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Exercise benefits a variety of organ systems in mammals, and some of the best-recognized effects of exercise on muscle are mediated by the transcriptional co-activator PPAR-γ co-activator-1 α (PGC1-α). Here we show in mouse that PGC1-α expression in muscle stimulates an increase in expression of FNDC5, a membrane protein that is cleaved and secreted as a newly identified hormone, irisin. Irisin acts on white adipose cells in culture and in vivo to stimulate UCP1 expression and a broad program of brown-fat-like development. Irisin is induced with exercise in mice and humans, and mildly increased irisin levels in the blood cause an increase in energy expenditure in mice with no changes in movement or food intake. This results in improvements in obesity and glucose homeostasis. Irisin could be therapeutic for human metabolic disease and other disorders that are improved with exercise. In mice, expression of PGC1-α in muscles is shown to stimulate expression of FNDC5, which is cleaved and secreted in the circulation as the newly identified hormone irisin; on exercise, this hormone stimulates browning of subcutaneous adipose tissue. Irisin counters diabetes and obesity Exercise is an effective therapy for obesity and type II diabetes. The transcriptional coactivator PGC1-α has been shown to mediate many of the effects of exercise in skeletal muscle, and here it is shown that PGC1-α expression in muscle stimulates the expression of the membrane protein FNDC5 in mice. FNDC5 is cleaved and secreted in the circulation as a previously unrecognized hormone, dubbed irisin, after Iris, the Greek messenger goddess. Irisin is elevated in the blood of humans and mice on exercising. It is a very powerful activator of a thermogenic program in primary white fat cells and causes a 'browning' of this cell type, including increased expression of UCP1 and enhanced respiration. These data identify irisin as a possible novel therapeutic for metabolic disorders.

The precise specification of left–right asymmetry is an essential process for patterning internal organs in vertebrates. In mouse embryonic development, the symmetry-breaking process in left–right determination is initiated by a leftward extraembryonic fluid flow on the surface of the ventral node. However, it is not known whether the signal transduction mechanism of this flow is chemical or mechanical. Here we show that fibroblast growth factor (FGF) signalling triggers secretion of membrane-sheathed objects 0.3–5 µm in diameter termed ‘nodal vesicular parcels’ (NVPs) that carry Sonic hedgehog and retinoic acid. These NVPs are transported leftward by the fluid flow and eventually fragment close to the left wall of the ventral node. The silencing effects of the FGF-receptor inhibitor SU5402 on NVP secretion and on a downstream rise in Ca 2+ were sufficiently reversed by exogenous Sonic hedgehog peptide or retinoic acid, suggesting that FGF-triggered surface accumulation of cargo morphogens may be essential for launching NVPs. Thus, we propose that NVP flow is a new mode of extracellular transport that forms a left–right gradient of morphogens. No left or right turn Though symmetrical from the outside, the body plan of vertebrates and other animals is far from symmetrical inside. By the time the human heart and lung develop in the embryo they are directed to the left and right of the body cavity. Much research has gone into establishing the genetics and signalling mechanisms that impose this asymmetry. But there is a catch: some tissues, chiefly the muscles and skeleton, must ignore or overrule the instruction if they are not to become asymmetric. Three papers in this issue, and a News and Views piece by Eran Hornstein and Clifford J. Tabin, address the fascinating question of how the somites, embryonic elements that give rise to symmetrical tissues, pull off that trick.

Natural competence for transformation is a common mode of horizontal gene transfer and contributes to bacterial evolution. Transformation occurs through the uptake of external DNA and its integration into the genome. Here we show that the type VI secretion system (T6SS), which serves as a predatory killing device, is part of the competence regulon in the naturally transformable pathogen Vibrio cholerae. The T6SS-encoding gene cluster is under the positive control of the competence regulators TfoX and QstR and is induced by growth on chitinous surfaces. Live-cell imaging revealed that deliberate killing of nonimmune cells via competence-mediated induction of T6SS releases DNA and makes it accessible for horizontal gene transfer in V. cholerae.

Type III secretion systems (T3SS) are specialized protein secretion systems that allow bacteria to inject toxins into eukaryotic cells. T3SS are important virulence factors, but their expression carries a fitness cost: they slow bacterial growth and make bacteria vulnerable to detection by the innate immune system. Some pathogens, like Pseudomonas aeruginosa, balance the costs and benefits of T3SS expression by restricting T3SS expression to a subset of cells. T3SS-ON cells arise from “primed” bacteria that express the transcriptional activator ExsA and respond immediately to T3SS activating signals. However, the mechanistic basis for priming is unknown. In this study, we tested whether expression of ExsA from a cAMP-dependent promoter could drive cells into the primed state and found this to be true. Whole-cell cryo-electron tomography demonstrated that primed bacteria assembled T3SS injectisomes. This work demonstrates how cAMP inputs into a bistable regulatory switch generate subpopulations of T3SS-primed cells.

