Targeting lymphoid-derived IL-17 signaling to delay skin aging

Skin aging is characterized by structural and functional changes that contribute to age-associated frailty. This probably depends on synergy between alterations in the local niche and stem cell-intrinsic changes, underscored by proinflammatory microenvironments that drive pleotropic changes. The nature of these age-associated inflammatory cues, or how they affect tissue aging, is unknown. Based on single-cell RNA sequencing of the dermal compartment of mouse skin, we show a skew towards an IL-17-expressing phenotype of T helper cells, γδ T cells and innate lymphoid cells in aged skin. Importantly, in vivo blockade of IL-17 signaling during aging reduces the proinflammatory state of the skin, delaying the appearance of age-related traits. Mechanistically, aberrant IL-17 signals through NF-κB in epidermal cells to impair homeostatic functions while promoting an inflammatory state. Our results indicate that aged skin shows signs of chronic inflammation and that increased IL-17 signaling could be targeted to prevent age-associated skin ailments.© 2023. The Author(s)

Dnmt3a and Dnmt3b Associate with Enhancers to Regulate Human Epidermal Stem Cell Homeostasis

The genome-wide localization and function of endogenous Dnmt3a and Dnmt3b in adult stem cells are unknown. Here, we show that in human epidermal stem cells, the two proteins bind in a histone H3K36me3-dependent manner to the most active enhancers and are required to produce their associated enhancer RNAs. Both proteins prefer super-enhancers associated to genes that either define the ectodermal lineage or establish the stem cell and differentiated states. However, Dnmt3a and Dnmt3b differ in their mechanisms of enhancer regulation: Dnmt3a associates with p63 to maintain high levels of DNA hydroxymethylation at the center of enhancers in a Tet2-dependent manner, whereas Dnmt3b promotes DNA methylation along the body of the enhancer. Depletion of either protein inactivates their target enhancers and profoundly affects epidermal stem cell function. Altogether, we reveal novel functions for Dnmt3a and Dnmt3b at enhancers that could contribute to their roles in disease and tumorigenesis.The S.A.B. laboratory research is supported by the European Research Council (ERC), the Worldwide Cancer Research Foundation, the Foundation La Marató de TV3, the Spanish Ministry of Economy and Development, the Foundation Vencer el Cancer (‘‘Beat Cancer’’), the Government of Cataluña (SGR and Mario Salvia’ grants), the Foundation Fundación Botín, and the Institute for Research in Biomedicine (IRB-Barcelona). L.R. is a La Caixa Foundation Ph.D. fellow. G.S. was supported by an AXA postdoctoral fellowship. IRB Barcelona is the recipient of a Severo Ochoa Award of Excellence from MINECO (Government of Spain). L.D.C. was supported by grants from the Spanish Ministerio de Educación y Ciencia (SAF2013-48926-P) and the European Commission’s 7th Framework Program 4DCellFate grant number 277899. We are grateful to the Common Fund’s Epigenomic Program from the NIH (USA) for providing the bisulphite whole genome sequencing data of human EpSC

β-catenina, una proteïna amb diverses funcions en l'acoblament de factors transcripcionals i factors d'unions adherents

