PAS Domain Research Articles

The Aer2 receptor from Vibrio cholerae is a dual PAS-heme oxygen sensor.

The diarrheal pathogen Vibrio cholerae navigates complex environments using three chemosensory systems and 44-45 chemoreceptors. Chemosensory cluster II modulates chemotaxis, whereas clusters I and III have unknown functions. Ligands have been identified for only five V. cholerae chemoreceptors. Here, we report that the cluster III receptor, VcAer2, binds and responds to O2 . VcAer2 is an ortholog of Pseudomonas aeruginosa Aer2 (PaAer2) but differs in that VcAer2 has two, rather than one, N-terminal PAS domain. We have determined that both PAS1 and PAS2 form homodimers and bind penta-coordinate b-type heme via an Eη-His residue. Heme binding to PAS1 required the entire PAS core, but receptor function also required the N-terminal cap. PAS2 functioned as an O2 -sensor [ , 19μM], utilizing the same Iβ Trp (W276) as PaAer2 to stabilize O2 . The crystal structure of PAS2-W276L was similar to that of PaAer2-PAS but resided in an active conformation mimicking the ligand-bound state, consistent with its signal-on phenotype. PAS1 also bound O2 [ , 12μM], although O2 binding was stabilized by either a Trp residue or Tyr residue. Moreover, PAS1 appeared to function as a signal modulator, regulating O2 -mediated signaling from PAS2 and resulting in activation of the cluster III chemosensory pathway.

Physical activity behaviours of persons living with HIV of low socioeconomic status: domain, intensity and sociodemographic correlates

ABSTRACTPhysical activity (PA) is healthy for People Living with HIV and AIDS (PLWHA). This study examined the PA profile of 978 PLWHA based on PA domains and PA intensity. The study also sought to determine whether employment status and level of education can predict PA among PLWHA of low SES. PA and sociodemographic data of 978 PLWHA of mean age 35 (8.77) years were collected using the global physical activity questionnaire and a sociodemographic questionnaire. Results showed that, participants engaged more in work-related PA [160.11 (346.95) min/wk], followed by transport related PA [115.21(SD = 142.04) min/wk], and lastly in leisure related PA [40.84 (SD = 110.37) min/wk]. Participants also engaged more in moderate PA [265.86 (SD = 335.45) min/wk], than in vigorous PA [50.29 (SD = 205.30) min/wk]. Employment was a significant predictor of overall PA controlling for age, CD4 count and education level. The model explained 2.5% of the variance (R2 = 0.025) on overall PA and tested significant at a 0.01 alpha level (p < 0.01). PA interventions for this population should be domain- and intensity-specific. Researchers promoting healthy lifestyle behaviour change can benefit from determining the factors that facilitate domain-specific PA.

Natural variation in genes potentially involved in plant architecture and adaptation in switchgrass (Panicum virgatum L.)

BackgroundAdvances in genomic technologies have expanded our ability to accurately and exhaustively detect natural genomic variants that can be applied in crop improvement and to increase our knowledge of plant evolution and adaptation. Switchgrass (Panicum virgatum L.), an allotetraploid (2n = 4× = 36) perennial C4 grass (Poaceae family) native to North America and a feedstock crop for cellulosic biofuel production, has a large potential for genetic improvement due to its high genotypic and phenotypic variation. In this study, we analyzed single nucleotide polymorphism (SNP) variation in 372 switchgrass genotypes belonging to 36 accessions for 12 genes putatively involved in biomass production to investigate signatures of selection that could have led to ecotype differentiation and to population adaptation to geographic zones.ResultsA total of 11,682 SNPs were mined from ~ 15 Gb of sequence data, out of which 251 SNPs were retained after filtering. Population structure analysis largely grouped upland accessions into one subpopulation and lowland accessions into two additional subpopulations. The most frequent SNPs were in homozygous state within accessions. Sixty percent of the exonic SNPs were non-synonymous and, of these, 45% led to non-conservative amino acid changes. The non-conservative SNPs were largely in linkage disequilibrium with one haplotype being predominantly present in upland accessions while the other haplotype was commonly present in lowland accessions. Tajima’s test of neutrality indicated that PHYB, a gene involved in photoperiod response, was under positive selection in the switchgrass population. PHYB carried a SNP leading to a non-conservative amino acid change in the PAS domain, a region that acts as a sensor for light and oxygen in signal transduction.ConclusionsSeveral non-conservative SNPs in genes potentially involved in plant architecture and adaptation have been identified and led to population structure and genetic differentiation of ecotypes in switchgrass. We suggest here that PHYB is a key gene involved in switchgrass natural selection. Further analyses are needed to determine whether any of the non-conservative SNPs identified play a role in the differential adaptation of upland and lowland switchgrass.