Pseudomonas aeruginosa is an opportunist bacterium that causes acute and chronic infections. During acute infections, the type III secretion system (T3SS) plays a pivotal role in allowing the bacteria to translocate effectors such as ExoS, ExoT, and ExoY into host cells for colonization. Previous research on the involvement of quorum sensing systems Las and Rhl in controlling the T3SS gene expression produced ambiguous results. In this study, we determined the role of the Las and Rhl systems and the PqsE protein on T3SS expression. Our results show that in the wild-type PAO1 strain, the deletion of lasR or pqsE do not affect the secretion of ExoS. However, rhlI inactivation increases the expression of T3SS genes. In contrast to the rhlI deletion, rhlR inactivation decreases both T3SS genes expression and ExoS secreted protein levels, and this phenotype is restored when this mutant is complemented with the exsA gene, which codes for the master regulator of the T3SS. Additionally, cytotoxicity is affected in the rhlR mutant strain compared with its PAO1 parental strain. Overall, our results indicate that neither the Las system nor PqsE are involved in regulating the T3SS. Moreover, the Rhl system components have opposite effects, RhlI participates in negatively controlling the T3SS expression, while RhlR does it in a positive way, and this regulation is independent of C4 or PqsE. Finally, we show that rhlR , rhlI , or pqsE inactivation abolished pyocyanin production in T3SS-induction conditions. The ability of RhlR to act as a positive T3SS regulator in the absence of its cognate autoinducer and PqsE shows that it is a versatile regulator that controls different virulence traits allowing P . aeruginosa to compete for a niche.

The type III secretion system (T3SS) is a pivotal virulence mechanism of many Gram-negative bacteria. During infection, the syringe-like T3SS injects cytotoxic proteins directly into the eukaryotic host cell cytoplasm. In Pseudomonas aeruginosa , expression of the T3SS is regulated by a signaling cascade involving the proteins ExsA, ExsC, ExsD, and ExsE. The AraC-type transcription factor ExsA activates transcription of all T3SS-associated genes. Prior to host cell contact, ExsA is inhibited through direct binding of the anti-activator protein ExsD. Host cell contact triggers secretion of ExsE and sequestration of ExsD by ExsC to cause the release of ExsA. ExsA does not bind ExsD through the canonical ligand binding pocket of AraC-type proteins. Using site-directed mutagenesis and a specific in vitro transcription assay, we have now discovered that backbone interactions between the amino terminus of ExsD and the ExsA beta barrel constitute a pivotal part of the ExsD-ExsA interface. Follow-up bacterial two-hybrid experiments suggest additional contacts create an even larger protein–protein interface. The discovered role of the amino terminus of ExsD in ExsA binding explains how ExsC might relieve the ExsD-mediated inhibition of T3SS gene expression, because the same region of ExsD interacts with ExsC following host cell contact.

Effector proteins secreted by bacteria that infect mammalian and plant cells often subdue eukaryotic host cell defenses by simultaneously affecting multiple targets. However, instances when a bacterial effector injected in the competing bacteria sabotage more than a single target have not been reported. Here, we demonstrate that the effector protein, LtaE, translocated by the type IV secretion system from the soil bacterium Lysobacter enzymogenes into the competing bacterium, Pseudomonas protegens, affects several targets, thus disabling the antibacterial defenses of the competitor. One LtaE target is the transcription factor, LuxR1, that regulates biosynthesis of the antimicrobial compound, orfamide A. Another target is the sigma factor, PvdS, required for biosynthesis of another antimicrobial compound, pyoverdine. Deletion of the genes involved in orfamide A and pyoverdine biosynthesis disabled the antibacterial activity of P. protegens, whereas expression of LtaE in P. protegens resulted in the near-complete loss of the antibacterial activity against L. enzymogenes. Mechanistically, LtaE inhibits the assembly of the RNA polymerase complexes with each of these proteins. The ability of LtaE to bind to LuxR1 and PvdS homologs from several Pseudomonas species suggests that it can sabotage defenses of various competitors present in the soil or on plant matter. Our study thus reveals that the multi-target effectors have evolved to subdue cell defenses not only in eukaryotic hosts but also in bacterial competitors.

