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Global analysis of genome, transcriptome and proteome reveals the response to aneuploidy in human cells.

Stingele S, Stoehr G, Peplowska K, Cox J, Mann M, Storchova Z - Mol. Syst. Biol. (2012)

Bottom Line: 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.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.

View Article: PubMed Central - PubMed

Affiliation: Group of Maintenance of Genome Stability, Max Planck Institute of Biochemistry, Martinsried, Germany.

ABSTRACT
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.

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Quantification of DNA, mRNA and protein abundance. (A) Ratios (log2) of DNA, mRNA and protein abundance changes between HCT116 5/4 and HCT116 aligned with respect to their chromosome position. Each dot represents abundance changes for one gene, the corresponding mRNA and the corresponding protein, respectively. CGH analysis revealed some deleted regions in chromosome 5 (upper panel); the disomic entries were omitted in all analyses (threshold 0.65 [log2]). (B) Overlays of mRNA and protein density histograms (HCT116 5/4 versus HCT116). The full line represents median of protein abundance changes, the dashed line median of mRNA abundance changes. Values of respective medians are plotted in the graph. The difference between distribution of proteins and mRNAs coded on chromosome 5 is statistically significant (Wilcoxon rank sum test). See also Supplementary Figure S2.
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f2: Quantification of DNA, mRNA and protein abundance. (A) Ratios (log2) of DNA, mRNA and protein abundance changes between HCT116 5/4 and HCT116 aligned with respect to their chromosome position. Each dot represents abundance changes for one gene, the corresponding mRNA and the corresponding protein, respectively. CGH analysis revealed some deleted regions in chromosome 5 (upper panel); the disomic entries were omitted in all analyses (threshold 0.65 [log2]). (B) Overlays of mRNA and protein density histograms (HCT116 5/4 versus HCT116). The full line represents median of protein abundance changes, the dashed line median of mRNA abundance changes. Values of respective medians are plotted in the graph. The difference between distribution of proteins and mRNAs coded on chromosome 5 is statistically significant (Wilcoxon rank sum test). See also Supplementary Figure S2.

Mentions: Using the series of isogenic diploid and aneuploid cell lines we asked how the mRNA and protein levels changed in response to the chromosome copy number changes. We examined the tetrasomic cell line HCT116 5/4, which guaranteed larger dynamic range of measured changes than in trisomic cells. To quantify the DNA levels, we used high-resolution CGH (Supplementary Table S1). To measure the corresponding mRNA, we determined the medians from microarray analysis of three independent samples (Supplementary Table S1). To compare the protein levels, we used stable isotope labeling with amino acids in cell culture (SILAC) followed by high-resolution mass spectrometry (Ong et al, 2002) and quantified the proteome to a depth of ∼6000 proteins (Supplementary Table S1). In total, we performed three biological replicates with six measurements for HCT116 5/4; all other cell lines were analyzed less extensively (up to three biological replicates, see Supplementary Table S1). The analysis of variability in technical and biological replicates demonstrated high quantification accuracy and reproducible measurements for all experiments including the reverse labeling, with Pearson correlation factors between 0.64 and 0.90 (Supplementary Figure S2A). We then determined the medians of quantitative protein changes. The log2 ratios of aneuploid-to-diploid abundance changes of DNA, the corresponding mRNAs and the proteins were matched directly using the annotated chromosomal positions (Figure 2A; Supplementary Figure S2B). In the case of equal levels in aneuploid and diploid cells, we expect the median aneuploid-to-diploid ratio (log2) to be 0 and this is observed for all DNA, mRNA and proteins coded on the disomes (DNA: 0.005, median mRNA: −0.03; proteins: −0.06; Figure 2B). The median ratio of mRNA encoded on the tetrasomic chromosome 5 is 1.09—very close to the expected value of 1.0—but in contrast, the median ratio of protein level changes is only 0.69 (Figure 2B). Indeed, 53 out of 197 proteins (27%) coded on the tetrasomic chromosome 5 are present at levels expected for disomes (median of diploid levels±twice the standard deviation). The calculated median of protein abundance of these 53 proteins is 0.26, whereas the median of the corresponding mRNAs is 1.16, further strengthening the idea that their expression is adjusted at the protein level. Similarly, 25% of proteins coded on chromosome 3 are present at diploid levels in HCT116 3/3 (Supplementary Table S1). We determined that the proteins coded on the multisomic chromosome are present at a lower level than expected in all analyzed cell lines (Supplementary Figure S3). In conclusion, whereas the mRNA expression corresponds to the increased chromosome copy numbers, approximately a quarter of the proteins are present at levels lower than expected and more similar to the disomic levels.