Structural insights and characterization of human Npas4 protein.

Npas4 is an activity dependent transcription factor which is responsible for gearing the expression of target genes involved in neuro-transmission. Despite the importance of Npas4 in many neuronal diseases, the tertiary structure of Npas4 protein along with its physico-chemical properties is limited. In the current study, first we perfomed the phylogenetic analysis of Npas4 and determined the content of hydrophobic, flexible and order-disorder promoting amino acids. The protein binding regions, post-translational modifications and crystallization propensity of Npas4 were predicted through different in-silico methods. The three dimensional model of Npas4 was predicted through LOMET, SPARSKS-X, I-Tasser, RaptorX, MUSTER and Pyhre and the best model was selected on the basis of Ramachandran plot, PROSA, and Qmean scores. The best model was then subjected to further refinement though MODREFINER. Finally the interacting partners of Npas4 were identified through STRING database. The phylogenetic analysis showed the human Npas4 gene to be closely related to other primates such as chimpanzees, monkey, gibbon. The physiochemical properties of Npas4 showed that it is an intrinsically disordered protein with N-terminal ordered region. The post-translational modification analyses indicated absence of acetylation and mannosylation sites. Three potential phosphorylation sites (S108, T130 and T136) were found in PAS A domain whilst a single phosphorylation site (S273) was present in PAS B domain. The predicted tertiary structure of Npas4 showed that bHLH domain and PAS domain possess tertiary structures while the rest of the protein exhibited disorder property. Protein-protein interaction analysis revealed NPas4 interaction with various proteins which are mainly involved in nuclear trafficking of proteins to cytoplasm, activity regulated gene transcription and neurodevelopmental disorders. Moreover the analysis also highlighted the direct relation to proteins involved in promoting neuronal survival, plasticity and cAMP responsive element binding protein proteins. The current study helps in understanding the physicochemical properties and reveals the neuro-modulatory role of Npas4 in crucial pathways involved in neuronal survival and neural signalling hemostasis.

The Protease ClpXP and the PAS Domain Protein DivL Regulate CtrA and Gene Transfer Agent Production in Rhodobacter capsulatus.

