Runx1 and p21 synergistically limit the extent of hair follicle stem cell quiescence in vivo.

Mechanisms of tissue stem cell (SC) quiescence control are important for normal homeostasis and for preventing cancer. Cyclin-dependent kinase inhibitors (CDKis) are known inhibitors of cell cycle progression. We document CDKis expression in vivo during hair follicle stem cell (HFSC) homeostasis and find p21 (cyclin-dependent kinase inhibitor 1a, Cdkn1a), p57, and p15 up-regulated at quiescence onset. p21 appears important for HFSC timely onset of quiescence. Conversely, we find that Runx1 (runt related transcription factor 1), which is known for promoting HFSC proliferation, represses p21, p27, p57, and p15 transcription in HFSC in vivo. Intriguingly, in cell culture, tumors, and normal homeostasis, Runx1 and p21 interplay modulates proliferation in opposing directions under the different conditions. Unexpectedly, Runx1 and p21 synergistically limit the extent of HFSC quiescence in vivo, which antagonizes the role of p21 as a cell cycle inhibitor. Importantly, we find in cultured keratinocytes that Runx1 and p21 bind to the p15 promoter and synergistically repress p15 mRNA transcription, thereby restraining cell cycle arrest. This documents a surprising ability of a CDKi (p21) to act as a direct transcriptional repressor of another CDKi (p15). We unveil a robust in vivo mechanism that enforces quiescence of HFSCs, and a context-dependent role of a CDKi (p21) to limit quiescence of SCs, potentially by directly down-regulating mRNA levels of (an)other CDKi(s).

Design, testing and validation of an innovative web-based instrument to evaluate school meal quality.

OBJECTIVE: To develop a feasible, valid, reliable web-based instrument to objectively evaluate school meal quality in Swedish primary schools. DESIGN: The construct 'school meal quality' was operationalized by an expert panel into six domains, one of which was nutritional quality. An instrument was drafted and pilot-tested. Face validity was evaluated by the panel. Feasibility was established via a large national study. Food-based criteria to predict the nutritional adequacy of school meals in terms of fat quality, iron, vitamin D and fibre content were developed. Predictive validity was evaluated by comparing the nutritional adequacy of school menus based on these criteria with the results from a nutritional analysis. Inter-rater reliability was also assessed. SETTING: The instrument was developed between 2010 and 2012. It is designed for use in all primary schools by school catering and/or management representatives. SUBJECTS: A pilot-test of eighty schools in Stockholm (autumn 2010) and a further test of feasibility in 191 schools nationally (spring 2011). RESULTS: The four nutrient-specific food-based criteria predicted nutritional adequacy with sensitivity ranging from 0.85 to 1.0, specificity from 0.45 to 1.0 and accuracy from 0.67 to 1.0. The sample in the national study was statistically representative and the majority of users rated the questionnaire positively, suggesting the instrument is feasible. The inter-rater reliability was fair to almost perfect for continuous variables and agreement was ≥ 67 % for categorical variables. CONCLUSIONS: An innovative web-based system to comprehensively monitor school meal quality across several domains, with validated questions in the nutritional domain, is available in Sweden for the first time.

Muscle progenitor specification and myogenic differentiation are associated with changes in chromatin topology.

Using Hi-C, promoter-capture Hi-C (pCHi-C), and other genome-wide approaches in skeletal muscle progenitors that inducibly express a master transcription factor, Pax7, we systematically characterize at high-resolution the spatio-temporal re-organization of compartments and promoter-anchored interactions as a consequence of myogenic commitment and differentiation. We identify key promoter-enhancer interaction motifs, namely, cliques and networks, and interactions that are dependent on Pax7 binding. Remarkably, Pax7 binds to a majority of super-enhancers, and together with a cadre of interacting transcription factors, assembles feed-forward regulatory loops. During differentiation, epigenetic memory and persistent looping are maintained at a subset of Pax7 enhancers in the absence of Pax7. We also identify and functionally validate a previously uncharacterized Pax7-bound enhancer hub that regulates the essential myosin heavy chain cluster during skeletal muscle cell differentiation. Our studies lay the groundwork for understanding the role of Pax7 in orchestrating changes in the three-dimensional chromatin conformation in muscle progenitors.

Epigenetic Silencing of CDR1as Drives IGF2BP3-Mediated Melanoma Invasion and Metastasis.

