Histone–GFP fusion protein enables sensitive analysis of chromosome dynamics in living mammalian cells

Kanda, Teru; Sullivan, Kevin F.; Wahl, Geoffrey M.

doi:10.1016/s0960-9822(98)70156-3

Cited by 909 publications

(796 citation statements)

References 42 publications

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“…The same protocol was used for control plasmids pEGFP-N1 (Clontech) and BOSH2BGFP-N1 (Kanda et al, 1998).…”

Section: Cell Culture and Cell-cycle Methodsmentioning

confidence: 99%

“…Also, of 20 randomly selected G418-resistant stable transfectants, none had any detectable expression of the PW29-GFP fusion protein. This effect was not due to the toxicity of the GFP molecule itself since, in control experiments with expression of GFP alone (which was partially localized in the nucleus) or expression of H2B-GFP (nuclear localization) (Kanda et al, 1998), no toxicity was detected. To investigate this phenomenon in more detail, we followed the fate of cells overexpressing PW29-GFP using real-time microscopy.…”

Section: Overexpression Of Pw29 Causes Arrest Of Cell Divisionmentioning

confidence: 95%

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Characterization of the components of the putative mammalian sister chromatid cohesion complex

Darwiche

Freeman

Strunnikov

1999

Gene

109

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Establishing and maintaining proper sister chromatid cohesion throughout the cell cycle are essential for maintaining genome integrity. To understand how sister chromatid cohesion occurs in mammals, we have cloned and characterized mouse orthologs of proteins known to be involved in sister chromatid cohesion in other organisms. The cDNAs for the mouse orthologs of SMC1 S.c. and SMC3 S.c. , mSMCB and mSMCD respectively, were cloned and the corresponding transcripts and proteins were characterized. mSMCB and mSMCD are transcribed at similar levels in adult mouse tissues except in testis, which has an excess of mSMCD transcripts. The mSMCB and mSMCD proteins, as well as the PW29 protein, a mouse homolog of Mcd1p S.c./ Rad21 S.p. , form a complex similar to cohesin in X. laevis. mSMCB, mSMCD and PW29 protein levels show no significant cellcycle dependence. The bulk of the mSMCB, mSMCD and PW29 proteins undergo redistribution from the chromosome vicinity to the cytoplasm during prometaphase and back to the chromatin in telophase. This pattern of intracellular localization suggests a complex role for this group of SMC proteins in chromosome dynamics. The PW29 protein and PCNA, which have both been implicated in sister chromatid cohesion, do not colocalize, indicating that these proteins may not function in the same cohesion pathway. Overexpression of a PW29-GFP fusion protein in mouse fibroblasts leads to inhibition of proliferation, implicating this protein and its complex with SMC proteins in the control of mitotic cycle progression.

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“…The same protocol was used for control plasmids pEGFP-N1 (Clontech) and BOSH2BGFP-N1 (Kanda et al, 1998).…”

Section: Cell Culture and Cell-cycle Methodsmentioning

confidence: 99%

Section: Overexpression Of Pw29 Causes Arrest Of Cell Divisionmentioning

confidence: 95%

Characterization of the components of the putative mammalian sister chromatid cohesion complex

Darwiche

Freeman

Strunnikov

1999

Gene

109

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“…Fluorescent tagged histone fusions can incorporate into chromatin without any adverse effects on cell viability in culture (Kanda et al, 1998). By comparison with reporters containing nuclear localization sequences (nls), histone fusions generate a superior signal-to-noise ratio.…”

Section: Introductionmentioning

confidence: 99%

Using a histone yellow fluorescent protein fusion for tagging and tracking endothelial cells in ES cells and mice

Fraser

Hadjantonakis

Sahr

et al. 2005

genesis

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SummaryWe report the first endothelial lineage-specific transgenic mouse allowing live imaging at subcellular resolution. We generated an H2B-EYFP fusion protein which can be used for fluorescent labeling of nucleosomes and used it to specifically label endothelial cells in mice and in differentiating embryonic stem (ES) cells. A fusion cDNA encoding a human histone H2B tagged at its C-terminus with enhanced yellow fluorescent protein (EYFP) was expressed under the control of an Flk1 promoter and intronic enhancer. The Flk1::H2B-EYFP transgenic mice are viable and high levels of chromatin-localized reporter expression are maintained in endothelial cells of developing embryos and in adult animals upon breeding. The onset of fluorescence in differentiating ES cells and in embryos corresponds with the beginning of endothelial cell specification. These transgenic lines permit real-time imaging in normal and pathological vasculogenesis and angiogenesis to track individual cells and mitotic events at a level of detail that is unprecedented in the mouse.

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“…The effects of CIN have been found to be strongly dependent on the rate and type of chromosome mis-segregation or mitotic defect. [14][15][16][17][18] CIN is often measured in a tissue culture setting, which we, for the purpose of this review, define as in vitro. CIN can be quantified via time-lapse microscopy or immunofluorescence staining of fixed cells in mitosis.…”

Section: How and Why Is Cin Measured In Vitro?mentioning

confidence: 99%

CIN and Aneuploidy: Different Concepts, Different Consequences

Schukken

Foijer

2017

BioEssays

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Chromosomal instability (CIN) and aneuploidy are similar concepts but not synonymous. CIN is the process that leads to chromosome copy number alterations, and aneuploidy is the result. While CIN and resulting aneuploidy often cause growth defects, they are also selected for in cancer cells. Although such contradicting fates may seem paradoxical at first, they can be better understood when CIN and aneuploidy are assessed separately, taking into account the in vitro or in vivo context, the rate of CIN, and severity of the aneuploid karyotype. As CIN can only be measured in living cells, which proves to be technically challenging in vivo, aneuploidy is more frequently quantified. However, CIN rates might be more predictive for tumor outcome than assessing aneuploidy rates alone. In reviewing the literature, we therefore conclude that there is an urgent need for new models in which we can monitor chromosome mis-segregation and its consequences in vivo. Also see the video abstract here: https://youtu.be/fL3LxZduchg

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Histone–GFP fusion protein enables sensitive analysis of chromosome dynamics in living mammalian cells

Abstract: The H2B-GFP system allows the high-resolution imaging of chromosomes, including DMs, without compromising nuclear and chromosomal structures and has revealed the distinctive clustering behavior of DMs in mitotic cells which contributes to their asymmetric distribution to daughter cells.

Cited by 909 publications

References 42 publications

Characterization of the components of the putative mammalian sister chromatid cohesion complex

Characterization of the components of the putative mammalian sister chromatid cohesion complex

Using a histone yellow fluorescent protein fusion for tagging and tracking endothelial cells in ES cells and mice

CIN and Aneuploidy: Different Concepts, Different Consequences

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