Several members of the Rhodobacterales (Alphaproteobacteria) produce a conserved horizontal gene transfer vector, called the gene transfer agent (GTA), that appears to have evolved from a bacteriophage. The model system used to study GTA biology is the Rhodobacter capsulatus GTA (RcGTA), a small, tailed bacteriophage-like particle produced by a subset of the cells in a culture. The response regulator CtrA is conserved in the Alphaproteobacteria and is an essential regulator of RcGTA production: it controls the production and maturation of the RcGTA particle and RcGTA release from cells. CtrA also controls the natural transformation-like system required for cells to receive RcGTA-donated DNA. Here, we report that dysregulation of the CckA-ChpT-CtrA phosphorelay either by the loss of the PAS domain protein DivL or by substitution of the autophosphorylation residue of the hybrid histidine kinase CckA decreased CtrA phosphorylation and greatly increased RcGTA protein production in R. capsulatus We show that the loss of the ClpXP protease or the three C-terminal residues of CtrA results in increased CtrA levels in R. capsulatus and identify ClpX(P) to be essential for the maturation of RcGTA particles. Furthermore, we show that CtrA phosphorylation is important for head spike production. Our results provide novel insight into the regulation of CtrA and GTAs in the RhodobacteralesIMPORTANCE Members of the Rhodobacterales are abundant in ocean and freshwater environments. The conserved GTA produced by many Rhodobacterales may have an important role in horizontal gene transfer (HGT) in aquatic environments and provide a significant contribution to their adaptation. GTA production is controlled by bacterial regulatory systems, including the conserved CckA-ChpT-CtrA phosphorelay; however, several questions about GTA regulation remain. Our identification that a short DivL homologue and ClpXP regulate CtrA in R. capsulatus extends the model of CtrA regulation from Caulobacter crescentus to a member of the Rhodobacterales We found that the magnitude of RcGTA production greatly depends on DivL and CckA kinase activity, adding yet another layer of regulatory complexity to RcGTA. RcGTA is known to undergo CckA-dependent maturation, and we extend the understanding of this process by showing that the ClpX chaperone is required for formation of tailed, DNA-containing particles.

Biochemical characterization of avian influenza viral polymerase containing PA or PB2 subunit from human influenza A virus

Adaptive mutations in viral polymerase, which is composed of PB1, PB2, and PA, of avian influenza viruses are major genetic determinants of the host range. In this study, to elucidate the molecular mechanism of mammalian adaptation of avian viral polymerase, we performed cell-based vRNP reconstitution assays and biochemical analyses using purified recombinant viral polymerase complexes. We found that avian viral polymerase from A/duck/Pennsylvania/10,218/84 (DkPen) enhances the viral polymerase activity in mammalian cells by replacing the PA or PB2 gene with that from human influenza virus A/WSN/33 (WSN). Chimeric constructs between DkPen PA and WSN PA showed that the N-terminal endonuclease domain of WSN PA was essential for the mammalian adaptation of DkPen viral polymerase. We also found that the cap-snatching activity of purified DkPen viral polymerase was more than 5 times weaker than that of WSN in vitro in a PB2 Glu627-dependent manner. However, the cap-snatching activity of DkPen viral polymerase was hardly increased by replacing DkPen PA to WSN PA. These results suggest that the activity of viral genome replication may be enhanced in the DkPen reassortant containing WSN PA.

Epidemiology Of Self-reported Physical Activity In Eight Latin American Countries

BACKGROUND: The prevalence of physical inactivity in Latin America was one of the highest reported worldwide. The purpose of this study was a comparative international study of population PA prevalence across eight countries from Latin American. METHODS: Data from the Latin American Study of Nutrition and Health (ELANS) were included in the analysis. The sample included 9,218 adolescents and adults aged 15-65 years. PA was assessed using the International Physical Activity Questionnaire. PA was compared among countries, gender, age group, socioeconomic, educational level and different PA domains. In addition to using the MET-minutes/week rank, we also used the rating in active or insufficiently active based in minutes/week. Individuals were categorized as active (mean ≥150 min/week) or insufficiently active (mean ≤150 min/week). RESULTS: The prevalence of physically active individuals was 52.5%. Men were more active than women in all countries. Only 13.4% of the population had high PA level and 27.4% showed moderate PA level. More than half (59.3%) of subjects exhibited low levels of PA. The prevalence of physically active individuals slightly, increased from low to high socioeconomic level. Regarding the educational level, the prevalence of physically active individuals was similar among those who have a lower educational levels, high school studies, and university degrees (52.4, 52.0, and 52.3%). The largest fraction of transportation time was explained by walking time (87%). Vigorous-intensity PA was the one that contributes the most for the total leisure time (52%). Recreation/sport time contributes with 25% to total leisure time. The total of minutes of PA is explained in 55% by leisure time and in 45% by transportation time. Most of the countries more than 80% of total MET-minutes/week were explained by walking (44%) and vigorous- intensity PA (39%). The only exceptions were Ecuador and Chile, the two countries with the highest levels of PA. CONCLUSIONS: The high percentage of Latin American subjects insufficiently active people and with low levels of PA should be of concern. Measures are needed to promote the practice of PA. If assessment methods are used consistently over time within this world region, trend data will inform countries about the success of their efforts to promote PA.