Metastasis is the primary cause of death of cancer patients. Dissecting mechanisms governing metastatic spread may uncover important tumor biology and/or yield promising therapeutic insights. Here, we investigated the role of circular RNAs (circRNA) in metastasis, using melanoma as a model aggressive tumor. We identified silencing of cerebellar degeneration-related 1 antisense (CDR1as), a regulator of miR-7, as a hallmark of melanoma progression. CDR1as depletion results from epigenetic silencing of LINC00632, its originating long non-coding RNA (lncRNA) and promotes invasion in vitro and metastasis in vivo through a miR-7-independent, IGF2BP3-mediated mechanism. Moreover, CDR1as levels reflect cellular states associated with distinct therapeutic responses. Our study reveals functional, prognostic, and predictive roles for CDR1as and expose circRNAs as key players in metastasis.

Pax7 remodels the chromatin landscape in skeletal muscle stem cells.

Pluripotent stem cells (PSC) hold great promise for the treatment of human skeletal muscle diseases. However, it remains challenging to convert PSC to skeletal muscle cells, and the mechanisms by which the master regulatory transcription factor, Pax7, promotes muscle stem (satellite) cell identity are not yet understood. We have taken advantage of PSC-derived skeletal muscle precursor cells (iPax7), wherein the induced expression of Pax7 robustly initiates the muscle program and enables the in vitro generation of precursors that seed the satellite cell compartment upon transplantation. Remarkably, we found that chromatin accessibility in myogenic precursors pre-figures subsequent activation of myogenic differentiation genes. We also found that Pax7 binding is generally restricted to euchromatic regions and excluded from H3K27 tri-methylated regions in muscle cells, suggesting that recruitment of this factor is circumscribed by chromatin state. Further, we show that Pax7 binding induces dramatic, localized remodeling of chromatin characterized by the acquisition of histone marks associated with enhancer activity and induction of chromatin accessibility in both muscle precursors and lineage-committed myoblasts. Conversely, removal of Pax7 leads to rapid reversal of these features on a subset of enhancers. Interestingly, another cluster of Pax7 binding sites is associated with a durably accessible and remodeled chromatin state after removal of Pax7, and persistent enhancer accessibility is associated with subsequent, proximal binding by the muscle regulatory factors, MyoD1 and myogenin. Our studies provide new insights into the epigenetic landscape of skeletal muscle stem cells and precursors and the role of Pax7 in satellite cell specification.

Signalling couples hair follicle stem cell quiescence with reduced histone H3 K4/K9/K27me3 for proper tissue homeostasis.

Mechanisms of plasticity to acquire different cell fates are critical for adult stem cell (SC) potential, yet are poorly understood. Reduced global histone methylation is an epigenetic state known to mediate plasticity in cultured embryonic SCs and T-cell progenitors. Here we find histone H3 K4/K9/K27me3 levels actively reduced in adult mouse skin and hair follicle stem cells (HFSCs) during G0 quiescence. The level of marks over specific gene promoters did not correlate to mRNA level changes in quiescent HFSCs. Skin hypomethylation during quiescence was necessary for subsequent progression of hair homeostasis (cycle). Inhibiting BMP signal, a known HFSC anti-proliferative factor, elevated HFSC methylation in vivo during quiescence prior to proliferation onset. Furthermore, removal of proliferation factors and addition of BMP4 reduced histone methylases and increased demethylases mRNAs in cultured skin epithelial cells. We conclude that signalling couples hair follicle stem cell quiescence with reduced H3 K4/K9/K27me3 levels for proper tissue homeostasis.

Runx1 modulates adult hair follicle stem cell emergence and maintenance from distinct embryonic skin compartments.

Runx1 controls hematopoietic stem cell emergence and hair follicle stem cell (HFSC) activation and proliferation in adult skin. Here we use lineage tracing and mouse genetic manipulation to address the role of Runx1 in the embryonic development of HFSCs. We find Runx1 is expressed in distinct classes of embryonic skin precursors for short-term HF progenitors, adult HFSCs, and mesenchymal progenitors. Runx1 acts in the embryonic epithelium for timely emergence of adult HFSCs and short-term progenitors, but is dispensable for both of them. In contrast, Runx1 is strictly needed in the embryonic mesenchyme for proper adult HFSC differentiation and long-term skin integrity. Our data implicate Runx1 in epithelial cell adhesion and migration and in regulation of paracrine epithelial-mesenchymal cross talk. The latter involves Lef1 and Wnt signaling modulation in opposing directions from two distinct skin compartments. Thus, a master regulator of hematopoiesis also controls HFSC emergence and maintenance via modulation of bidirectional cross talking between nascent stem cells and their niche.