Promyelocytic leukemia nuclear bodies (PML-NBs) are PML-based nuclear structures that regulate various cellular processes. SUMOylation, the process of covalently conjugating small ubiquitin-like modifiers (SUMOs), is required for both the formation and the disruption of PML-NBs. However, detailed mechanisms of how SUMOylation regulates these processes remain unknown. Here we report that SUMO5, a novel SUMO variant, mediates the growth and disruption of PML-NBs. PolySUMO5 conjugation of PML at lysine 160 facilitates recruitment of PML-NB components, which enlarges PMLNBs. SUMO5 also increases polySUMO2/3 conjugation of PML, resulting in RNF4-mediated disruption of PML-NBs. The acute promyelocytic leukemia oncoprotein PML-RARα blocks SUMO5 conjugation of PML, causing cytoplasmic displacement of PML and disruption of PML-NBs. Our work not only identifies a new member of the SUMO family but also reveals the mechanistic basis of the PML-NB life cycle in human cells.Promyelocytic leukemia nuclear bodies (PML-NBs) are non-membrane-bound domains in the cell nucleus that regulate transcription, antiviral response, DNA repair, apoptosis, senescence, and tumor suppression 1 . PML-NBs require PML to form 2 . Mature PML-NBs also contain other components such as SP100, HIPK2 and Daxx, which depends on the cellular environment 2-4 and results in heterogeneity in PML-NBs. Proper generation of PML-NBs is critical to the individual as an absence or delocalization of PML-NBs results in several pathological conditions, including polyglutamine repeat neurodegenerative diseases 5 and acute promyelocytic leukemia (APL) 6 . Arsenic trioxide (ATO) induces the formation of PML-NBs through a two-step process: first, oxidized PML dimerizes using the BRCC domain 7,8 to form a ring-like structure of PML shells. This step has been referred to as nucleation of PML-NBs 9 . PML shells then start to mature by recruiting additional PML-NB components 1 . In cells without oxidative stress, PML-NBs become prominent when cells enter G1 phase. During mitosis, PML-NBs fall apart and PML forms aggregates. PML-NB components start to enter the bodies to colocalize with PML after mitotic exit 10,11 , suggesting that the initiation of PML-NB formation happens at the end of mitosis. Taken together, current evidence suggests that the biogenesis of PML-NBs, which finishes in G1, needs at least two steps: nucleation through PML dimerization and the recruitment of PML-NB components.The recruitment of PML-NB components depends on SUMOylation 2,12 as loss of UBC9, an E2 ligase for SUMOylation, blocks the formation of PML-NBs 13 . PML can be conjugated by SUMO1 on three lysines: K65, K160, and K490 12,14-17 . PML also contains a SUMO interaction domain (SIM), which mediates interaction with SUMOs. As PML-NB components are also SUMO1-conjugated or SIM-containing proteins 18 , it was proposed that PML-NBs mature through SUMO-SIM interaction networks that recruit PML-NB components, causing PML shells to enlarge 19 . However, a recent report showing that loss of SUMO1...

Keloids are wounding‐induced fibroproliferative human tumor‐like skin scars of complex genetic makeup and poorly defined pathogenesis. To reveal dynamic epigenetic and transcriptome changes of keloid fibroblasts, we performed RNA‐seq and ATAC‐seq analysis on an early passage keloid fibroblast cell strain and its paired normal control fibroblasts. This keloid strain produced keloid‐like scars in a plasma clot‐based skin equivalent humanized keloid animal model. RNA‐seq analysis reveals gene ontology terms including hepatic fibrosis, Wnt‐β‐catenin, TGF‐β, regulation of epithelial‐mesenchymal transition (EMT), STAT3 and adherens junction. ATAC‐seq analysis suggests STAT3 signalling is the most significantly enriched gene ontology term in keloid fibroblasts, followed by Wnt signalling (Wnt5) and regulation of the EMT pathway. Immunohistochemistry confirms that STAT3 (Tyr705 phospho‐STAT3) is activated and β‐catenin is up‐regulated in the dermis of keloid clinical specimens and keloid skin equivalent implants from the humanized mouse model. A non‐linear dose‐response of cucurbitacin I, a selective JAK2/STAT3 inhibitor, in collagen type I expression of keloid‐derived plasma clot‐based skin equivalents implicates a likely role of STAT3 signalling in keloid pathogenesis. This work also demonstrates the utility of the recently established humanized keloid mouse model in exploring the mechanism of keloid formation.