Identification and Characterization of c-di-GMP Metabolic Enzymes of Leptospira interrogans and c-di-GMP Fluctuations After Thermal Shift and Infection.

Leptospirosis is a widespread zoonotic disease caused by pathogenic Leptospira species. The most common species, Leptospira interrogans, can transfer from contaminated soil or water to the human body. It is able to survive these changing environments through sensing and responding to the changes of environmental cues. Cyclic di-GMP (c-di-GMP) is a special secondary messenger in bacteria, which can respond to the environment and regulate diverse bacterial behaviors. The c-di-GMP levels in bacterial cells are regulated by diguanylatecyclases (DGC) and phosphodiesterases (PDE), which are responsible for synthesizing or hydrolyzing c-di-GMP, respectively. In this study, distribution and phylogenetics of c-di-GMP metabolic genes among 15 leptospiral species were systematically analyzed. Bioinformatics analysis revealed that leptospiral species contain a multitude of c-di-GMP metabolic genes. C-di-GMP metabolic genes in L. interrogans strain Lai 56601 were further analyzed and the results showed that these genes have very diverse expression patterns. Most of the putative DGCs and PDEs possess enzymatic activities, as determined by riboswitch-based dual-fluorescence reporters in vivo or HPLC in vitro. Furtherer analysis of subdomains from GGDEF-containing proteins revealed that the ability to synthesize c-di-GMP was lost when the GAF domain from LA1483 and PAS domain from LA2932 were deleted, while deletion of the REC domain from LA2528 did not affect its ability to synthesize c-di-GMP. Furthermore, high temperatures generally resulted in low c-di-GMP concentrations in L. interrogans and most of the c-di-GMP metabolic genes exhibited differential temperature regulation. Also, infection of murine J774A.1 cells resulted in reduced c-di-GMP levels, while no significant change of c-di-GMP metabolic genes on transcriptional levels were observed during the infection of J774A.1 cells. Taken together, these results provide a basic platform for future studies of c-di-GMP signaling pathways in Leptospira.

Synergistic stabilisation of NOsGC by cinaciguat and non-hydrolysable nucleotides: Evidence for sGC activator-induced communication between the heme-binding and catalytic domains

Nitric oxide sensitive guanylyl cyclase (NOsGC) is a heterodimeric enzyme consisting of one α and one β subunit. Each subunit consists of four domains: the N-terminal heme-nitric oxide oxygen binding (HNOX) domain, a PAS domain, a coiled-coil domain and the C-terminal catalytic domain. Upon activation by the endogenous ligand NO or activating drugs, NOsGC catalyses the conversion of GTP to cGMP. Although several crystal structures of the isolated domains are known, the structure of the full-length enzyme and the interdomain conformational changes during activation remain unsolved to date. In the current study, we performed protein thermal shift assays of purified NOsGC to identify discrete conformational states amenable to further analysis e.g. by crystallisation. A non-hydrolysable substrate analogue binding to the catalytic domain led to a subtle change in melting temperature. An activator drug binding to the HNOX domain led to a small increase. However, the combination of substrate analogue and activator drug led to a marked synergistic increase from 51 °C to 60 °C. This suggests reciprocal communication between HNOX domain and catalytic domain and formation of a stable activated conformation amenable to further biophysical characterization.

ResDE Two-Component Regulatory System Mediates Oxygen Limitation-Induced Biofilm Formation by Bacillus amyloliquefaciens SQR9.

