Human TLR8 senses UR/URR motifs in bacterial and mitochondrial RNA

Toll‐like receptor (TLR) 13 and TLR2 are the major sensors of Gram‐positive bacteria in mice. TLR13 recognizes Sa19, a specific 23S ribosomal (r) RNA‐derived fragment and bacterial modification of Sa19 ablates binding to TLR13, and to antibiotics such as erythromycin. Similarly, RNase A‐treated Staphylococcus aureus activate human peripheral blood mononuclear cells (PBMCs) only via TLR2, implying single‐stranded (ss) RNA as major stimulant. Here, we identify human TLR8 as functional TLR13 equivalent that promiscuously senses ssRNA. Accordingly, Sa19 and mitochondrial (mt) 16S rRNA sequence‐derived oligoribonucleotides (ORNs) stimulate PBMCs in a MyD88‐dependent manner. These ORNs, as well as S. aureus‐, Escherichia coli‐, and mt‐RNA, also activate differentiated human monocytoid THP‐1 cells, provided they express TLR8. Moreover, Unc93b1 ^−/−‐ and Tlr8 ^−/−‐THP‐1 cells are refractory, while endogenous and ectopically expressed TLR8 confers responsiveness in a UR/URR RNA ligand consensus motif‐dependent manner. If TLR8 function is inhibited by suppression of lysosomal function, antibiotic treatment efficiently blocks bacteria‐driven inflammatory responses in infected human whole blood cultures. Sepsis therapy might thus benefit from interfering with TLR8 function.     

Progranulin deficiency leads to severe inflammation,lung injury and cell death in a mouse model of endotoxic shock

Progranulin (PGRN) is a crucial secreted growth factor involved in various kinds of physiologic and disease processes and often has a protective role in inflammatory diseases. This study was designed to investigate the protective effects of PGRN on endotoxic shock in a mouse model of PGRN deficiency. After lipopolysaccharide (LPS) injection to induce endotoxic shock in mice, PGRN levels were induced in wild‐type (WT) mice at 6 and 24 hrs. Survival rate analysis, haematoxylin and eosin staining, immunohistochemical staining, enzyme‐linked immunosorbent assay and in situ terminal deoxynucleotidyl transferase–mediated uridine triphosphate nick‐end labelling assay were used to reveal the susceptibility, lung injury, inflammatory cell infiltration, production of inflammatory mediators and lung cell death in mice after LPS injection. PGRN‐deficient (Grn ^−/−) mice were highly susceptible to LPS‐induced endotoxic shock, with decreased survival, severe lung injury, increased production of pro‐inflammatory mediators, and inflammatory cell infiltration and apoptotic death in the lung. Additionally, recombinant PGRN (rPGRN) administration before LPS stimulation ameliorated the survival of and abnormalities in both WT and Grn ^−/− mice. Altogether, these findings indicate that PGRN may be a novel biologic agent with therapeutic potential for endotoxic shock probably by inhibiting LPS‐induced systemic and local inflammation in mice for treating endotoxic shock.     

Molecular architecture of the ribosome‐bound Hepatitis C Virus internal ribosomal entry site RNA

Internal ribosomal entry sites (IRESs) are structured cis‐acting RNAs that drive an alternative, cap‐independent translation initiation pathway. They are used by many viruses to hijack the translational machinery of the host cell. IRESs facilitate translation initiation by recruiting and actively manipulating the eukaryotic ribosome using only a subset of canonical initiation factor and IRES transacting factors. Here we present cryo‐EM reconstructions of the ribosome 80S‐ and 40S‐bound Hepatitis C Virus (HCV) IRES. The presence of four subpopulations for the 80S•HCV IRES complex reveals dynamic conformational modes of the complex. At a global resolution of 3.9 Å for the most stable complex, a derived atomic model reveals a complex fold of the IRES RNA and molecular details of its interaction with the ribosome. The comparison of obtained structures explains how a modular architecture facilitates mRNA loading and tRNA binding to the P‐site. This information provides the structural foundation for understanding the mechanism of HCV IRES RNA‐driven translation initiation.     

