Suchergebnisse

The relationship between the 4D folding of the genome and its function is an outstanding question in biology. A range of methods that probe the folding of the genome in space and time with unprecedented resolution have been developed. These methods, including chromosome conformation capture and high-resolution light and electron microscopy, are shedding new light on genome architecture and function. Here, we review the emerging picture of genome organization revealed by super-resolution and live-cell imaging. We compare and contrast population-based chromosome conformation capture approaches and imaging-based approaches and highlight future challenges. ; This work was supported by the University of Pennsylvania Epigenetics Pilot Award (to M.L.), the Center for Engineering and Mechanobiology (CEMB), an NSF Science and Technology Center Pilot Award under grant agreement CMMI 15-48571 (to M.L.), a Linda Pechenik Montague Investigator Award (to M.L.), the European Union's Horizon 2020 Research and Innovation Programme (CellViewer No 686637 to M.L. and M.P.C.), Ministerio de Ciencia e Innovación, grant BFU2017-86760-P (AEI/FEDER, UE), and an AGAUR grant from Secretaria d'Universitats i Recerca del Departament d'Empresa i Coneixement de la Generalitat de Catalunya (2017 SGR 689 to M.P.C.). We acknowledge the support of the Spanish Ministry of Science and Innovation to the EMBL partnership, the Centro de Excelencia Severo Ochoa and the CERCA Programme / Generalitat de Catalunya

Chromatin fiber organization is essential for gene function in all cell types. Moreover it helps to determine cell fate in embryonic/adult stem cells and in somatic cells undergoing reprogramming to pluripotency. Until now the diffraction limit of light has limited the inspection of the chromatin fiber organization to a level sufficient to understand how it impacts gene function. The development of advanced microscopy methods, such as single molecule localization microscopy, has largely opened a new field of research providing us with the tools to visualize and quantitatively analyze chromatin fiber organization and thus gene activity at nanoscale resolution in single cells. ; We are grateful for support from Ministerio de Economia y Competitividad and FEDER funds (SAF2011-28580, BFU2014-54717-P, and BFU2015-71984-ERC to MPC, FIS2015-63550-R to ML, BFU2013-49867-EXP to MPC and ML), ERC 337191-MOTORS to ML, AGAUR grant (2014 SGR1137 to MPC), the European Union's Horizon 2020 research and innovation programme under grant agreement CellViewer No. 686637 (to MPC and ML), Spanish Ministry of Economy and Competitiveness, 'Centro de Excelencia Severo Ochoa 2013–2017 and the CERCA Programme/Generalitat de Catalunya (to MPC).

Homotypic and heterotypic cell-to-cell fusion are key processes during development and tissue regeneration. Nevertheless, aberrant cell fusion can contribute to tumour initiation and metastasis. Additionally, a form of cell-in-cell structure called entosis has been observed in several human tumours. Here we investigate cell-to-cell interaction between mouse mesenchymal stem cells (MSCs) and embryonic stem cells (ESCs). MSCs represent an important source of adult stem cells since they have great potential for regenerative medicine, even though they are also involved in cancer progression. We report that MSCs can either fuse forming heterokaryons, or be invaded by ESCs through entosis. While entosis-derived hybrids never share their genomes and induce degradation of the target cell, fusion-derived hybrids can convert into synkaryons. Importantly we show that hetero-to-synkaryon transition occurs through cell division and not by nuclear membrane fusion. Additionally, we also observe that the ROCK-actin/myosin pathway is required for both fusion and entosis in ESCs but only for entosis in MSCs. Overall, we show that MSCs can undergo fusion or entosis in culture by generating distinct functional cellular entities. These two processes are profoundly different and their outcomes should be considered given the beneficial or possible detrimental effects of MSC-based therapeutic applications. ; We are grateful for support from an ERC grant (242630-RERE to M.P.C.), Ministerio de Economia y Competitividad and FEDER funds (BFU2014-54717-P, and BFU2015-71984-ERC to M.P.C.), AGAUR grant (2014 SGR1137 to M.P.C.), the European Union's Horizon 2020 research and innovation programme under grant agreement CellViewer No 686637 (to M.P.C.), La Caixa international PhD fellowship (to F.S.), People Programme Marie Curie Actions of the European Union's Seventh Framework Programme (FP7/2007-2013/, n° 290123 to I.T.) and Ministerio de Ciencia e Innovacio´ FPI (to F.A.). We acknowledge support of the Spanish Ministry of Economy and Competitiveness, 'Centro de Excelencia Severo Ochoa 2013–2017'.

