scholarly journals Monitoring cell cycle distributions in living cells by videomicrofluorometry and discriminant factorial analysis

Cytometry ◽  
2003 ◽  
Vol 56A (1) ◽  
pp. 8-14 ◽  
Author(s):  
Julien Savatier ◽  
J. Vigo ◽  
J.-M. Salmon
Keyword(s):  
Cell Cycle ◽  
Living Cells ◽  
Factorial Analysis ◽  
Discriminant Factorial Analysis

scholarly journals Target Gene Specificity of E2F and Pocket Protein Family Members in Living Cells

2000 ◽  
Vol 20 (16) ◽  
pp. 5797-5807 ◽  
Author(s):  
Julie Wells ◽  
Kathryn E. Boyd ◽  
Christopher J. Fry ◽  
Stephanie M. Bartley ◽  
Peggy J. Farnham
Keyword(s):  
Cell Cycle ◽  
Target Gene ◽  
Target Genes ◽  
Protein Complexes ◽  
Current Model ◽  
Family Members ◽  
Living Cells ◽  
Gene Promoters ◽  
Pocket Protein ◽  
Protein Dna Interactions

ABSTRACT E2F-mediated transcription is thought to involve binding of an E2F-pocket protein complex to promoters in the G0 phase of the cell cycle and release of the pocket protein in late G1, followed by release of E2F in S phase. We have tested this model by monitoring protein-DNA interactions in living cells using a formaldehyde cross-linking and immunoprecipitation assay. We find that E2F target genes are bound by distinct E2F-pocket protein complexes which change as cells progress through the cell cycle. We also find that certain E2F target gene promoters are bound by pocket proteins when such promoters are transcriptionally active. Our data indicate that the current model applies only to certain E2F target genes and suggest that Rb family members may regulate transcription in both G0 and S phases. Finally, we find that a given promoter can be bound by one of several different E2F-pocket protein complexes at a given time in the cell cycle, suggesting that cell cycle-regulated transcription is a stochastic, not a predetermined, process.

scholarly journals Use of the Ki67 promoter to label cell cycle entry in living cells

2010 ◽  
Vol 77A (6) ◽  
pp. 564-570 ◽  
Author(s):  
Alexander C. Zambon
Keyword(s):  
Cell Cycle ◽  
Label Cell ◽  
Living Cells ◽  
Cell Cycle Entry

scholarly journals Quantifying Tubulin Concentration and Microtubule Number Throughout the Fission Yeast Cell Cycle

Biomolecules ◽  
2019 ◽  
Vol 9 (3) ◽  
pp. 86 ◽  
Author(s):  
Isabelle Loiodice ◽  
Marcel Janson ◽  
Penny Tavormina ◽  
Sebastien Schaub ◽  
Divya Bhatt ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Fission Yeast ◽  
Genetic Model ◽  
Spindle Pole Body ◽  
Model Organism ◽  
Spindle Pole ◽  
Living Cells ◽  
Fluorescent Imaging ◽  
Mother And Daughter ◽  
In The Wild

The fission yeast Schizosaccharomyces pombe serves as a good genetic model organism for the molecular dissection of the microtubule (MT) cytoskeleton. However, analysis of the number and distribution of individual MTs throughout the cell cycle, particularly during mitosis, in living cells is still lacking, making quantitative modelling imprecise. We use quantitative fluorescent imaging and analysis to measure the changes in tubulin concentration and MT number and distribution throughout the cell cycle at a single MT resolution in living cells. In the wild-type cell, both mother and daughter spindle pole body (SPB) nucleate a maximum of 23 ± 6 MTs at the onset of mitosis, which decreases to a minimum of 4 ± 1 MTs at spindle break down. Interphase MT bundles, astral MT bundles, and the post anaphase array (PAA) microtubules are composed primarily of 1 ± 1 individual MT along their lengths. We measure the cellular concentration of αβ-tubulin subunits to be ~5 µM throughout the cell cycle, of which one-third is in polymer form during interphase and one-quarter is in polymer form during mitosis. This analysis provides a definitive characterization of αβ-tubulin concentration and MT number and distribution in fission yeast and establishes a foundation for future quantitative comparison of mutants defective in MTs.

scholarly journals Dynamics of inner kinetochore assembly and maintenance in living cells

2008 ◽  
Vol 180 (6) ◽  
pp. 1101-1114 ◽  
Author(s):  
Peter Hemmerich ◽  
Stefanie Weidtkamp-Peters ◽  
Christian Hoischen ◽  
Lars Schmiedeberg ◽  
Indri Erliandri ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Binding Sites ◽  
Living Cells ◽  
S Phase ◽  
Quantitative Microscopy ◽  
Diffusion Behavior ◽  
Kinetochore Assembly ◽  
Centromere Proteins ◽  
Wide Range ◽  
Dynamic Exchange

To investigate the dynamics of centromere organization, we have assessed the exchange rates of inner centromere proteins (CENPs) by quantitative microscopy throughout the cell cycle in human cells. CENP-A and CENP-I are stable centromere components that are incorporated into centromeres via a “loading-only” mechanism in G1 and S phase, respectively. A subfraction of CENP-H also stays stably bound to centromeres. In contrast, CENP-B, CENP-C, and some CENP-H and hMis12 exhibit distinct and cell cycle–specific centromere binding stabilities, with residence times ranging from seconds to hours. CENP-C and CENP-H are immobilized at centromeres specifically during replication. In mitosis, all inner CENPs become completely immobilized. CENPs are highly mobile throughout bulk chromatin, which is consistent with a binding-diffusion behavior as the mechanism to scan for vacant high-affinity binding sites at centromeres. Our data reveal a wide range of cell cycle–specific assembly plasticity of the centromere that provides both stability through sustained binding of some components and flexibility through dynamic exchange of other components.

