CTCF supports preferentially short lamina-associated domains

Kaczmarczyk, Lukasz; Levi, Nehora; Segal, Tamar; Salmon‐Divon, Mali; Gerlitz, Gabi

doi:10.1007/s10577-022-09686-5

Cited by 8 publications

(9 citation statements)

References 68 publications

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“…Identifying the importance of CTCF in mitosis requires a CTCF knockdown (KD) cell line. Previously we generated CRISPR-based knockdown of CTCF in B16-F1 mouse melanoma cells to decrease the total CTCF protein levels by 60-70% while the remaining CTCF was truncated with reduced chromatin binding capabilities (Kaczmarczyk et al 2022). Complete knockdown of CTCF was shown to be lethal (Wan et al 2008; Moore et al 2012; Nora et al 2017), thus preventing the study of cell proliferation and mitosis.…”

Section: Resultsmentioning

confidence: 99%

“…Specifically, the cut phenotype, first detailed in yeast (Samejima et al 1993), where non-segregating chromosome mass in anaphase is split by cytokinesis was reported to be due to failure of cohesin cleavage by separase (Hauf et al 2001). It is possible that the loss of the N-terminal domain in these CTCF knockdowns (Kaczmarczyk et al 2022) disrupts important interactions for both cohesion retention and chromatin loops (Pugacheva et al 2020). The cut phenotype is one of the prominent displayed by both CTCF knockdowns.…”

Section: Discussionmentioning

confidence: 99%

“…To determine if CTCF has a role in mitotic fidelity, we used CTCF CRSIPR-based knockdown (KD) clones in B16 mouse melanoma cell line (Kaczmarczyk et al 2022). We first live-cell imaged mitotic behavior by tracking SiR-DNA labeled in wild type and two different CTCF knockdowns.…”

Section: Introductionmentioning

confidence: 99%

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CTCF is essential for proper mitotic spindle structure and anaphase segregation

Chiu

Berrada

Eskndir

et al. 2023

Preprint

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Mitosis is an essential process in which the duplicate genome is segregated equally into two daughter cells. CTCF has been reported to be present in mitosis but its importance for mitotic fidelity remains to be determined. To evaluate the importance of CTCF in mitosis, we tracked mitotic behaviors in wild type and two different CTCF CRISPR-based genetic knockdowns. We find that knockdown of CTCF results in prolonged mitoses and failed anaphase segregation via time lapse imaging of SiR-DNA. CTCF knockdown did not alter cell cycling or the mitotic checkpoint, which was activated upon nocodazole treatment. Immunofluorescence imaging of the mitotic spindle in CTCF knockdowns revealed disorganization via tri/tetrapolar spindles and chromosomes behind the spindle pole. Imaging of interphase nuclei showed that nuclear size increased drastically, consistent with failure to divide the duplicated genome in anaphase. Population measurements of nuclear shape in CTCF knockdowns do not display decreased circularity or increased nuclear blebbing relative to wild type. However, failed mitoses do display abnormal nuclear morphologies relative to successful mitoses, suggesting population images do not capture individual behaviors. Thus, CTCF is important for both proper metaphase organization and anaphase segregation which impacts the size and shape of the interphase nucleus.

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Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

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CTCF is essential for proper mitotic spindle structure and anaphase segregation

Chiu

Berrada

Eskndir

et al. 2023

Preprint

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“…To evaluate the effect of CTCF on cell migration we used the mouse melanoma B16-F1 cells in which we reduced CTCF expression levels by the CRISPR-Cas9 system 46 . Targeting of Cas9 to the beginning of the CTCF gene led to elimination of the full-length form of CTCF and the appearance of truncated forms of CTCF at low levels that are probably formed from downstream AUGs.…”

Section: Resultsmentioning

confidence: 99%

The CTCF-H3K27me3 axis supports melanoma cell migration by repressing cholesterol biosynthesis

Kaczmarczyk

Levi

Salmon‐Divon

et al. 2023

Preprint

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CCCTC-binding factor (CTCF) is a zinc finger protein that regulates correct three-dimensional chromatin folding in addition to activation and repression of transcription and splicing. CTCF was found to affect tumor cell migration, however the molecular mechanism is still obscure. To identify the molecular mechanism by which CTCF supports melanoma cell migration we used mouse melanoma cells with a partial loss of function (pLoF) of CTCF generated by the CRISPR-Cas9 system. Here we show that CTCF pLoF inhibited cell migration rate. Using RNA sequencing we identified an increase in the expression of multiple enzymes in the cholesterol biosynthesis pathway along with an increase in the cellular cholesterol levels. Inhibition of cholesterol synthesis in CTCF pLoF cells by Fatostatin restored the cellular migration rate, suggesting that CTCF supports cell migration by inhibition of cholesterol synthesis. Detailed analysis of the promoter of Hmgcs1, an upstream enzyme in the cholesterol synthesis pathway, revealed that CTCF pLoF led to reduced H3K27me3 levels in parallel to increased binding of the transcriptional activator SREBP2. Moreover, inhibition of H3K27 methylation in wildtype cells led to increased SREBP2 binding to this promoter. Taken together, our results suggest that CTCF represses transcription of Hmgc1 by promoting H3K27 methylation to suppress SREBP2 binding. By that, CTCF fine tune the cellular cholesterol levels to support cell migration.

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“…Locally fitting attraction potentials to these regions as a function of gene activity or epigenomic states should predict minimal conditions required to promote and maintain weaker NL-chromatin interactions. In this context, adding “binders” based on experimental evidence of factors involved in the modulation of chromatin-NL interactions, either as LADs or focal domains in LADs (e.g., CTCF) ( van Schaik et al, 2021 ; Kaczmarczyk et al, 2022 ), should enable advances in our mechanistic understanding of these interactions. Modeling the behavior of a chromatin polymer with various thicknesses ( Buckle et al, 2018 ) may also predict the dynamics of chromatin domains with varying compaction levels at the NL.…”

Section: Perspectivesmentioning

confidence: 99%

Biology and Model Predictions of the Dynamics and Heterogeneity of Chromatin-Nuclear Lamina Interactions

Madsen-Østerbye

Bellanger

Galigniana

et al. 2022

Front. Cell Dev. Biol.

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Associations of chromatin with the nuclear lamina, at the nuclear periphery, help shape the genome in 3 dimensions. The genomic landscape of lamina-associated domains (LADs) is well characterized, but much remains unknown on the physical and mechanistic properties of chromatin conformation at the nuclear lamina. Computational models of chromatin folding at, and interactions with, a surface representing the nuclear lamina are emerging in attempts to characterize these properties and predict chromatin behavior at the lamina in health and disease. Here, we highlight the heterogeneous nature of the nuclear lamina and LADs, outline the main 3-dimensional chromatin structural modeling methods, review applications of modeling chromatin-lamina interactions and discuss biological insights inferred from these models in normal and disease states. Lastly, we address perspectives on future developments in modeling chromatin interactions with the nuclear lamina.

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CTCF supports preferentially short lamina-associated domains

Cited by 8 publications

References 68 publications

CTCF is essential for proper mitotic spindle structure and anaphase segregation

CTCF is essential for proper mitotic spindle structure and anaphase segregation

The CTCF-H3K27me3 axis supports melanoma cell migration by repressing cholesterol biosynthesis

Biology and Model Predictions of the Dynamics and Heterogeneity of Chromatin-Nuclear Lamina Interactions

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