The live cell DNA stain SiR-Hoechst induces DNA damage responses and impairs cell cycle progression

Sen, Onur; Saurin, Adrian T.; Higgins, Jonathan M.G.

doi:10.1038/s41598-018-26307-6

Cited by 36 publications

(31 citation statements)

References 51 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…It is worth noting that the cell lines we employed stably express the proteins of interest at close to physiological levels; thus, our data does not suffer from often-encountered protein overexpression artifacts [ 21 ]. Moreover, the cells were never pretreated with DNA damage-sensitizing agents (such as Hoechst), a common practice in micro-irradiation experiments, which could lead to significant deviations in cell physiology and, consequently, in the kinetic behavior of the repair proteins [ 22 ]. This makes our data prone to rapid and straightforward comparisons, a unique attribute of the dataset featured in DNArepairK compared to similar databases [ 23 ].…”

Section: Resultsmentioning

confidence: 99%

DNArepairK: An Interactive Database for Exploring the Impact of Anticancer Drugs onto the Dynamics of DNA Repair Proteins

et al. 2021

View full text Add to dashboard Cite

Cells are constantly exposed to numerous mutagens that produce diverse types of DNA lesions. Eukaryotic cells have evolved an impressive array of DNA repair mechanisms that are able to detect and repair these lesions, thus preventing genomic instability. The DNA repair process is subjected to precise spatiotemporal coordination, and repair proteins are recruited to lesions in an orderly fashion, depending on their function. Here, we present DNArepairK, a unique open-access database that contains the kinetics of recruitment and removal of 70 fluorescently tagged DNA repair proteins to complex DNA damage sites in living HeLa Kyoto cells. An interactive graphical representation of the data complemented with live cell imaging movies facilitates straightforward comparisons between the dynamics of proteins contributing to different DNA repair pathways. Notably, most of the proteins included in DNArepairK are represented by their kinetics in both nontreated and PARP1/2 inhibitor-treated (talazoparib) cells, thereby providing an unprecedented overview of the effects of anticancer drugs on the regular dynamics of the DNA damage response. We believe that the exclusive dataset available in DNArepairK will be of value to scientists exploring the DNA damage response but, also, to inform and guide the development and evaluation of novel DNA repair-targeting anticancer drugs.

show abstract

Section: Resultsmentioning

confidence: 99%

DNArepairK: An Interactive Database for Exploring the Impact of Anticancer Drugs onto the Dynamics of DNA Repair Proteins

et al. 2021

View full text Add to dashboard Cite

show abstract

“…These observations underline that apoptosis is a dynamic process that occurs during a specific period of time and is concentrationdependent [15]. One of the main benefits of Hoechst 33342 is that it can be used on live cells at low concentrations [16]. Therefore, the authors measured apoptosis in nonfixed cells.…”

Section: Autoptosis For Detection Of Apoptosismentioning

confidence: 91%

Apoptosis Assessment in High-Content and High-Throughput Screening Assays

Rens

2021

BioTechniques

View full text Add to dashboard Cite

Here the authors describe the development of AUTOptosis, an economical and rapid apoptosis monitoring method suitable for high-content and high-throughput screening assays. AUTOptosis is based on the quantification of nuclei intensity via staining with Hoechst 33342. First, the authors calibrated the method using standard apoptosis inducers in multiple cell lines. Next, the authors validated the applicability of this approach to high-content screening using a small library of compounds and compared it with the terminal deoxynucleotidyl transferase dUTP nick end labeling gold standard. Finally, the authors demonstrated the specificity of the method by using AUTOposis to detect apoptosis triggered by Mycobacterium tuberculosis intracellular infections.

show abstract

“…A drawback of using Hoechst dye is its short excitation wavelength (ultraviolet to 405 nm) so that phototoxicity from repeated excitation could cause apoptosis [ 8 ]. Hoechst-tagged long-wavelength excitable dyes have been developed [ 9 , 10 ], although the use of these dyes still needs caution to avoid any toxicity [ 11 ].…”

Section: Visualizing Chromatin In Living Cellsmentioning

confidence: 99%

Live-cell imaging probes to track chromatin modification dynamics

2021

View full text Add to dashboard Cite

The spatiotemporal organization of chromatin is regulated at different levels in the nucleus. Epigenetic modifications such as DNA methylation and histone modifications are involved in chromatin regulation and play fundamental roles in genome function. While the one-dimensional epigenomic landscape in many cell types has been revealed by chromatin immunoprecipitation and sequencing, the dynamic changes of chromatin modifications and their relevance to chromatin organization and genome function remain elusive. Live-cell probes to visualize chromatin and its modifications have become powerful tools to monitor dynamic chromatin regulation. Bulk chromatin can be visualized both by small fluorescent dyes and fluorescent proteins, and specific endogenous genomic loci have been detected by adapting genome-editing tools. To track chromatin modifications in living cells, various types of probes have been developed. Protein domains that bind to specific modifications weakly, such as chromodomains for histone methylation, can be repeated to create a tighter binding probe that can then be tagged with a fluorescent protein. It has also been demonstrated that antigen-binding fragments and single-chain variable fragments from modification-specific antibodies can serve as binding probes without disturbing cell division, development and, differentiation. These modification-binding modules are used in modification sensors based on fluorescence/Förster resonance energy transfer to measure the intramolecular conformation changes triggered by modifications. Other probes can be created using a bivalent binding system, such as fluorescence complementation, or luciferase chemiluminescence. Live-cell chromatin modification imaging using these probes will address dynamic chromatin regulation and will be useful for assaying and screening effective epigenome drugs in cells and organisms.

show abstract

The live cell DNA stain SiR-Hoechst induces DNA damage responses and impairs cell cycle progression

Cited by 36 publications

References 51 publications

DNArepairK: An Interactive Database for Exploring the Impact of Anticancer Drugs onto the Dynamics of DNA Repair Proteins

DNArepairK: An Interactive Database for Exploring the Impact of Anticancer Drugs onto the Dynamics of DNA Repair Proteins

Apoptosis Assessment in High-Content and High-Throughput Screening Assays

Live-cell imaging probes to track chromatin modification dynamics

Contact Info

Product

Resources

About