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Many cnidarians have been reported in association with hermit crabs from shallow waters to the deep sea. Some of these actiniarians and zoantharians produce a carcinoecium, a chitin-like pseudo-she...

Sympatric speciation has been demonstrated in few empirical case studies, despite intense searches, because of difficulties in testing the criteria for this mode of speciation. Here, we report a possible case of sympatric speciation in ricefishes of the genus Oryzias on Sulawesi, an island of Wallacea. Three species of Oryzias are known to be endemic to Lake Poso, an ancient tectonic lake in central Sulawesi. Phylogenetic analyses using RAD-seq-derived single nucleotide polymorphisms (SNPs) revealed that these species are monophyletic. We also found that the three species are morphologically distinguishable and clearly separated by population-structure analyses based on the SNPs, suggesting that they are reproductively isolated from each other. A mitochondrial DNA chronogram suggested that their speciation events occurred after formation of the tectonic lake, and existence of a historical allopatric phase was not supported by coalescent-based demographic inference. Demographic inference also suggested introgressive hybridization from an outgroup population. However, differential admixture among the sympatric species was not supported by any statistical tests. These results all concur with criteria necessary to demonstrate sympatric speciation. Ricefishes in this Wallacean lake provide a promising new model system for the study of sympatric speciation.

Antivirals are now more important than ever. To efficiently inhibit virus replication, antiviral multivalent strategies need sufficient affinity to overcome the excellent matching between the virus and its receptor. This report highlights a phage capsid scaffold strategy that can be used to precisely position sialic acid moieties to inhibit influenza A virus replication.

In this work we report a rational design strategy for the identification of new peptide prototypes for the non-disruptive supramolecular permeation of membranes and the transport of different macromolecular giant cargos. The approach targets a maximal enhancement of helicity in the presence of membranes with sequences bearing the minimal number of cationic and hydrophobic moieties. The here reported folding enhancement in membranes allowed the selective non-lytic translocation of different macromolecular cargos including giant proteins. The transport of different high molecular weight polymers and functional proteins was demonstrated in vesicles and in cells with excellent efficiency and optimal viability. As a proof of concept, functional monoclonal antibodies were transported for the first time into different cell lines and cornea tissues by exploiting the helical control of a short peptide sequence. This work introduces a rational design strategy that can be employed to minimize the number of charges and hydrophobic residues of short peptide carriers to achieve non-destructive transient membrane permeation and transport of different macromolecules., The helical enhancement of a short oligoalanine peptide scaffold in anionic membranes triggered the supramolecular assembly of a nanopore, which allowed the transport and release of proteins in the cytosol of cells and tissues.

This is the peer reviewed version of the following article: S. Díaz, I. Insua, G. Bhak, J. Montenegro, Chem. Eur. J. 2020, 26, 14765, which has been published in final form at https://doi.org/10.1002/chem.202003265. This article may be used for non-commercial purposes in accordance with Wiley Terms and Conditions for Use of Self-Archived Versions. The inherent ability of peptides to self-assemble with directional and rationally predictable interactions has fostered a plethora of synthetic two-dimensional (2D) supramolecular biomaterials. However, the design of peptides with hierarchical assembly in different dimensions across mesoscopic lengths remains a challenging task. We here describe the structural exploration of a d/l-alternating cyclic octapeptide capable of assembling one-dimensional (1D) nanotubes in water, which subsequently pack laterally to form giant 2D nanosheets up to 500 μm long with a constant 3.2 nm thickness. Specific amino acid mutations allowed the mapping of structure–assembly relationships that determine 2D self-assembly. Nine peptide modifications were studied, revealing key features in the peptide sequence that nanosheets tolerated, while a total of three peptide variants included modifications that compromised their 2D arrangement. These lessons will serve as guide and inspiration for new 2D supramolecular peptide designs This work was partially supported by the Spanish Agencia Estatal de Investigación (AEI) [SAF2017-89890-R], Xunta de Galicia (AD031 2016, ED431C 2017/25 and ED431G2019/03) and the European Commission (EC) (European Regional Development Fund-ERDF) Instituto de Salud Carlos III (COV20/00297). I.I. thanks the European Commission (EC) and AEI for MSCA-IF (2018-843332) and Juan de la Cierva (FJCI-2017-31795) fellowships, respectively. J.M. thanks the Ramón y Cajal (RYC-2013-13784), ERC- STG (DYNAP, 2016-677786), ERC-POC (TraffikGene, 2019-838002) and Human Frontier Science Programme Young Investigator Grant (RGY0066/2017) for funding SI

The recent advances in the supramolecular polymerization of synthetic building blocks in aqueous conditions has given rise to new artificial and biocompatible functional materials. However, despite the importance of spatially resolved self‐assembly for natural and artificial molecular machines, the spatial control of supramolecular polymerization with synthetic monomers has not been experimentally established yet. Here we describe a microfluidic‐regulated tandem process of supramolecular polymerization and droplet encapsulation to control the position of self‐assembled microfibrillar bundles of cyclic peptide nanotubes in water droplets. This method allowed the precise preferential localization of the fibres either at the interface or into the core of the droplets. UV absorbance, circular dichroism and fluorescence microscopy indicated that the microfluidic control of the stimuli (changes in pH or ionic strength) can be employed to adjust the packing degree and the spatial position of microfibrillar bundles of cyclic peptide nanotubes. Additionally, this spatially organized supramolecular polymerization of peptide nanotubes was applied in the assembly of highly ordered two‐dimensional droplet networks This work was partially supported by the Spanish Agencia Estatal de Investigación (AEI) [SAF2017-89890-R, CTQ2016-78423-R], the Xunta de Galicia (ED431G/09, ED431C 2017/25 and 2016-AD031) and the ERDF. A. M.-A. received a MCIF from the EC (GLYCONANOPEP-750248). J. M. holds a Ramón y Cajal (RYC-2013-13784), an ERC-Stg (DYNAP-677786) and a Young Investigator Grant from the HFSP (RGY0066/2017). J. R. G. thanks to the mobility program (PRX17/00147) SI