Objective: Accumulating evidence raises the hypothesis that dysregulation of intrinsic clock mechanisms are involved in the development of the metabolic syndrome, type 2 diabetes mellitus and cardiovascular disease. The aim of the present study was to investigate the relationship between three known common polymorphisms in the Clock gene and features of the metabolic syndrome in man. Methods: Genotype and haplotype analysis was carried out in a cohort of 537 individuals from 89 families characterized for inflammatory, atherothrombotic and metabolic risk associated with insulin resistance. Results: Heritability of the metabolic syndrome, defined according to International Diabetes Federation criteria, was 0.40. Haplotype analysis indicated three common haplotypes: CAT, TGT and CGC (rs4864548-rs3736544-rs1801260) with frequencies of 31, 33 and 28%, respectively. The CGC haplotype was less prevalent in subjects with the metabolic syndrome (P=0.0015) and was associated with lower waist circumference (P=0.007), lower hip circumference (P=0.023), lower body mass index (P=0.043) and lower leptin levels (P=0.028). The CAT haplotype was significantly associated with the presence of the metabolic syndrome (P=0.020). Conclusions: These findings suggest that the Clock gene CGC haplotype may be protective for the development of obesity and support the hypothesis that genetic variation in the Clock gene may play a role in the development of the metabolic syndrome, type 2 diabetes and cardiovascular disease.

All pancreatic endocrine cell types arise from a common endocrine precursor cell population, yet the molecular mechanisms that establish and maintain the unique gene expression programs of each endocrine cell lineage have remained largely elusive. Such knowledge would improve our ability to correctly program or reprogram cells to adopt specific endocrine fates. Here, we show that the transcription factor Nkx6.1 is both necessary and sufficient to specify insulin-producing beta cells. Heritable expression of Nkx6.1 in endocrine precursors of mice is sufficient to respecify non-beta endocrine precursors towards the beta cell lineage, while endocrine precursor- or beta cell-specific inactivation of Nkx6.1 converts beta cells to alternative endocrine lineages. Remaining insulin(+) cells in conditional Nkx6.1 mutants fail to express the beta cell transcription factors Pdx1 and MafA and ectopically express genes found in non-beta endocrine cells. By showing that Nkx6.1 binds to and represses the alpha cell determinant Arx, we identify Arx as a direct target of Nkx6.1. Moreover, we demonstrate that Nkx6.1 and the Arx activator Isl1 regulate Arx transcription antagonistically, thus establishing competition between Isl1 and Nkx6.1 as a critical mechanism for determining alpha versus beta cell identity. Our findings establish Nkx6.1 as a beta cell programming factor and demonstrate that repression of alternative lineage programs is a fundamental principle by which beta cells are specified and maintained. Given the lack of Nkx6.1 expression and aberrant activation of non-beta endocrine hormones in human embryonic stem cell (hESC)-derived insulin(+) cells, our study has significant implications for developing cell replacement therapies.

In healthy people, blood glucose levels are maintained within a narrow range by several physiological mechanisms. Key among them is the release of the hormone insulin by pancreatic β cells, which occurs when glucose levels rise after a meal. In response to insulin, blood glucose is taken up by tissues that need fuel, such as muscle. β cells can anticipate the body's varying demand for insulin throughout the 24-hour day because they have their own circadian clock. How this clock controls insulin release has been unclear. Perelis et al. now show that the activity of transcriptional enhancers specific to β cells regulates the rhythmic expression of genes involved in the assembly and trafficking of insulin secretory vesicles (see the Perspective by Dibner and Schibler). Science , this issue p. 10.1126/science.aac4250 ; see also p. 628 Circadian control of insulin release is mediated by transcriptional enhancers active specifically in pancreatic β cells. [Also see Perspective by Dibner and Schibler ] The mammalian transcription factors CLOCK and BMAL1 are essential components of the molecular clock that coordinate behavior and metabolism with the solar cycle. Genetic or environmental perturbation of circadian cycles contributes to metabolic disorders including type 2 diabetes. To study the impact of the cell-autonomous clock on pancreatic β cell function, we examined pancreatic islets from mice with either intact or disrupted BMAL1 expression both throughout life and limited to adulthood. We found pronounced oscillation of insulin secretion that was synchronized with the expression of genes encoding secretory machinery and signaling factors that regulate insulin release. CLOCK/BMAL1 colocalized with the pancreatic transcription factor PDX1 within active enhancers distinct from those controlling rhythmic metabolic gene networks in liver. We also found that β cell clock ablation in adult mice caused severe glucose intolerance. Thus, cell type–specific enhancers underlie the circadian control of peripheral metabolism throughout life and may help to explain its dysregulation in diabetes.