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Micronuclei-based model system reveals functional consequences of chromothripsis in human cells
(2019)
Cancer cells often harbor chromosomes in abnormal numbers and with aberrant structure. The consequences of these chromosomal aberrations are difficult to study in cancer, and therefore several model systems have been developed in recent years. We show that human cells with extra chromosome engineered via microcell-mediated chromosome transfer often gain massive chromosomal rearrangements. The rearrangements arose by chromosome shattering and rejoining as well as by replication-dependent mechanisms. We show that the isolated micronuclei lack functional lamin B1 and become prone to envelope rupture, which leads to DNA damage and aberrant replication. The presence of functional lamin B1 partly correlates with micronuclei size, suggesting that the proper assembly of nuclear envelope might be sensitive to membrane curvature. The chromosomal rearrangements in trisomic cells provide growth advantage compared to cells without rearrangements. Our model system enables to study mechanisms of massive chromosomal rearrangements of any chromosome and their consequences in human cells.
Background: Aneuploidy, or abnormal chromosome numbers, severely alters cell physiology and is widespread in
cancers and other pathologies. Using model cell lines engineered to carry one or more extra chromosomes, it has
been demonstrated that aneuploidy per se impairs proliferation, leads to proteotoxic as well as replication stress
and triggers conserved transcriptome and proteome changes.
Results: In this study, we analysed for the first time miRNAs and demonstrate that their expression is altered in
response to chromosome gain. The miRNA deregulation is independent of the identity of the extra chromosome
and specific to individual cell lines. By cross-omics analysis we demonstrate that although the deregulated miRNAs
differ among individual aneuploid cell lines, their known targets are predominantly associated with cell development,
growth and proliferation, pathways known to be inhibited in response to chromosome gain. Indeed, we show that up
to 72% of these targets are downregulated and the associated miRNAs are overexpressed in aneuploid cells, suggesting
that the miRNA changes contribute to the global transcription changes triggered by aneuploidy. We identified
hsa-miR-10a-5p to be overexpressed in majority of aneuploid cells. Hsa-miR-10a-5p enhances translation of a
subset of mRNAs that contain so called 5’TOP motif and we show that its upregulation in aneuploids provides
resistance to starvation-induced shut down of ribosomal protein translation.
Conclusions: Our work suggests that the changes of the microRNAome contribute on one hand to the adverse
effects of aneuploidy on cell physiology, and on the other hand to the adaptation to aneuploidy by supporting
translation under adverse conditions.
Keywords: Aneuploidy, Cancer, miRNA, miR-10a-5p, Trisomy
Phospho-regulation of the Shugoshin - Condensin interaction at the centromere in budding yeast
(2020)
Correct bioriented attachment of sister chromatids to the mitotic spindle is essential for chromosome segregation. In budding yeast, the conserved protein shugoshin (Sgo1) contributes to biorientation by recruiting the protein phosphatase PP2A-Rts1 and the condensin complex to centromeres. Using peptide prints, we identified a Serine-Rich Motif (SRM) of Sgo1 that mediates the interaction with condensin and is essential for centromeric condensin recruitment and the establishment of biorientation. We show that the interaction is regulated via phosphorylation within the SRM and we determined the phospho-sites using mass spectrometry. Analysis of the phosphomimic and phosphoresistant mutants revealed that SRM phosphorylation disrupts the shugoshin–condensin interaction. We present evidence that Mps1, a central kinase in the spindle assembly checkpoint, directly phosphorylates Sgo1 within the SRM to regulate the interaction with condensin and thereby condensin localization to centromeres. Our findings identify novel mechanisms that control shugoshin activity at the centromere in budding yeast.
Loss of USP28 and SPINT2 expression promotes cancer cell survival after whole genome doubling
(2021)
Background
Whole genome doubling is a frequent event during cancer evolution and shapes the cancer genome due to the occurrence of chromosomal instability. Yet, erroneously arising human tetraploid cells usually do not proliferate due to p53 activation that leads to CDKN1A expression, cell cycle arrest, senescence and/or apoptosis.
Methods
To uncover the barriers that block the proliferation of tetraploids, we performed a RNAi mediated genome-wide screen in a human colorectal cancer cell line (HCT116).
Results
We identified 140 genes whose depletion improved the survival of tetraploid cells and characterized in depth two of them: SPINT2 and USP28. We found that SPINT2 is a general regulator of CDKN1A transcription via histone acetylation. Using mass spectrometry and immunoprecipitation, we found that USP28 interacts with NuMA1 and affects centrosome clustering. Tetraploid cells accumulate DNA damage and loss of USP28 reduces checkpoint activation, thus facilitating their proliferation.
Conclusions
Our results indicate three aspects that contribute to the survival of tetraploid cells: (i) increased mitogenic signaling and reduced expression of cell cycle inhibitors, (ii) the ability to establish functional bipolar spindles and (iii) reduced DNA damage signaling.