Despite recent developments in two-dimensional self-assembly, most supramolecular 2D materials are assembled by tedious methodologies, with complex surface chemistry and small sizes. We here report d/l-alternating cyclic peptides that undergo one-dimensional self-assembly into amphiphilic nanotubes, which subsequently arrange as tubular bilayers to form giant nanosheets in the mesoscale. Reversible transitions between the assembled, dispersed, and aggregated states of these nanosheets can be triggered by external stimuli. The characteristic flexibility, defined chemical topology, and length scale of these nanosheets set a clear distinction between this new supramolecular architecture and previously reported 2D nanostructures. The sequential 1D-to-2D self-assembly of peptides described here provides a conceptually new approach to achieve two-dimensional materials with hierarchical organization. These giant nanosheets represent one of the largest 2D supramolecular materials ever made, with potential application as long-range molecular transporters, responsive surfaces, and (bio)sensors This work was partially supported by the Spanish Agencia Estatal de Investigación (AEI) [SAF2017-89890-R], the Xunta de Galicia (ED431C 2017/25, 2016-AD031 and Centro Singular de Investigación de Galicia accreditation 2016-2019, ED431G/09), the ISCIII (RD16/0008/003), and the European Union (European Regional Development Fund, ERDF). I.I. thanks the European Commission for a Marie Curie fellowship (MSCA-IF-2018-843332) and the Spanish AEI for a Juan de la Cierva—Formación Fellowship (FJCI-2017-31795). J.M. received a Ramón y Cajal (RYC-2013-13784), an ERC Starting Investigator Grant (DYNAP-677786), and a Young Investigator Grant from the HFSP (RGY0066/2017). We thank Dr. Eugenio Solla, Prof. Juan Granja, Dr. Julian Bergueiro, Dr. Andreas Vargas Jentzsch, and Dr. Mark J. van Raaij for helpful discussions. We also thank Dr. Bergueiro for Figure 1 and Dr. Vargas Jentzsch for assistance with DFT calculations SI

This is the Accepted Manuscript of the following article: Méndez-Ardoy, A., Granja, J., & Montenegro, J. (2018). pH-Triggered self-assembly and hydrogelation of cyclic peptide nanotubes confined in water micro-droplets. Nanoscale Horizons. http://dx.doi.org/10.1039/c8nh00009c The controlled one-dimensional supramolecular polymerization of synthetic building blocks in confined spaces constitutes a key challenge to simplify the understanding of the fundamental physical principles behind the behavior of more complex encapsulated polymer networks. Cyclic peptide nanotubes constitute an optimal scaffold for the fabrication of hierarchical one-dimensional self-assembled architectures. Herein we report the pH-controlled nanotube formation and fibrillation of supramolecular cyclic peptides in confined aqueous droplets. The externally triggered self-assembly of these peptides gave rise to viscoelastic hydrogels in which the one-dimensional molecular arrangement was perfectly preserved from the nano- to the micro-scale. The cyclic peptide building blocks were confined inside water microdroplets and the base-triggered supramolecular polymerization was externally triggered and followed by confocal microscopy showing that the confined fibrillation spanned and affected the shape of the droplet micro container This work was partially supported by the Spanish Agencia Estatal de Investigación (AEI) [SAF2017-89890-R, CTQ2014-59646-R and CTQ2016-78423-R], the Xunta de Galicia (ED431G/09, ED431C 2017/25 and 2016-AD031) and the ERDF. A. M.-A. received a MCIF from the EC (GLYCONANOPEP-750248). J. M. holds a Ramón y Cajal (RYC-2013-13784), an ERC-Stg (DYNAP-677786) and a Young Investigator Grant from the HFSP (RGY0066/2017) SI

Many straits in the Japanese archipelago have been proposed as biogeographical boundaries, but there is disagreement regarding their importance as historic barriers against dispersal of terrestrial and freshwater taxa. Mitochondrial DNA haplotype and phylogenetic analyses of Lefua nikkonis, a primary freshwater fish inhabiting northern Japan and descendent from Siberia, revealed that the species is genetically structured within its geographic range, but that two major haplotypes are widely distributed across the Ishikari Lowland of Hokkaido Island as well as across the Tsugaru Strait between Hokkaido and Honshu Islands, two well-known biogeographical boundaries of northern Japan. The two major haplotypes were separated from each other by only one mutational step, and many other haplotypes, including those endemic to the region south of these barriers, have diverged from the major haplotypes, suggesting rapid range expansion and local differentiation. Divergence-time estimates, based on vicariance of the Honshu endemic congener L. echigonia via uplift of the Central Highlands, demonstrated that the southward dispersal of L. nikkonis from Hokkaido Island to Honshu Island occurred less than 0.08–0.19 Mya, suggesting that a land bridge emerged at the Tsugaru Strait during the Riss glaciation. Given that other freshwater taxa crossed the strait earlier (during the Middle Pleistocene), it is likely that land bridges in the strait have repeatedly emerged. The fact that L. nikkonis invaded only the northern part of Honshu, and that many other freshwater species also have the limit of their distribution ranges in this area as well, indicates that a faunal transition zone might persist even without the Tsugaru Strait. Thus, straits and lowlands in northern Japan are likely to have been less important as dispersal barriers to freshwater taxa than is currently thought.

The Red Sea was one of the first areas of the Indo-Pacific to be investigated by marine taxonomists, and the literature on suborder Brachycnemina (Anthozoa: Hexacorallia: Zoantharia) for this region dates from the early nineteenth century. However, in the last 100 years there has been only one focused study on this group in the Red Sea. In the present study, specimens collected from the Saudi Arabian coast of the northern half of the Red Sea were phylogenetically analyzed by sequencing nuclear internal transcribed spacer regions of ribosomal DNA (ITS-rDNA), mitochondrial cytochrome oxidase subunit I (COI), and 16S ribosomal DNA (16S–rDNA). The new results were compared with historical data in the literature and recent results from the Persian Gulf and the southeastern coast of Africa. Results show the presence of six to seven potential Brachycnemina species in the Red Sea; five to six Palythoa species (Palythoa mutuki, P. tuberculosa, P. cf. heliodiscus, P. aff. heliodiscus, and one to two species within the P. sp. “sakurajimensis” group) together with Zoanthus sansibaricus. While P. mutuki, P. tuberculosa, and Z. sansibaricus are known to be widely distributed in the Indo-Pacific, P. cf. heliodiscus and P. sp. “sakurajimensis” have not been reported from the Persian Gulf or the southeastern coast of Africa, and the current results represent large range extensions for these two species. Only one of the observed species, P. aff. heliodiscus, is potentially endemic to the Red Sea, further demonstrating the generally wide distributions of most zooxanthellate Brachycnemina species.