Imprinted genes control several cellular and metabolic processes in embryonic and adult tissues. In particular, paternally expressed gene-3 (Peg3) is active in the adult stem cell population and during muscle and neuronal lineage development. Here we have investigated the role of Peg3 in mouse embryonic stem cells (ESCs) and during the process of somatic cell reprogramming towards pluripotency. Our data show that Peg3 knockdown increases expression of pluripotency genes in ESCs and enhances reprogramming efficiency of both mouse embryonic fibroblasts and neural stem cells. Interestingly, we observed that altered activity of Peg3 correlates with major perturbations of mitochondrial gene expression and mitochondrial function, which drive metabolic changes during somatic cell reprogramming. Overall, our study shows that Peg3 is a regulator of pluripotent stem cells and somatic cell reprogramming. ; We thank Marie Victoire Neguembor, Martina Pesaresi and Sergi Angel Bonilla Pons for suggestions on the manuscript and Umberto Di Vicino for technical support. We thank Angelique di Domenico and the group of Antonella Consiglio for providing us the OPA1, DRP1 and VDAC1 antibodies. We thank Mari Carmen Ortells and the group of Bill Keyes for providing us the OXPHOS antibody. The pInducer10-mir-RUP-PheS plasmid was a gift from Stephen Elledge (Addgene plasmid # 44011). We are grateful for support from Ministerio de Economia y Competitividad and FEDER funds (SAF2011-28580, BFU2014-54717-P, and BFU2015-71984-ERC to M.P.C.), Secretaria d'Universitats i Recerca del Departament d'Economia i Coneixement de la Generalitat de Catalunya (2014 SGR1137 to M.P.C.), the European Union's Horizon 2020 research and innovation programme under grant agreement CellViewer No 686637 (to M.P.C.), Spanish Ministry of Economy and Competitiveness, 'Centro de Excelencia Severo Ochoa 2013-2017 and the CERCA Programme/Generalitat de Catalunya (to M.P.C.), People Programme Marie Curie Actions of the European Union's Seventh Framework Programme (FP7/2007-2013/, n° 290123 to I.T.), La Caixa international PhD fellowship (to F.S.), Ministerio de Ciencia e Innovació FPI (to F.A.) and Ministerio de Ciencia e Innovación FPI-Severo Ochoa (to A.C.G).

Here, we describe three complementary microscopy-based approaches to quantify morphological changes of chromatin organization in cultured adherent cells: the analysis of the coefficient of variation of DNA, the measurement of DNA-free nuclear areas, and the quantification of chromatin-associated proteins at the nuclear edge. These approaches rely on confocal imaging and stochastic optical reconstruction microscopy and allow a fast and robust quantification of chromatin compaction. These approaches circumvent inter-variability between imaging conditions and apply to every type of adherent cells. For complete details on the use and execution of this protocol, please refer to Neguembor et al. (2021). ; The authors acknowledge the support from European Union's Horizon 2020 Research and Innovation Programme (CellViewer no. 686637 to M.P.C. and M.L.); Ministerio de Ciencia e Innovación, grant (BFU2017-86760-P [AEI/FEDER, UE] to M.P.C.), and an AGAUR grant from Secretaria d'Universitats i Recerca del Departament d'Empresa i Coneixement de la Generalitat de Catalunya ([2017 SGR 689] to M.P.C.); Centro de Excelencia Severo Ochoa (2013–2017 to M.P.C.); CERCA Programme/Generalitat de Catalunya (to M.P.C.); the Spanish Ministry of Science and Innovation to the EMBL partnership (to M.P.C.); National Natural Science Foundation of China (31971177 to M.P.C.); Innovative Team Program of Guangzhou Regenerative Medicine and Health Guangdong Laboratory (2018GZR110103001 to M.P.C.); People Program (Marie Curie Actions) FP7/2007–2013 under REA grant 608959 (to M.V.N.); Juan de la Cierva-Incorporación 2017 (to M.V.N.); grant for the recruitment of early-stage research staff FI-2020 (Operational Program of Catalonia 2014-2020 CCI 2014ES05SFOP007 of the European Social Fund to L.M.); "La Caixa" Foundation Fellowship (ID 100010434, #LCF/BQ/DR20/11790016) (to L.M.); and "La Caixa-Severo Ochoa" pre-doctoral fellowship (to A.C.-G.)