Efficient biofilm formation and root colonization capabilities facilitate the ability of beneficial plant rhizobacteria to promote plant growth and antagonize soilborne pathogens. Biofilm formation by plant-beneficial Bacillus strains is triggered by environmental cues, including oxygen deficiency, but the pathways that sense these environmental signals and regulate biofilm formation have not been thoroughly elucidated. In this study, we showed that the ResDE two-component regulatory system in the plant growth-promoting rhizobacterium Bacillus amyloliquefaciens strain SQR9 senses the oxygen deficiency signal and regulates biofilm formation. ResE is activated by sensing the oxygen limitation-induced reduction of the NAD+/NADH pool through its PAS domain, stimulating its kinase activity, and resulting in the transfer of a phosphoryl group to ResD. The phosphorylated ResD directly binds to the promoter regions of the qoxABCD and ctaCDEF operons to improve the biosynthesis of terminal oxidases, which can interact with KinB to activate biofilm formation. These results not only revealed the novel regulatory function of the ResDE two-component system but also contributed to the understanding of the complicated regulatory network governing Bacillus biofilm formation. This research may help to enhance the root colonization and the plant-beneficial efficiency of SQR9 and other Bacillus rhizobacteria used in agriculture.IMPORTANCEBacillus spp. are widely used as bioinoculants for plant growth promotion and disease suppression. The exertion of their plant-beneficial functions is largely dependent on their root colonization, which is closely related to their biofilm formation capabilities. On the other hand, Bacillus is the model bacterium for biofilm study, and the process and molecular network of biofilm formation are well characterized (B. Mielich-Süss and D. Lopez, Environ Microbiol 17:555-565, 2015, https://doi.org/10.1111/1462-2920.12527; L. S. Cairns, L. Hobley, and N. R. Stanley-Wall, Mol Microbiol 93:587-598, 2014, https://doi.org/10.1111/mmi.12697; H. Vlamakis, C. Aguilar, R. Losick, and R. Kolter, Genes Dev 22:945-953, 2008, https://doi.org/10.1101/gad.1645008; S. S. Branda, A. Vik, L. Friedman, and R. Kolter, Trends Microbiol 13:20-26, 2005, https://doi.org/10.1016/j.tim.2004.11.006; C. Aguilar, H. Vlamakis, R. Losick, and R. Kolter, Curr Opin Microbiol 10:638-643, 2007, https://doi.org/10.1016/j.mib.2007.09.006; S. S. Branda, J. E. González-Pastor, S. Ben-Yehuda, R. Losick, and R. Kolter, Proc Natl Acad Sci U S A 98:11621-11626, 2001, https://doi.org/10.1073/pnas.191384198). However, the identification and sensing of environmental signals triggering Bacillus biofilm formation need further research. Here, we report that the oxygen deficiency signal inducing Bacillus biofilm formation is sensed by the ResDE two-component regulatory system. Our results not only revealed the novel regulatory function of the ResDE two-component regulatory system but also identified the sensing system of a biofilm-triggering signal. This knowledge can help to enhance the biofilm formation and root colonization of plant-beneficial Bacillus strains and also provide new insights of bacterial biofilm formation regulation.

PASsing on signals to COMPASS: A novel intramolecular interactions in PAS Kinase controls the stem cell fate via regulating COMPASS protein complexes.

