Here we design and optimize a genetically encoded fluorescent indicator, iAChSnFR, for the ubiquitous neurotransmitter acetylcholine, based on a bacterial periplasmic binding protein. iAChSnFR shows large fluorescence changes, rapid rise and decay kinetics, and insensitivity to most cholinergic drugs. iAChSnFR revealed large transients in a variety of slice and in vivo preparations in mouse, fish, fly and worm. iAChSnFR will be useful for the study of acetylcholine in all animals. IntroductionAcetylcholine (ACh) is a critical neurotransmitter in all animals. Among invertebrates, it is the most prevalent excitatory transmitter in the brain, sensory ganglia, and frequently the neuromuscular junction (NMJ). Among vertebrates, only a minority of neurons release ACh, but these signals play varying key roles. For instance, ACh signals at the NMJ, in the autonomic nervous system, and in subsets of the central nervous system, particularly projections arising from the brainstem and basal forebrain. Other cholinergic neuron populations in the brain include striatal interneurons, the stria vascularis-medial habenula-interpeduncular nucleus pathway, and sparse, incompletely characterized cell types such as intrinsic cholinergic interneurons in cortex 1 and hippocampus 2 . ACh helps to regulate attention 3 and wakefulness 4 , and participates in memory formation and consolidation 5 . ACh is also an important transmitter in glia, and between the nervous and immune systems 6 .Acetylcholine is synthesized pre-synaptically from choline and acetyl-CoA by choline acetyltransferase (ChAT), then packaged into synaptic vesicles by the vesicular acetylcholine transporter (VAChT). A key, partially understood aspect of cholinergic signaling is co-release with other neurotransmitters, including GABA, ATP, and glutamate 7,8 . To understand the role of co-release, one must measure ACh release alongside emerging measurements of other neurotransmitters.Acetylcholine receptors are among the most diverse neurotransmitter receptor families. Humans possess five muscarinic G protein-coupled receptors (GPCRs) for ACh (mAChRs) with diverse expression in the brain and smooth, cardiac, and skeletal muscle. Vertebrate nicotinic ACh receptors (nAChRs) are pentameric ligand-gated cation channels. Humans have a total of 17 nAChR subunit genes, in five classes: 10 a, 4 b, and one each of g, d, and e. nAChRs occur with many subunit combinations 9 , and others may be undiscovered. Invertebrates also have AChgated chloride channels. On neurons, receptors can be localized pre-, post-, and extrasynaptically, often with different isoforms in each place 10

Aim: Understanding the variation in biodiversity and its underlying drivers and mechanisms is a core task in biogeography and ecology. We examined (a) the relative contributions of species replacement (i.e., turnover) and richness difference (i.e., nestedness) to taxonomically and trait-based β-diversity of stream benthic algae; (b) whether these two facets of β-diversity are correlated with each other; and (c) the relative contributions of local environmental, geo-climatic and spatial factors to the two facets of β-diversity and their components. Location: Hun-Tai River Basin, northeastern China. Taxon: Stream benthic algae. Methods: A total of 157 sites were sampled. Mantel tests were used to examine the complementarities between the two facets of β-diversity and their components. Distance-based redundancy analysis and variation partitioning were utilized to investigate the relative contributions of local environmental, geo-climatic and spatial factors to each facet of β-diversity and their components. Results: Weak correlations between taxonomically and trait-based β-diversity and their components were detected, which indicated complementarity of ecological information. Taxonomically based total β-diversity was largely driven by turnover, whereas trait-based total β-diversity was more driven by nestedness. Variation partitioning results indicated that local environmental and spatial factors contributed more than geo-climatic variables to the total explained variation in taxonomically and traitbased β-diversity. Main conclusions: Different facets of β-diversity and their decomposition are important for understanding diversity patterns of benthic algae relative to abiotic factors. A high level of trait-based convergence among benthic algae communities, despite high taxonomic divergence, demonstrated turnover of species with similar biological traits across our study region. Our study provides a trait-based insight into stream benthic algae communities, which was less documented by previous freshwater studies that focused on regions undergoing recovery following human disturbances.| 1843 WU et al.

