Versatile protein tagging in cells with split fluorescent protein

被引：277

作者：

Kamiyama, Daichi ^{[1
]}

Sekine, Sayaka ^{[1
]}

Barsi-Rhyne, Benjamin ^{[2
]}

Hu, Jeffrey ^{[1
]}

Chen, Baohui ^{[1
]}

Gilbert, Luke A. ^{[3
]}

Ishikawa, Hiroaki ^{[4
]}

Leonetti, Manuel D. ^{[3
]}

Marshall, Wallace F. ^{[4
]}

Weissman, Jonathan S. ^{[3
,5
]}

Huang, Bo ^{[1
,4
]}

机构：

[1] Univ Calif San Francisco, Dept Pharmaceut Chem, San Francisco, CA 94143 USA

[2] Univ Calif San Francisco, Tetrad Grad Program, San Francisco, CA 94143 USA

[3] Univ Calif San Francisco, Dept Cellular & Mol Pharmacol, San Francisco, CA 94143 USA

[4] Univ Calif San Francisco, Dept Biochem & Biophys, San Francisco, CA 94143 USA

[5] Howard Hughes Med Inst, San Francisco, CA 94143 USA

来源：

NATURE COMMUNICATIONS | 2016年 / 7卷

基金：

日本学术振兴会;

关键词：

GLOBAL ANALYSIS; GFP; LOCALIZATION; EXPRESSION; SECRETION; SYSTEM;

D O I：

10.1038/ncomms11046

中图分类号：

O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];

学科分类号：

07 ; 0710 ; 09 ;

摘要：

In addition to the popular method of fluorescent protein fusion, live cell protein imaging has now seen more and more application of epitope tags. The small size of these tags may reduce functional perturbation and enable signal amplification. To address their background issue, we adapt self-complementing split fluorescent proteins as epitope tags for live cell protein labelling. The two tags, GFP11 and sfCherry11 are derived from the eleventh beta-strand of super-folder GFP and sfCherry, respectively. The small size of FP11-tags enables a cost-effective and scalable way to insert them into endogenous genomic loci via CRISPR-mediated homology-directed repair. Tandem arrangement FP11-tags allows proportional enhancement of fluorescence signal in tracking intraflagellar transport particles, or reduction of photobleaching for live microtubule imaging. Finally, we show the utility of tandem GFP11-tag in scaffolding protein oligomerization. These experiments illustrate the versatility of FP11-tag as a labelling tool as well as a multimerization-control tool for both imaging and non-imaging applications.

引用

页数：9

共 34 条

[1] Mechanism and energetics of green fluorescent protein chromophore synthesis revealed by trapped intermediate structures [J].

Barondeau, DP ;

Putnam, CD ;

Kassmann, CJ ;

Tainer, JA ;

Getzoff, ED .

PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 2003, 100 (21) :12111-12116

[2] Protein tagging and detection with engineered self-assembling fragments of green fluorescent protein [J].

Cabantous, S ;

Terwilliger, TC ;

Waldo, GS .

NATURE BIOTECHNOLOGY, 2005, 23 (01) :102-107

[3] Fast live simultaneous multiwavelength four-dimensional optical microscopy [J].

Carlton, Peter M. ;

Boulanger, Jerome ;

Kervrann, Charles ;

Sibarita, Jean-Baptiste ;

Salamero, Jean ;

Gordon-Messer, Susannah ;

Bressan, Debra ;

Haber, James E. ;

Haase, Sebastian ;

Shao, Lin ;

Winoto, Lukman ;

Matsuda, Atsushi ;

Kner, Peter ;

Uzawa, Satoru ;

Gustafsson, Mats ;

Kam, Zvi ;

Agard, David A. ;

Sedat, John W. .

PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 2010, 107 (37) :16016-16022

[4] Fluorescent fusion protein knockout mediated by anti-GFP nanobody [J].

Caussinus, Emmanuel ;

Kanca, Oguz ;

Affolter, Markus .

NATURE STRUCTURAL & MOLECULAR BIOLOGY, 2012, 19 (01) :117-U142

[5]

Chavez A, 2015, NAT METHODS, V12, P326, DOI [10.1038/nmeth.3312, 10.1038/NMETH.3312]

[6] Dynamic Imaging of Genomic Loci in Living Human Cells by an Optimized CRISPR/Cas System [J].

Chen, Baohui ;

Gilbert, Luke A. ;

Cimini, Beth A. ;

Schnitzbauer, Joerg ;

Zhang, Wei ;

Li, Gene-Wei ;

Park, Jason ;

Blackburn, Elizabeth H. ;

Weissman, Jonathan S. ;

Qi, Lei S. ;

Huang, Bo .

CELL, 2013, 155 (07) :1479-1491

[7] Spatially resolved, highly multiplexed RNA profiling in single cells [J].

Chen, Kok Hao ;

Boettiger, Alistair N. ;

Moffitt, Jeffrey R. ;

Wang, Siyuan ;

Zhuang, Xiaowei .

SCIENCE, 2015, 348 (6233)

[8] Split mCherry as a new red bimolecular fluorescence complementation system for visualizing protein-protein interactions in living cells [J].

Fan, Jin-Yu ;

Cui, Zong-Qiang ;

Wei, Hong-Ping ;

Zhang, Zhi-Ping ;

Zhou, Ya-Feng ;

Wang, Yun-Peng ;

Zhang, Xian-En .

BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, 2008, 367 (01) :47-53

[9] GFP reconstitution across synaptic partners (GRASP) defines cell contacts and Synapses in living nervous systems [J].

Feinberg, Evan H. ;

VanHoven, Miri K. ;

Bendesky, Andres ;

Wang, George ;

Fetter, Richard D. ;

Shen, Kang ;

Bargmannl, Cornelia I. .

NEURON, 2008, 57 (03) :353-363

[10] Fluorescent probes for super-resolution imaging in living cells [J].

Fernandez-Suarez, Marta ;

Ting, Alice Y. .

NATURE REVIEWS MOLECULAR CELL BIOLOGY, 2008, 9 (12) :929-943

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