Here we present the synthesis and post-polymerisation modification of poly(acryloyl hydrazide), a versatile scaffold for the preparation of functional polymers: poly(acryloyl hydrazide) was prepared from commercially available starting materials in a three step synthesis on a large scale, in good yields and high purity. Our synthetic approach included the synthesis of a Boc-protected acryloyl hydrazide, the preparation of polymers via RAFT polymerisation and the deprotection of the corresponding Boc-protected poly(acryloyl hydrazide). Post-polymerisation modification of poly(acryloyl hydrazide) was then demonstrated using a range of conditions for both hydrophilic and hydrophobic aldehydes. These experiments demonstrate the potential of poly(acryloyl hydrazide) as a scaffold in the synthesis of functional polymers, in particular those applications where in situ screening of the activity of the functionalised polymers may be required (e.g. biological applications) This work was supported by the Royal Society, U.K (IE130688) and the Wellcome Trust (177ISSFPP). F. F.-T. thanks the Birmingham Science City and the European Regional Development Fund, the Royal Society (RG140273), and the University of Birmingham (John Evans Fellowship). J. M. thanks funding from MINECO (CTQ2014-59646-R, RYC-2013-1378) the Xunta de Galicia (ED431G/09 and 2016-AD031) and the ERC (Stg-DYNAP-677786) SI

Tracking the pH with spatiotemporal resolution is a critical challenge for synthetic chemistry, chemical biology and beyond. Over the last decade different small probes and supramolecular systems have emerged for in celluloor in vivo pH tracking. However, pH reporting still presents critical limitations such as background reduction, sensor improved stability, cell targeting, endosomal escape, near and far infrared ratiometric pH tracking, adaptation to the new imaging techniques (i.e. super‐resolution), etc. These challenges will demand the combined efforts of synthetic and supramolecular chemistry working together to develop a next generation of smart materials that will resolve the current limitations. In this review we describe the recent advances in the synthesis of small fluorescent probes together with new supramolecular functional systems employed for pH tracking with emphasis in ratiometric probes. The combination of organic synthesis and stimuli‐responsive supramolecular functional materials will be essential to solve future challenges of pH tracking such as the improved signal to noise ratio, on target activation and microenvironment reporting This work was partially supported by the Spanish Agencia Estatal de Investigación (AEI) [SAF2017-89890-R], the Xunta de Galicia (ED431C 2017/25, 2016-AD031 and Centro Singular de Investigación de Galicia accreditation 2016–2019, ED431G/09), the ISCIII (RD16/0008/003), and the European Union (European Regional Development Fund –ERDF). A.M. received a Marie Curie fellowship (GLYCONANOPEP-750248). J.M. received a Ramón y Cajal (RYC-2013-13784), an ERC Starting Investigator Grant (DYNAP-677786) and a Young Investigator Grant from the Human Frontier Science Research Program (RGY0066/2017) SI

Although there are many examples of color evolution potentially driven by sensory drive, only few studies have examined whether distinct species inhabiting the same environments evolve similar body colors via shared sensory mechanisms. In this study, we tested whether two sympatric freshwater fish taxa, halfbeaks of the genus Nomorhamphus and ricefishes of the genus Oryzias in Sulawesi Island, converge in both body color and visual sensitivity. After reconstructing the phylogeny separately for Nomorhamphus and Oryzias using transcriptome‐wide sequences, we demonstrated positive correlations of body redness between these two taxa across environments, even after phylogenetic corrections, which support convergent evolution. However, substantial differences were observed in the expression profiles of opsin genes in the eyes between Nomorhamphus and Oryzias. Particularly, the expression levels of the long wavelength‐sensitive genes were negatively correlated between the taxa, indicating that they have different visual sensitivities despite living in similar light environments. Thus, the convergence of body colorations between these two freshwater fish taxa was not accompanied by convergence in opsin sensitivities. This system presents a case in which body color convergence can occur between sympatric species via different mechanisms.

Diversity surveys in isolated sites, such as oceanic islands, provide biogeographic data that can improve our analyses and knowledge of evolutionary processes in the oceans. Zoantharians (Cnidaria: Anthozoa) are common and widespread components of shallow-water reefs, but distributional analyses are scarce for this group. In this study, we collected Zoantharia specimens from around Saint Helena Island (STH) in the mid-Atlantic and identified species using external morphology and molecular data. Moreover, we compiled and analysed the most comprehensive distributional data for zoantharian species in the subtropical and tropical Atlantic Ocean to date. Our results show eight zoantharian species in STH, which includes seven new records for STH waters. Furthermore, all families and genera of the suborder Brachycnemina recorded are widespread in the Atlantic Ocean, including at least four amphi-Atlantic species. The Caribbean is the richness centre in the Atlantic Ocean for zoantharian species, a pattern similar to that observed for many other subtropical/tropical marine taxa. However, Zoantharia may have a lower endemism rate in some areas than other common reef animals, for example zooxanthellate scleractinian corals and reef fishes. Moreover, zoantharian species have a more extensive distribution than close-related taxa such as zooxanthellate scleractinian corals and hydrocorals in the Atlantic Ocean. Japanese Government (MEXT)Ministry of Education, Culture, Sports, Science and Technology, Japan (MEXT) JSPSMinistry of Education, Culture, Sports, Science and Technology, Japan (MEXT)Japan Society for the Promotion of Science

31 p.-6 fig.-14 fig. supl., The cell membrane regulates the exchange of molecules and information with the external environment. However, this control barrier hinders the delivery of exogenous bioactive molecules that can be applied to correct cellular malfunctions. Therefore, the traffic of macromolecules across the cell membrane represents a great challenge for the development of the next generation of therapies and diagnostic methods. Cell‐penetrating peptides are short peptide sequences capable of delivering a broad range of biomacromolecules across the cellular membrane. However, penetrating peptides still suffer from limitations, mainly related to their lack of specificity and potential toxicity. Glycosylation has emerged as a potential promising strategy for the biological improvement of synthetic materials. In this work we have developed a new convergent strategy for the synthesis of penetrating peptides functionalized with glycan residues by an oxime bond connection. The uptake efficiency and intracellular distribution of these glycopeptides have been systematically characterized by means of flow cytometry and confocal microscopy and in zebrafish animal models. The incorporation of these glycan residues into the peptide structure influenced the internalization efficiency and cellular toxicity of the resulting glycopeptide hybrids in the different cell lines tested. The results reported herein highlight the potential of the glycosylation of penetrating peptides to modulate their activity., We acknowledge funding from the Spanish Agencia Estatal de Investigación (AEI; CTQ2014‐59646‐R, SAF2017‐89890‐R, CTQ2016‐76263‐P, CTQ2015‐64597‐C2‐2P), the Xunta de Galicia (ED431G/09, ED431C 2017/25, and 2016‐AD031), and the ERDF. I.G. received a predoctoral fellowship from the Xunta de Galicia (ED481A‐2018/116). A.R. received a predoctoral fellowship from the Fundación Segundo Gil Dávila. J.G.‐V. and L.S. acknowledge financial support received from the Xunta de Galicia (Galicia, Spain) under the Grupos de Referencia Competitiva Programme: Project GRC2014/010. J.M. received a Ramón y Cajal (RYC‐2013‐13784), an ERC Starting Investigator Grant (DYNAP‐677786) and a Young Investigator Grant from the HFSP (RGY0066/2017).