Stem cell fate in the tissue niche is intimately connected with intra- and extra-cellular metabolic state and the hormonal stimulations. We have shown that a sensory kinase, PAS domain Kinase (PASK) phosphorylates Wdr5, a member of COMPASS family of histone methyltransferases, to activate the stem cell differentiation program in multiple paradigms (elife, 2016). PASK is expressed highly and exclusively in stem cells, yet the differentiation signaling cues are required to activate PASK and its downstream functions, suggesting PASK remains inaccessible to bind Wdr5 in proliferating stem cells. Our recent data showed that the mechanistic Target of Rapamycin (mTOR) activates PASK-Wdr5 signaling pathway in response to the differentiation signaling cues such as nutrients and insulin (in peer-review). mTOR induces multi-site phosphorylation on PASK to stimulate the recruitment of Wdr5 by PASK, resulting in Wdr5 phosphorylation at Ser49. Phosphorylation of Wdr5 stimulates the histone trimethylation at lysine 4 (H3K4me3) at the Myogenin promoter to activate the muscle stem cell differentiation program. How mTOR phosphorylation stimulates PASK-Wdr5 interactions remains unknown but is a key mechanistic question that could help understand how nutrient and metabolic signaling could acutely control stem cell differentiation on demand. By using multi-disciplinary approaches, we show that the PAS domain of PASK negatively regulates PASK-Wdr5 interaction by occupying the Wdr5 interacting surface onto PASK. Interestingly, we have mapped mTOR stimulated phosphorylation sites on PASK adjacent to a region where both PAS and Wdr5 binds. We show that mTOR phosphorylation serves to disrupt intra-molecular inhibition of PAS domain, allowing for Wdr5 binding and phosphorylation by PASK. As PAS domains are versatile sensors of the metabolic environment and bind diverse sets of ligands such as heme, oxygen, blue light and organic small molecules, our data also suggest a possibility of additional allosteric regulation of PASK activity via PAS domains which could impact stem cell fate. In proliferating stem cells, PASK expression is high, but it is kept inaccessible to bind Wdr5 by intramolecular association between PAS domain and PAS interacting motif (PIM). PIM is situated adjacent to Wdr5 binding motif and mTOR phosphorylation sites. Upon differentiation signaling, mTOR mediated phosphorylation induces local conformation change allowing PIM to be dissociated from PAS domain resulting in Wdr5 binding onto PASK, its phosphorylation and the onset of differentiation. This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Expression and Purification of human Neuronal PAS domain protein 2 (hNPAS2)

The circadian rhythm is an internal clock responsible for a variety of physiological processes occurring within a 24‐hour cycle. Synchronized by a master clock in the brain, the circadian rhythm consists of a transcriptional‐translational feedback loop (TTFL) capable of inducing oscillating gene expression in all the cells throughout the organism. The TTFL begins with the heterodimerization of two transcription factors, BMAL1 and CLOCK, resulting in the transcription and translation of many circadian‐related genes. The target protein of interest, human neuronal PAS domain protein 2 (hNPAS2) is a paralog of CLOCK that can serve as a positive regulator of the circadian rhythm by heterodimerizing with BMAL1. Previous research suggests that disruption in the expression of hNPAS2 may result in improper circadian functioning and development of sleeping and mental disorders. In this study, the full‐length eukaryotic form of hNPAS2 was cloned into a bacterial expression vector. However, bacterial expression has led to misfolding and aggregation of the recombinant hNPAS2, becoming the central issue of the study. In an effort to obtain soluble and homogeneous hNPAS2, multiple expression and purification methods have been developed. The resultant purified soluble protein will enable future structural and functional studies, which can bring new insights to the regulatory mechanisms of the circadian rhythm and understanding of circadian‐related disorders. Additionally, a portion of this research was integrated into a teaching module delivered by the leading student researcher to a senior undergraduate advanced biochemistry course. Multiple novel pedagogic techniques were employed in the teaching module with the objective of raising the students' interests to research as well as their abilities to apply the principles and concepts of biochemistry learned in the course in a real scientific study.Support or Funding InformationResearch reported in this poster was supported by the National Institute of General medical Sciences of the National Institutes of Health under linked Award Numbers RL5GM118969, TL4GM118971, AND UL1GM118970. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health Research reported in this poster was supported by the National Research of General Medical Sciences of NIH under award number SC3GM109870 Research reported in this poster was supported has used core facility at BBRC supported by National Institute of Minority Health and Health Disparities under award number 5G12MD007592This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

A Miniaturized Escherichia coli Green Light Sensor with High Dynamic Range.