Animal skin pigment patterns are excellent models to study the mechanism of biological self-organization. Theoretical approaches developed mathematical models of pigment patterning and molecular genetics have brought progress; however, the responsible cellular mechanism is not fully understood. One long unsolved controversy is whether the patterning information is autonomously determined by melanocytes or nonautonomously determined from the environment. Here, we transplanted purified melanocytes and demonstrated that melanocytes could form periodic pigment patterns cell autonomously. Results of heterospecific transplantation among quail strains are consistent with this finding. Further, we observe that developing melanocytes directly connect with each other via filopodia to form a network in culture and in vivo. This melanocyte network is reminiscent of zebrafish pigment cell networks, where connexin is implicated in stripe formation via genetic studies. Indeed, we found connexin40 (cx40) present on developing melanocytes in birds. Stripe patterns can form in quail skin explant cultures. Several calcium channel modulators can enhance or suppress pigmentation globally, but a gap junction inhibitor can change stripe patterning. Most interestingly, in ovo, misexpression of dominant negative cx40 expands the black region, while overexpression of cx40 expands the yellow region. Subsequently, melanocytes instruct adjacent dermal cells to express agouti signaling protein (ASIP), the regulatory factor for pigment switching, which promotes pheomelanin production. Thus, we demonstrate Japanese quail melanocytes have an autonomous periodic patterning role during body pigment stripe formation. We also show dermal agouti stripes and how the coupling of melanocytes with dermal cells may confer stable and distinct pigment stripe patterns.Japanese quail | stripe pattern | melanocytes | ASIP | gap junction A nimal skin pigment patterns, such as periodic leopard spots and zebra stripes, represent some of the most amazing phenomena observed in nature, which have fascinated biologists and nonbiologists. The mechanisms of pigment patterning have been studied by mathematical and empirical approaches. Studies on zebrafish stripe patterning have led investigators to propose that the pattern is formed by pigment cell interactions that satisfy a Turing-type model (1,2). Mammalian genetic studies performed on horses, zebras, cheetahs, and chipmunks have identified some of the molecules involved in this process (3-5). Although theoretical models and the genetic backgrounds in the pigment patterning are well studied, how the pigment-related genes control the cell-cell interactions that generate the pigment pattern is largely unknown. Avian species present an excellent model system to answer these questions because of their extraordinarily diverse micropigment patterns within feathers (6) and embryonic manipulability that allows analyses of cell-cell interactions leading to macropigment patterning throughout the body. Japanese quail (JQ), a m...

Highlights d Two epigenetic modes control two b-keratin gene clusters during skin specification d One enhancer co-activates all subclustered Keratin genes on the Chr25 cluster d 3D chromatin looping mediates differential Keratin expression of the Chr27 cluster d 3-factor model explains how to establish region-specific chromatin configuration

Tissue regeneration is a process that recapitulates and restores organ structure and function. Although previous studies have demonstrated wound-induced hair neogenesis (WIHN) in laboratory mice (Mus), the regeneration is limited to the center of the wound unlike those observed in African spiny (Acomys) mice. Tissue mechanics have been implicated as an integral part of tissue morphogenesis. Here, we use the WIHN model to investigate the mechanical and molecular responses of laboratory and African spiny mice, and report these models demonstrate opposing trends in spatiotemporal morphogenetic field formation with association to wound stiffness landscapes. Transcriptome analysis and K14-Cre-Twist1 transgenic mice show the Twist1 pathway acts as a mediator for both epidermal-dermal interactions and a competence factor for periodic patterning, differing from those used in development. We propose a Turing model based on tissue stiffness that supports a two-scale tissue mechanics process: (1) establishing a morphogenetic field within the wound bed (mm scale) and (2) symmetry breaking of the epidermis and forming periodically arranged hair primordia within the morphogenetic field (μm scale). Thus, we delineate distinct chemo-mechanical events in building a Turing morphogenesis-competent field during WIHN of laboratory and African spiny mice and identify its evo-devo advantages with perspectives for regenerative medicine.

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Background: Animals develop skin regional specificities to best adapt to their environments. Birds are excellent models in which to study the epigenetic mechanisms that facilitate these adaptions. Patients suffering from SATB2 mutations exhibit multiple defects including ectodermal dysplasia-like changes. The preferential expression of SATB2, a chromatin regulator, in feather-forming compared to scale-forming regions, suggests it functions in regional specification of chicken skin appendages by acting on either differentiation or morphogenesis.Results: Retrovirus mediated SATB2 misexpression in developing feathers, beaks, and claws causes epidermal differentiation abnormalities (e.g. knobs, plaques) with few organ morphology alterations. Chicken β-keratins are encoded in 5 sub-clusters (Claw, Feather, Feather-like, Scale, and Keratinocyte) on Chromosome 25 and a large Feather keratin cluster on Chromosome 27.Type I and II α-keratin clusters are located on Chromosomes 27 and 33, respectively. Transcriptome analyses showed these keratins (1) are often tuned up or down collectively as a sub-cluster, and (2) these changes occur in a temporospatial specific manner.Conclusions: These results suggest an organizing role of SATB2 in clusterlevel gene co-regulation during skin regional specification.

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