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FireDB (http://firedb.bioinfo.cnio.es) is a curated inventory of catalytic and biologically relevant small ligand-binding residues culled from the protein structures in the Protein Data Bank. Here we present the important new additions since the publication of FireDB in 2007. The database now contains an extensive list of manually curated biologically relevant compounds. Biologically relevant compounds are informative because of their role in protein function, but they are only a small fraction of the entire ligand set. For the remaining ligands, the FireDB provides cross-references to the annotations from publicly available biological, chemical and pharmacological compound databases. FireDB now has external references for 95% of contacting small ligands, making FireDB a more complete database and providing the scientific community with easy access to the pharmacological annotations of PDB ligands. In addition to the manual curation of ligands, FireDB also provides insights into the biological relevance of individual binding sites. Here, biological relevance is calculated from the multiple sequence alignments of related binding sites that are generated from all-against-all comparison of each FireDB binding site. The database can be accessed by RESTful web services and is available for download via MySQL. ; The Spanish Ministry of Economy and Competitiveness (BIO2012-40205); technical assistance was provided by the Spanish Bioinformatics Institute (INB), a platform of the ISCII; European Union [FP7-REGPOT-2012-CT2012-31637-IMBRAIN to L.G.L.]. Funding for open access charge: Spanish Ministry of Economy and Competitiveness [grant: BIO2012-40205] ; Sí

The RNA polymerase II-associated protein 1 (RPAP1) is conserved across metazoa and required for stem cell differentiation in plants; however, very little is known about its mechanism of action or its role in mammalian cells. Here, we report that RPAP1 is essential for the expression of cell identity genes and for cell viability. Depletion of RPAP1 triggers cell de-differentiation, facilitates reprogramming toward pluripotency, and impairs differentiation. Mechanistically, we show that RPAP1 is essential for the interaction between RNA polymerase II (RNA Pol II) and Mediator, as well as for the recruitment of important regulators, such as the Mediator-specific RNA Pol II factor Gdown1 and the C-terminal domain (CTD) phosphatase RPAP2. In agreement, depletion of RPAP1 diminishes the loading of total and Ser5-phosphorylated RNA Pol II on many genes, with super-enhancer-driven genes among the most significantly downregulated. We conclude that Mediator/RPAP1/RNA Pol II is an ancient module, conserved from plants to mammals, critical for establishing and maintaining cell identity. ; We are grateful to Elisa Varela for assistance with morula and blastocyst fixa- tion. Work in the laboratory of M.S. is funded by the CNIO and the IRB and by grants from the Spanish Ministry of Economy co-funded by the European Regional Development Fund (ERDF) (SAF2013-48256-R), the European Research Co uncil (ERC-2014-AdG/66 9622), the Region al Government of Ma- drid co-funded by the Euro pean Social Fund (ReCaRe project), the Euro pean Union (RISK-IR project), the Botin Foundation and Banco Santander (Santander Universities Glo bal Division), the Ramon Areces Found ation, and the AXA Foundation. S.R. was funded by a contract from the Ramon y Cajal Program(RYC-2011-09242) and by the Spanish Ministry of Economy co- funded by the ERDF (SAF2013-49147- P and SAF2016-80874-P ; Sí

Triple-negative breast cancer (TNBC) lacks prognostic and predictive markers. Here, we use high-throughput phosphoproteomics to build a functional TNBC taxonomy. A cluster of 159 phosphosites is upregulated in relapsed cases of a training set (n = 34 patients), with 11 hyperactive kinases accounting for this phosphoprofile. A mass-spectrometry-to-immunohistochemistry translation step, assessing 2 independent validation sets, reveals 6 kinases with preserved independent prognostic value. The kinases split the validation set into two patterns: one without hyperactive kinases being associated with a >90% relapse-free rate, and the other one showing ≥1 hyperactive kinase and being associated with an up to 9.5-fold higher relapse risk. Each kinase pattern encompasses different mutational patterns, simplifying mutation-based taxonomy. Drug regimens designed based on these 6 kinases show promising antitumour activity in TNBC cell lines and patient-derived xenografts. In summary, the present study elucidates phosphosites and kinases implicated in TNBC and suggests a target-based clinical classification system for TNBC. ; We are grateful to the Spanish Biobanks integrated within the Spanish Network of National Biobanks for the tumor samples used in our investigations and to the Breast Cancer Group at VHIO for providing study materials. The present study was funded by FIS PI10/00288, FIS PI13/00430, and AECC Scientific Foundation "Beca de Retorno 2010"awarded to MQF; FIS PI11-00832, FIS PI14-00726, and ISCIII PIE15/00068 awarded to RC. I.Z. is a recipient of a La Caixa PhD Fellowship, 2011. S.F. is a recipient of a FPI grant (SEV-2015-0510-16-6). V.S. is supported by the Miguel Servet Program (CP14/00228). This work was also supported by the PRIME-XS project, Grant Agree- ment Number 262067, funded by the European Union Seventh Framework Program and the project Proteins At Work (project 184.032.201), a program of the Netherlands Proteomics Centre financed by the Netherlands Organisation for Scientific Research (NWO). ...