Genetically engineered photoreceptors enable unrivaled control over gene expression. Previously, we ported the Synechocystis PCC 6803 CcaSR two-component system, which is activated by green light and deactivated by red, into Escherichia coli, resulting in a sensor with a sixfold dynamic range. Later, we optimized pathway protein expression levels and the output promoter sequence to decrease transcriptional leakiness and to increase the dynamic range to approximately 120-fold. These CcaSR v 1.0 and v 2.0 systems have been used for precise quantitative, temporal, and spatial control of gene expression for a variety of applications. Recently, other workers deleted two PAS domains of unknown function from the CcaS sensor histidine kinase in a system similar to CcaSR v 1.0. Here we apply these deletions to CcaSR v 2.0, resulting in a v 3.0 light sensor with an output four times less leaky and a dynamic range of nearly 600-fold. We demonstrate that the PAS domain deletions have no deleterious effect on CcaSR green light sensitivity or response dynamics. CcaSR v 3.0 is the best-performing engineered bacterial green light sensor available, and should have broad applications in fundamental and synthetic biology studies.

Protective Role of Neuronal PAS Domain Protein 3 NPAS3 Gene in the Survival of Human Brain Stem Cell

Protective Role of Neuronal PAS Domain Protein 3 NPAS3 Gene in the Survival of Human Brain Stem Cell

Phyletic Distribution and Lineage-Specific Domain Architectures of Archaeal Two-Component Signal Transduction Systems.

ABSTRACTThe two-component signal transduction (TCS) machinery is a key mechanism of sensing environmental changes in the prokaryotic world. TCS systems have been characterized thoroughly in bacteria but to a much lesser extent in archaea. Here, we provide an updated census of more than 2,000 histidine kinases and response regulators encoded in 218 complete archaeal genomes, as well as unfinished genomes available from metagenomic data. We describe the domain architectures of the archaeal TCS components, including several novel output domains, and discuss the evolution of the archaeal TCS machinery. The distribution of TCS systems in archaea is strongly biased, with high levels of abundance in haloarchaea and thaumarchaea but none detected in the sequenced genomes from the phyla Crenarchaeota, Nanoarchaeota, and Korarchaeota. The archaeal sensor histidine kinases are generally similar to their well-studied bacterial counterparts but are often located in the cytoplasm and carry multiple PAS and/or GAF domains. In contrast, archaeal response regulators differ dramatically from the bacterial ones. Most archaeal genomes do not encode any of the major classes of bacterial response regulators, such as the DNA-binding transcriptional regulators of the OmpR/PhoB, NarL/FixJ, NtrC, AgrA/LytR, and ActR/PrrA families and the response regulators with GGDEF and/or EAL output domains. Instead, archaea encode multiple copies of response regulators containing either the stand-alone receiver (REC) domain or combinations of REC with PAS and/or GAF domains. Therefore, the prevailing mechanism of archaeal TCS signaling appears to be via a variety of protein-protein interactions, rather than direct transcriptional regulation.IMPORTANCE Although the Archaea represent a separate domain of life, their signaling systems have been assumed to be closely similar to the bacterial ones. A study of the domain architectures of the archaeal two-component signal transduction (TCS) machinery revealed an overall similarity of archaeal and bacterial sensory modules but substantial differences in the signal output modules. The prevailing mechanism of archaeal TCS signaling appears to involve various protein-protein interactions rather than direct transcription regulation. The complete list of histidine kinases and response regulators encoded in the analyzed archaeal genomes is available online at http://www.ncbi.nlm.nih.gov/Complete_Genomes/TCSarchaea.html.

Phenolic Pyrogallol Fluorogen for Red Fluorescence Development in a PAS Domain Protein

Phenolic Pyrogallol Fluorogen for Red Fluorescence Development in a PAS Domain Protein

Transcriptional response of Methoprene-tolerant (Met) gene to three insect growth disruptors in Leptinotarsa decemlineata (Say)

