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SETD2 loss-of-function promotes renal cancer branched evolution through replication stress and impaired DNA repair.

Kanu N, Grönroos E, Martinez P, Burrell RA, Yi Goh X, Bartkova J, Maya-Mendoza A, Mistrík M, Rowan AJ, Patel H, Rabinowitz A, East P, Wilson G, Santos CR, McGranahan N, Gulati S, Gerlinger M, Birkbak NJ, Joshi T, Alexandrov LB, Stratton MR, Powles T, Matthews N, Bates PA, Stewart A, Szallasi Z, Larkin J, Bartek J, Swanton C - Oncogene (2015)

Bottom Line: Distinct SETD2 mutations have been identified in spatially separated regions in ccRCC, indicative of intratumour heterogeneity.In this study, we have addressed the consequences of SETD2 loss-of-function through an integrated bioinformatics and functional genomics approach.Consistent with these data, we observe chromosomal breakpoint locations are biased away from H3K36me3 sites in SETD2 wild-type ccRCCs relative to tumours with bi-allelic SETD2 aberrations and that H3K36me3-negative ccRCCs display elevated DNA damage in vivo.

View Article: PubMed Central - PubMed

Affiliation: UCL Cancer Institute, Paul O'Gorman Building, London, UK.

ABSTRACT
Defining mechanisms that generate intratumour heterogeneity and branched evolution may inspire novel therapeutic approaches to limit tumour diversity and adaptation. SETD2 (Su(var), Enhancer of zeste, Trithorax-domain containing 2) trimethylates histone-3 lysine-36 (H3K36me3) at sites of active transcription and is mutated in diverse tumour types, including clear cell renal carcinomas (ccRCCs). Distinct SETD2 mutations have been identified in spatially separated regions in ccRCC, indicative of intratumour heterogeneity. In this study, we have addressed the consequences of SETD2 loss-of-function through an integrated bioinformatics and functional genomics approach. We find that bi-allelic SETD2 aberrations are not associated with microsatellite instability in ccRCC. SETD2 depletion in ccRCC cells revealed aberrant and reduced nucleosome compaction and chromatin association of the key replication proteins minichromosome maintenance complex component (MCM7) and DNA polymerase δ hindering replication fork progression, and failure to load lens epithelium-derived growth factor and the Rad51 homologous recombination repair factor at DNA breaks. Consistent with these data, we observe chromosomal breakpoint locations are biased away from H3K36me3 sites in SETD2 wild-type ccRCCs relative to tumours with bi-allelic SETD2 aberrations and that H3K36me3-negative ccRCCs display elevated DNA damage in vivo. These data suggest a role for SETD2 in maintaining genome integrity through nucleosome stabilization, suppression of replication stress and the coordination of DNA repair.

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SETD2 mutation exposes H3K36me3 sites to chromosome breakage. (a) Schematic illustrating the mapping of chromosomal breakpoint regions to sites of H3K36me3/H3K27me3 (data from the ENCODE consortium) in both SETD2LOH and SETD2mut. Breakpoint regions, identified from SNP 6.0 array data, are the regions between two segments of a chromosome present at different allele-specific copy numbers. (b) Representative plots of observed H3K36me3 frequencies against expected frequencies in SETD2LOH and SETD2mut tumours, using a minimum segment length of 10 Mb and a maximum breakpoint region length of 20 kb.
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fig2: SETD2 mutation exposes H3K36me3 sites to chromosome breakage. (a) Schematic illustrating the mapping of chromosomal breakpoint regions to sites of H3K36me3/H3K27me3 (data from the ENCODE consortium) in both SETD2LOH and SETD2mut. Breakpoint regions, identified from SNP 6.0 array data, are the regions between two segments of a chromosome present at different allele-specific copy numbers. (b) Representative plots of observed H3K36me3 frequencies against expected frequencies in SETD2LOH and SETD2mut tumours, using a minimum segment length of 10 Mb and a maximum breakpoint region length of 20 kb.

Mentions: Chromosomal breakpoints were identified using single-nucleotide polymorphism (SNP) array data, the resolution of which only permits identification of breakpoint regions (between two genomic segments present at different copy numbers), rather than specific sites of breakage. Breakpoint regions were then defined using 16 different threshold combinations for minimum flanking segment length and maximum breakpoint region length, to take into account the occurrence of false positives in the predicted chromosomal aberrations (Supplementary Table S3 and Supplementary Methods). Using this cohort, we assessed the location of breakpoint regions, with respect to H3K36me3 sites in normal adult kidney, in the 210 SETD2LOH tumours and the 29 SETD2mut tumours (Figure 2a and Supplementary Figure S1a). We generated 5000 random breakpoint region profiles to estimate the expected background frequency distributions of H3K36me3 sites within breakpoint regions for SETD2mut versus SETD2LOH TCGA tumours, for each threshold combination.