Global analysis of genome, transcriptome and proteome reveals the response to aneuploidy in human cells.

Stingele S, Stoehr G, Peplowska K, Cox J, Mann M, Storchova Z - Mol. Syst. Biol. (2012)

Quantification of DNA, mRNA and protein abundance. (A) Ratios (log2) of DNA, mRNA and protein abundance changes between HCT116 5/4 and HCT116 aligned with respect to their chromosome position. Each dot represents abundance changes for one gene, the corresponding mRNA and the corresponding protein, respectively. CGH analysis revealed some deleted regions in chromosome 5 (upper panel); the disomic entries were omitted in all analyses (threshold 0.65 [log2]). (B) Overlays of mRNA and protein density histograms (HCT116 5/4 versus HCT116). The full line represents median of protein abundance changes, the dashed line median of mRNA abundance changes. Values of respective medians are plotted in the graph. The difference between distribution of proteins and mRNAs coded on chromosome 5 is statistically significant (Wilcoxon rank sum test). See also Supplementary Figure S2.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC3472693&req=5

f2: Quantification of DNA, mRNA and protein abundance. (A) Ratios (log2) of DNA, mRNA and protein abundance changes between HCT116 5/4 and HCT116 aligned with respect to their chromosome position. Each dot represents abundance changes for one gene, the corresponding mRNA and the corresponding protein, respectively. CGH analysis revealed some deleted regions in chromosome 5 (upper panel); the disomic entries were omitted in all analyses (threshold 0.65 [log2]). (B) Overlays of mRNA and protein density histograms (HCT116 5/4 versus HCT116). The full line represents median of protein abundance changes, the dashed line median of mRNA abundance changes. Values of respective medians are plotted in the graph. The difference between distribution of proteins and mRNAs coded on chromosome 5 is statistically significant (Wilcoxon rank sum test). See also Supplementary Figure S2.
Mentions: Using the series of isogenic diploid and aneuploid cell lines we asked how the mRNA and protein levels changed in response to the chromosome copy number changes. We examined the tetrasomic cell line HCT116 5/4, which guaranteed larger dynamic range of measured changes than in trisomic cells. To quantify the DNA levels, we used high-resolution CGH (Supplementary Table S1). To measure the corresponding mRNA, we determined the medians from microarray analysis of three independent samples (Supplementary Table S1). To compare the protein levels, we used stable isotope labeling with amino acids in cell culture (SILAC) followed by high-resolution mass spectrometry (Ong et al, 2002) and quantified the proteome to a depth of ∼6000 proteins (Supplementary Table S1). In total, we performed three biological replicates with six measurements for HCT116 5/4; all other cell lines were analyzed less extensively (up to three biological replicates, see Supplementary Table S1). The analysis of variability in technical and biological replicates demonstrated high quantification accuracy and reproducible measurements for all experiments including the reverse labeling, with Pearson correlation factors between 0.64 and 0.90 (Supplementary Figure S2A). We then determined the medians of quantitative protein changes. The log2 ratios of aneuploid-to-diploid abundance changes of DNA, the corresponding mRNAs and the proteins were matched directly using the annotated chromosomal positions (Figure 2A; Supplementary Figure S2B). In the case of equal levels in aneuploid and diploid cells, we expect the median aneuploid-to-diploid ratio (log2) to be 0 and this is observed for all DNA, mRNA and proteins coded on the disomes (DNA: 0.005, median mRNA: −0.03; proteins: −0.06; Figure 2B). The median ratio of mRNA encoded on the tetrasomic chromosome 5 is 1.09—very close to the expected value of 1.0—but in contrast, the median ratio of protein level changes is only 0.69 (Figure 2B). Indeed, 53 out of 197 proteins (27%) coded on the tetrasomic chromosome 5 are present at levels expected for disomes (median of diploid levels±twice the standard deviation). The calculated median of protein abundance of these 53 proteins is 0.26, whereas the median of the corresponding mRNAs is 1.16, further strengthening the idea that their expression is adjusted at the protein level. Similarly, 25% of proteins coded on chromosome 3 are present at diploid levels in HCT116 3/3 (Supplementary Table S1). We determined that the proteins coded on the multisomic chromosome are present at a lower level than expected in all analyzed cell lines (Supplementary Figure S3). In conclusion, whereas the mRNA expression corresponds to the increased chromosome copy numbers, approximately a quarter of the proteins are present at levels lower than expected and more similar to the disomic levels.

Bottom Line: 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.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.

View Article: PubMed Central - PubMed

Affiliation: Group of Maintenance of Genome Stability, Max Planck Institute of Biochemistry, Martinsried, Germany.

ABSTRACT
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.

Show MeSH