Abstract Some insect growth disruptors (IGDs), such as pyriproxyfen and halofenozide, may be used to control Leptinotarsa decemlineata. However, their mechanism of action remains elusive. Methoprene-tolerant (Met) mediates juvenile hormone (JH) signal to control numerous essential physiological processes. In the present paper, we identified a Met gene (LdMet). LdMet protein was a typical basic helix-loop-helix/Per-Arnt-Sim (bHLH-PAS) transcription factor with a bHLH domain, two PAS domains (PAS-A and PAS-B) and a region called PAS associated C terminal (PAC). Eight conserved amino acids critical for JH binding were located in PAS-B and PAC domains. The temporal expression pattern of LdMet was in accordance with the variation of circulating JH titers. Feeding of juvenoid methoprene or pyriproxyfen, or provide for JH dose-dependently stimulated the expression of LdMet. RNA interference-mediated knockdown of two JH degradation genes increased the transcription of LdMet, while silencing of a JH biosynthesis gene repressed the transcription. Conversely, ingestion of an ecdysteroid agonist halofenozide or 20-hydroxyecdysone (20E) reduced the mRNA levels of LdMet, in a dose-dependent manner, whereas knockdown of either ecdysteroidogenesis or 20E signaling genes increased the mRNA accumulation. Providing that the expression of LdMet can be disturbed by methoprene, pyriproxyfen and halofenozide, LdMet may be a potential target of these IGDs in L. decemlineata larvae.

Identification of influenza polymerase inhibitors targeting C-terminal domain of PA through surface plasmon resonance screening

Currently, many strains of influenza A virus have developed resistance against anti-influenza drugs, and it is essential to find new chemicals to combat this virus. The influenza polymerase with three proteins, PA, PB1 and PB2, is a crucial component of the viral ribonucleoprotein (RNP) complex. Here, we report the identification of a hit compound 221 by surface plasmon resonance (SPR) direct binding screening on the C-terminal of PA (PAC). Compound 221 can subdue influenza RNP activities and attenuate influenza virus replication. Its analogs were subsequently investigated and twelve of them could attenuate RNP activities. One of the analogs, compound 312, impeded influenza A virus replication in Madin-Darby canine kidney cells with IC50 of 27.0 ± 16.8 μM. In vitro interaction assays showed that compound 312 bound directly to PAC with Kd of about 40 μM. Overall, the identification of novel PAC-targeting compounds provides new ground for drug design against influenza virus in the future.

The hERG PAS Domain Facilitates Gating Charge Deactivation at Physiological Temperature

The hERG PAS Domain Facilitates Gating Charge Deactivation at Physiological Temperature

Socio-economic determinants of physical activity across the life course: A "DEterminants of DIet and Physical ACtivity" (DEDIPAC) umbrella literature review.

BackgroundTo date, the scientific literature on socioeconomic correlates and determinants of physical activity behaviours has been dispersed throughout a number of systematic reviews, often focusing on one factor (e.g. education or parental income) in one specific age group (e.g. pre-school children or adults). The aim of this umbrella review is to provide a comprehensive and systematic overview of the scientific literature from previously conducted research by summarising and synthesising the importance and strength of the evidence related to socioeconomic correlates and determinants of PA behaviours across the life course.MethodsMedline, Embase, ISI Web of Science, Scopus and SPORTDiscus were searched for systematic literature reviews and meta-analyses of observational studies investigating the association between socioeconomic determinants of PA and PA itself (from January 2004 to September 2017). Data extraction evaluated the importance of determinants, strength of evidence, and methodological quality of the selected papers. The full protocol is available from PROSPERO (PROSPERO2014:CRD42015010616).ResultsNineteen reviews were included. Moderate methodological quality emerged. For adults, convincing evidence supports a relationship between PA and socioeconomic status (SES), especially in relation to leisure time (positive relationship) and occupational PA (negative relationship). Conversely, no association between PA and SES or parental SES was found for pre-school, school-aged children and adolescents.ConclusionsAvailable evidence on the socioeconomic determinants of PA behaviour across the life course is probable (shows fairly consistent associations) at best. While some evidence is available for adults, less was available for youth. This is mainly due to a limited quantity of primary studies, weak research designs and lack of accuracy in the PA and SES assessment methods employed. Further PA domain specific studies using longitudinal design and clear measures of SES and PA assessment are required.