This is the Accepted Manuscript of the following article: Juanes, M., Creese, O., Fernández-Trillo, P., & Montenegro, J. (2019). Messenger RNA delivery by hydrazone-activated polymers. Medchemcomm, 10(7), 1138-1144. doi: 10.1039/c9md00231f The intracellular delivery of DNA and RNA therapeutics requires the assistance of vectors and/or nucleotide modifications to protect the nucleic acids against host nucleases and promote cellular internalization and release. Recently, messenger RNA (mRNA) has attracted much attention due to its transient activity and lack of genome permanent recombination and persistent expression. Therefore, there is a strong interest in the development of conceptually new non-viral vectors with low toxicity that could improve mRNA transfection efficiency. We have recently introduced the potential of polyhydrazones and the importance of the degree of polymerization for the delivery of siRNA and plasmid DNA. Here, we demonstrate that this technology can be easily adapted to the more interesting complexation and delivery inside living cells of mRNA. The polyplexes resulting from the combination of the amphiphilic polyhydrazone were characterized and the transfection efficiency and cell viability were studied for a discrete collection of functionalized polyhydrazones. The results obtained demonstrated the versatility of these polymeric vectors as excellent candidates for the delivery of mRNA and validate the easy adaptability of the technology to more sensitive and therapeutically relevant nucleic acids This work has received financial support from Spanish Agencia Estatal de Investigación (AEI) [CTQ2014-59646-R, SAF2017-89890-R], the Xunta de Galicia (ED431C 2017/25, 2016-AD031 and Centro singular de investigación de Galicia accreditation 2016–2019, ED431G/09) and the European Union (European Regional Development Fund - ERDF). M.J. received a F.P.I. fellowship from MINECO (CTQ2014-59646- R). J.M. received a Ramón y Cajal (RYC-2013-13784), an ERC Starting Investigator Grant (DYNAP- 677786) and a Young Investigator Grant from the Human Frontier Science Research Program (RGY0066/2017). F.F.T. thanks the Birmingham Science City and the European Regional Development Fund, and the University of Birmingham (John Evans Fellowship). O.C. thanks the Midlands Integrative Biosciences Training Partnership (MIBTP) for the PhD scholarship SI

The formation of oligomeric soluble aggregates is related to the toxicity of amyloid peptides and proteins. In this manuscript, we report the use of a ruthenium polypyridyl complex ([Ru(bpy)2(dpqp)]2+) to track the formation of amyloid oligomers at different times using photoluminescence anisotropy. This technique is sensitive to the rotational correlation time of the molecule under study, which is consequently related to the size of the molecule. [Ru(bpy)2(dpqp)]2+ presents anisotropy values of zero when free in solution (due to its rapid rotation and long lifetime) but larger values as the size and concentration of amyloid-β (Aβ) oligomers increase. Our assays show that Aβ forms oligomers immediately after the assay is started, reaching a steady state at ∼48 h. SDS–PAGE, DLS, and TEM were used to confirm and characterize the formation of oligomers. Our experiments show that the rate of formation for Aβ oligomers is temperature dependent, with faster rates as the temperature of the assay is increased. The probe was also effective in monitoring the formation of α-synuclein oligomers at different times AAM thanks the Welch Foundation (Grant C-1743) and JM thanks AEI (SAF2017-89890-R), ERC (DYNAP-677786) and HFSP (RGY0066/2017) for financial support SI

Linear peptide amphiphiles are excellent biocompatible scaffolds for the hierarchical self-assembly of one dimensional nanostructures in aqueous media. However, their structural exploration and screening of self-assembling properties is often limited by time-consuming synthesis and purification steps. We here describe the application of the oxime bond as a powerful synthetic tool towards the conjugation of peptide heads bearing a hydroxylamine group with hydrophobic aldehyde tails. This methodology allowed the quick prepraration of a small library of oxime-connected peptide amphiphiles, whose supramolecular screening revealed nano-to-micro fibrillation with dependency on their chemical structure. These results demonstrate the simplicity and the synthetic potential of the oxime conjugation for the preparation of peptide amphiphiles with improved self-assembling capabilities This work was partially supported by the Spanish Agencia Estatal de Investigación (AEI) [CTQ2014-59646-R, SAF2017-89890-R], the Xunta de Galicia (ED431G/09, ED431C 2017/25 and 2016-AD031) and the ERDF. I.I. thanks the Spanish AEI for a Juan de la Cierva -Formación fellowship (FJCI-2017-31795). J.M. received a Ramón y Cajal (RYC-2013-13784), an ERC Starting Investigator Grant (DYNAP-677786) and a Young Investigator Grant from the Human Frontier Science Research Program (RGY0066/2017) SI

The internalisation and delivery of active substances into cells is a field of growing interest for chemical biology and therapeutics. As we move from small-molecule-based drugs towards bigger cargos, such as antibodies, enzymes, nucleases or nucleic acids, the development of efficient delivery systems becomes critical for their practical application. Different strategies and synthetic carriers have been developed; these include cationic lipids, gold nanoparticles, polymers, cell-penetrating peptides (CPPs), protein surface modification etc. However, all of these methodologies still present limitations relating to the precise targeting of the different intracellular compartments and, in particular, difficulties in access to the cellular cytosol. Additionally, the precise quantification of the cellular uptake of a compound is not enough to demonstrate delivery and/or functional activity. Therefore, methods to determine cellular distributions of cargos and carriers are of critical importance for identifying the barriers that are blocking the activity. Herein we survey the different techniques that can currently be used to track and to monitor the subcellular localisation of the synthetic compounds that we deliver inside cells.

