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Many scientists spend unnecessary time reformatting papers to submit them to different journals. We propose a uniform submission format that we hope journals will include in their options for submission. Widespread adoption of this uniform submission format could shorten the submission and publishing process, freeing up time for research.

Ultraviolet (UV) light and incompletely understood genetic and epigenetic variations determine skin color. Here we describe an UV- and microphthalmia-associated transcription factor (MITF)-independent mechanism of skin pigmentation. Targeting the mitochondrial redox-regulating enzyme nicotinamide nucleotide transhydrogenase (NNT) resulted in cellular redox changes that affect tyrosinase degradation. These changes regulate melanosome maturation and, consequently, eumelanin levels and pigmentation. Topical application of small-molecule inhibitors yielded skin darkening in human skin, and mice with decreased NNT function displayed increased pigmentation. Additionally, genetic modification of NNT in zebrafish alters melanocytic pigmentation. Analysis of four diverse human cohorts revealed significant associations of skin color, tanning, and sun protection use with various single-nucleotide polymorphisms within NNT. NNT levels were independent of UVB irradiation and redox modulation. Individuals with postinflammatory hyperpigmentation or lentigines displayed decreased skin NNT levels, suggesting an NNT-driven, redox-dependent pigmentation mechanism that can be targeted with NNT-modifying topical drugs for medical and cosmetic purposes.

Establishment and maintenance of the blood system relies on self-renewing hematopoietic stem cells (HSCs) that normally reside in small numbers in the bone marrow niche of adult mammals. This Review describes the developmental origins of HSCs and the molecular mechanisms that regulate lineage-specific differentiation. Studies of hematopoiesis provide critical insights of general relevance to other areas of stem cell biology including the role of cellular interactions in development and tissue homeostasis, lineage programming and reprogramming by transcription factors, and stage- and age-specific differences in cellular phenotypes.

In this issue of Cell, a study by N. Cho and coworkers provides insight into the role of vascular endothelial growth factor (VEGF) signaling in Drosophila hematopoiesis. Their work suggests that an ancestral function of VEGF was to guide blood cell migration and highlights the conservation of at least one aspect of VEGF signaling during evolution.

In this issue of Cell, a study by Adolfsson and coworkers (Adolfsson et al., 2005) provides insight into the early lineage commitment events of multipotent hematopoietic stem cells (HSCs). These studies demonstrate the importance of the Flt3 receptor tyrosine kinase as the earliest marker of hematopoietic cell fate commitment in that erythrocyte and megakaryocyte potentials are lost first as HSCs differentiate to lymphocyte progenitors.

SummaryEx vivo expansion of satellite cells and directed differentiation of pluripotent cells to mature skeletal muscle have proved difficult challenges for regenerative biology. Using a zebrafish embryo culture system with reporters of early and late skeletal muscle differentiation, we examined the influence of 2,400 chemicals on myogenesis and identified six that expanded muscle progenitors, including three GSK3β inhibitors, two calpain inhibitors, and one adenylyl cyclase activator, forskolin. Forskolin also enhanced proliferation of mouse satellite cells in culture and maintained their ability to engraft muscle in vivo. A combination of bFGF, forskolin, and the GSK3β inhibitor BIO induced skeletal muscle differentiation in human induced pluripotent stem cells (iPSCs) and produced engraftable myogenic progenitors that contributed to muscle repair in vivo. In summary, these studies reveal functionally conserved pathways regulating myogenesis across species and identify chemical compounds that expand mouse satellite cells and differentiate human iPSCs into engraftable muscle.

SummaryDNA methylation at the 5 position of cytosine (5-mC) is a key epigenetic mark that is critical for various biological and pathological processes. 5-mC can be converted to 5-hydroxymethylcytosine (5-hmC) by the ten-eleven translocation (TET) family of DNA hydroxylases. Here, we report that “loss of 5-hmC” is an epigenetic hallmark of melanoma, with diagnostic and prognostic implications. Genome-wide mapping of 5-hmC reveals loss of the 5-hmC landscape in the melanoma epigenome. We show that downregulation of isocitrate dehydrogenase 2 (IDH2) and TET family enzymes is likely one of the mechanisms underlying 5-hmC loss in melanoma. Rebuilding the 5-hmC landscape in melanoma cells by reintroducing active TET2 or IDH2 suppresses melanoma growth and increases tumor-free survival in animal models. Thus, our study reveals a critical function of 5-hmC in melanoma development and directly links the IDH and TET activity-dependent epigenetic pathway to 5-hmC-mediated suppression of melanoma progression, suggesting a new strategy for epigenetic cancer therapy.

