Drug-induced cell cycle modulation leading to cell-cycle arrest, nuclear mis-segregation, or endoreplication

Sakaue-Sawano, Asako; Kobayashi, Tamiyo; Ohtawa, Kenji; Miyawaki, Atsushi

doi:10.1186/1471-2121-12-2

Cited by 134 publications

(139 citation statements)

References 35 publications

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“…This is of particular importance when studying apoptosis in the context of cell cycle progression. Similar conclusions were presented in a recent work, demonstrating that combining cytometry analysis with the FUCCI probe may contribute to development of an assay that integrates the complexity of cell cycle regulation into drug discovery screens (10). …”

Section: Resultssupporting

confidence: 63%

Analysis of apoptosis in FUCCI HeLa cells

Malka¹,

Eden²

2011

Cytometry Pt A

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Section: Resultssupporting

confidence: 63%

Analysis of apoptosis in FUCCI HeLa cells

Malka¹,

Eden²

2011

Cytometry Pt A

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“…To monitor cell cycle progression and paclitaxel-induced apoptosis in living cells, we first established a HeLa-based cell line that stably co-expresses the GMNN(Geminin)-Venus 22,23 and DIABLO(SMAC)-mCherry 24 fusion proteins, which function as reporters of the G2/M cell cycle phase and MOMP, respectively (HeLa-GI23 line, Fig. S1).…”

Section: Construction Of Apoptosis-prone or -Reluctant Hela Cell Linementioning

confidence: 99%

Paclitaxel-induced aberrant mitosis and mitotic slippage efficiently lead to proliferative death irrespective of canonical apoptosis and p53

et al. 2016

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Spindle poisons elicit various cellular responses following metaphase arrest, but how they relate to longterm clonogenicity has remained unclear. We prepared several HeLa lines in which the canonical apoptosis pathway was attenuated, and compared their acute responses to paclitaxel, as well as long-term fate, with the parental line. Three-nanomolar paclitaxel induced brief metaphase arrest (<5 h) often followed by aberrant mitosis, and about 90% of the cells of each line had lost their clonogenicity after 48 h of the treatment. A combination of the same concentration of paclitaxel with the kinesin-5 inhibitor, S-trityl-L-cysteine (STLC), at 1 mM led to much longer arrest (»20 h) and predominance of subsequent linespecific responses: mitochondrial outer membrane permeabilization (MOMP) in the apoptosis-prone line, or mitotic slippage without obvious MOMP in the apoptosis-reluctant lines. In spite of this, combination with STLC did not lead to a marked difference in clonogenicity between the apoptosis-prone and -reluctant lines, and intriguingly resulted in slightly better clonogenicity than that of cells treated with 3 nM paclitaxel alone. This indicates that changes in the short-term response within 3 possible scenarios -acute MOMP, mitotic slippage or aberrant mitosis ¡ has only a weak impact on clonogenicity. Our results suggest that once cells have committed to slippage or aberrant mitosis they eventually undergo proliferative death irrespective of canonical apoptosis or p53 function. Consistent with this, cells with irregular DNA contents originating from mitotic slippage or aberrant mitosis were mostly eliminated from the population within several rounds of division after the drug treatment.

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“…12 A number of Fucci mouse lines exist. CAG-Fucci is not inducible and is composed of 2 lines; CAG-mKO2-hCdt1(30/ 120) and CAG-mAG-hGem(1/110) generated by random transgenesis.…”

Section: Introductionmentioning

confidence: 99%

Fucci2a:A bicistronic cell cycle reporter that allows Cre mediated tissue specific expression in mice

et al. 2014

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Markers of cell cycle stage allow estimation of cell cycle dynamics in cell culture and during embryonic development. The Fucci system incorporates genetically encoded probes that highlight G1 and S/G2/M phases of the cell cycle allowing live imaging. However the available mouse models that incorporate Fucci are beset by problems with transgene inactivation, varying expression level, lack of conditional potential and/or the need to maintain separate transgenes-there is no transgenic mouse model that solves all these problems. To address these shortfalls we reengineered the Fucci system to create 2 bicistronic Fucci variants incorporating both probes fused using the Thosea asigna virus 2A (T2A) self cleaving peptide. We characterize these variants in stable 3T3 cell lines. One of the variants (termed Fucci2a) faithfully recapitulated the nuclear localization and cell cycle stage specific florescence of the original Fucci system. We go on to develop a conditional mouse allele (R26Fucci2aR) carefully designed for high, inducible, ubiquitous expression allowing investigation of cell cycle status in single cell lineages within the developing embryo. We demonstrate the utility of R26Fucci2aR for live imaging by using high resolution confocal microscopy of ex vivo lung, kidney and neural crest development. Using our 3T3 system we describe and validate a method to estimate cell cycle times from relatively short time-lapse sequences that we then apply to our neural crest data. The Fucci2a system and the R26Fucci2aR mouse model are compelling new tools for the investigation of cell cycle dynamics in cell culture and during mouse embryonic development.

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Drug-induced cell cycle modulation leading to cell-cycle arrest, nuclear mis-segregation, or endoreplication

Cited by 134 publications

References 35 publications

Analysis of apoptosis in FUCCI HeLa cells

Analysis of apoptosis in FUCCI HeLa cells

Paclitaxel-induced aberrant mitosis and mitotic slippage efficiently lead to proliferative death irrespective of canonical apoptosis and p53

Fucci2a:A bicistronic cell cycle reporter that allows Cre mediated tissue specific expression in mice

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