RRC ID 49548
Author Sakaue-Sawano A, Yo M, Komatsu N, Hiratsuka T, Kogure T, Hoshida T, Goshima N, Matsuda M, Miyoshi H, Miyawaki A.
Title Genetically Encoded Tools for Optical Dissection of the Mammalian Cell Cycle.
Journal Mol Cell
Abstract Eukaryotic cells spend most of their life in interphase of the cell cycle. Understanding the rich diversity of metabolic and genomic regulation that occurs in interphase requires the demarcation of precise phase boundaries in situ. Here, we report the properties of two genetically encoded fluorescence sensors, Fucci(CA) and Fucci(SCA), which enable real-time monitoring of interphase and cell-cycle biology. We re-engineered the Cdt1-based sensor from the original Fucci system to respond to S phase-specific CUL4Ddb1-mediated ubiquitylation alone or in combination with SCFSkp2-mediated ubiquitylation. In cultured cells, Fucci(CA) produced a sharp triple color-distinct separation of G1, S, and G2, while Fucci(SCA) permitted a two-color readout of G1 and S/G2. Fucci(CA) applications included tracking the transient G1 phase of rapidly dividing mouse embryonic stem cells and identifying a window for UV-irradiation damage in S phase. These results show that Fucci(CA) is an essential tool for quantitative studies of interphase cell-cycle regulation.
Volume 68(3)
Pages 626-640.e5
Published 2017-11-2
DOI 10.1016/j.molcel.2017.10.001
PII S1097-2765(17)30750-5
PMID 29107535
IF 15.584
Times Cited 11
DNA material mCherry-hCdt1(1/100)Cy(-)/pcDNA3 (RDB15459) mCherry-hCdt1(1/100)Cy(-)/pCSII-EF (RDB15441) mCherry-hCdt1(1/100)/pCSII-EF (RDB15442) tFucci(SA)2/pCSII-EF (RDB15443) tFucci(SA)2.1/pCSII-EF (RDB15444) tFucci(SA)2.2/pCSII-EF (RDB15445) tFucci(CA)2/pCSII-EF (RDB15446) tFucci(CA)2.1/pCSII-EF (RDB15447) tFucci(CA)2.2/pCSII-EF (RDB15448) tFucci(SA)2/pCSII-CMV (RDB15449) tFucci(SA)2.1/pCSII-CMV (RDB15450) tFucci(SA)2.2/pCSII-CMV (RDB15451) tFucci(CA)2/pCSII-CMV (RDB15452) tFucci(CA)2.1/pCSII-CMV (RDB15453) tFucci(CA)2.2/pCSII-CMV (RDB15454)