The straightforward preparation of glycosyl neoconjugates by oxime (or hydrazone) bond formation represents a key bioorthogonal tool in chemical biology. However, when this strategy is employed by reacting the reducing end of the glycan moiety, the configuration and the stereochemical information is lost due to partial (or complete) opening of the glycan cyclic hemiacetal and the formation of the corresponding opened tautomers. We have completed the synthesis of a library of glycosyl aldehydes to be used as scaffold for the synthesis of neoglycoconjugates via oxime bond formation. These glycosyl aldehydes constitute a simple and accessible alternative to avoid loss of chiral information when conjugating, by oxime (or hydrazone) bonds, the aldehyde functionality present at the reducing end of natural carbohydrates This work was partially supported by the Spanish Agencia Estatal de Investigación (AEI) [CTQ2014–59646-R], the Xunta de Galicia (ED431G/09, ED431C 2017/25 and 2016-AD031) and the ERDF. J.M. received a Ramón y Cajal (RYC-2013–13784), an ERC-Stg (DYNAP- 677786) and a Young Investigator Grant from the Human Frontier Science Research Program (RGY0066/2017) SI

This manuscript has been accepted after peer review and appears as anAccepted Article online prior to editing, proofing, and formal publicationof the final Version of Record (VoR). This work is currently citable byusing the Digital Object Identifier (DOI) given below. The VoR will bepublished online in Early View as soon as possible and may be differentto this Accepted Article as a result of editing. Readers should obtainthe VoR from the journal website shown below when it is publishedto ensure accuracy of information. The authors are responsible for thecontent of this Accepted Article The intracellular transport of exogenous proteins has emerged as one of the most promising methodologies for biotechnology and chemical biology. Current protein delivery is mainly approached by liposome encapsulation, translational fusion and ionic/hydrophobic non‐covalent aggregation with transporting molecular vehicles. We here introduce the concept of supramolecular recognition and selective transport of proteins by peptide hybrid materials. We have designed a helical amphiphilic cationic peptide that bears two orthogonal alkoxyamines for the precise anchoring of protein ligands. After the attachment of these protein ligands, the peptide showed a high binding affinity for its protein target (i.e. mannose/Concanavalin A, Biotin/Streptavidin). The resulting peptide/protein hybrids were taken up by human cells such as HeLa and HepG2. The concept described in this manuscript could potentially be adapted, through the appropriate choice of ligands, to the transport of other proteins with suitable supramolecular binding motifs This work was partially supported by the Spanish Agencia Estatal de Investigación (AEI) [CTQ2014-59646-R, CTQ2013-43264-R, SAF2017-89890-R], the Xunta de Galicia (ED431G/09, ED431C 2017/25 and 2016-AD031) and the ERDF. We thank Pili Canoa (CACTI, UVIGO) for assistance with the SPR measurements. M.J. received a F.P.I. fellowship from MINECO (CTQ2014-59646-R). J.M. received a Ramón y Cajal (RYC-2013-13784), an ERC Starting Investigator Grant (DYNAP- 677786) and a Young Investigator Grant from the Human Frontier Science Research Program (RGY0066/2017) SI

Binding constants, Log K ≈ 6.6 M-1, and NMR characterization of the complexes formed by sulfobetaines and cucurbit[7]uril (CB7) support the electrostatic interaction as major driving force. This very strong binding motif is cross-linked by additional CB7 molecules resulting in the formation of supramolecular nanoparticles (SNPs) with an average diameter of 172 nm and negative surface potential. The time course evolution of the particle size and the surface potential suggests the very fast formation of an amorphous aggregate that absorbs additional amount of sulfobetaine. These aggregates afford very stable (more than two weeks) nanoparticles in aqueous dispersion. The reversibility of the sulfobetaine/CB7 host:guest complexes allows SNPs disaggregation by adding a competitive guest as shown by treatment with tetraethylammonium chloride. The addition of this competitive cation triggers a SNPs to micelle transition. The potential application of these nanoparticles as drug delivery vehicles was investigated by using carboxyfluorescein. These experiments revealed that upon externally induced disruption of the SNPs (by tetraethylammonium chloride) the fluorescent dye was trapped into micellar aggregates that can be further disrupted by cyclodextrin additionhttp://www.usc.es/ Financial support from Ministerio de Economia y Competitividad of Spain (projects CTQ2014-55208-P, CTQ2017-84354-P, CTQ2014-59646-R and MAT2015-67458-P), Xunta de Galicia (GR 2007/085; IN607C 2016/03 and Centro singular de investigación de Galicia accreditation 2016-2019, ED431G/09) and the European Union (European Regional Development Fund – ERDF), is gratefully acknowledged. J.M. received a Ramón y Cajal (RYC-2013- 13784) and a starting grant from the ERC (DYNAP677786) SI

The genus Bergia was established by Duchassaing de Fonbressin and Michelotti in 1860, for two species, Bergia catelunaris and B. vialactea. Subsequently, in 1903 Duerden recognized these two species as conspecific, and used B. catelunaris in favour of B. vialactea, and transferred B. catelunaris to the genus Parazoanthus. However, over the last decade, it has been found that the genus Parazoanthus is actually polyphyletic and therefore it has gradually been divided and redefined. Based on phylogenetic analyses, Parazoanthus sensu stricto was recently limited to species which form associations with sponges, but it still comprised of three distinctive and monophyletic subclades. Of these clades, one Parazoanthus clade contains the type species for Parazoanthus, P. axinellae, while another clade was recently described as the genus Umimayanthus based on mitochondrial 16S-rDNA sequences. However, the other remaining Parazoanthus clade contains P. catenularis, the original type species of the genus Bergia. Base...

The objective of this Feature Article is to reflect on the importance of established and emerging principles of supramolecular organic chemistry to address one of the most persistent problems in life sciences. The main topic is dynamic covalent chemistry on cell surfaces, particularly disulfide exchange for thiol-mediated uptake. Examples of boronate and hydrazone exchange are added for contrast, comparison and completion. Of equal importance are the discussions of proximity effects in polyions and counterion hopping, and more recent highlights on ring tension and ion pair-π interactions. These lessons from supramolecular organic chemistry apply to cell-penetrating peptides, particularly the origin of "arginine magic" and the "pyrenebutyrate trick," and the currently emerging complementary "disulfide magic" with cell-penetrating poly(disulfide)s. They further extend to the voltage gating of neuronal potassium channels, gene transfection, and the delivery of siRNA. The collected examples illustrate that the input from conceptually innovative chemistry is essential to address the true challenges in biology beyond incremental progress and random screening.

