Micronuclei-based model system reveals functional consequences of chromothripsis in human cells
- TU Kaiserslautern, Germany
- University Medical Center, Netherlands
- Max Planck Institute of Biochemistry, Germany
- Cold Spring Harbor Laboratory, United States
Figures
Figure 1 with 3 supplements
Unique de novo CNAs specifically affect the transferred chromosome and resemble chromothripsis.
(a) Detection of CNAs in HCT116- and RPE1-derived cell lines with extra chromosome. A total of 201 (HCT116) and 50 (RPE1) CNAs were found. (b) Boxplot showing CNA sizes on the transferred versus the …
Figure 1—figure supplement 1
SNP and SMASH array analysis shows stable addition of the extra chromosome to HCT116 acceptor cells.
Each column represents one chromosome, each row one dataset from SNP array (black) or SMASH (red) analyses. The chromosome copy number is colour-coded. Only autosomes are visualized.
Figure 1—figure supplement 2
SNP and SMASH array analysis shows stable addition of the extra chromosome to RPE1 acceptor cells.
Each column represents one chromosome, each row one dataset from SNP array (black) or SMASH (red) analyses. The chromosome copy number is colour-coded. Only autosomes are visualized. Please note …
Figure 1—figure supplement 3
Copy number and structural rearrangement breakpoint junctions for cell lines analyzed by whole genome sequencing.
Plots show the breakpoint junctions (solid lines) and CNAs (black dots) detected by WGS. Lines are colored according to the orientation of the breakpoint junction, from low to high chromosomal …
Figure 2 with 1 supplement
DNA damage in isolated micronuclei after mitotic slippage.
(a) Schematic depiction of MMCT. (b) Murine A9 cell before micronucleation. (c) Formation of micronuclei in the A9 donor cell upon colchicine treatment. (d) Microcells with micronucleus isolated via …
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Figure 2—source data 1
Source data for Figure 2.
- https://cdn.elifesciences.org/articles/50292/elife-50292-fig2-data1-v2.xlsx
Figure 2—figure supplement 1
Characteristics of micronuclei.
(a) Cells after fusion with isolated MNs were immunolabelled with anti-EdU (red) to detect DNA replication. They were categorized in four groups: non-replicating DNA in primary nuclei (PN) and MN …
Figure 3
Micronuclei frequently lack lamin B1 in the nuclear envelope.
(a) Schematic depiction of marker proteins with nuclear localization (red) and cytoplasmic localization (green) after nuclear envelope rupture. (b) Localization of NLS-RFP and CLS-GFP in untreated …
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Figure 3—source data 1
Source data for Figure 3.
- https://cdn.elifesciences.org/articles/50292/elife-50292-fig3-data1-v2.xlsx
Figure 4
DNA damage occurs predominantly in lamin B1 negative MN.
(a) Immunofluorescence staining of isolated MN with lamin B1 antibody combined with TUNEL assay. The lamin B1 negative MN is highlighted by yellow arrowhead, the lamin B1 positive MN is indicated by …
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Figure 4—source data 1
Source data for Figure 4.
- https://cdn.elifesciences.org/articles/50292/elife-50292-fig4-data1-v2.xlsx
Figure 5 with 2 supplements
Chromothripsis provides functional advantages to the cells.
(a) Example of soft agar assay to determine colony formation ability in parental cell line HCT116, trisomy with a highly rearranged extra chromosome 5 (Htr5-16) and trisomy with a nearly intact …
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Figure 5—source data 1
Source data for Figure 5.
- https://cdn.elifesciences.org/articles/50292/elife-50292-fig5-data1-v2.xlsx
Figure 5—figure supplement 1
Gene-specific changes.
(a) Percentage of TSGs and OGs as defined by the TUSON explorer with a TUSON p value < 0.05 that overlap with the CAN regions on the transferred chromosome of the samples Htr13-02, Htr13-03, …
Figure 5—figure supplement 2
Correlation of colony forming capacity.
(a) The normalized colony forming capacity of the cell lines relative to the percentage of MN in trisomic and tetrasomic samples. (b) The amount of additional genomic material in trisomic and …
Additional files
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Supplementary file 1
Cell lines used in the study.
The cell lines were created by microcell-mediated chromosome transfer. The % of cells with whole chromosome aneuploidy (WCA) shows the % of metaphase spreads that scored positive for the expected WCA.
- https://cdn.elifesciences.org/articles/50292/elife-50292-supp1-v2.docx
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Supplementary file 2
List of all identified copy number aberrations.
- https://cdn.elifesciences.org/articles/50292/elife-50292-supp2-v2.xlsx
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Supplementary file 3
The number and percentage of informative SNPs per chromosomal region used for determination of the chromosome affected by unique CNAs; regions are determined per clone, by copy number.
- https://cdn.elifesciences.org/articles/50292/elife-50292-supp3-v2.docx
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Supplementary file 4
Breakpoint junctions detected by WGS that were selected for validation by PCR, Sanger sequencing and de novo assembly.
- https://cdn.elifesciences.org/articles/50292/elife-50292-supp4-v2.xlsx
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Supplementary file 5
Overview of informative SNPs deduced from whole genome sequencing reads, allowing determination of the affected chromosome.
- https://cdn.elifesciences.org/articles/50292/elife-50292-supp5-v2.docx
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Transparent reporting form
- https://cdn.elifesciences.org/articles/50292/elife-50292-transrepform-v2.docx
