Clustered mutation signatures reveal that error-prone DNA repair targets mutations to active genes

Many processes can cause the same nucleotide change in a genome, making the identification of the mechanisms causing mutations a difficult challenge. Here, we show that clustered mutations provide a more precise fingerprint of mutagenic processes. Of nine clustered mutation signatures identified from >1,000 tumor genomes, three relate to variable APOBEC activity and three are associated with tobacco smoking. An additional signature matches the spectrum of translesion DNA polymerase eta (POLH). In lymphoid cells, these mutations target promoters, consistent with AID-initiated somatic hypermutation. In solid tumors, however, they are associated with UV exposure and alcohol consumption and target the H3K36me3 chromatin of active genes in a mismatch repair (MMR)-dependent manner. These regions normally have a low mutation rate because error-free MMR also targets H3K36me3 chromatin. Carcinogens and error-prone repair therefore redistribute mutations to the more important regions of the genome, contributing a substantial mutation load in many tumors, including driver mutations.This work was supported by grants from the ERC (616434), MINECO (BFU2011-26206 and SEV-2012-0208), AXA Research Fund (AXA Chair in Risk Prediction in Age-related Diseases), Bettencourt Schueller Foundation, AGAUR (2014 SGR 831), FP7 (projects 4DCellFate 277899, MAESTRA ICT-2013-612944, and InnoMol FP7-REGPOT-2012-2013-1-316289), the EMBL-CRG Systems Biology Program, and the CERCA progra

Cancer type-dependent genetic interactions between cancer driver alterations indicate plasticity of epistasis across cell types

Cancers, like many diseases, are normally caused by combinations of genetic alterations rather than by changes affecting single genes. It is well established that the genetic alterations that drive cancer often interact epistatically, having greater or weaker consequences in combination than expected from their individual effects. In a stringent statistical analysis of data from > 3,000 tumors, we find that the co-occurrence and mutual exclusivity relationships between cancer driver alterations change quite extensively in different types of cancer. This cannot be accounted for by variation in tumor heterogeneity or unrecognized cancer subtypes. Rather, it suggests that how genomic alterations interact cooperatively or partially redundantly to driver cancer changes in different types of cancers. This re-wiring of epistasis across cell types is likely to be a basic feature of genetic architecture, with important implications for understanding the evolution of multicellularity and human genetic diseases. In addition, if this plasticity of epistasis across cell types is also true for synthetic lethal interactions, a synthetic lethal strategy to kill cancer cells may frequently work in one type of cancer but prove ineffective in another.S.P. was funded by a Postdoctoral Fellowship from Novartis and by the Juan de la Cierva program (MINECO). Work in the laboratory of B.L. is funded by a European Research Council (ERC) Consolidator Grant (IR-DC, GA616434), Plan Nacional Grant BFU2011-26206 (MINECO), AGAUR (2009 SGR 112), the EMBO Young Investigator Program (MINECO), EU Framework 7 project 277899 4DCellFate, the EMBLCRG Systems Biology Program, the Severo Ochoa Program, and the AXA Research Fund

Human genes with CpG island promoters have a distinct transcription-associated chromatin organization

Background: More than 50% of human genes initiate transcription from CpG dinucleotide-rich regions referred to as CpG islands. These genes show differences in their patterns of transcription initiation, and have been reported to have higher levels of some activation-associated chromatin modifications. Results: Here we report that genes with CpG island promoters have a characteristic transcription-associated chromatin organization. This signature includes high levels of the transcription elongation-associated histone modifications H4K20me1, H2BK5me1 and H3K79me1/2/3 in the 5' end of the gene, depletion of the activation marks H2AK5ac, H3K14ac and H3K23ac immediately downstream of the transcription start site (TSS), and characteristic epigenetic asymmetries around the TSS. The chromosome organization factor CTCF may be bound upstream of RNA polymerase in most active CpG island promoters, and an unstable nucleosome at the TSS may be specifically marked by H4K20me3, the first example of such a modification. H3K36 monomethylation is only detected as enriched in the bodies of active genes that have CpG island promoters. Finally, as expression levels increase, peak modification levels of the histone methylations H3K9me1, H3K4me1, H3K4me2 and H3K27me1 shift further away from the TSS into the gene body. Conclusions: These results suggest that active genes with CpG island promoters have a distinct step-like series of modified nucleosomes after the TSS. The identity, positioning, shape and relative ordering of transcription-associated histone modifications differ between genes with and without CpG island promoters. This supports a model where chromatin organization reflects not only transcription activity but also the type of promoter in which transcription initiates.TV is funded by MICINN grant BFU2011-30246, Ramon y Cajal grant RYC-2010-07114, European Commission Framework 7 European Re-integration grant PERG08-GA-2010-276741, and by the Institute of Predictive and Personalized Medicine of Cancer. BL is funded by an ERC Starting Grant, ERASysBio+ ERANET, MICINN grant BFU2008-00365, AGAUR, the EMBO Young Investigator Program, European Commission Framework 7 integrated project 4DCellFate, and by the EMBL-CRG Systems Biology Progra