The strong wind that was blowing in Brunnen on the 4th of May 2017 was prophetic of the storm of ideas and creativity that would later fall over the participants of the 52nd edition of the Bürgenstock conference. The Bürgenstock conference is a truly unique event. A meeting held in a beautiful and relaxing landscape where the Alps merge with the crystal clear water of the Luzern Lake, researchers from the best Universities, Institutes and companies came to Brunnen for the 52nd Bu¨rgenstock conference from all over the globe R. J. P. and J. M. are very grateful to the organising committee of the 2017 Bürgenstock conference for the award of JSP fellowships. R. J. P is grateful to the Royal Society for a University Research Fellowship. J. M. is grateful to MINECO for a Ramón y Cajal fellowship SI

The Oryzias woworae species group, composed of O. asinua, O. wolasi, and O. woworae, is widely distributed in southeastern Sulawesi, an island in the Indo-Australian Archipelago. Deep-elongated body shape divergence is evident among these three species to the extent that it is used as a species-diagnostic character. These fishes inhabit a variety of habitats, ranging from upper streams to ponds, suggesting that the body shape divergence among the three species may reflect adaptation to local environments. First, our geometric morphometrics among eight local populations of this species group revealed that the three species cannot be separated by body shape and that riverine populations had more elongated bodies and longer caudal parts than lacustrine populations. Second, their phylogenetic relationships did not support the presence of three species; phylogenies using mitochondrial DNA and genomic data obtained from RNA-Seq revealed that the eight populations could not be sorted into three different clades representing three described species. Third, phylogenetic corrections of body shape variations and ancestral state reconstruction of body shapes demonstrated that body shape divergence between riverine and lacustrine populations persisted even if the phylogenies were considered and that body shape evolved rapidly irrespective of phylogeny. Sexual dimorphism in body shape was also evident, but the degree of dimorphism did not significantly differ between riverine and lacustrine populations after phylogenetic corrections, suggesting that sexual selection may not substantially contribute to geographical variations in body shape. Overall, these results indicate that the deep-elongated body shape divergence of the O. woworae species group evolved locally in response to habitat environments, such as water currents, and that a thorough taxonomic reexamination of the O. woworae species group may be necessary.

Very few studies have been conducted on the long-term effects of typhoon damage on mesophotic coral reefs. This study investigates the long-term community dynamics of damage from Typhoon 17 (Jelawat) in 2012 on the coral community of the upper mesophotic Ryugu Reef in Okinawa, Japan. A shift from foliose to bushy coral morphologies between December 2012 and August 2015 was documented, especially on the area of the reef that was previously recorded to be poor in scleractinian genera diversity and dominated by foliose corals. Comparatively, an area with higher diversity of scleractinian coral genera was observed to be less affected by typhoon damage with more stable community structure due to less change in dominant coral morphologies. Despite some changes in the composition of dominant genera, the generally high coverage of the mesophotic coral community is facilitating the recovery of Ryugu Reef after typhoon damage.

We here describe the application of oxime bond formation between a peptide scaffold and different aldehydes to modify the transporting capabilities of penetrating peptides. We show that bonds such as oximes offer a great synthetic advantage for the modification of the properties of model penetrating peptides. We believe that this approach will allow the development of improved intracellular delivery vehicles This work was supported by the Spanish Ministry of Economy and Competitivity (MINECO) [CTQ2014-59646-R], the Xunta de Galicia (ED431G/09) and the ERDF. J.P. received an F.P.I. fellowship from MINECO. J.M. received a Ramon y Cajal (RYC-2013-13784) and an ERC Starting Investigator Grant (DYNAP-677786) SI

Gene transfection continues to be a major challenge in chemistry, biology and materials sciences. New methodologies and recent breakthroughs have renewed the interest in the discovery and development of new tools for efficient gene transfection. Hydrazone formation between a cationic head and hydrophobic tails has emerged as one of the most promising techniques for nucleotide delivery. In this contribution, we have exploited hydrazone formation to modulate the transfection activity of a parent linear peptide in combination with a plasmid DNA cargo. This strategy allowed the straightforward preparation, under physiologically compatible conditions, of a discrete library of amphiphilic modulated penetrating peptides. Without the requirement of any isolation or purification steps, these modulated amphiphilic peptides were combined with a plasmid DNA and screened in transfection experiments of human HeLa cells. Three of these hydrazone-conjugated peptides were identified as excellent vectors for plasmid delivery with comparable, or even higher, efficiencies and lower toxicity than the commercial reagents employed in routine transfection assays We are thankful to Dr. Irene Lostalé-Seijo for cell culture assistance and discussions. We acknowledge funding from the Spanish Government MINECO: [CTQ2014-59646-R] and [CTQ2015-74621-JIN], the Xunta de Galicia (ED431G/09), the ERDF and the CESGA. R. G.-F. received a FCT Investigator Grant from Portugal (IF/01133/2015). J.M. received a Ramon y Cajal (RYC-2013-13784) and an ERC Starting Investigator Grant (DYNAP-677786) SI

NOTICE: This is the peer reviewed version of the following article: Fuertes A, Juanes M, Granja J, Montenegro J. Supramolecular functional assemblies: dynamic membrane transporters and peptide nanotubular composites. Chem Commun. 2017;53(56):7861-7871. [doi: 10.1039/C7CC02997G]. This article may be used for non-commercial purposes in accordance with RSC Terms and Conditions for self-archiving The fabrication of functional molecular devices constitutes one of the most important current challenges for chemical sciences. The complex processes accomplished by living systems continuously demand the assistance of non-covalent interactions between molecular building blocks. Additionally, these building blocks (proteins, membranes, nucleotides) are also constituted by self-assembled structures. Therefore, supramolecular chemistry is the discipline required to understand the properties of the minimal self-assembled building blocks of living systems and to develop new functional smart materials. In the first part of this feature article, we highlight selected examples of the preparation of supramolecular membrane transporters with special emphasis in the application of dynamic covalent bonds. In the second section of the paper we review recent breakthroughs in the preparation of peptide nanotubes hybrids with functional applications. The development of these devices constitutes an exciting process from where we can learn how to understand and manipulate supramolecular functional assemblies We acknowledge funding from the Spanish Agencia Estatal de Investigación (AEI) (CTQ2014-59646-R and CTQ2016-78423-R), the Xunta de Galicia (ED431G/09 and 2016-AD031) and the ERDF. A. F. and M. J. received F. P. U. and F. P. I. fellowships from MECD and MINECO. J. M. received a Ramon y Cajal Grant (RYC-2013-13784), an ERC Starting Investigator Grant (DYNAP-677786) and a Young Investigator Grant from the Human Frontier Research Program (RGY0066/2017) SI