CRISPR/dCas9-based labeling has allowed direct visualization of genomic regions in living cells. However, poor labeling efficiency and signal-to-background ratio have limited its application to visualize genome organization using super-resolution microscopy. We developed (Po)STAC (Polycistronic SunTAg modified CRISPR) by combining CRISPR/dCas9 with SunTag labeling and polycistronic vectors. (Po)STAC enhances both labeling efficiency and fluorescence signal detected from labeled loci enabling live cell imaging as well as super-resolution fixed-cell imaging of multiple genes with high spatiotemporal resolution. ; European Union's Horizon 2020 Research and Innovation Programme [CellViewer No 686637 to M.L., M.P.C.]; Ministerio de Economia y Competitividad [BFU2013–49867-EXP to M.L., M.P.C.]; Fundació Cellex Barcelona (to M.L); European Union Seventh Framework Programme under the European Research Council Grants [337191-MOTORS to M.L.]; 'Severo Ochoa' Programme for Centres of Excellence in R&D [SEV-2015- 0522 to M.L.]; Ministerio de Economia y Competitividad and FEDER Funds [BFU2014–54717-P, BFU2015–71984-ERC to M.P.C.]; AGAUR Grant [2014 SGR1137 to M.P.C.]; Spanish Ministry of Economy and Competitiveness (to M.P.C.); Centro de Excelencia Severo Ochoa [2013–2017 to M.P.C.]; CERCA Programme/Generalitat de Catalunya (to M.P.C); Ministerio de Ciencia e Innovacion FPI (to F.A.); People Program (Marie Curie Actions) FP7/2007–2013 under REA grant [608959 to M.V.N.]. Funding for open access charge: European Union's Horizon 2020 Research and Innovation Programme [CellViewer No 686637].

The Wnt/β-catenin signaling pathway is a key regulator of embryonic stem cell (ESC) self-renewal and differentiation. Constitutive activation of this pathway has been shown to increase mouse ESC (mESC) self-renewal and pluripotency gene expression. In this study, we generated a novel β-catenin knockout model in mESCs to delete putatively functional N-terminally truncated isoforms observed in previous knockout models. We showed that aberrant N-terminally truncated isoforms are not functional in mESCs. In the generated knockout line, we observed that canonical Wnt signaling is not active, as β-catenin ablation does not alter mESC transcriptional profile in serum/LIF culture conditions. In addition, we observed that Wnt signaling activation represses mESC spontaneous differentiation in a β-catenin-dependent manner. Finally, β-catenin (ΔC) isoforms can rescue β-catenin knockout self-renewal defects in mESCs cultured in serum-free medium and, albeit transcriptionally silent, cooperate with TCF1 and LEF1 to inhibit mESC spontaneous differentiation in a GSK3-dependent manner. ; The authors would like to thank the Genomics Unit at the CRG for assistance with the mRNA-seq (library prep and sequencing), and CRG and the Bristol University flow-cytometry facilities. This work was supported by the European Union's Horizon2020 Research and Innovation Programme (CellViewer No 686637 to M.P.C.), Ministerio de Ciencia e Innovación, grant BFU2017-86760-P (AEI/FEDER, UE), and an AGAUR grant from Secretaria d'Universitats i Recerca del Departament d'Empresa I Coneixement de la Generalitat de Catalunya (2017 SGR 689 to M.P.C.).