Determining protein structures using deep mutagenesis

Determining the three-dimensional structures of macromolecules is a major goal of biological research, because of the close relationship between structure and function; however, thousands of protein domains still have unknown structures. Structure determination usually relies on physical techniques including X-ray crystallography, NMR spectroscopy and cryo-electron microscopy. Here we present a method that allows the high-resolution three-dimensional backbone structure of a biological macromolecule to be determined only from measurements of the activity of mutant variants of the molecule. This genetic approach to structure determination relies on the quantification of genetic interactions (epistasis) between mutations and the discrimination of direct from indirect interactions. This provides an alternative experimental strategy for structure determination, with the potential to reveal functional and in vivo structures.This work was supported by a European Research Council (ERC) Consolidator grant (616434), the Spanish Ministry of Economy, Industry and Competitiveness (MEIC; BFU2017-89488-P), the AXA Research Fund, the Bettencourt Schueller Foundation, Agencia de Gestio d’Ajuts Universitaris i de Recerca (AGAUR, 2017 SGR 1322), the EMBL-CRG Systems Biology Program and the CERCA Program/Generalitat de Catalunya. J.M.S. was supported by an EMBO Long-Term Fellowship (ALTF 857-2016). This project has received funding from the European Union’s Horizon 2020 research and innovation program under the Marie Skłodowska-Curie grant agreement 752809 (J.M.S.). We acknowledge support from the Spanish Ministry of Economy, Industry and Competitiveness (MEIC) to the EMBL partnership and the Centro de Excelencia Severo Ochoa

Clustered mutation signatures reveal that error-prone DNA repair targets mutations to active genes

Many processes can cause the same nucleotide change in a genome, making the identification of the mechanisms causing mutations a difficult challenge. Here, we show that clustered mutations provide a more precise fingerprint of mutagenic processes. Of nine clustered mutation signatures identified from >1,000 tumor genomes, three relate to variable APOBEC activity and three are associated with tobacco smoking. An additional signature matches the spectrum of translesion DNA polymerase eta (POLH). In lymphoid cells, these mutations target promoters, consistent with AID-initiated somatic hypermutation. In solid tumors, however, they are associated with UV exposure and alcohol consumption and target the H3K36me3 chromatin of active genes in a mismatch repair (MMR)-dependent manner. These regions normally have a low mutation rate because error-free MMR also targets H3K36me3 chromatin. Carcinogens and error-prone repair therefore redistribute mutations to the more important regions of the genome, contributing a substantial mutation load in many tumors, including driver mutations.This work was supported by grants from the ERC (616434), MINECO (BFU2011-26206 and SEV-2012-0208), AXA Research Fund (AXA Chair in Risk Prediction in Age-related Diseases), Bettencourt Schueller Foundation, AGAUR (2014 SGR 831), FP7 (projects 4DCellFate 277899, MAESTRA ICT-2013-612944, and InnoMol FP7-REGPOT-2012-2013-1-316289), the EMBL-CRG Systems Biology Program, and the CERCA progra

The genetic landscape of a physical interaction

A key question in human genetics and evolutionary biology is how mutations in different genes combine to alter phenotypes. Efforts to systematically map genetic interactions have mostly made use of gene deletions. However, most genetic variation consists of point mutations of diverse and difficult to predict effects. Here, by developing a new sequencing-based protein interaction assay - deepPCA - we quantified the effects of >120,000 pairs of point mutations on the formation of the AP-1 transcription factor complex between the products of the FOS and JUN proto-oncogenes. Genetic interactions are abundant both in cis (within one protein) and trans (between the two molecules) and consist of two classes - interactions driven by thermodynamics that can be predicted using a three-parameter global model, and structural interactions between proximally located residues. These results reveal how physical interactions generate quantitatively predictable genetic interactions.This work was supported by a European Research Council (ERC) Consolidator grant (616434), the Spanish Ministry of Economy and Competitiveness (BFU2011-26206 and ‘Centro de Excelencia Severo Ochoa' SEV-2012–0208), the AXA Research Fund, the Bettencourt Schueller Foundation, Agencia de Gestio d’Ajuts Universitaris i de Recerca (AGAUR, SGR-831), the EMBL-CRG Systems Biology Program, and the CERCA Program/Generalitat de Catalunya. GD is a non-stipendiary EMBO Fellow and a Marie-Curie Fellow under grant agreement 608959

Differential DNA mismatch repair underlies mutation rate variation across the human genome

Cancer genome sequencing has revealed considerable variation in somatic mutation rates across the human genome, with mutation rates elevated in heterochromatic late replicating regions and reduced in early replicating euchromatin. Multiple mechanisms have been suggested to underlie this, but the actual cause is unknown. Here we identify variable DNA mismatch repair (MMR) as the basis of this variation. Analysing ∼17 million single-nucleotide variants from the genomes of 652 tumours, we show that regional autosomal mutation rates at megabase resolution are largely stable across cancer types, with differences related to changes in replication timing and gene expression. However, mutations arising after the inactivation of MMR are no longer enriched in late replicating heterochromatin relative to early replicating euchromatin. Thus, differential DNA repair and not differential mutation supply is the primary cause of the large-scale regional mutation rate variation across the human genome.This work was supported by grants from the Spanish Ministry of Economy and Competitiveness (BFU2011-26206 and ‘Centro de Excelencia Severo Ochoa 2013-2017’ SEV-2012-0208), a European Research Council Consolidator grant IR-DC (616434), Agència de Gestió d’Ajuts Universitaris i de Recerca (AGAUR),the EMBOYoung Investigator Program, the EMBL-CRG Systems Biology Program, FP7 project 4DCellFate (277899),FP7 project MAESTRA (ICT-2013-612944)and by Marie Curie Actions

Harmonious genetic combinations rewire regulatory networks and flip gene essentiality

We lack an understanding of how the full range of genetic variants that occur in individuals can interact. To address this shortcoming, here we combine diverse mutations between genes in a model regulatory network, the galactose (GAL) switch of budding yeast. The effects of thousands of pairs of mutations fall into a limited number of phenotypic classes. While these effects are mostly predictable using simple rules that capture the 'stereotypical' genetic interactions of the network, some double mutants have unexpected outcomes including constituting alternative functional switches. Each of these 'harmonious' genetic combinations exhibits altered dependency on other regulatory genes. These cases illustrate how both pairwise and higher epistasis determines gene essentiality and how combinations of mutations rewire regulatory networks. Together, our results provide an overview of how broad spectra of mutations interact, how these interactions can be predicted, and how diverse genetic solutions can achieve 'wild-type' phenotypic behavior.This work was supported by a European Research Council (ERC) Consolidator grant (616434), the Spanish Ministry of Economy and Competitiveness (BFU2017–89488-P and SEV-2012–0208), the AXA Research Fund, the Bettencourt Schueller Foundation, Agencia de Gestio d’Ajuts Universitaris i de Recerca (AGAUR, 2017 SGR 1322.), and the CERCA Program/Generalitat de Catalunya. A.M. New was supported in part by fellowships from EMBO ALTF 505–2014 and the Spanish Ministry of Economy and Competitiveness Juan de la Cierva fund

Harmonious genetic combinations rewire regulatory networks and flip gene essentiality

We lack an understanding of how the full range of genetic variants that occur in individuals can interact. To address this shortcoming, here we combine diverse mutations between genes in a model regulatory network, the galactose (GAL) switch of budding yeast. The effects of thousands of pairs of mutations fall into a limited number of phenotypic classes. While these effects are mostly predictable using simple rules that capture the 'stereotypical' genetic interactions of the network, some double mutants have unexpected outcomes including constituting alternative functional switches. Each of these 'harmonious' genetic combinations exhibits altered dependency on other regulatory genes. These cases illustrate how both pairwise and higher epistasis determines gene essentiality and how combinations of mutations rewire regulatory networks. Together, our results provide an overview of how broad spectra of mutations interact, how these interactions can be predicted, and how diverse genetic solutions can achieve 'wild-type' phenotypic behavior.This work was supported by a European Research Council (ERC) Consolidator grant (616434), the Spanish Ministry of Economy and Competitiveness (BFU2017–89488-P and SEV-2012–0208), the AXA Research Fund, the Bettencourt Schueller Foundation, Agencia de Gestio d’Ajuts Universitaris i de Recerca (AGAUR, 2017 SGR 1322.), and the CERCA Program/Generalitat de Catalunya. A.M. New was supported in part by fellowships from EMBO ALTF 505–2014 and the Spanish Ministry of Economy and Competitiveness Juan de la Cierva fund

Intergenerational and transgenerational epigenetic inheritance in animals

Animals transmit not only DNA but also other molecules, such as RNA, proteins and metabolites, to their progeny via gametes. It is currently unclear to what extent these molecules convey information between generations and whether this information changes according to their physiological state and environment. Here, we review recent work on the molecular mechanisms by which 'epigenetic' information is transmitted between generations over different timescales, and the importance of this information for development and physiology.This work was supported by a European Research Council (ERC) consolidator grant (616434), the Spanish Ministry of Economy and Competitiveness (BFU2017-89488-P and SEV-2012-0208), the AXA Research Fund, the Bettencourt Schueller Foundation, Agencia de Gestio d’Ajuts Universitaris i de Recerca (AGAUR; SGR-831), the EMBL-CRG Systems Biology Program and the CERCA Program/Generalitat de Catalunya