title: Versatile protein tagging in cells with split fluorescent protein.
abstract: 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 β-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.
authors.affiliation: Department of Pharmaceutical Chemistry, University of California, San Francisco, California 94143, USA.
authors.forename: Daichi
authors.initials: D
authors.lastname: Kamiyama
authors.affiliation: Department of Pharmaceutical Chemistry, University of California, San Francisco, California 94143, USA.
authors.forename: Sayaka
authors.initials: S
authors.lastname: Sekine
authors.affiliation: Tetrad Graduate Program, University of California, San Francisco, California 94143, USA.
authors.forename: Benjamin
authors.initials: B
authors.lastname: Barsi-Rhyne
authors.affiliation: Department of Pharmaceutical Chemistry, University of California, San Francisco, California 94143, USA.
authors.forename: Jeffrey
authors.initials: J
authors.lastname: Hu
authors.affiliation: Department of Pharmaceutical Chemistry, University of California, San Francisco, California 94143, USA.
authors.forename: Baohui
authors.initials: B
authors.lastname: Chen
authors.affiliation: Department of Cellular and Molecular Pharmacology, University of California, San Francisco, California 94143, USA.
authors.forename: Luke A
authors.initials: LA
authors.lastname: Gilbert
authors.affiliation: Department of Biochemistry and Biophysics, University of California, San Francisco, California 94143, USA.
authors.forename: Hiroaki
authors.initials: H
authors.lastname: Ishikawa
authors.affiliation: Department of Cellular and Molecular Pharmacology, University of California, San Francisco, California 94143, USA.
authors.forename: Manuel D
authors.initials: MD
authors.lastname: Leonetti
authors.affiliation: Department of Biochemistry and Biophysics, University of California, San Francisco, California 94143, USA.
authors.forename: Wallace F
authors.initials: WF
authors.lastname: Marshall
authors.affiliation: Department of Cellular and Molecular Pharmacology, University of California, San Francisco, California 94143, USA.|Howard Hughes Medical Institute, San Francisco, California 94143, USA.
authors.forename: Jonathan S
authors.initials: JS
authors.lastname: Weissman
authors.affiliation: Department of Pharmaceutical Chemistry, University of California, San Francisco, California 94143, USA.|Department of Biochemistry and Biophysics, University of California, San Francisco, California 94143, USA.
authors.forename: Bo
authors.initials: B
authors.lastname: Huang
mesh_terms: D002470:Cell Survival; D005453:Fluorescence; D049452:Green Fluorescent Proteins / Q000378:metabolism*; D006801:Humans; D055503:Protein Multimerization; D011993:Recombinant Fusion Proteins / Q000378:metabolism*; D013194:Staining and Labeling
chemical_list: D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins