The consequences of aneuploidy in human cells

Aneuploidy is a change in number or structure of one or more chromosomes that are not a multiple of the whole chromosome set. One of the best known pathological aneuploidies is trisomy 21 (Down syndrome), with chromosome 21 present in three instead of two copies. Patients with Down syndrome display severe mental retardation and growth defects. In fact, most abnormal aneuploid karyotypes lead to spontaneous abortions during embryogenesis, indicating that aneuploidy is not well tolerated in humans. Aneuploidy was also shown to be a common hallmark of cancer tissues; however, the debate is ongoing whether aneuploidy is rather a by-product or a trigger of tumorigenesis. Even though aneuploid karyotypes were already identified more than 100 years ago little is understood about cellular physiology of aneuploidy cells, especially in humans. To uncover the consequences of numerical aneuploidy in human cells, I generated aneuploid cell lines derived from the human cell lines HCT116 and RPE-1 hTERT. First, we showed that aneuploid cells proliferate slower compared to their disomic counterparts. A detailed cell cycle analysis revealed that this delay was due to a prolonged G1 and S phase, whereas G2 and M phase remained unperturbed. Furthermore, we conducted an in depth genome wide comparison of DNA, mRNA and protein levels in aneuploid cells. Using CGH, mRNA array and SILAC technology, we quantified the changes in DNA, mRNA and protein abundance. We revealed that extra chromosomes are actively transcribed and translated. However, the abundance of some proteins, particularly subunits of protein complexes and protein kinases, are adjusted towards disomic levels. Additionally, we asked how the cellular physiology is affected by the addition of a specific chromosome. Two scenarios are possible: either the cellular response depends on the additional chromosomes or all aneuploid cells show the same changes of cellular physiology. Indeed, we found that all aneuploid cell lines show similar physiological responses, irrespective of the type of additional chromosome. All aneuploid cell lines down-regulate DNA and RNA metabolism and up-regulate among others energy metabolism, lysosome function and membrane biosynthesis pathways. Lysosomes which are involved in autophagy are besides the ubiquitin-proteasome system important for cellular protein turn over. We found p62-dependent selective autophagy increased in all analyzed cell lines with extra chromosomes suggesting a role of p62-dependent selective autophagy in maintenance of protein homeostasis upon expression of extra protein in these cell lines

Mol. Syst. Biol.

Extra chromosome copies markedly alter the physiology of eukaryotic cells, but the underlying reasons are not well understood. We created human trisomic and tetrasomic cell lines and determined the quantitative changes in their transcriptome and proteome in comparison with their diploid counterparts. We found that whereas transcription levels reflect the chromosome copy number changes, the abundance of some proteins, such as subunits of protein complexes and protein kinases, is reduced toward diploid levels. Furthermore, using the quantitative data we investigated the changes of cellular pathways in response to aneuploidy. This analysis revealed specific and uniform alterations in pathway regulation in cells with extra chromosomes. For example, the DNA and RNA metabolism pathways were downregulated, whereas several pathways such as energy metabolism, membrane metabolism and lysosomal pathways were upregulated. In particular, we found that the p62-dependent selective autophagy is activated in the human trisomic and tetrasomic cells. Our data present the first broad proteomic analysis of human cells with abnormal karyotypes and suggest a uniform cellular response to the presence of an extra chromosome

Unique features of the transcriptional response to model aneuploidy in human cells

