白成核和延伸事件进行直接可视化和定量分析。
Direct Visualization and Quantification of the Actin Nucleation andElongation Events by TIRF Microscopy.
作者信息
Jiang Yuxiang, Huang Shanjin
机构信息
Center for Plant Biology, School of Life Sciences, Tsinghua University, Beijing, China.
Institute of Botany, Chinese Academy of Sciences, Beijing, China.
出版信息
Bio Protoc. 2017 Mar 5;7(5):e2146. doi: 10.21769/BioProtoc.2146.
Abstract
Total internal reflection fluorescence (TIRF) microscopy is a powerful tool for visualizing the dynamics of actin filaments at single-filament resolution . Thanks to the development of various fluorescent probes, we can easily monitor all kinds of events associated with actin dynamics, including nucleation, elongation, bundling, fragmentation and monomer dissociation. Here we present a detailed protocol regarding the visualization and quantification of actin nucleation and filament elongation events by TIRF microscopy , which is based on the methods previously reported ( Liu , 2015 ; Yang , 2011 ).
摘要
全内反射荧光（TIRF）显微镜是一种强大的工具，可在单丝分辨率下可视化肌动蛋白丝的动力学。由于各种荧光探针的发展，我们可以轻松监测与肌动蛋白动力学相关的各种事件，包括成核、伸长、成束、断裂和单体解离。在这里，我们基于先前报道的方法（Liu，2015；Yang，2011），给出了一份关于通过TIRF显微镜可视化和定量肌动蛋白成核和丝伸长事件的详细方案。
相似文献
1
Direct Visualization and Quantification of the Actin Nucleation andElongation Events by TIRF Microscopy.通过全内反射荧光显微镜对肌动蛋白成核和延伸事件进行直接可视化和定量分析。
Bio Protoc. 2017 Mar 5;7(5):e2146. doi: 10.21769/BioProtoc.2146.
2
The role of the FH1 domain and profilin in formin-mediated actin-filament elongation and nucleation.FH1结构域和丝切蛋白在formin介导的肌动蛋白丝延伸和成核中的作用。
Curr Biol. 2008 Jan 8;18(1):9-19. doi: 10.1016/j.cub.2007.11.062. Epub 2007 Dec 20.
3
Tropomyosin and Profilin Cooperate to Promote Formin-Mediated Actin Nucleation and Drive Yeast Actin Cable Assembly.原肌球蛋白和丝切蛋白协同促进formin介导的肌动蛋白成核并驱动酵母肌动蛋白电缆组装。
Curr Biol. 2016 Dec 5;26(23):3230-3237. doi: 10.1016/j.cub.2016.09.053. Epub 2016 Nov 17.
4
Dissection of two parallel pathways for formin-mediated actin filament elongation.解析两个平行通路在成核延伸肌动蛋白丝中的作用。
J Biol Chem. 2018 Nov 16;293(46):17917-17928. doi: 10.1074/jbc.RA118.004845. Epub 2018 Sep 28.
5
Visualizing the in vitro assembly of tropomyosin/actin filaments using TIRF microscopy.使用全内反射荧光显微镜观察原肌球蛋白/肌动蛋白丝的体外组装。
Biophys Rev. 2020 Aug;12(4):879-885. doi: 10.1007/s12551-020-00720-6. Epub 2020 Jul 7.
6
A Processive Arabidopsis Formin Modulates Actin Filament Dynamics in Association with Profilin.一个连续的拟南芥形成蛋白与 Profilin 一起调节肌动蛋白丝动力学。
Mol Plant. 2016 Jun 6;9(6):900-10. doi: 10.1016/j.molp.2016.03.006. Epub 2016 Mar 17.
7
Profilin-Dependent Nucleation and Assembly of Actin Filaments Controls Cell Elongation in Arabidopsis.肌动蛋白丝的肌动蛋白单体结合蛋白依赖性成核与组装控制拟南芥细胞伸长
Plant Physiol. 2016 Jan;170(1):220-33. doi: 10.1104/pp.15.01321. Epub 2015 Nov 16.