Parkinson's disease is a common neurodegenerative disorder, which is due to the loss of dopaminergic neurons in the substantia nigra pars compacta (SNpc) and for which no definitive cure is currently available. Cellular functions in mouse and human tissues can be restored after fusion of bone marrow (BM)-derived cells with a variety of somatic cells. Here, after transplantation of hematopoietic stem and progenitor cells (HSPCs) in the SNpc of two different mouse models of Parkinson's disease, we significantly ameliorated the dopaminergic neuron loss and function. We show fusion of transplanted HSPCs with neurons and with glial cells in the ventral midbrain of Parkinson's disease mice. Interestingly, the hybrids can undergo reprogramming in vivo and survived up to 4 weeks after transplantation, while acquiring features of mature astroglia. These newly generated astroglia produced Wnt1 and were essential for functional rescue of the dopaminergic neurons. Our data suggest that glial-derived hybrids produced upon fusion of transplanted HSPCs in the SNpc can rescue the Parkinson's disease phenotype via a niche-mediated effect, and can be exploited as an efficient cell-therapy approach. ; We are grateful for the support from a Fundació La Marató de TV3 grant (120530 to M.P.C. and M.V.), an ERC grant (242630-RERE to M.P.C.), Ministerio de Ciencia e Inovación grants (SAF2011-28580, 2014SGR1137 and BFU2015-71984-ERC to M.P.C.), an AGAUR grant (2014 SGR 780 to M.P.C.), Fondo de Investigación Sanitaria-Instituto de Salud Carlos III grants (PI13/01897 to M.V.; CP11/00229 to J.B.), an AGAUR-GRC grant (2014-SGR-1609 to M.V.), European Union Seventh Framework Programme (FP7/2007-2013) under the Marie Curie Intra-European Fellowship (274882 to W.A.X) and Juan de la Cierva fellowship (to W.A.X), Spanish Ministry of Economy and Competitiveness,'Centro de Excelencia Severo Ochoa 2013-2017', SEV-2012-0208.

Cells with high ploidy content are common in mammalian extraembryonic and adult tissues. Cell-to-cell fusion generates polyploid cells during mammalian development and tissue regeneration. However, whether increased ploidy can be occasionally tolerated in embryonic lineages still remains largely unknown. Here, we show that pluripotent, fusion-derived tetraploid cells, when injected in a recipient mouse blastocyst, can generate diploid cells upon ploidy reduction. The generated diploid cells form part of the adult tissues in mouse chimeras. Parental chromosomes in pluripotent tetraploid cells are segregated through tripolar mitosis both randomly and nonrandomly and without aneuploidy. Tetraploid-derived diploid cells show a differentiated phenotype. Overall, we discovered an unexpected process of controlled genome reduction in pluripotent tetraploid cells. This mechanism can ultimately generate diploid cells during mouse embryo development and should also be considered for cell fusion-mediated tissue regeneration approaches. ; This work was supported by the Ministerio de Ciencia, Innovación y Universidades, (BFU2017-86760-P to M.P.C.) (AEI/FEDER, UE) and the AGAUR grant from Secretaria d'Universitats i Recerca del Departament d'Empresa i Coneixement de la Generalitat de Catalunya, (2017 SGR 689 to M.P.C.). Work in M.P.C. lab was supported by funding from the European Union's Horizon 2020 research and innovation programme under grant agreement no. 686637 (CellViewer). We acknowledge support of the Spanish Ministry of Economy, Industry and Competitiveness (MEIC) to the EMBL partnership, the Centro de Excelencia Severo Ochoa and the CERCA Programme / Generalitat de Catalunya. KU Leuven C1 funds (C14/16/078 to F.L.), FWO national grants (G097618N to F.L.), Instituto de Salud Carlos III (CP10/00445 to F.L.), a "CRG-Severo Ochoa" predoctoral fellowship (to P.C.), and a grant from the Fundação para a Ciência e a Tecnologia (FCT) (SFRH/BD/76068/2011 to J.F.).

Transcription and genome architecture are interdependent, but it is still unclear how nucleosomes in the chromatin fiber interact with nascent RNA, and which is the relative nuclear distribution of these RNAs and elongating RNA polymerase II (RNAP II). Using super-resolution (SR) microscopy, we visualized the nascent transcriptome, in both nucleoplasm and nucleolus, with nanoscale resolution. We found that nascent RNAs organize in structures we termed RNA nanodomains, whose characteristics are independent of the number of transcripts produced over time. Dual-color SR imaging of nascent RNAs, together with elongating RNAP II and H2B, shows the physical relation between nucleosome clutches, RNAP II, and RNA nanodomains. The distance between nucleosome clutches and RNA nanodomains is larger than the distance measured between elongating RNAP II and RNA nanodomains. Elongating RNAP II stands between nascent RNAs and the small, transcriptionally active, nucleosome clutches. Moreover, RNA factories are small and largely formed by few RNAP II. Finally, we describe a novel approach to quantify the transcriptional activity at an individual gene locus. By measuring local nascent RNA accumulation upon transcriptional activation at single alleles, we confirm the measurements made at the global nuclear level. ; Innovative Team Program of Guangzhou Regenerative Medicine and Health Guangdong Laboratory [2018GZR110103001 to M.P.C.]; Guangzhou Key Projects of Brain Science and Brain-Like Intelligence Technology [20200730009 to M.P.C.]; National Natural Science Foundation of China [31971177 to M.P.C.]; Science and Technology Program of Guangzhou, China [202002030146 to M.P.C.]; European Union's Horizon 2020 Research and Innovation Programme [CellViewer No. 686637 to M.P.C. and M.L.]; Ministerio de Ciencia e Innovación [BFU2017-86760-P (AEI/FEDER, UE) to M.P.C.]; AGAUR grant from Secretaria d'Universitats i Recerca del Departament d'Empresa I Coneixement de la Generalitat de Catalunya [2017 SGR 689 to M.P.C.]; Centro de Excelencia Severo Ochoa [2013–2017 to M.P.C.]; CERCA Programme/Generalitat de Catalunya [to M.P.C.]; People Program (Marie Curie Actions) FP7/2007–2013 under REA grant [608959 to M.V.N.]; Spanish Ministry of Science and Innovation to the European Molecular Biology Laboratory (EMBL) partnership [to M.P.C.]; Juan de la Cierva-Incorporación 2017 [to M.V.N.]. Funding for open access charge: National Natural Science Foundation of China (NSFC) grant [31971177 to M.P.C].