Abstract 3387 BMP and Wnt signaling pathways control essential cellular responses through activation of the transcription factors SMAD (BMP) and TCF (Wnt). Here, we have evaluated their function during hematopoietic regeneration after irradiation. Using heat-shock inducible transgenic zebrafish lines that overexpress BMP2 or Wnt8, we demonstrated accelerated marrow recovery following irradiation. Heat-shock induced overexpression of the respective inhibitors Chordin and DKK1 blunted the recovery. Surprisingly, gene expression profiling after induction of BMP or Wnt signaling in zebrafish marrow cells post-irradiation revealed increased expression of the key hematopoietic genes scl, runx1, and gata2. To determine if the effect of BMP and Wnt signaling on hematopoietic genes during regeneration was direct, we performed ChIP-PCR for Smad1 and the hematopoietic regulator Gata2 in murine lineage-negative progenitors seven days after a sublethal irradiation. We found that Smad1 and Gata2 co-occupy hematopoietic genes including Cd9, Il13, Mapk6, and Meis1. To examine the binding of SMAD1 and TCF7L2 throughout the genome of hematopoietic cells, we employed ChIP-seq in human erythroid and myeloid leukemia cell lines, K562 and U937, respectively. More than 70% of the genes bound by SMAD1 and TCF7L2 were co-occupied with the lineage transcription factors GATA1 and GATA2 in erythroid cells, and with C/EBPα in myeloid cells. This finding suggests that signaling transcription factors control hematopoietic gene programs by binding DNA adjacent to lineage-specific transcription factors. The transcriptional output of BMP and Wnt activity was tested on an LMO2 enhancer reporter construct. Expression of SMAD1 or TCF7L2 alone had little effect, but markedly increased reporter activity in conjunction with GATA2, indicating that BMP and Wnt signaling cooperate with lineage regulators to enhance transcription of cell-type specific target genes. To establish the order of transcription factor occupancy, we utilized estrogen-inducible C/EBPα-ER in K562 cells or GATA1 induction in murine G1ER cell lines, and assessed SMAD1 occupancy before and after induction of each respective lineage regulator. Induction of the myeloid lineage regulator C/EBPα in K562 cells shifted binding of SMAD1, such that SMAD1 co-occupancy with C/EBPα changed from 6% to 15% of C/EBPα targets. In contrast, expression of the erythroid regulator GATA1 promoted loss of SMAD1 on 82% of its targets, and restricted more than 98% of the remaining SMAD1 sites to erythroid targets adjacent to GATA1. Co-occupancy of signaling factors and lineage regulators was further tested in primary human CD34+ multipotent hematopoietic progenitors and CD34+ cells directed to the erythroid lineage. Both SMAD1 and TCF7L2 co-localized with GATA2 on greater than 75% of bound genes in multipotent CD34+ progenitor cells. Similar to our results following GATA1 induction in G1ER cells, SMAD1 occupancy shifted to 65% erythroid targets upon differentiation of progenitors to the erythroid lineage. These data provide strong evidence that the binding of signaling factors follows the genomic occupancy of the dominant lineage regulator during differentiation. Together, our findings demonstrate that hematopoietic regeneration is driven by collaboration of master regulators and signaling transcription factors to control the entire hematopoietic program. Disclosures: Daley: Verastem, Inc: Equity Ownership, Membership on an entity's Board of Directors or advisory committees; iPierian, Inc: Equity Ownership, Membership on an entity's Board of Directors or advisory committees; Epizyme, Inc: Equity Ownership, Membership on an entity's Board of Directors or advisory committees; Solasia, KK: Equity Ownership, Membership on an entity's Board of Directors or advisory committees; MPM Capital, Inc: Consultancy, Membership on an entity's Board of Directors or advisory committees; Johnson & Johnson: Membership on an entity's Board of Directors or advisory committees. Zon:Fate Therapeutics:; Stemgent: Consultancy.

In this issue of Cell, a study by N. Cho and coworkers provides insight into the role of vascular endothelial growth factor (VEGF) signaling in Drosophila hematopoiesis. Their work suggests that an ancestral function of VEGF was to guide blood cell migration and highlights the conservation of at least one aspect of VEGF signaling during evolution.

Summary Recent genome-wide studies have demonstrated that pausing of RNA polymerase II (Pol II) occurred on many vertebrate genes. By genetic studies in the zebrafish tif1γ mutant moonshine we found that loss of function of Pol II-associated factors PAF or DSIF rescued erythroid gene transcription in tif1γ -deficient animals. Biochemical analysis established physical interactions among TIF1γ, the blood-specific SCL transcription complex, and the positive elongation factors p-TEFb and FACT. Chromatin immunoprecipitation assays in human CD34 + cells supported a TIF1γ-dependent recruitment of positive elongation factors to erythroid genes to promote transcription elongation by counteracting Pol II pausing. Our study establishes a mechanism for regulating tissue cell fate and differentiation through transcription elongation.

SummaryHematopoietic stem and progenitor cells (HSPCs) can reconstitute and sustain the entire blood system. We generated a highly specific transgenic reporter of HSPCs in zebrafish. This allowed us to perform high-resolution live imaging on endogenous HSPCs not currently possible in mammalian bone marrow. Using this system, we have uncovered distinct interactions between single HSPCs and their niche. When an HSPC arrives in the perivascular niche, a group of endothelial cells remodel to form a surrounding pocket. This structure appears conserved in mouse fetal liver. Correlative light and electron microscopy revealed that endothelial cells surround a single HSPC attached to a single mesenchymal stromal cell. Live imaging showed that mesenchymal stromal cells anchor HSPCs and orient their divisions. A chemical genetic screen found that the compound lycorine promotes HSPC-niche interactions during development and ultimately expands the stem cell pool into adulthood. Our studies provide evidence for dynamic niche interactions upon stem cell colonization.PaperFlick