Two of the main types of nanotubular architectures are the single-walled carbon nanotubes (SWCNTs) and the self-assembling cyclic peptide nanotubes (SCPNs). We here report the preparation of the dual composite resulting from the ordered combination of both tubular motifs. In the resulting architecture, the SWCNTs can act as templates for the assembly of SCPNs that engage the carbon nanotubes noncovalently via pyrene "paddles", each member of the resulting hybrid stabilizing the other in aqueous solution. The particular hybrids obtained in the present study formed highly ordered oriented arrays and display complementary properties such as electrical conductivity. Furthermore, a self-sorting of the cyclic peptides toward semiconducting rather than metallic SWCNTs is also observed in the aqueous dispersions. It is envisaged that a broad range of exploitable properties may be achieved and/or controlled by varying the cyclic peptide components of similar SWCNT/SCPN hybrids.

NOTICE: This is the peer reviewed version of the following article: Salvadó, I, Gamba, I, Montenegro J, Martínez-Costas J, Brea JM, Loza MI, Vázquez López M, Vázquez M.E. Membrane-disrupting iridium(III) oligocationic organometallopeptides. Chem. Commun., 2016,52, 11008-11011. DOI: 10.1039/C6CC05537K. This article may be used for non-commercial purposes in accordance with RSC Terms and Conditions for self-archiving A series of oligoarginine peptide derivatives containing cyclometallated iridium(III) units display remarkable cytotoxicity, comparable to that of cisplatin. In vitro studies with unilamellar vesicles support a membrane-disrupting mechanism of action We are thankful for the support given by the Spanish grants SAF2013-41943-R, CTQ2015-70698-R, CTQ2013-49317-EXP,CTQ2014-59646-R, and BFU2013-43513-R, and the Xunta de Galicia GRC2013-041. Support from COST Action CM1105 and the orfeo-cinqa network (CTQ2014-51912-REDC) is kindly acknowledged SI

Palytoxin (PLTX) is a lethal natural toxin often found in Palythoa zoantharians that, together with its congeners, may induce adverse effects in humans after inhalation of toxic aerosols both in open-air and domestic environments, namely in the vicinity of public and private aquaria. In this study, we describe a poisoning of an aquarium hobbyist who was hospitalized after handling a PLTXs-containing zoantharian hexacoral. Furthermore, we provide evidence for water detoxification. The zoantharian was morphologically and genetically identified as Palythoa cf. toxica (Cnidaria: Anthozoa). Palytoxin itself and two new PLTX congeners, a hydroxyPLTX and a deoxyPLTX, were detected and structurally identified by liquid chromatography high resolution multiple stage mass spectrometry (LC-HRMSn, n = 1, 2). Total and individual toxins were quantified by LC-HRMS and sandwich ELISA both in the zoantharian (93.4 and 96.80 μg/g, respectively) and in the transport water (48.3 and 42.56 μg/mL, respectively), with an excellent mean bias of 1.3% between the techniques. Activated carbon adsorbed 99.7% of PLTXs contained in the seawater and this represents a good strategy for preventing aquarium hobbyist poisonings.

NOTICE:This is the peer-reviewed version of the following article:Juan M. Priegue, Daniel N. Crisan, José Martínez-Costas, Juan R. Granja, Francisco Fernandez-Trillo, and Javier Montenegro (2016),“In situ Functionalized Polymers for siRNA Delivery ; Angew. Chem. Int. Ed., 55, 7492–7495 [doi: 10.1002/anie.201601441]. This article may be used for non-commercial purposes in accordance with Wiley Terms and Conditions for self-archiving A new method is reported herein for screening the biological activity of functional polymers across a consistent degree of polymerization and in situ, that is, under aqueous conditions and without purification/isolation of candidate polymers. In brief, the chemical functionality of a poly(acryloyl hydrazide) scaffold was activated under aqueous conditions using readily available aldehydes to obtain amphiphilic polymers. The transport activity of the resulting polymers can be evaluated in situ using model membranes and living cells without the need for tedious isolation and purification steps. This technology allowed the rapid identification of a supramolecular polymeric vector with excellent efficiency and reproducibility for the delivery of siRNA into human cells (HeLa-EGFP). The reported method constitutes a blueprint for the high-throughput screening and future discovery of new polymeric functional materials with important biological applications Royal Society. Grant Number: IE130688 Spanish Ministry of Economy and Competitiveness. Grant Numbers: CTQ2014-59646-R, CTQ2013-43264-R, BFU2013-43513-R Birmingham Science City European Regional Development Fund Royal Society. Grant Number: RG140273 University of Birmingham MEC MINECO ERC. Grant Number: DYNAP-677786 SI

The objective of this account is to summarize our recent progress with functional biosupramolecular systems concisely. The functions covered are artificial photosynthesis, anion transport, and sensing in lipid bilayer membranes. With artificial photosynthesis, the current emphasis is on the construction of ordered and oriented architectures on solid surfaces. Recent examples include the zipper assembly of photosystems with supramolecular n/p-heterojunctions and oriented antiparallel redox gradients. Current transport systems in lipid bilayers reveal new interactions at work. Examples include anion−macrodipole or anion−π interactions. Current attention with membrane-based sensing systems shifts from biosensor approaches with enzymatic signal generation to aptamers (i.e., the DNA version of immunosensing) and differential sensing with dynamic polyion−counterion transporters. The functional diversity accessible with biosupramolecular systems is highlighted, as is the critical importance of cross-fertilization at intertopical convergence zones.

The design, synthesis, and evaluation of small peptides with one to three negative charges and one to three hydrazides as key components of membrane-based synthetic sensing systems are reported. Their spontaneous reaction with hydrophobic aldehydes or ketones gives rapid access to small collections of amphiphilic anions. These anionic amphiphiles can activate polycations as anion transporters in lipid-bilayer membranes. Odorants are used as representative hydrophobic aldehydes and ketones, and cell-penetrating peptides (CPPs) as polycationic transporters in fluorogenic vesicles. Different activities obtained with different counterion activators are used to generate multidimensional patterns that can be recognized by principal component and hierarchical cluster analysis to extract unique “fingerprints” for individual analytes (including enantiomers, cis–trans isomers or perfumes as illustrative analyte mixtures). Comparison of the peptide activators reveals that carboxylates perform better than phosphonates. Gemini-like activators containing two carboxylates and two hydrophobic hydrazone tails are best, whereas excessive charges and tails give weaker activities. This result differs from cationic activators of polyanionic transporters such as DNA, which worked best with octopus amphiphiles with one cationic head and four hydrophobic tentacles.