Fas-apoptosis inhibitory molecule (FAIM) is inducibly expressed in B lymphocytes and had been shown to antagonize Fas-mediated killing of B-cell lines in vitro. However, its mechanism and role in vivo are unknown. We have generated faim À/À mice and found these mutants to be viable. In contrast to fas À/À mice, faim À/À mice have normal B-and T-cell populations. However, faim À/À B cells and thymocytes show increased sensitivity to Fas-triggered apoptosis in vitro, and faim À/À mice suffer greater mortality and exhibit exacerbated liver damage in response to Fas (CD95) engagement in vivo. The lack of FAIM results in greater activation of caspase-8 and -3 in Fas-stimulated thymocytes. CD95 or Fas (APO-1/TNFRSF6) is a member of the tumor necrosis factor (TNF) receptor superfamily and is capable of inducing apoptosis in a variety of cell-types. 1,2 The physiological ligand for Fas is CD178 (FasL), which is expressed mainly by activated T cells. 3 In mice, mutation in fas (lpr mice) or fasl (gld mice) leads to lymphoproliferation, the accumulation of abnormal lymphocytes, autoantibody production, and this ultimately results in systemic autoimmunity. 3,4 Thus, Fasmediated apoptosis is critical for regulating the function and homeostasis of lymphocytes.Fas-signaling is triggered upon the binding of FasL, which leads to the trimerization of Fas and the formation of the downstream death-inducing signaling complex (DISC) that comprises the cytosolic adapter protein FADD and procaspase-8. 5 Procaspase-8 undergoes auto-cleavage to generate active caspase-8, which in turn leads to the activation of caspase-3 and execution of apoptosis. As apoptosis plays an important role in physiology, its activation process must be tightly regulated. Hence, other than activators, negative regulators of Fas-signaling also exist. One of these inhibitors of Fas-mediated apoptosis is the cellular FLICE-inhibitory protein (c-FLIP) which is known to antagonize Fas-signaling by interfering with the recruitment of procaspase-8 to the DISC. 6 Fas-apoptosis inhibitory molecule (FAIM) was also cloned as an inducibly expressed, anti-apoptotic protein that antagonized Fas-triggered cell death of B-cell lines in vitro. 7 However, its mode of action is unknown. FAIM is highly conserved in evolution and widely expressed in all tissues although it bears no homology to other known proteins. 7 Although most tissues express the short isoform of the protein (FAIM-S), neuronal cells express both FAIM-S and a long isoform, FAIM-L. 8 FAIM-S was shown to promote neurite outgrowth through a mechanism involving NF-kB and Ras-ERK activation, 8 whereas FAIM-L was demonstrated to protect neurons from cell death in response to Fas and TNFR1 engagement. 9 Apart from its role in protecting B-lymphocytes and neuronal cells from death receptor-induced apoptosis, 7,9 FAIM was also shown to improve the survival of HEK-293 cells in serum-free media 10 and CHO cells in fed-batch cultures. 11 This suggests a potential application of FAIM in biotechnology through its role in e...

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The aim of the study was to assess the association between total flavonoids/flavonoid subclasses intake and prostate cancer risk. Several databases were searched to select eligible studies with predefined criteria. Risk ratios (RRs) with 95% confidence intervals (CIs) were used as the effect size. Publication bias and sensitivity analysis were performed. A total of five studies including four prospective cohort studies and one case-control study were included in the meta-analysis. The pooled result demonstrated a significantly increased risk of prostate cancer with higher intake of total flavonoids (RR = 1.12, 95% CI: 1.02-1.23, P = 0.013). However, sensitivity analysis indicated that there lacked a significant association after removing the study of Wang et al. (RR = 1.17, 95% CI: 0.94-1.46). Subgroup analysis stratified by flavonoids subclasses found that higher intake of anthocyanidins and flavan-3-ols were significantly associated with increased prostate cancer risk (RR = 1.12, 95% CI: 1.03-1.21, P = 0.011; RR = 1.21, 95% CI: 1.10-1.32, P < 0.001). Sensitivity analysis also indicated that after removing Wang's study, no significant association between anthocyanidins intake and prostate cancer risk was detected (RR = 1.22, 95% CI: 0.97-1.54). In conclusion, higher intake of flavonoids may not be associated with prostate cancer risk.

Although trees are key to ecosystem functioning, many forests and tree species across the globe face strong threats. Preserving areas of high biodiversity is a core priority for conservation; however, different dimensions of biodiversity and varied conservation targets make it difficult to respond effectively to this challenge. Here, we (i) identify priority areas for global tree conservation using comprehensive coverage of tree diversity based on taxonomy, phylogeny, and functional traits; and (ii) compare these findings to existing protected areas and global biodiversity conservation frameworks. We find that ca. 51% of the top-priority areas for tree biodiversity are located in current protected areas. The remaining half top-priority areas are subject to moderate to high human pressures, indicating conservation actions are needed to mitigate these human impacts. Our findings emphasize the effectiveness of using tree conservation priority areas for future global conservation planning.

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