Large Advanced Space Systems (LASS) computer-aided design program additions

NASA Technical Reports Server (NTRS)

Farrell, C. E.

1982-01-01

The LSS preliminary and conceptual design requires extensive iteractive analysis because of the effects of structural, thermal, and control intercoupling. A computer aided design program that will permit integrating and interfacing of required large space system (LSS) analyses is discussed. The primary objective of this program is the implementation of modeling techniques and analysis algorithms that permit interactive design and tradeoff studies of LSS concepts. Eight software modules were added to the program. The existing rigid body controls module was modified to include solar pressure effects. The new model generator modules and appendage synthesizer module are integrated (interfaced) to permit interactive definition and generation of LSS concepts. The mass properties module permits interactive specification of discrete masses and their locations. The other modules permit interactive analysis of orbital transfer requirements, antenna primary beam n, and attitude control requirements.

Formulation of advanced consumables management models: Environmental control and electrical power system performance models requirements

NASA Technical Reports Server (NTRS)

Daly, J. K.; Torian, J. G.

1979-01-01

Software design specifications for developing environmental control and life support system (ECLSS) and electrical power system (EPS) programs into interactive computer programs are presented. Specifications for the ECLSS program are at the detail design level with respect to modification of an existing batch mode program. The FORTRAN environmental analysis routines (FEAR) are the subject batch mode program. The characteristics of the FEAR program are included for use in modifying batch mode programs to form interactive programs. The EPS program specifications are at the preliminary design level. Emphasis is on top-down structuring in the development of an interactive program.

Gcn4-Mediator Specificity Is Mediated by a Large and Dynamic Fuzzy Protein-Protein Complex.

PubMed

Tuttle, Lisa M; Pacheco, Derek; Warfield, Linda; Luo, Jie; Ranish, Jeff; Hahn, Steven; Klevit, Rachel E

2018-03-20

Transcription activation domains (ADs) are inherently disordered proteins that often target multiple coactivator complexes, but the specificity of these interactions is not understood. Efficient transcription activation by yeast Gcn4 requires its tandem ADs and four activator-binding domains (ABDs) on its target, the Mediator subunit Med15. Multiple ABDs are a common feature of coactivator complexes. We find that the large Gcn4-Med15 complex is heterogeneous and contains nearly all possible AD-ABD interactions. Gcn4-Med15 forms via a dynamic fuzzy protein-protein interface, where ADs bind the ABDs in multiple orientations via hydrophobic regions that gain helicity. This combinatorial mechanism allows individual low-affinity and specificity interactions to generate a biologically functional, specific, and higher affinity complex despite lacking a defined protein-protein interface. This binding strategy is likely representative of many activators that target multiple coactivators, as it allows great flexibility in combinations of activators that can cooperate to regulate genes with variable coactivator requirements. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

Dpb11 may function with RPA and DNA to initiate DNA replication.

PubMed

Bruck, Irina; Dhingra, Nalini; Martinez, Matthew P; Kaplan, Daniel L

2017-01-01

Dpb11 is required for the initiation of DNA replication in budding yeast. We found that Dpb11 binds tightly to single-stranded DNA (ssDNA) or branched DNA structures, while its human homolog, TopBP1, binds tightly to branched-DNA structures. We also found that Dpb11 binds stably to CDK-phosphorylated RPA, the eukaryotic ssDNA binding protein, in the presence of branched DNA. A Dpb11 mutant specifically defective for DNA binding did not exhibit tight binding to RPA in the presence of DNA, suggesting that Dpb11-interaction with DNA may promote the recruitment of RPA to melted DNA. We then characterized a mutant of Dpb11 that is specifically defective in DNA binding in budding yeast cells. Expression of dpb11-m1,2,3,5,ΔC results in a substantial decrease in RPA recruitment to origins, suggesting that Dpb11 interaction with DNA may be required for RPA recruitment to origins. Expression of dpb11-m1,2,3,5,ΔC also results in diminished GINS interaction with Mcm2-7 during S phase, while Cdc45 interaction with Mcm2-7 is like wild-type. The reduced GINS interaction with Mcm2-7 may be an indirect consequence of diminished origin melting. We propose that the tight interaction between Dpb11, CDK-phosphorylated RPA, and branched-DNA may be required for the essential function of stabilizing melted origin DNA in vivo. We also propose an alternative model, wherein Dpb11-DNA interaction is required for some other function in DNA replication initiation, such as helicase activation.