Role of Kir2.1 in human monocyte‐derived foam cell maturation

The role of K⁺ channels in macrophage immunomodulation has been well‐established. However, it remains unclear whether K⁺ channels are involved in the lipid uptake of macrophages. The expression and function of the inward rectifier potassium channel (Kir2.1, KCNJ2) in Human acute monocytic leukemia cell line (THP‐1) cells and human monocytes derived macrophages (HMDMs) were investigated using RT‐PCR and western blotting, and patch clamp technique. The expression of scavenger receptors in THP‐1–derived macrophages was detected using western blotting. Expressions of Kir2.1 mRNA and protein in HMDMs were significantly decreased by 60% (P < 0.05) and 90% (P < 0.001) on macrophage maturation, but overexpressed by approximately 1.3 (P > 0.05) and 3.8 times (P = 0.001) after foam cell formation respectively. Concurrently, the Kir2.1 peak current density in HMDMs, mature macrophages and foam cells, measured at −150 mV, were −22.61 ± 2.1 pA/pF, −7.88 ± 0.60 pA/pF and −13.39 ± 0.80 pA/pF respectively (P < 0.05). In association with an up‐regulation of Kir2.1 in foam cells, the SR‐A protein level was significantly increased by over 1.5 times compared with macrophages (P < 0.05). THP‐1 cells contained much less lipids upon Kir2.1 knockdown and cholesterol ester/total cholesterol ratio was 29.46 ± 2.01% (P < 0.05), and the SR‐BI protein level was increased by over 6.2 times, compared to that of macrophages (P < 0.001). Kir2.1 may participate in macrophage maturation and differentiation, and play a key role in lipid uptake and foam cell formation through modulating the expression of scavenger receptors.     

Correlation between endogenous polyamines in human cardiac tissues and clinical parameters in patients with heart failure

Polyamines contribute to several physiological and pathological processes, including cardiac hypertrophy in experimental animals. This involves an increase in ornithine decarboxylase (ODC) activity and intracellular polyamines associated with cyclic adenosine monophosphate (cAMP) increases. The aim of the study was to establish the role of these in the human heart in living patients. For this, polyamines (by high performance liquid chromatography) and the activity of ODC and N¹‐acetylpolyamine oxidases (APAO) were determined in the right atrial appendage of 17 patients undergoing extracorporeal circulation to correlate with clinical parameters. There existed enzymatic activity associated with the homeostasis of polyamines. Left atria size was positively associated with ODC (r = 0.661, P = 0.027) and negatively with APAO‐N¹‐acetylspermine (r = −0.769, P = 0.026), suggesting that increased levels of polyamines are associated with left atrial hemodynamic overload. Left ventricular ejection fraction (LVEF) and heart rate were positively associated with spermidine (r = 0.690, P = 0.003; r = 0.590, P = 0.021) and negatively with N¹‐acetylspermidine (r = −0.554, P = 0.032; r = −0.644, P = 0.018). LVEF was negatively correlated with cAMP levels (r = −0.835, P = 0.001) and with cAMP/ODC (r = −0.794, P = 0.011), cAMP/spermidine (r = −0.813, P = 0.001) and cAMP/spermine (r = −0.747, P = 0.003) ratios. Abnormal LVEF patients showed decreased ODC activity and spermidine, and increased N¹‐acetylspermidine, and cAMP. Spermine decreased in congestive heart failure patients. The trace amine isoamylamine negatively correlated with septal wall thickness (r = −0.634, P = 0.008) and was increased in cardiac heart failure. The results indicated that modifications in polyamine homeostasis might be associated with cardiac function and remodelling. Increased cAMP might have a deleterious effect on function. Further studies should confirm these findings and the involvement of polyamines in different stages of heart failure.     

miR‐9‐5p suppresses pro‐fibrogenic transformation of fibroblasts and prevents organ fibrosis by targeting NOX4 and TGFBR2