Synchrotron Fourier transform infrared (FTIR) analysis of single living cells progressing through the cell cycle

The Analyst ◽  
2013 ◽  
Vol 138 (14) ◽  
pp. 3891 ◽  
Author(s):  
Donna R. Whelan ◽  
Keith R. Bambery ◽  
Ljiljana Puskar ◽  
Don McNaughton ◽  
Bayden R. Wood
Keyword(s):  
Cell Cycle ◽  
Fourier Transform ◽  
Fourier Transform Infrared ◽  
Living Cells ◽  
Ftir Analysis ◽  
Single Living Cells ◽  
Single Living

scholarly journals Classic “broken cell” techniques and newer live cell methods for cell cycle assessment

2013 ◽  
Vol 304 (10) ◽  
pp. C927-C938 ◽  
Author(s):  
Lindsay Henderson ◽  
Dante S. Bortone ◽  
Curtis Lim ◽  
Alexander C. Zambon
Keyword(s):  
Cell Cycle ◽  
Cell Division ◽  
Single Cell ◽  
Living Cells ◽  
Cell Division Cycle ◽  
Live Cell ◽  
Nucleotide Analogs ◽  
Protein Ubiquitination ◽  
Cell Cycle Pathway ◽  
Cell Niche

Many common, important diseases are either caused or exacerbated by hyperactivation (e.g., cancer) or inactivation (e.g., heart failure) of the cell division cycle. A better understanding of the cell cycle is critical for interpreting numerous types of physiological changes in cells. Moreover, new insights into how to control it will facilitate new therapeutics for a variety of diseases and new avenues in regenerative medicine. The progression of cells through the four main phases of their division cycle [G0/G1, S (DNA synthesis), G2, and M (mitosis)] is a highly conserved process orchestrated by several pathways (e.g., transcription, phosphorylation, nuclear import/export, and protein ubiquitination) that coordinate a core cell cycle pathway. This core pathway can also receive inputs that are cell type and cell niche dependent. “Broken cell” methods (e.g., use of labeled nucleotide analogs) to assess for cell cycle activity have revealed important insights regarding the cell cycle but lack the ability to assess living cells in real time (longitudinal studies) and with single-cell resolution. Moreover, such methods often require cell synchronization, which can perturb the pathway under study. Live cell cycle sensors can be used at single-cell resolution in living cells, intact tissue, and whole animals. Use of these more recently available sensors has the potential to reveal physiologically relevant insights regarding the normal and perturbed cell division cycle.

scholarly journals DNA Damage Stress: Cui Prodest?

2019 ◽  
Vol 20 (5) ◽  
pp. 1073 ◽  
Author(s):  
Nagendra Verma ◽  
Matteo Franchitto ◽  
Azzurra Zonfrilli ◽  
Samantha Cialfi ◽  
Rocco Palermo ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Death ◽  
Genomic Stability ◽  
Living Cells ◽  
Cancer Development ◽  
Adaptive Mechanism ◽  
Cellular Life ◽  
Checkpoint Adaptation ◽  
Multicellular Organisms ◽  
Damaged Dna

DNA is an entity shielded by mechanisms that maintain genomic stability and are essential for living cells; however, DNA is constantly subject to assaults from the environment throughout the cellular life span, making the genome susceptible to mutation and irreparable damage. Cells are prepared to mend such events through cell death as an extrema ratio to solve those threats from a multicellular perspective. However, in cells under various stress conditions, checkpoint mechanisms are activated to allow cells to have enough time to repair the damaged DNA. In yeast, entry into the cell cycle when damage is not completely repaired represents an adaptive mechanism to cope with stressful conditions. In multicellular organisms, entry into cell cycle with damaged DNA is strictly forbidden. However, in cancer development, individual cells undergo checkpoint adaptation, in which most cells die, but some survive acquiring advantageous mutations and selfishly evolve a conflictual behavior. In this review, we focus on how, in cancer development, cells rely on checkpoint adaptation to escape DNA stress and ultimately to cell death.

The Effect of Enucleation on Flagellar Regeneration in the Protozoon Peranema Trichophorum

1969 ◽  
Vol 4 (1) ◽  
pp. 171-178
Author(s):  
S. L. TAMM
Keyword(s):  
Cell Cycle ◽  
Mitotic Cycle ◽  
Living Cells ◽  
Regeneration Capacity ◽  
Flagellar Regeneration ◽  
Single Living Cells ◽  
Single Living

A rotocompressor was used to enucleate the flagellate protozoon Peranema trichophorum at known stages in the mitotic cycle. This new enucleation technique, combined with recently devised methods for amputating the flagellum and recording its regeneration in single living cells, permitted the investigation of the role of the nucleus in flagellar regeneration at different cell ages. The flagellar regeneration capacity of an enucleate Peranema depended on the stage in the cell cycle when the nucleus was removed. Post-division enucleate cells regenerated about half the length reached by sham-operated controls, and at slower rates, while predivision enucleate cells regenerated flagella equally as well as the controls. Therefore, the nucleus is making an immediate contribution to flagellar regeneration early in the cell cycle, but not late in the cell cycle.

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