8
Visualization of Actin Assembly and Filament Turnover by In Vitro Multicolor TIRF Microscopy.通过体外多色全内反射荧光显微镜观察肌动蛋白组装和丝状体周转
Methods Mol Biol. 2016;1407:287-306. doi: 10.1007/978-1-4939-3480-5_21.
9
The role of formin tails in actin nucleation, processive elongation, and filament bundling.formin 尾部在肌动蛋白成核、延伸和纤维束形成中的作用。
J Biol Chem. 2014 Oct 31;289(44):30602-30613. doi: 10.1074/jbc.M114.588368. Epub 2014 Sep 22.
10
Using Microfluidics Single Filament Assay to Study Formin Control of Actin Assembly.使用微流控单丝分析法研究formin对肌动蛋白组装的控制。
Methods Mol Biol. 2018;1805:75-92. doi: 10.1007/978-1-4939-8556-2_4.
引用本文的文献
1
Maspin/SerpinB5 is a cytoskeleton-binding protein that regulates epithelial cell shape.乳脂肪球表皮生长因子8/丝氨酸蛋白酶抑制剂B5是一种调节上皮细胞形状的细胞骨架结合蛋白。
Commun Biol. 2025 Aug 22;8(1):1262. doi: 10.1038/s42003-025-08688-3.
2
Differential sensitivity of ADF isovariants to a pH gradient promotes pollen tube growth.ADF 同工型对 pH 梯度的差异敏感性促进花粉管生长。
J Cell Biol. 2023 Nov 6;222(11). doi: 10.1083/jcb.202206074. Epub 2023 Aug 23.
3
In vitro Reconstitution of Phase-separated p62 Bodies on the Arp2/3-derived Actin Network.在源自Arp2/3的肌动蛋白网络上对相分离的p62小体进行体外重构。
Bio Protoc. 2023 Apr 20;13(8):e4656. doi: 10.21769/BioProtoc.4656.
4
Lipid Species Dependent Vesicles Clustering Caused by alpha-Synuclein as Revealed by Single-Vesicle Imaging with Total Internal Reflection Fluorescence Microscopy.通过全内反射荧光显微镜单囊泡成像揭示的由α-突触核蛋白引起的脂质种类依赖性囊泡聚集
Biophys Rep. 2021 Dec;7(6):437-448. doi: 10.52601/bpr.2021.210020.
本文引用的文献
1
Profilin Regulates Apical Actin Polymerization to Control Polarized Pollen Tube Growth.丝状肌动蛋白调节蛋白调控顶端肌动蛋白聚合以控制极性花粉管生长。
Mol Plant. 2015 Dec 7;8(12):1694-709. doi: 10.1016/j.molp.2015.09.013. Epub 2015 Oct 1.
2
The functionally distinct fission yeast formins have specific actin-assembly properties.功能不同的裂殖酵母formin 具有特定的肌动蛋白组装特性。
Mol Biol Cell. 2011 Oct;22(20):3826-39. doi: 10.1091/mbc.E11-06-0492. Epub 2011 Aug 24.
3
BENT UPPERMOST INTERNODE1 encodes the class II formin FH5 crucial for actin organization and rice development.BENT UPPERMOST INTERNODE1 编码了 FH5 这个 II 类formin，它对肌动蛋白组织和水稻发育至关重要。
Plant Cell. 2011 Feb;23(2):661-80. doi: 10.1105/tpc.110.081802. Epub 2011 Feb 9.
4
Real-time measurements of actin filament polymerization by total internal reflection fluorescence microscopy.通过全内反射荧光显微镜对肌动蛋白丝聚合进行实时测量。
Biophys J. 2005 Feb;88(2):1387-402. doi: 10.1529/biophysj.104.047399. Epub 2004 Nov 19.
5
Direct real-time observation of actin filament branching mediated by Arp2/3 complex using total internal reflection fluorescence microscopy.使用全内反射荧光显微镜对由Arp2/3复合体介导的肌动蛋白丝分支进行直接实时观察。
Proc Natl Acad Sci U S A. 2001 Dec 18;98(26):15009-13. doi: 10.1073/pnas.211556398. Epub 2001 Dec 11.
Zotero 插件