Uncontrolled extracellular matrix (ECM) production by fibroblasts in response to injury contributes to fibrotic diseases, including idiopathic pulmonary fibrosis (IPF). Reactive oxygen species (ROS) generation is involved in the pathogenesis of IPF. Transforming growth factor‐β1 (TGF‐β1) stimulates the production of NADPH oxidase 4 (NOX4)‐dependent ROS, promoting lung fibrosis (LF). Dysregulation of microRNAs (miRNAs) has been shown to contribute to LF. To identify miRNAs involved in redox regulation relevant for IPF, we performed arrays in human lung fibroblasts exposed to ROS. miR‐9‐5p was selected as the best candidate and we demonstrate its inhibitory effect on TGF‐β receptor type II (TGFBR2) and NOX4 expression. Increased expression of miR‐9‐5p abrogates TGF‐β1‐dependent myofibroblast phenotypic transformation. In the mouse model of bleomycin‐induced LF, miR‐9‐5p dramatically reduces fibrogenesis and inhibition of miR‐9‐5p and prevents its anti‐fibrotic effect both in vitro and in vivo. In lung specimens from patients with IPF, high levels of miR‐9‐5p are found. In omentum‐derived mesothelial cells (MCs) from patients subjected to peritoneal dialysis (PD), miR‐9‐5p also inhibits mesothelial to myofibroblast transformation. We propose that TGF‐β1 induces miR‐9‐5p expression as a self‐limiting homeostatic response.     

Human haematopoietic stem/progenitor cells express several functional sex hormone receptors

Evidence has accumulated that murine haematopoietic stem/progenitor cells (HSPCs) share several markers with the germline, a connection supported by recent reports that pituitary and gonadal sex hormones (SexHs) regulate development of murine HSPCs. It has also been reported that human HSPCs, like their murine counterparts, respond to certain SexHs (e.g. androgens). However, to better address the effects of SexHs, particularly pituitary SexHs, on human haematopoiesis, we tested for expression of receptors for pituitary SexHs, including follicle‐stimulating hormone (FSH), luteinizing hormone (LH), and prolactin (PRL), as well as the receptors for gonadal SexHs, including progesterone, oestrogens, and androgen, on HSPCs purified from human umbilical cord blood (UCB) and peripheral blood (PB). We then tested the functionality of these receptors in ex vivo signal transduction studies and in vitro clonogenic assays. In parallel, we tested the effect of SexHs on human mesenchymal stromal cells (MSCs). Finally, based on our observation that at least some of the UCB‐derived, CD45⁻ very small embryonic‐like stem cells (VSELs) become specified into CD45⁺ HSPCs, we also evaluated the expression of pituitary and gonadal SexH receptors on these cells. We report for the first time that human HSPCs and VSELs, like their murine counterparts, express pituitary and gonadal SexH receptors at the mRNA and protein levels. Most importantly, SexH if added to suboptimal doses of haematopoietic cytokines and growth factors enhance clonogenic growth of human HSPCs as well as directly stimulate proliferation of MSCs.     

Tree species richness decreases while species evenness increases with disturbance frequency in a natural boreal forest landscape

Understanding species diversity and disturbance relationships is important for biodiversity conservation in disturbance‐driven boreal forests. Species richness and evenness may respond differently with stand development following fire. Furthermore, few studies have simultaneously accounted for the influences of climate and local site conditions on species diversity. Using forest inventory data, we examined the relationships between species richness, Shannon''s index, evenness, and time since last stand‐replacing fire (TSF) in a large landscape of disturbance‐driven boreal forest. TSF has negative effect on species richness and Shannon''s index, and a positive effect on species evenness. Path analysis revealed that the environmental variables affect richness and Shannon''s index only through their effects on TSF while affecting evenness directly as well as through their effects on TSF. Synthesis and applications. Our results demonstrate that species richness and Shannon''s index decrease while species evenness increases with TSF in a boreal forest landscape. Furthermore, we show that disturbance frequency, local site conditions, and climate simultaneously influence tree species diversity through complex direct and indirect effects in the studied boreal forest.     

Macroscale intraspecific variation and environmental heterogeneity: analysis of cold and warm zone abundance,mortality, and regeneration distributions of four eastern US tree species

