Bright and photostable yellow fluorescent proteins for extended imaging
Fluorescent proteins are indispensable molecular tools for visualizing biological structures and processes, but their limited photostability restricts the duration of dynamic imaging experiments. Yellow fluorescent proteins (YFPs), in particular, photobleach rapidly. Here, we introduce mGold2s and mGold2t, YFPs with up to 25-fold greater photostability than mVenus and mCitrine, two commonly used YFPs, while maintaining comparable brightness. These variants were identified using a high-throughput pooled single-cell platform, simultaneously screening for high brightness and photostability. Compared with our previous benchmark, mGold, the mGold2 variants display a ~4-fold increase in photostability without sacrificing brightness. mGold2s and mGold2t extend imaging durations across diverse modalities, including widefield, total internal reflection fluorescence (TIRF), super-resolution, single-molecule, and laser-scanning confocal microscopy. When incorporated into fluorescence resonance energy transfer (FRET)-based biosensors, the proposed YFPs enable more reliable, prolonged imaging of dynamic cellular processes. Overall, the enhanced photostability of mGold2s and mGold2t enables high-sensitivity imaging of subcellular structures and cellular activity over extended periods, broadening the scope and precision of biological imaging.
Authors
- Francisco A. Blanco (ORCID: https://orcid.org/0000-0002-1228-8625)
- Shujuan Lai (ORCID: https://orcid.org/0000-0003-1249-4258)
- Gerard G. Lambert (ORCID: https://orcid.org/0000-0002-1607-1124)
- François St-Pierre (ORCID: https://orcid.org/0000-0001-8618-4135)
- Liangyi Chen (ORCID: https://orcid.org/0000-0003-1270-7321)
- John F. Ahrens (ORCID: https://orcid.org/0000-0002-1251-1153)
- Anne C. Lyons (ORCID: https://orcid.org/0009-0006-4685-1136)
- Kimberley F. Tolias (ORCID: https://orcid.org/0000-0002-2092-920X)
- Shuyuan Yang (ORCID: https://orcid.org/0000-0001-5977-9782)
- Shiqun Zhao
- Nathan C. Shaner (ORCID: https://orcid.org/0000-0002-0148-0769)
- Taekjip Ha (ORCID: https://orcid.org/0000-0003-2195-6258)
- Zhuohe Liu (ORCID: https://orcid.org/0000-0002-1818-646X)
- Jin Zhang (ORCID: https://orcid.org/0000-0001-7145-7823)
- Jihwan Lee (ORCID: https://orcid.org/0000-0002-5426-2765)
- Haixin Liu (ORCID: https://orcid.org/0000-0003-3217-856X)
- N. Nguyen
- Raquel Merino‐Urteaga (ORCID: https://orcid.org/0009-0002-9367-0435)
Institutions
- Northwestern University (US)
- Boston Children's Hospital (US)
- The University of Texas MD Anderson Cancer Center (US)
- Howard Hughes Medical Institute (US)
- Harvard University (US)
- Johns Hopkins University (US)
- Baylor College of Medicine (US)
- Peking University (CN)
- University of California San Diego (US)
- Chinese Institute for Brain Research (CN)
- La Jolla Bioengineering Institute (US)
- Rice University (US)
Publication Details
- Journal
- Nature Communications
- Published
- 2025-04-04
- DOI
- https://doi.org/10.1038/s41467-025-58223-5
- Citations
- 13
- Primary Topic
- Advanced Fluorescence Microscopy Techniques
- Type
- article
- Field-Weighted Citation Impact
- 21.24
Funders
- National Science Foundation
- American Heart Association
- Cancer Prevention and Research Institute of Texas
- Rice University
- National Natural Science Foundation of China
- National Institutes of Health
- University of Texas MD Anderson Cancer Center