Probing fibronectin–antibody interactions using AFM force spectroscopy and lateral force microscopy

PubMed Central

Kulik, Andrzej J; Lee, Kyumin; Pyka-Fościak, Grazyna; Nowak, Wieslaw

2015-01-01

Summary The first experiment showing the effects of specific interaction forces using lateral force microscopy (LFM) was demonstrated for lectin–carbohydrate interactions some years ago. Such measurements are possible under the assumption that specific forces strongly dominate over the non-specific ones. However, obtaining quantitative results requires the complex and tedious calibration of a torsional force. Here, a new and relatively simple method for the calibration of the torsional force is presented. The proposed calibration method is validated through the measurement of the interaction forces between human fibronectin and its monoclonal antibody. The results obtained using LFM and AFM-based classical force spectroscopies showed similar unbinding forces recorded at similar loading rates. Our studies verify that the proposed lateral force calibration method can be applied to study single molecule interactions. PMID:26114080

STAT3 or USF2 Contributes to HIF Target Gene Specificity

PubMed Central

Pawlus, Matthew R.; Wang, Liyi; Murakami, Aya; Dai, Guanhai; Hu, Cheng-Jun

2013-01-01

The HIF1- and HIF2-mediated transcriptional responses play critical roles in solid tumor progression. Despite significant similarities, including their binding to promoters of both HIF1 and HIF2 target genes, HIF1 and HIF2 proteins activate unique subsets of target genes under hypoxia. The mechanism for HIF target gene specificity has remained unclear. Using siRNA or inhibitor, we previously reported that STAT3 or USF2 is specifically required for activation of endogenous HIF1 or HIF2 target genes. In this study, using reporter gene assays and chromatin immuno-precipitation, we find that STAT3 or USF2 exhibits specific binding to the promoters of HIF1 or HIF2 target genes respectively even when over-expressed. Functionally, HIF1α interacts with STAT3 to activate HIF1 target gene promoters in a HIF1α HLH/PAS and N-TAD dependent manner while HIF2α interacts with USF2 to activate HIF2 target gene promoters in a HIF2α N-TAD dependent manner. Physically, HIF1α HLH and PAS domains are required for its interaction with STAT3 while both N- and C-TADs of HIF2α are involved in physical interaction with USF2. Importantly, addition of functional USF2 binding sites into a HIF1 target gene promoter increases the basal activity of the promoter as well as its response to HIF2+USF2 activation while replacing HIF binding site with HBS from a HIF2 target gene does not change the specificity of the reporter gene. Importantly, RNA Pol II on HIF1 or HIF2 target genes is primarily associated with HIF1α or HIF2α in a STAT3 or USF2 dependent manner. Thus, we demonstrate here for the first time that HIF target gene specificity is achieved by HIF transcription partners that are required for HIF target gene activation, exhibit specific binding to the promoters of HIF1 or HIF2 target genes and selectively interact with HIF1α or HIF2α protein. PMID:23991099

Phosphorylation regulates the Star-PAP-PIPKIα interaction and directs specificity toward mRNA targets

PubMed Central

Mohan, Nimmy; AP, Sudheesh; Francis, Nimmy; Anderson, Richard; Laishram, Rakesh S.

2015-01-01

Star-PAP is a nuclear non-canonical poly(A) polymerase (PAP) that shows specificity toward mRNA targets. Star-PAP activity is stimulated by lipid messenger phosphatidyl inositol 4,5 bisphoshate (PI4,5P2) and is regulated by the associated Type I phosphatidylinositol-4-phosphate 5-kinase that synthesizes PI4,5P2 as well as protein kinases. These associated kinases act as coactivators of Star-PAP that regulates its activity and specificity toward mRNAs, yet the mechanism of control of these interactions are not defined. We identified a phosphorylated residue (serine 6, S6) on Star-PAP in the zinc finger region, the domain required for PIPKIα interaction. We show that S6 is phosphorylated by CKIα within the nucleus which is required for Star-PAP nuclear retention and interaction with PIPKIα. Unlike the CKIα mediated phosphorylation at the catalytic domain, Star-PAP S6 phosphorylation is insensitive to oxidative stress suggesting a signal mediated regulation of CKIα activity. S6 phosphorylation together with coactivator PIPKIα controlled select subset of Star-PAP target messages by regulating Star-PAP-mRNA association. Our results establish a novel role for phosphorylation in determining Star-PAP target mRNA specificity and regulation of 3′-end processing. PMID:26138484