I test for macroscale intraspecific variation of abundance, mortality, and regeneration of four eastern US tree species (Tsuga canadensis,Betula lenta,Liriodendron tulipifera, and Quercus prinus) by splitting them into three climatic zones based on plant hardiness zones (PHZs). The primary goals of the analysis are to assess the differences in environmental heterogeneity and demographic responses among climatic zones, map regional species groups based on decision tree rules, and evaluate univariate and multivariate patterns of species demography with respect to environmental variables. I use the Forest Inventory Analysis (FIA) data to derive abundance, mortality, and regeneration indices and split the range into three climatic zones based on USDA PHZs: (1) cold adapted, leading region; (2) middle, well‐adapted region; and (3) warm adapted, trailing region. I employ decision tree ensemble methods to assess the importance of environmental predictors on the abundance of the species between the cold and warm zones and map zonal variations in species groups. Multivariate regression trees are used to simultaneously explore abundance, mortality, and regeneration in tandem to assess species vulnerability. Analyses point to the relative importance of climate in the warm adapted, trailing zone (especially moisture) compared to the cold adapted, leading zone. Higher mortality and lower regeneration patterns in the warm trailing zone point to its vulnerability to growing season temperature and precipitation changes that could figure more prominently in the future. This study highlights the need to account for intraspecific variation of demography in order to understand environmental heterogeneity and differential adaptation. It provides a methodology for assessing the vulnerability of tree species by delineating climatic zones based on easily available PHZ data, and FIA derived abundance, mortality, and regeneration indices as a proxy for overall growth and fitness. Based on decision tree rules, ecologically meaningful variations in species abundance among the climatic zones can be related to environmental variability and mapped.     

The C9orf72 protein interacts with Rab1a and the ULK1 complex to regulate initiation of autophagy

A GGGGCC hexanucleotide repeat expansion in the C9orf72 gene is the most common genetic cause of amyotrophic lateral sclerosis and frontotemporal dementia (C9ALS/FTD). C9orf72 encodes two C9orf72 protein isoforms of unclear function. Reduced levels of C9orf72 expression have been reported in C9ALS/FTD patients, and although C9orf72 haploinsufficiency has been proposed to contribute to C9ALS/FTD, its significance is not yet clear. Here, we report that C9orf72 interacts with Rab1a and the Unc‐51‐like kinase 1 (ULK1) autophagy initiation complex. As a Rab1a effector, C9orf72 controls initiation of autophagy by regulating the Rab1a‐dependent trafficking of the ULK1 autophagy initiation complex to the phagophore. Accordingly, reduction of C9orf72 expression in cell lines and primary neurons attenuated autophagy and caused accumulation of p62‐positive puncta reminiscent of the p62 pathology observed in C9ALS/FTD patients. Finally, basal levels of autophagy were markedly reduced in C9ALS/FTD patient‐derived iNeurons. Thus, our data identify C9orf72 as a novel Rab1a effector in the regulation of autophagy and indicate that C9orf72 haploinsufficiency and associated reductions in autophagy might be the underlying cause of C9ALS/FTD‐associated p62 pathology.     

MicroRNA‐455 regulates brown adipogenesis via a novel HIF1an‐AMPK‐PGC1α signaling network

Brown adipose tissue (BAT) dissipates chemical energy as heat and can counteract obesity. MicroRNAs are emerging as key regulators in development and disease. Combining microRNA and mRNA microarray profiling followed by bioinformatic analyses, we identified miR‐455 as a new regulator of brown adipogenesis. miR‐455 exhibits a BAT‐specific expression pattern and is induced by cold and the browning inducer BMP7. In vitro gain‐ and loss‐of‐function studies show that miR‐455 regulates brown adipocyte differentiation and thermogenesis. Adipose‐specific miR‐455 transgenic mice display marked browning of subcutaneous white fat upon cold exposure. miR‐455 activates AMPKα1 by targeting HIF1an, and AMPK promotes the brown adipogenic program and mitochondrial biogenesis. Concomitantly, miR‐455 also targets the adipogenic suppressors Runx1t1 and Necdin, initiating adipogenic differentiation. Taken together, the data reveal a novel microRNA‐regulated signaling network that controls brown adipogenesis and may be a potential therapeutic target for human metabolic disorders.     

