At the body surface, skin's stratified squamous epithelium is challenged by environmental extremes. The surface of the skin is composed of enucleated, flattened surface squames. They derive from ...underlying, transcriptionally active keratinocytes that display filaggrin-containing keratohyalin granules (KGs) whose function is unclear. Here, we found that filaggrin assembles KGs through liquid-liquid phase separation. The dynamics of phase separation governed terminal differentiation and were disrupted by human skin barrier disease-associated mutations. We used fluorescent sensors to investigate endogenous phase behavior in mice. Phase transitions during epidermal stratification crowded cellular spaces with liquid-like KGs whose coalescence was restricted by keratin filament bundles. We imaged cells as they neared the skin surface and found that environmentally regulated KG phase dynamics drive squame formation. Thus, epidermal structure and function are driven by phase-separation dynamics.
The skin barrier is the body's first line of defence against environmental assaults, and is maintained by epithelial stem cells (EpSCs). Despite the vulnerability of EpSCs to inflammatory pressures, ...neither the primary response to inflammation nor its enduring consequences are well understood. Here we report a prolonged memory to acute inflammation that enables mouse EpSCs to hasten barrier restoration after subsequent tissue damage. This functional adaptation does not require skin-resident macrophages or T cells. Instead, EpSCs maintain chromosomal accessibility at key stress response genes that are activated by the primary stimulus. Upon a secondary challenge, genes governed by these domains are transcribed rapidly. Fuelling this memory is Aim2, which encodes an activator of the inflammasome. The absence of AIM2 or its downstream effectors, caspase-1 and interleukin-1β, erases the ability of EpSCs to recollect inflammation. Although EpSCs benefit from inflammatory tuning by heightening their responsiveness to subsequent stressors, this enhanced sensitivity probably increases their susceptibility to autoimmune and hyperproliferative disorders, including cancer.
The gain of eccrine sweat glands in hairy body skin has empowered humans to run marathons and tolerate temperature extremes. Epithelial-mesenchymal cross-talk is integral to the diverse patterning of ...skin appendages, but the molecular events underlying their specification remain largely unknown. Using genome-wide analyses and functional studies, we show that sweat glands are specified by mesenchymal-derived bone morphogenetic proteins (BMPs) and fibroblast growth factors that signal to epithelial buds and suppress epithelial-derived sonic hedgehog (SHH) production. Conversely, hair follicles are specified when mesenchymal BMP signaling is blocked, permitting SHH production. Fate determination is confined to a critical developmental window and is regionally specified in mice. In contrast, a shift from hair to gland fates is achieved in humans when a spike in BMP silences SHH during the final embryonic wave(s) of bud morphogenesis.
Increasing evidence suggests that microRNAs may play important roles in regulating self-renewal and differentiation in mammalian stem cells (SCs). Here, we explore this issue in skin. We first ...characterize microRNA expression profiles of skin SCs versus their committed proliferative progenies and identify a microRNA subset associating with “stemness.” Of these, miR-125b is dramatically downregulated in early SC progeny. We engineer an inducible mice system and show that when miR-125b is sustained in SC progenies, tissue balance is reversibly skewed toward stemness at the expense of epidermal, oil-gland, and HF differentiation. Using gain- and loss-of-function in vitro, we further implicate miR-125b as a repressor of SC differentiation. In vivo, transcripts repressed upon miR-125b induction are enriched >700% for predicted miR-125b targets normally downregulated upon SC-lineage commitment. We verify some of these miR-125b targets, and show that
Blimp1 and
VDR in particular can account for many tissue imbalances we see when miR-125b is deregulated.
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► A distinct microRNA signature is associated with “stemness” in the skin ► miR-125b regulates the balance of stemness and lineage commitment in Hair follicle SC ► miR-125b targets Blimp1 and Vdr to regulate the growth of SG and hair, respectively
Adult stem cells occur in niches that balance self-renewal with lineage selection and progression during tissue homeostasis. Following injury, culture or transplantation, stem cells outside their ...niche often display fate flexibility. Here we show that super-enhancers underlie the identity, lineage commitment and plasticity of adult stem cells in vivo. Using hair follicle as a model, we map the global chromatin domains of hair follicle stem cells and their committed progenitors in their native microenvironments. We show that super-enhancers and their dense clusters ('epicentres') of transcription factor binding sites undergo remodelling upon lineage progression. New fate is acquired by decommissioning old and establishing new super-enhancers and/or epicentres, an auto-regulatory process that abates one master regulator subset while enhancing another. We further show that when outside their niche, either in vitro or in wound-repair, hair follicle stem cells dynamically remodel super-enhancers in response to changes in their microenvironment. Intriguingly, some key super-enhancers shift epicentres, enabling their genes to remain active and maintain a transitional state in an ever-changing transcriptional landscape. Finally, we identify SOX9 as a crucial chromatin rheostat of hair follicle stem cell super-enhancers, and provide functional evidence that super-enhancers are dynamic, dense transcription-factor-binding platforms which are acutely sensitive to pioneer master regulators whose levels define not only spatial and temporal features of lineage-status but also stemness, plasticity in transitional states and differentiation.