La β-catenina és una proteïna acobladora de factors transcripcionals i també de factors involucrats en la formació de les unions adherents en cèl·lules epitelials. En la primera part del treball es presenta un estudi de β-catenina i placoglobina,en el que s'ha caracteritzat la interacció d'aquestes proteïnes amb els seus cofactors. La β-catenina i la placoglobina estan composades per dos extrems N-i C-terminals i un domini central de repeticions armadillo, per on interaccionen amb la majoria de cofactors. De la mateixa manera com passa amb la β-catenina, els extrems de placoglobina també interaccionen amb el domini armadillo i regulen la capacitat de la placoglobina per unir-se als diferents factors. A més, els extrems terminals, i no el domini armadillo, són els que confereixen l'especificitat d'interacció a la β-catenina i a la placoglobina amb els seus cofactors. En la segona part de la tesi s'ha aprofundit en la interrelació entre dues vies de senyalització, la de β-catenina i la ruta de NF-κB. S'havia descrit una regulació negativa de β-catenina sobre l'activitat transcripcional del factor NF-κB. Per primera vegada s'ha detectat la formació de complexes de NF-κB amb E-cadherina i amb altres components implicats en adhesió cel·lular, amb una colocalització d'aquests factors en la membrana plasmàtica de la cèl·lula. De manera similar a com passa amb β-catenina, l'activitat transcripcional de NF-κB també estaria regulada per E-cadherina, evitant així la transcripció de gens mesenquimals.β-catenin is a coupling protein for transcription factors and proteins involved in adherent junctions formation in epithelial cells. In the first chapter a study of β-catenin and plakoglobin is presented. The interaction of these two proteins with their cofactors has been characterized. βcatenin and plakoglobin are composed of two N-and C-terminal tails and an armadillo-like repeat containing central domain, surface that interacts with most of the cofactors. As it has been described for β-catenin, plakoglobin terminal tails interact with the armadillo domain and regulate the binding with different factors. Moreover, the terminal tails, and not the armadillo domain, are the domains responsible for determining β-catenin and plakoglobin specificity of interaction with their cofactors. In the second chapter of this thesis the crosstalk between β-catenin and NF-κB signaling pathways has been studied in deep. For the first time the complexes formed by NF-κB and E-cadherin, among other intercellular adhesion molecules, have been detected. These complexes were localized at the cellular membrane. As it has been described for β-catenin, NF-κB transcriptional activity is controlled by E-cadherin, avoiding the expression of mesenchymal genes

E-cadherin controls β-catenin and NF-κB transcriptional activity in mesenchymal gene expression

11 pages, 7 figures.-- et al.E-cadherin and its transcriptional repressor Snail1 (Snai1) are two factors that control epithelial phenotype. Expression of Snail1 promotes the conversion of epithelial cells to mesenchymal cells, and occurs concomitantly with the downregulation of E-cadherin and the upregulation of expression of mesenchymal genes such as those encoding fibronectin and LEF1. We studied the molecular mechanism controlling the expression of these genes in mesenchymal cells. Forced expression of E-cadherin strongly downregulated fibronectin and LEF1 RNA levels, indicating that E-cadherin-sensitive factors are involved in the transcription of these genes. E-cadherin overexpression decreased the transcriptional activity of the fibronectin promoter and reduced the interaction of beta-catenin and NF-kappaB with this promoter. Similar to beta-catenin, NF-kappaB was found, by co-immunoprecipitation and pull-down assays, to be associated with E-cadherin and other cell-adhesion components. Interaction of the NF-kappaB p65 subunit with E-cadherin or beta-catenin was reduced when adherens junctions were disrupted by K-ras overexpression or by E-cadherin depletion using siRNA. These conditions did not affect the association of p65 with the NF-kappaB inhibitor IkappaBalpha. The functional significance of these results was stressed by the stimulation of NF-kappaB transcriptional activity, both basal and TNF-alpha-stimulated, induced by an E-cadherin siRNA. Therefore, these results demonstrate that E-cadherin not only controls the transcriptional activity of beta-catenin but also that of NF-kappaB. They indicate too that binding of this latter factor to the adherens junctional complex prevents the transcription of mesenchymal genes.G. S. was supported by a fellowship from the UAB; M.E. and D.C. from the Ministerio de Educación; C.A., from the FIS (Fondo de Investigaciones Sanitarias), and M.P. from the AGAUR (Agència de Gestió d’Ajuts Universitaris i de Recerca). S.P. was supported by a La Cierva contract. This research was funded by FIS grants 01/3060 and 03/0925 to J.B., SAF2006-00339 to A.G.H., BFU2006-03203 to M.D., and SAF2004-01015 to A.M. Partial support through grants from the Instituto Carlos III (RTICCC, C03710), Comunidad de Madrid (S-GEN-0266-2006) and the Generalitat de Catalunya (2005SGR00970) is also appreciated.Peer reviewe