Altres ajuts: We are grateful for the support from a Fundació La Marató de TV3 grant (120530 to M.P.C. and M.V.), an ERC grant (242630-RERE to M.P.C.),European Union Seventh Framework Programme (FP7/2007-2013) under the Marie Curie IntraEuropean Fellowship (274882 to W.A.X). SEV-2012-0208. ; Parkinson's disease is a common neurodegenerative disorder, which is due to the loss of dopaminergic neurons in the substantia nigra pars compacta (SNpc) and for which no definitive cure is currently available. Cellular functions in mouse and human tissues can be restored after fusion of bone marrow (BM)-derived cells with a variety of somatic cells. Here, after transplantation of hematopoietic stem and progenitor cells (HSPCs) in the SNpc of two different mouse models of Parkinson's disease, we significantly ameliorated the dopaminergic neuron loss and function. We show fusion of transplanted HSPCs with neurons and with glial cells in the ventral midbrain of Parkinson's disease mice. Interestingly, the hybrids can undergo reprogramming in vivo and survived up to 4 weeks after transplantation, while acquiring features of mature astroglia. These newly generated astroglia produced Wnt1 and were essential for functional rescue of the dopaminergic neurons. Our data suggest that glial-derived hybrids produced upon fusion of transplanted HSPCs in the SNpc can rescue the Parkinson's disease phenotype via a niche-mediated effect, and can be exploited as an efficient cell-therapy approach. A definitive therapy for Parkinson's disease is not available. Here, we transplanted hematopoietic stem and progenitor cells into the substantia nigra of brains of two different mouse models of Parkinson's disease. These transplanted cells fused with neurons and glial cells of the recipient mice. Four weeks after transplantation, the hybrids acquired features of mature astroglia, secreted Wnt1, and functionally ameliorated dopaminergic neuron loss. Current cell therapy approaches are being pursued in the striatum with the aim to increase dopamine ...

Nucleosomes form heterogeneous groups in vivo, named clutches. Clutches are smaller and less dense in mouse embryonic stem cells (ESCs) compared to neural progenitor cells (NPCs). Using coarse-grained modeling of the pluripotency Pou5f1 gene, we show that the genome-wide clutch differences between ESCs and NPCs can be reproduced at a single gene locus. Larger clutch formation in NPCs is associated with changes in the compaction and internucleosome contact probability of the Pou5f1 fiber. Using single-molecule tracking (SMT), we further show that the core histone protein H2B is dynamic, and its local mobility relates to the structural features of the chromatin fiber. H2B is less stable and explores larger areas in ESCs compared to NPCs. The amount of linker histone H1 critically affects local H2B dynamics. Our results have important implications for how nucleosome organization and H2B dynamics contribute to regulate gene activity and cell identity. ; This work was supported by a University of Pennsylvania Epigenetics Pilot Award (M.L.); an NSF Center for Engineering and Mechanobiology (CEMB) Pilot Award (M.L.); a Linda Pechenik Montague Investigator Award (M.L.); the European Union's Horizon 2020 Research and Innovation Programme (CellViewer grant 686637 to M.L., M.P.C., and E.M.); Ministerio de Ciencia, Innovación y Universidades ( BFU2017-86760-P [AEI/FEDER, UE]) and Secretaria d'Universitats i Recerca del Departament d'Empresa i Coneixement de la Generalitat de Catalunya (AGAUR grant 2017 SGR 689 to M.P.C.); National NSFC grant 319711771003712 (M.P.C.); National Institutes of Health , National Institute of General Medical Sciences awards R01-GM055264 and R35-GM122562 ; People Program (Marie Curie Actions) FP7/2007–2013 under REA grant 608959 (M.V.N.); and Juan de la Cierva-Incorporación 2017 (M.V.N.)