Thermal tolerance and climate warming sensitivity in tropical snails

Tropical ectotherms are predicted to be especially vulnerable to climate change because their thermal tolerance limits generally lie close to current maximum air temperatures. This prediction derives primarily from studies on insects and lizards and remains untested for other taxa with contrasting ecologies. We studied the HCT (heat coma temperatures) and ULT (upper lethal temperatures) of 40 species of tropical eulittoral snails (Littorinidae and Neritidae) inhabiting exposed rocky shores and shaded mangrove forests in Oceania, Africa, Asia and North America. We also estimated extremes in animal body temperature at each site using a simple heat budget model and historical (20 years) air temperature and solar radiation data. Phylogenetic analyses suggest that HCT and ULT exhibit limited adaptive variation across habitats (mangroves vs. rocky shores) or geographic locations despite their contrasting thermal regimes. Instead, the elevated heat tolerance of these species (HCT = 44.5 ± 1.8°C and ULT = 52.1 ± 2.2°C) seems to reflect the extreme temperature variability of intertidal systems. Sensitivity to climate warming, which was quantified as the difference between HCT or ULT and maximum body temperature, differed greatly between snails from sunny (rocky shore; Thermal Safety Margin, TSM = −14.8 ± 3.3°C and −6.2 ± 4.4°C for HCT and ULT, respectively) and shaded (mangrove) habitats (TSM = 5.1 ± 3.6°C and 12.5 ± 3.6°C). Negative TSMs in rocky shore animals suggest that mortality is likely ameliorated during extreme climatic events by behavioral thermoregulation. Given the low variability in heat tolerance across species, habitat and geographic location account for most of the variation in TSM and may adequately predict the vulnerability to climate change. These findings caution against generalizations on the impact of global warming across ectothermic taxa and highlight how the consideration of nonmodel animals, ecological transitions, and behavioral responses may alter predictions of studies that ignore these biological details.     

Integrative genomics positions MKRN1 as a novel ribonucleoprotein within the embryonic stem cell gene regulatory network

In embryonic stem cells (ESCs), gene regulatory networks (GRNs) coordinate gene expression to maintain ESC identity; however, the complete repertoire of factors regulating the ESC state is not fully understood. Our previous temporal microarray analysis of ESC commitment identified the E3 ubiquitin ligase protein Makorin‐1 (MKRN1) as a potential novel component of the ESC GRN. Here, using multilayered systems‐level analyses, we compiled a MKRN1‐centered interactome in undifferentiated ESCs at the proteomic and ribonomic level. Proteomic analyses in undifferentiated ESCs revealed that MKRN1 associates with RNA‐binding proteins, and ensuing RIP‐chip analysis determined that MKRN1 associates with mRNAs encoding functionally related proteins including proteins that function during cellular stress. Subsequent biological validation identified MKRN1 as a novel stress granule‐resident protein, although MKRN1 is not required for stress granule formation, or survival of unstressed ESCs. Thus, our unbiased systems‐level analyses support a role for the E3 ligase MKRN1 as a ribonucleoprotein within the ESC GRN.     

Molecular and iridescent feather reflectance data reveal recent genetic diversification and phenotypic differentiation in a cloud forest hummingbird

The present day distribution and spatial genetic diversity of Mesoamerican biota reflects a long history of responses to habitat change. The hummingbird Lampornis amethystinus is distributed in northern Mesoamerica, with geographically disjunct populations. Based on sampling across the species range using mitochondrial DNA (mtDNA) sequences and nuclear microsatellites jointly analysed with phenotypic and climatic data, we (1) test whether the fragmented distribution is correlated with main evolutionary lineages, (2) assess body size and plumage color differentiation of populations in geographic isolation, and (3) evaluate a set of divergence scenarios and demographic patterns of the hummingbird populations. Analysis of genetic variation revealed four main groups: blue‐throated populations (Sierra Madre del Sur); two groups of amethyst‐throated populations (Trans‐Mexican Volcanic Belt and Sierra Madre Oriental); and populations east of the Isthmus of Tehuantepec (IT) with males showing an amethyst throat. The most basal split is estimated to have originated in the Pleistocene, 2.39–0.57 million years ago (MYA), and corresponded to groups of populations separated by the IT. However, the estimated recent divergence time between blue‐ and amethyst‐throated populations does not correspond to the 2‐MY needed to be in isolation for substantial plumage divergence, likely because structurally iridescent colors are more malleable than others. Results of species distribution modeling and Approximate Bayesian Computation analysis fit a model of lineage divergence west of the Isthmus after the Last Glacial Maximum (LGM), and that the species’ suitable habitat was disjunct during past and current conditions. These results challenge the generality of the contraction/expansion glacial model to cloud forest‐interior species and urges management of cloud forest, a highly vulnerable ecosystem to climate change and currently facing destruction, to prevent further loss of genetic diversity or extinction.