Pharmacokinetic Modeling of JP-8 Jet Fuel Components: II. A Conceptual Framework

DTIC Science & Technology

2003-12-01

example, a single type of (simple) binary interaction between 300 components would require the specification of some 105 interaction coefficients . One...individual substances, via binary mechanisms, is enough to predict the interactions present in the mixture. Secondly, complex mixtures can often be...approximated as pseudo- binary systems, consisting of the compound of interest plus a single interacting complex vehicle with well-defined, composite

Specific Non-Local Interactions Are Not Necessary for Recovering Native Protein Dynamics

PubMed Central

Dasgupta, Bhaskar; Kasahara, Kota; Kamiya, Narutoshi; Nakamura, Haruki; Kinjo, Akira R.

2014-01-01

The elastic network model (ENM) is a widely used method to study native protein dynamics by normal mode analysis (NMA). In ENM we need information about all pairwise distances, and the distance between contacting atoms is restrained to the native value. Therefore ENM requires O(N2) information to realize its dynamics for a protein consisting of N amino acid residues. To see if (or to what extent) such a large amount of specific structural information is required to realize native protein dynamics, here we introduce a novel model based on only O(N) restraints. This model, named the ‘contact number diffusion’ model (CND), includes specific distance restraints for only local (along the amino acid sequence) atom pairs, and semi-specific non-local restraints imposed on each atom, rather than atom pairs. The semi-specific non-local restraints are defined in terms of the non-local contact numbers of atoms. The CND model exhibits the dynamic characteristics comparable to ENM and more correlated with the explicit-solvent molecular dynamics simulation than ENM. Moreover, unrealistic surface fluctuations often observed in ENM were suppressed in CND. On the other hand, in some ligand-bound structures CND showed larger fluctuations of buried protein atoms interacting with the ligand compared to ENM. In addition, fluctuations from CND and ENM show comparable correlations with the experimental B-factor. Although there are some indications of the importance of some specific non-local interactions, the semi-specific non-local interactions are mostly sufficient for reproducing the native protein dynamics. PMID:24625758

Development of a functional cell-based assay that probes the specific interaction between influenza A virus NP and its packaging signal sequence RNA.

PubMed

Woo, Jiwon; Yu, Kyung Lee; Lee, Sun Hee; You, Ji Chang

2015-02-06

Although cis-acting packaging signal RNA sequences for the influenza virus NP encoding vRNA have been identified recently though genetic studies, little is known about the interaction between NP and the vRNA packaging signals either in vivo or in vitro. Here, we provide evidence that NP is able to interact specifically with the vRNA packaging sequence RNA within living cells and that the specific RNA binding activity of NP in vivo requires both the N-terminal and central region of the protein. This assay established would be a valuable tool for further detailed studies of the NP-packaging signal RNA interaction in living cells. Copyright © 2014 Elsevier Inc. All rights reserved.

Distinct TERB1 Domains Regulate Different Protein Interactions in Meiotic Telomere Movement.

PubMed

Zhang, Jingjing; Tu, Zhaowei; Watanabe, Yoshinori; Shibuya, Hiroki

2017-11-14

Meiotic telomeres attach to the nuclear envelope (NE) and drive the chromosome movement required for the pairing of homologous chromosomes. The meiosis-specific telomere proteins TERB1, TERB2, and MAJIN are required to regulate these events, but their assembly processes are largely unknown. Here, we developed a germ-cell-specific knockout mouse of the canonical telomere-binding protein TRF1 and revealed an essential role for TRF1 in directing the assembly of TERB1-TERB2-MAJIN. Further, we identified a TERB2 binding (T2B) domain in TERB1 that is dispensable for the TRF1-TERB1 interaction but is essential for the subsequent TERB1-TERB2 interaction and therefore for telomere attachment to the NE. Meanwhile, cohesin recruitment at telomeres, which is required for efficient telomere movement, is mediated by the MYB-like domain of TERB1, but not by TERB2-MAJIN. Our results reveal distinct protein interactions through various domains of TERB1, which enable the sequential assembly of the meiotic telomere complex for their movements. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