Aging manifests with architectural alteration and functional decline of multiple organs throughout an organism. In mammals, aged skin is accompanied by a marked reduction in hair cycling and ...appearance of bald patches, leading researchers to propose that hair follicle stem cells (HFSCs) are either lost, differentiate, or change to an epidermal fate during aging. Here, we employed single-cell RNA-sequencing to interrogate aging-related changes in the HFSCs. Surprisingly, although numbers declined, aging HFSCs were present, maintained their identity, and showed no overt signs of shifting to an epidermal fate. However, they did exhibit prevalent transcriptional changes particularly in extracellular matrix genes, and this was accompanied by profound structural perturbations in the aging SC niche. Moreover, marked age-related changes occurred in many nonepithelial cell types, including resident immune cells, sensory neurons, and arrector pili muscles. Each of these SC niche components has been shown to influence HF regeneration. When we performed skin injuries that are known to mobilize young HFSCs to exit their niche and regenerate HFs, we discovered that aged skin is defective at doing so. Interestingly, however, in transplantation assays in vivo, aged HFSCs regenerated HFs when supported with young dermis, while young HFSCs failed to regenerate HFs when combined with aged dermis. Together, our findings highlight the importance of SC:niche interactions and favor a model where youthfulness of the niche microenvironment plays a dominant role in dictating the properties of its SCs and tissue health and fitness.
Hair follicle stem cells (HFSCs) regenerate hair in response to Wnt signalling. Here, we unfold genome-wide transcriptional and chromatin landscapes of β-catenin-TCF3/4-TLE circuitry, and genetically ...dissect their biological roles within the native HFSC niche. We show that during HFSC quiescence, TCF3, TCF4 and TLE (Groucho) bind coordinately and transcriptionally repress Wnt target genes. We also show that β-catenin is dispensable for HFSC viability, and if TCF3/4 levels are sufficiently reduced, it is dispensable for proliferation. However, β-catenin is essential to activate genes that launch hair follicle fate and suppress sebocyte fate determination. TCF3/4 deficiency mimics Wnt-β-catenin-dependent activation of these hair follicle fate targets; TCF3 overexpression parallels their TLE4-dependent suppression. Our studies unveil TCF3/4-TLE histone deacetylases as a repressive rheostat, whose action can be relieved by Wnt-β-catenin signalling. When TCF3/4 and TLE levels are high, HFSCs can maintain stemness, but remain quiescent. When these levels drop or when Wnt-β-catenin levels rise, this balance is shifted and hair regeneration initiates.
During embryogenesis, stem cells are set aside to fuel the postnatal hair cycle and repair the epidermis after injury. To define how hair follicle stem cells are specified and maintained in an ...undifferentiated state, we developed a strategy to isolate and transcriptionally profile embryonic hair progenitors in mice. We identified Lhx2 as a transcription factor positioned downstream of signals necessary to specify hair follicle stem cells, but upstream from signals required to drive activated stem cells to terminally differentiate. Using gain- and loss-of-function studies, we uncovered a role for Lhx2 in maintaining the growth and undifferentiated properties of hair follicle progenitors.
Immune and tissue stem cells retain an epigenetic memory of inflammation that intensifies sensitivity to future encounters. We investigated whether and to what consequence stem cells possess and ...accumulate memories of diverse experiences. Monitoring a choreographed response to wounds, we found that as hair follicle stem cells leave their niche, migrate to repair damaged epidermis, and take up long-term foreign residence there, they accumulate long-lasting epigenetic memories of each experience, culminating in post-repair epigenetic adaptations that sustain the epidermal transcriptional program and surface barrier. Each memory is distinct, separable, and has its own physiological impact, collectively endowing these stem cells with heightened regenerative ability to heal wounds and broadening their tissue-regenerating tasks relative to their naïve counterparts.
In adult skin, each hair follicle contains a reservoir of stem cells (the bulge), which can be mobilized to regenerate the new follicle with each hair cycle and to reepithelialize epidermis during ...wound repair. Here we report new methods that permit their clonal analyses and engraftment and demonstrate the two defining features of stem cells, namely self-renewal and multipotency. We also show that, within the bulge, there are two distinct populations, one of which maintains basal lamina contact and temporally precedes the other, which is suprabasal and arises only after the start of the first postnatal hair cycle. This spatial distinction endows them with discrete transcriptional programs, but surprisingly, both populations are growth inhibited in the niche but can self-renew in vitro and make epidermis and hair when grafted. These findings suggest that the niche microenvironment imposes intrinsic “stemness” features without restricting the establishment of epithelial polarity and changes in gene expression.
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