Dynamic amphiphiles are amphiphiles with dynamic covalent bridges between their hydrophilic heads and their hydrophobic tails. Their usefulness to activate ion transporters for odorant release and for differential sensing of odorants and perfumes has been demonstrated recently. Here we report that the same "fragrant" dynamic amphiphiles are ideal to screen for new siRNA transfection agents. The advantages of this approach include rapid access to fairly large libraries of complex structures and possible transformation en route to assist uptake and minimize toxicity. We report single component systems that exceed the best commercially available multicomponent cocktails with regard to both efficiency and velocity of EGFP knockdown in HeLa cells. In human primary fibroblasts siRNA mediated enzyme knockdown nearly doubled from >30 for Lipofectamine to >60 for our best hit. The identified structures were predictable neither from literature nor from results in fluorogenic vesicles and thus support the importance of conceptually innovative screening approaches. © 2013 American Chemical Society.

Dynamic amphiphiles have a “bridge” between their charged head and their hydrophobic tails. The presence of dynamic covalent bonds is of interest for differential and biosensing applications as well as for rapid access to the libraries needed to screen for gene delivery or cellular uptake of siRNA. However, efforts to develop libraries have so far concentrated on hydrazone bridges to monocationic heads. Here, we report synthesis efforts to enlarge this focused library with oxime and disulfide bridges and dynamic amphiphiles with more than one positive charge. Evaluation in fluorogenic vesicles reveals best activation of DNA as ion transporters by dynamic amphiphiles with dendritic scaffolds, doubly charged heads and four tails. Moreover, oximes, contrary to hydrazones, remain active under acidic conditions. Linear elongation of dendritic head-groups seems to cause increasing detergent effects and should therefore be avoided.

This article summarizes the background and a few preliminary results concerning project 7 of the NCCR Chemical Biology. The general objective is to explore new concepts for cellular uptake, membrane tunneling, sensing and labeling. Emphasis is on the use of dynamic covalent chemistry for counterion activation, slow release of polyions and fluorescent probes, and the generation of activator libraries and polyions that grow and shrink.

Transport across the membranes of polymersomes remains difficult in part due to the great thickness of the polymer bilayers. Here, we report that dynamic polyion-counterion transport systems are active in fluorogenic polymersomes composed of poly(dimethylsiloxane)-b-poly(2-methyloxazoline) (PDMS-PMOXA). These results suggest that counterion-activated calf-thymus DNA can act as cation carrier that moves not only across lipid bilayer and bulk chloroform membranes but also across the "plastic" membranes of polymersomes. Compared to egg yolk phosophatidylcholine (EYPC) lipsosomes, activities and activator scope in PDMS-PMOXA polymersomes are clearly reduced. Embedded in agar gel matrices, fluorogenic PDMS-PMOXA polymersomes respond reliably to polyion-counterion transporters, with high contrast, high stability and preserved selectivity. Compared to standard EYPC liposomes, it cannot be said that PDMS-PMOXA polymersomes are better. However, they are different, and this difference could be interesting for the development of sensing devices.

This critical review covers progress with synthetic transport systems, particularly ion channels and pores, between January 2006 and December 2009 in a comprehensive manner. This is the third part of a series launched in the year 2000, covering a rich collection of structural and functional motifs that should appeal to a broad audience of non-specialists, including to organic, biological, supramolecular and polymer chemists. Impressive breakthroughs have been achieved over the past four years in part because of a fruitful expansion toward new types of interactions, including metal–organic, π–π, aromatic electron donor–acceptor, anion–π or anion–macrodipole interactions as well as dynamic covalent bonds (169 references).

Dynamic octopus amphiphiles contain one charged "head," here a guanidinium cation, together several hydrophobic "tails" (or "tentacles") that can be attached and exchanged in situ by reversible hydrazone formation. Quite surprisingly, their ability to activate DNA as transporters in lipid bilayer membranes was found to increase with the number of tails (up to four) as well as with their length (up to eight carbons). Both encouraged and puzzled by these results, we decided that a comprehensive screening of octopus amphiphiles with regard to number (from one to six) and length (from three to eighteen carbons) of their tails would be appropriate at this point. For this purpose, we here report the synthesis of cationic hexahydrazide peptide dendrons together with that of aldehydes with long, saturated, unsaturated and branched hydrophobic tails. Comprehensive screening of the completed collection of tails and heads reveals that the ability of octopus amphiphiles to activate DNA transporters shifts with increasing number of tails to decreasing length of the tails. Moreover, cis-alkenyl and branched alkyl tails are more active than their linear analogs, branched aromatic tails are best. These overall very meaningful trends for octopus amphiphiles will be of importance for sensing applications and fragrant cellular uptake.

The cytosolic delivery of hydrophilic, anionic molecular probes and therapeutics is a major challenge in chemical biology and medicine. Herein, we describe the design and synthesis of peptide–cage hybrids that allow an efficient supramolecular binding, cell membrane translocation and cytosolic delivery of a number of anionic dyes, including pyranine, carboxyfluorescein and several sulfonate-containing Alexa dyes. This supramolecular caging strategy is successful in different cell lines, and the dynamic carrier mechanism has been validated by U-tube experiments. The high efficiency of the reported approach allowed intracellular pH tracking by exploiting the ratiometric excitation of the pyranine fluorescent probe We acknowledge funding from the Spanish Agencia Estatal de Investigación (AEI) [CTQ2014-59646-R, SAF2017-89890-R, and SAF2016-76689-R], the Xunta de Galicia (ED431G/09, 2015-CP082, ED431C 2017/19, 2016-AD031, and Centro Singular de Investigación de Galicia accreditation 2016–2019, and ED431G/09) and the European Union (European Regional Development Fund – ERDF), and the European Research Council (Advanced Grant No. 340055 for JLM and Starting Grant for J. Montenegro (DYNAP-677786)). H. F.-C. thanks Xunta de Galicia (Predoctoral fellowship: ED481A-2017/047). J. Montenegro received a RyC (RYC-2013-13784) and a Young Investigator Grant from the HFSP (RGY0066/2017). J. M. and M. M.-C. thank MINECO for JdC fellowships (FJCI-2015-25080 and IJCI-2014-19326) SI