SETD2 loss-of-function promotes renal cancer branched evolution through replication stress and impaired DNA repair.

Kanu N, Grönroos E, Martinez P, Burrell RA, Yi Goh X, Bartkova J, Maya-Mendoza A, Mistrík M, Rowan AJ, Patel H, Rabinowitz A, East P, Wilson G, Santos CR, McGranahan N, Gulati S, Gerlinger M, Birkbak NJ, Joshi T, Alexandrov LB, Stratton MR, Powles T, Matthews N, Bates PA, Stewart A, Szallasi Z, Larkin J, Bartek J, Swanton C - Oncogene (2015)

SETD2 mutation exposes H3K36me3 sites to chromosome breakage. (a) Schematic illustrating the mapping of chromosomal breakpoint regions to sites of H3K36me3/H3K27me3 (data from the ENCODE consortium) in both SETD2LOH and SETD2mut. Breakpoint regions, identified from SNP 6.0 array data, are the regions between two segments of a chromosome present at different allele-specific copy numbers. (b) Representative plots of observed H3K36me3 frequencies against expected frequencies in SETD2LOH and SETD2mut tumours, using a minimum segment length of 10 Mb and a maximum breakpoint region length of 20 kb.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4660036&req=5

fig2: SETD2 mutation exposes H3K36me3 sites to chromosome breakage. (a) Schematic illustrating the mapping of chromosomal breakpoint regions to sites of H3K36me3/H3K27me3 (data from the ENCODE consortium) in both SETD2LOH and SETD2mut. Breakpoint regions, identified from SNP 6.0 array data, are the regions between two segments of a chromosome present at different allele-specific copy numbers. (b) Representative plots of observed H3K36me3 frequencies against expected frequencies in SETD2LOH and SETD2mut tumours, using a minimum segment length of 10 Mb and a maximum breakpoint region length of 20 kb.
Mentions: Chromosomal breakpoints were identified using single-nucleotide polymorphism (SNP) array data, the resolution of which only permits identification of breakpoint regions (between two genomic segments present at different copy numbers), rather than specific sites of breakage. Breakpoint regions were then defined using 16 different threshold combinations for minimum flanking segment length and maximum breakpoint region length, to take into account the occurrence of false positives in the predicted chromosomal aberrations (Supplementary Table S3 and Supplementary Methods). Using this cohort, we assessed the location of breakpoint regions, with respect to H3K36me3 sites in normal adult kidney, in the 210 SETD2LOH tumours and the 29 SETD2mut tumours (Figure 2a and Supplementary Figure S1a). We generated 5000 random breakpoint region profiles to estimate the expected background frequency distributions of H3K36me3 sites within breakpoint regions for SETD2mut versus SETD2LOH TCGA tumours, for each threshold combination.

Bottom Line: Distinct SETD2 mutations have been identified in spatially separated regions in ccRCC, indicative of intratumour heterogeneity.In this study, we have addressed the consequences of SETD2 loss-of-function through an integrated bioinformatics and functional genomics approach.Consistent with these data, we observe chromosomal breakpoint locations are biased away from H3K36me3 sites in SETD2 wild-type ccRCCs relative to tumours with bi-allelic SETD2 aberrations and that H3K36me3-negative ccRCCs display elevated DNA damage in vivo.

View Article: PubMed Central - PubMed

Affiliation: UCL Cancer Institute, Paul O'Gorman Building, London, UK.

ABSTRACT
Defining mechanisms that generate intratumour heterogeneity and branched evolution may inspire novel therapeutic approaches to limit tumour diversity and adaptation. SETD2 (Su(var), Enhancer of zeste, Trithorax-domain containing 2) trimethylates histone-3 lysine-36 (H3K36me3) at sites of active transcription and is mutated in diverse tumour types, including clear cell renal carcinomas (ccRCCs). Distinct SETD2 mutations have been identified in spatially separated regions in ccRCC, indicative of intratumour heterogeneity. In this study, we have addressed the consequences of SETD2 loss-of-function through an integrated bioinformatics and functional genomics approach. We find that bi-allelic SETD2 aberrations are not associated with microsatellite instability in ccRCC. SETD2 depletion in ccRCC cells revealed aberrant and reduced nucleosome compaction and chromatin association of the key replication proteins minichromosome maintenance complex component (MCM7) and DNA polymerase δ hindering replication fork progression, and failure to load lens epithelium-derived growth factor and the Rad51 homologous recombination repair factor at DNA breaks. Consistent with these data, we observe chromosomal breakpoint locations are biased away from H3K36me3 sites in SETD2 wild-type ccRCCs relative to tumours with bi-allelic SETD2 aberrations and that H3K36me3-negative ccRCCs display elevated DNA damage in vivo. These data suggest a role for SETD2 in maintaining genome integrity through nucleosome stabilization, suppression of replication stress and the coordination of DNA repair.

Show MeSH
Related in: MedlinePlus