Background: Aneuploidy, a karyotype deviating from multiples of a haploid chromosome set, affects the physiology of eukaryotes. In humans, aneuploidy is linked to pathological defects such as developmental abnormalities, mental retardation or cancer, but the underlying mechanisms remain elusive. There are many different types and origins of aneuploidy, but whether there is a uniform cellular response to aneuploidy in human cells has not been addressed so far. Results: Here we evaluate the transcription profiles of eleven trisomic and tetrasomic cell lines and two cell lines with complex aneuploid karyotypes. We identify a characteristic aneuploidy response pattern defined by upregulation of genes linked to endoplasmic reticulum, Golgi apparatus and lysosomes, and downregulation of DNA replication, transcription as well as ribosomes. Strikingly, complex aneuploidy elicits the same transcriptional changes as trisomy. To uncover the triggers of the response, we compared the profiles with transcription changes in human cells subjected to stress conditions. Interestingly, we found an overlap only with the response to treatment with the autophagy inhibitor bafilomycin A1. Finally, we identified 23 genes whose expression is significantly altered in all aneuploids and which may thus serve as aneuploidy markers. Conclusions: Our analysis shows that despite the variability in chromosome content, aneuploidy triggers uniform transcriptional response in human cells. A common response independent of the type of aneuploidy might be exploited as a novel target for cancer therapy. Moreover, the potential aneuploidy markers identified in our analysis might represent novel biomarkers to assess the malignant potential of a tumor

A non‐proteolytic release mechanism for HMCES‐DNA‐protein crosslinks

The conserved protein HMCES crosslinks to abasic (AP) sites in ssDNA to prevent strand scission and the formation of toxic dsDNA breaks during replication. Here, we report a non‐proteolytic release mechanism for HMCES‐DNA‐protein crosslinks (DPCs), which is regulated by DNA context. In ssDNA and at ssDNA‐dsDNA junctions, HMCES‐DPCs are stable, which efficiently protects AP sites against spontaneous incisions or cleavage by APE1 endonuclease. In contrast, HMCES‐DPCs are released in dsDNA, allowing APE1 to initiate downstream repair. Mechanistically, we show that release is governed by two components. First, a conserved glutamate residue, within HMCES' active site, catalyses reversal of the crosslink. Second, affinity to the underlying DNA structure determines whether HMCES re‐crosslinks or dissociates. Our study reveals that the protective role of HMCES‐DPCs involves their controlled release upon bypass by replication forks, which restricts DPC formation to a necessary minimum

mTORC1 activity is supported by spatial association with focal adhesions

The mammalian target of rapamycin complex 1 (mTORC1) integrates mitogenic and stress signals to control growth and metabolism. Activation of mTORC1 by amino acids and growth factors involves recruitment of the complex to the lysosomal membrane and is further supported by lysosome distribution to the cell periphery. Here, we show that translocation of lysosomes toward the cell periphery brings mTORC1 into proximity with focal adhesions (FAs). We demonstrate that FAs constitute discrete plasma membrane hubs mediating growth factor signaling and amino acid input into the cell. FAs, as well as the translocation of lysosome-bound mTORC1 to their vicinity, contribute to both peripheral and intracellular mTORC1 activity. Conversely, lysosomal distribution to the cell periphery is dispensable for the activation of mTORC1 constitutively targeted to FAs. This study advances our understanding of spatial mTORC1 regulation by demonstrating that the localization of mTORC1 to FAs is both necessary and sufficient for its activation by growth-promoting stimuli

mTORC1 activity is supported by spatial association with focal adhesions

The mammalian target of rapamycin complex 1 (mTORC1) integrates mitogenic and stress signals to control growth and metabolism. Activation of mTORC1 by amino acids and growth factors involves recruitment of the complex to the lysosomal membrane and is further supported by lysosome distribution to the cell periphery. Here, we show that translocation of lysosomes toward the cell periphery brings mTORC1 into proximity with focal adhesions (FAs). We demonstrate that FAs constitute discrete plasma membrane hubs mediating growth factor signaling and amino acid input into the cell. FAs, as well as the translocation of lysosome-bound mTORC1 to their vicinity, contribute to both peripheral and intracellular mTORC1 activity. Conversely, lysosomal distribution to the cell periphery is dispensable for the activation of mTORC1 constitutively targeted to FAs. This study advances our understanding of spatial mTORC1 regulation by demonstrating that the localization of mTORC1 to FAs is both necessary and sufficient for its activation by growth-promoting stimuli