Cell Mol Life Sci, 2021, 78(7):3467-3476

FUCCI-Red: a single-color cell cycle indicator for fluorescence lifetime imaging.

The phase of the cell cycle determines numerous aspects of cancer cell behaviour including invasiveness, ability to migrate and responsiveness to cytotoxic drugs. To non-invasively monitor progression of cell cycle in vivo, a family of genetically encoded fluorescent indicators, FUCCI (fluorescent ubiquitination-based cell cycle indicator), has been developed. Existing versions of FUCCI are based on fluorescent proteins of two or more different colors fused to cell-cycle-dependent degradation motifs. Thus, FUCCI-expressing cells emit light of different colors in different phases providing a robust way to monitor cell cycle progression by fluorescence microscopy and flow cytometry but limiting the possibility to simultaneously visualize other markers. To overcome this limitation, we developed a single-color variant of FUCCI, called FUCCI-Red, which utilizes two red fluorescent proteins with distinct fluorescence lifetimes, mCherry and mKate2. Similarly to FUCCI, these proteins carry cell cycle-dependent degradation motifs to resolve G1 and S/G2/M phases. We showed utility of FUCCI-Red by visualizing cell cycle progression of cancer cells in 2D and 3D cultures and monitoring development of tumors in vivo by confocal and fluorescence lifetime imaging microscopy (FLIM). Single-channel registration and red-shifted spectra make FUCCI-Red sensor a promising instrument for multiparameter in vivo imaging applications, which was demonstrated by simultaneous detection of cellular metabolic state using endogenous fluorescence in the blue range.

Shirmanova MV, Gorbachev DA, Sarkisyan KS, Parnes AP, Gavrina AI, Polozova AV, Kovaleva TF, Snopova LB, Dudenkova VV, Zagaynova EV, Lukyanov KA

IBCH: 9086
Ссылка на статью в журнале: http://link.springer.com/10.1007/s00018-020-03712-7
Кол-во цитирований на 04.2024: 9
Данные статьи проверены модераторами 2021-02-12

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