Stat1 and Stat3 are latent transcriptional factors activated initially through phosphorylation on single tyrosine residues induced by cytokine and growth factor occupation of cell surface receptors. Here we show that phosphorylation on a single serine (residue 727) in each protein is also required for maximal transcriptional activity. Both cytokines and growth factors are capable of inducing the serine phosphorylation of Stat1 and Stat3. These experiments show that gene activation by Stat1 and Stat3, which obligatorily require tyrosine phosphorylation to become active, also depends for maximal activation on one or more of the many serine kinases.

The STAT family of proteins carries out a dual function: signal transduction and activation of transcription. A new family member, Stat3, becomes activated through phosphorylation on tyrosine as a DNA binding protein in response to epidermal growth factor (EGF) and interleukin-6 (IL-6) but not interferon gamma (IFN-gamma). It is likely that this phosphoprotein forms homodimers as well as heterodimers with the first described member of the STAT family, Stat91 (renamed Stat1 alpha), which is activated by the IFNs and EGF. Differential activation of different STAT proteins in response to different ligands should help to explain specificity in nuclear signaling from the cell surface.

Microglia are the primary immune cells in the brain. Under physiological conditions, they typically stay in a "resting" state, with ramified processes continuously extending to and retracting from surrounding neural tissues. Whether and how such highly dynamic resting microglia functionally interact with surrounding neurons are still unclear. Using in vivo time-lapse imaging of both microglial morphology and neuronal activity in the optic tectum of larval zebrafish, we found that neuronal activity steers resting microglial processes and facilitates their contact with highly active neurons. This process requires the activation of pannexin-1 hemichannels on neurons. Reciprocally, such resting microglia-neuron contact reduces both spontaneous and visually evoked activities of contacted neurons. Our findings reveal an instructive role for neuronal activity in resting microglial motility and suggest the function for microglia in homeostatic regulation of neuronal activity in the healthy brain.

Statl and Stat3 are two members of the ligand-activated transcription factor family that serve the dual functions of signal transducers and activators of transcription. Whereas the two proteins select very similar (not identical) optimum binding sites from random oligonucleotides, differences in their binding affinity were readily apparent with natural STAT-binding sites. To take advantage of these different affinities, chimeric Statl:Stat3 molecules were used to locate the amino acids that could discriminate a general binding site from a specific binding site. The amino acids between residues -400 and -500 of these -750-amino-acid-long proteins determine the DNA-binding site specificity. Mutations within this region result in Stat proteins that are activated normally by tyrosine phosphorylation and that dimerize but have greatly reduced DNA-binding affinities.

In the course of molecular studies on the requirements for cell growth and mitogenesis, those polypeptide ligands and their receptors that activate cell proliferation were first identified and more intensively studied than proteins that restrict cell growth. The recognition of the existence of tumor suppressor genes and the recent recognition of small proteins that inhibit cell cycle progression have created great interest in the balance that must exist between proteins that promote or restrict the cell cycle (1). The interferons (IFNs) were among the earliest polypeptides found to exert growth inhibition in cultured cells (2)(3)(4). After cells are treated with IFN-a or IFN-,y, new transcriptional programs are instituted, and recently the molecular mechanism by which IFN-a and IFN--y effect immediate transcriptional changes has been uncovered (5-10). Latent transcriptional factors termed signal transducers and activators of transcription (STATs) are activated by phosphorylation on tyrosine, dimerization occurs, nuclear translocation follows, and site-specific DNA binding leads to gene activation.

Fsp27, a member of the Cide family proteins, was shown to localize to lipid droplet and promote lipid storage in adipocytes. We aimed to understand the biological role of Fsp27 in regulating adipose tissue differentiation, insulin sensitivity and energy balance. Fsp27 −/− mice and Fsp27/lep double deficient mice were generated and we examined the adiposity, whole body metabolism, BAT and WAT morphology, insulin sensitivity, mitochondrial activity, and gene expression changes in these mouse strains. Furthermore, we isolated mouse embryonic fibroblasts (MEFs) from wildtype and Fsp27 −/− mice, followed by their differentiation into adipocytes in vitro. We found that Fsp27 is expressed in both brown adipose tissue (BAT) and white adipose tissue (WAT) and its levels were significantly elevated in the WAT and liver of leptin-deficient ob/ob mice. Fsp27 −/− mice had increased energy expenditure, lower levels of plasma triglycerides and free fatty acids. Furthermore, Fsp27 −/− and Fsp27/lep double-deficient mice are resistant to diet-induced obesity and display increased insulin sensitivity. Moreover, white adipocytes in Fsp27 −/− mice have reduced triglycerides accumulation and smaller lipid droplets, while levels of mitochondrial proteins, mitochondrial size and activity are dramatically increased. We further demonstrated that BAT-specific genes and key metabolic controlling factors such as FoxC2, PPAR and PGC1α were all markedly upregulated. In contrast, factors inhibiting BAT differentiation such as Rb, p107 and RIP140 were down-regulated in the WAT of Fsp27 −/− mice. Remarkably, Fsp27 −/− MEFs differentiated in vitro show many brown adipocyte characteristics in the presence of the thyroid hormone triiodothyronine (T3). Our data thus suggest that Fsp27 acts as a novel regulator in vivo to control WAT identity, mitochondrial activity and insulin sensitivity.

Background: Macrophages and neutrophils are key phagocytes in regeneration. Results: Neutrophils are the primary phagocytes in the inflammatory stage and are dispensable for zebrafish fin regeneration, whereas macrophages mainly function in the resolution stage and are required for fin regeneration. Conclusion: Macrophages and neutrophils behave differently during zebrafish fin injury and regeneration. Significance: Our study documents that macrophages and neutrophils play distinct functions in tissue regeneration.

Transcription factor p53 forms a network with associated factors to regulate the cell cycle and apoptosis in response to environmental stresses. However, there is currently no direct genetic evidence to show if or how the p53 pathway functions during organogenesis. Here we present evidence to show that the zebrafish def (digestive-organ expansion factor) gene encodes a novel pan-endoderm-specific factor. A loss-of-function mutation in def confers hypoplastic digestive organs and selectively up-regulates the expression of ⌬113p53, counterpart to a newly identified isoform of p53 produced by an alternative internal promoter in intron 4 of the p53 gene in human. The increased ⌬113p53 expression is limited to within the mutant digestive organs, and this increase selectively induces the expression of p53-responsive genes to trigger the arrest of the cell cycle but not apoptosis, resulting in compromised organ growth in the mutant. Our data demonstrate that, while induction of expression of p53 and/or its isoforms is crucial to suppress abnormal cell growth, ⌬113p53 is tightly regulated by an organ/tissue-specific factor Def, especially during organogenesis, to prevent adverse inhibition of organ/tissue growth.[Keywords: Def (digestive-organ expansion factor); endoderm organogenesis; p53; zebrafish] Supplemental material is available at http://www.genesdev.org.