β-catenina, una proteïna amb diverses funcions en l'acoblament de factors transcripcionals i factors d'unions adherents

La β-catenina és una proteïna acobladora de factors transcripcionals i també de factors involucrats en la formació de les unions adherents en cèl·lules epitelials. En la primera part del treball es presenta un estudi de β-catenina i placoglobina,en el que s'ha caracteritzat la interacció d'aquestes proteïnes amb els seus cofactors. La β-catenina i la placoglobina estan composades per dos extrems N-i C-terminals i un domini central de repeticions armadillo, per on interaccionen amb la majoria de cofactors. De la mateixa manera com passa amb la β-catenina, els extrems de placoglobina també interaccionen amb el domini armadillo i regulen la capacitat de la placoglobina per unir-se als diferents factors. A més, els extrems terminals, i no el domini armadillo, són els que confereixen l'especificitat d'interacció a la β-catenina i a la placoglobina amb els seus cofactors. En la segona part de la tesi s'ha aprofundit en la interrelació entre dues vies de senyalització, la de β-catenina i la ruta de NF-κB. S'havia descrit una regulació negativa de β-catenina sobre l'activitat transcripcional del factor NF-κB. Per primera vegada s'ha detectat la formació de complexes de NF-κB amb E-cadherina i amb altres components implicats en adhesió cel·lular, amb una colocalització d'aquests factors en la membrana plasmàtica de la cèl·lula. De manera similar a com passa amb β-catenina, l'activitat transcripcional de NF-κB també estaria regulada per E-cadherina, evitant així la transcripció de gens mesenquimals.β-catenin is a coupling protein for transcription factors and proteins involved in adherent junctions formation in epithelial cells. In the first chapter a study of β-catenin and plakoglobin is presented. The interaction of these two proteins with their cofactors has been characterized. βcatenin and plakoglobin are composed of two N-and C-terminal tails and an armadillo-like repeat containing central domain, surface that interacts with most of the cofactors. As it has been described for β-catenin, plakoglobin terminal tails interact with the armadillo domain and regulate the binding with different factors. Moreover, the terminal tails, and not the armadillo domain, are the domains responsible for determining β-catenin and plakoglobin specificity of interaction with their cofactors. In the second chapter of this thesis the crosstalk between β-catenin and NF-κB signaling pathways has been studied in deep. For the first time the complexes formed by NF-κB and E-cadherin, among other intercellular adhesion molecules, have been detected. These complexes were localized at the cellular membrane. As it has been described for β-catenin, NF-κB transcriptional activity is controlled by E-cadherin, avoiding the expression of mesenchymal genes

Loss of dnmt3a and dnmt3b does not affect epidermal homeostasis but promotes squamous transformation through PPAR-g

The DNA methyltransferase Dnmt3a suppresses tumorigenesis in models of leukemia and lung cancer. Conversely, deregulation of Dnmt3b is thought to generally promote tumorigenesis. However, the role of Dnmt3a and Dnmt3b in many types of cancer remains undefined. Here, we show that Dnmt3a and Dnmt3b are dispensable for homeostasis of the murine epidermis. However, loss of Dnmt3a-but not Dnmt3b-increases the number of carcinogen-induced squamous tumors, without affecting tumor progression. Only upon combined deletion of Dnmt3a and Dnmt3b, squamous carcinomas become more aggressive and metastatic. Mechanistically, Dnmt3a promotes the expression of epidermal differentiation genes by interacting with their enhancers and inhibits the expression of lipid metabolism genes, including PPAR-γ, by directly methylating their promoters. Importantly, inhibition of PPAR-γ partially prevents the increase in tumorigenesis upon deletion of Dnmt3a. Altogether, we demonstrate that Dnmt3a and Dnmt3b protect the epidermis from tumorigenesis and that squamous carcinomas are sensitive to inhibition of PPAR-γ.The Spanish Ministry of Economy and Development (MINECO), and the Institute supported this project in the laboratory of SAB for Research in Biomedicine (IRB-Barcelona). IRB Barcelona is the recipient of a Severo Ochoa Award of Excellence from MINECO (Government of Spain). We are very grateful to the laboratories of Rudolph Jaenisch and Rafii Ahmed for providing us the Dnmt3a and Dnmt3b flox/flox animals. LR was sponsored by La Caixa International PhD fellowship. We thank all the core facilities at the IRB-Barcelona for their assistance in our work, and Veronica Raker for editing the manuscript. The raw data for every dataset included in the manuscript can be found at GEO (GSE87412)