Bi-species, fusion-mediated, somatic cell reprogramming allows precise, organism-specific tracking of unknown lineage drivers. The fusion of Tcf7l1-/- murine embryonic stem cells with EBV-transformed human B cell lymphocytes, leads to the generation of bi-species heterokaryons. Human mRNA transcript profiling at multiple time points permits the tracking of the reprogramming of B cell nuclei to a multipotent state. Interrogation of a human B cell regulatory network with gene expression signatures identifies 8 candidate master regulator proteins. Of these 8 candidates, ectopic expression of BAZ2B, from the bromodomain family, efficiently reprograms hematopoietic committed progenitors into a multipotent state and significantly enhances their long-term clonogenicity, stemness, and engraftment in immunocompromised mice. Unbiased systems biology approaches let us identify the early driving events of human B cell reprogramming. ; This work was supported by Human Frontier Science Program Grant 2010 (to M.P.C. and A.C.); the European Union's Horizon 2020 Research and Innovation Programme (CellViewer no. 686637 to M.P.C); Ministerio de Ciencia e Innovación grant no. BFU2017-86760-P (AEI/FEDER, UE); AGAUR grant from Secretaria d'Universitats i Recerca del Departament d'Empresa i Coneixement de la Generalitat de Catalunya ( 2017 SGR 689 to M.P.C.); Guangzhou Key Projects of Brain Science and Brain-Like Intelligence Technology ( 20200730009 to M.P.C.); Juan de la Cierva Fellowship (to K.A.); BIST Master Fellowship (to X.T.); and the R35CA197745 outstanding NCI investigator award to A.C. We acknowledge the support of the Spanish Ministry of Science and Innovation to the EMBL partnership, the Centro de Excelencia Severo Ochoa , and the CERCA Programme (to M.P.C.), and the two instrumentation grants S10OD012351 and S10OD021764 to A.C. supporting the analytical work

Mouse embryonic stem cells (mESCs) are pluripotent and can differentiate into cells belonging to the three germ layers of the embryo. However, mESC pluripotency and genome stability can be compromised in prolonged in vitro culture conditions. Several factors control mESC pluripotency, including Wnt/β-catenin signaling pathway, which is essential for mESC differentiation and proliferation. Here we show that the activity of the Wnt/β-catenin signaling pathway safeguards normal DNA methylation of mESCs. The activity of the pathway is progressively silenced during passages in culture and this results into a loss of the DNA methylation at many imprinting control regions (ICRs), loss of recruitment of chromatin repressors, and activation of retrotransposons, resulting into impaired mESC differentiation. Accordingly, sustained Wnt/β-catenin signaling maintains normal ICR methylation and mESC homeostasis and is a key regulator of genome stability. ; We acknowledge the support from Spanish Ministry of Economy and Competitiveness and FEDER funds (SAF2011-28580, BFU2014-54717-P, BFU2017-86760-P and BFU2015-71984-ERC to M.P.C.), as well as 'Centro de Excelencia Severo Ochoa 2013–2017', Secretaria d'Universitats i Recerca del Departament d'Economia i Coneixement de la Generalitat de Catalunya grant (2014 SGR1137 to M.P.C.), AGAUR (SGR 2017-2019 to M.P.C) and the CERCA Programme/Generalitat de Catalunya (to M.P.C.), the European Union's Horizon 2020 research and innovation programme under grant agreement CellViewer No 686637 (to M.P.C.), People Programme Marie Curie Actions of the European Union's Seventh Framework Programme (FP7/2007–2013/, n° 290123 to I.T.), La Caixa international PhD fellowship (to F.S.) and Ministerio de Ciencia e Innovació FPI (to F.A.) and People Program (Marie Curie Actions) FP7/2007–2013 under REA grant (608959 to M.V.N.).