Targeting lymphoid-derived IL-17 signaling to delay skin aging

Skin aging is characterized by structural and functional changes that contribute to age-associated frailty. This probably depends on synergy between alterations in the local niche and stem cell-intrinsic changes, underscored by proinflammatory microenvironments that drive pleotropic changes. The nature of these age-associated inflammatory cues, or how they affect tissue aging, is unknown. Based on single-cell RNA sequencing of the dermal compartment of mouse skin, we show a skew towards an IL-17-expressing phenotype of T helper cells, γδ T cells and innate lymphoid cells in aged skin. Importantly, in vivo blockade of IL-17 signaling during aging reduces the proinflammatory state of the skin, delaying the appearance of age-related traits. Mechanistically, aberrant IL-17 signals through NF-κB in epidermal cells to impair homeostatic functions while promoting an inflammatory state. Our results indicate that aged skin shows signs of chronic inflammation and that increased IL-17 signaling could be targeted to prevent age-associated skin ailments

Circadian Reprogramming in the Liver Identifies Metabolic Pathways of Aging

The process of aging and circadian rhythms are intimately intertwined, but how peripheral clocks involved in metabolic homeostasis contribute to aging remains unknown. Importantly, caloric restriction (CR) extends lifespan in several organisms and rewires circadian metabolism. Using young versus old mice, fed ad libitum or under CR, we reveal reprogramming of the circadian transcriptome in the liver. These age-dependent changes occur in a highly tissue-specific manner, as demonstrated by comparing circadian gene expression in the liver versus epidermal and skeletal muscle stem cells. Moreover, de novo oscillating genes under CR show an enrichment in SIRT1 targets in the liver. This is accompanied by distinct circadian hepatic signatures in NAD+-related metabolites and cyclic global protein acetylation. Strikingly, this oscillation in acetylation is absent in old mice while CR robustly rescues global protein acetylation. Our findings indicate that the clock operates at the crossroad between protein acetylation, liver metabolism, and aging

Long-lived force patterns and deformation waves at repulsive epithelial boundaries

\u3cp\u3eFor an organism to develop and maintain homeostasis, cell types with distinct functions must often be separated by physical boundaries. The formation and maintenance of such boundaries are commonly attributed to mechanisms restricted to the cells lining the boundary. Here we show that, besides these local subcellular mechanisms, the formation and maintenance of tissue boundaries involves long-lived, long-ranged mechanical events. Following contact between two epithelial monolayers expressing, respectively, EphB2 and its ligand ephrinB1, both monolayers exhibit oscillatory patterns of traction forces and intercellular stresses that tend to pull cell-matrix adhesions away from the boundary. With time, monolayers jam, accompanied by the emergence of deformation waves that propagate away from the boundary. This phenomenon is not specific to EphB2/ephrinB1 repulsion but is also present during the formation of boundaries with an inert interface and during fusion of homotypic epithelial layers. Our findings thus unveil a global physical mechanism that sustains tissue separation independently of the biochemical and mechanical features of the local tissue boundary.\u3c/p\u3