Mass spectrometric identification of proteins that interact through specific domains of the poly(A) binding protein

PubMed Central

Zhang, Chongxu; Nielsen, Maria E. O.; Chiang, Yueh-Chin; Kierkegaard, Morten; Wang, Xin; Lee, Darren J.; Andersen, Jens S.; Yao, Gang

2013-01-01

Poly(A) binding protein (PAB1) is involved in a number of RNA metabolic functions in eukaryotic cells and correspondingly is suggested to associate with a number of proteins. We have used mass spectrometric analysis to identify 55 non-ribosomal proteins that specifically interact with PAB1 from Saccharomyces cerevisiae. Because many of these factors may associate only indirectly with PAB1 by being components of the PAB1-mRNP structure, we additionally conducted mass spectrometric analyses on seven metabolically defined PAB1 deletion derivatives to delimit the interactions between these proteins and PAB1. These latter analyses identified 13 proteins whose associations with PAB1 were reduced by deleting one or another of PAB1’s defined domains. Included in this list of 13 proteins were the translation initiation factors eIF4G1 and eIF4G2, translation termination factor eRF3, and PBP2, all of whose previously known direct interactions with specific PAB1 domains were either confirmed, delimited, or extended. The remaining nine proteins that interacted through a specific PAB1 domain were CBF5, SLF1, UPF1, CBC1, SSD1, NOP77, yGR250c, NAB6, and GBP2. In further study, UPF1, involved in nonsense-mediated decay, was confirmed to interact with PAB1 through the RRM1 domain. We additionally established that while the RRM1 domain of PAB1 was required for UPF1-induced acceleration of deadenylation during nonsense-mediated decay, it was not required for the more critical step of acceleration of mRNA decapping. These results begin to identify the proteins most likely to interact with PAB1 and the domains of PAB1 through which these contacts are made. PMID:22836166

Mass spectrometric identification of proteins that interact through specific domains of the poly(A) binding protein.

PubMed

Richardson, Roy; Denis, Clyde L; Zhang, Chongxu; Nielsen, Maria E O; Chiang, Yueh-Chin; Kierkegaard, Morten; Wang, Xin; Lee, Darren J; Andersen, Jens S; Yao, Gang

2012-09-01

Poly(A) binding protein (PAB1) is involved in a number of RNA metabolic functions in eukaryotic cells and correspondingly is suggested to associate with a number of proteins. We have used mass spectrometric analysis to identify 55 non-ribosomal proteins that specifically interact with PAB1 from Saccharomyces cerevisiae. Because many of these factors may associate only indirectly with PAB1 by being components of the PAB1-mRNP structure, we additionally conducted mass spectrometric analyses on seven metabolically defined PAB1 deletion derivatives to delimit the interactions between these proteins and PAB1. These latter analyses identified 13 proteins whose associations with PAB1 were reduced by deleting one or another of PAB1's defined domains. Included in this list of 13 proteins were the translation initiation factors eIF4G1 and eIF4G2, translation termination factor eRF3, and PBP2, all of whose previously known direct interactions with specific PAB1 domains were either confirmed, delimited, or extended. The remaining nine proteins that interacted through a specific PAB1 domain were CBF5, SLF1, UPF1, CBC1, SSD1, NOP77, yGR250c, NAB6, and GBP2. In further study, UPF1, involved in nonsense-mediated decay, was confirmed to interact with PAB1 through the RRM1 domain. We additionally established that while the RRM1 domain of PAB1 was required for UPF1-induced acceleration of deadenylation during nonsense-mediated decay, it was not required for the more critical step of acceleration of mRNA decapping. These results begin to identify the proteins most likely to interact with PAB1 and the domains of PAB1 through which these contacts are made.

Dpb11 may function with RPA and DNA to initiate DNA replication

PubMed Central

Bruck, Irina; Dhingra, Nalini; Martinez, Matthew P.

2017-01-01

Dpb11 is required for the initiation of DNA replication in budding yeast. We found that Dpb11 binds tightly to single-stranded DNA (ssDNA) or branched DNA structures, while its human homolog, TopBP1, binds tightly to branched-DNA structures. We also found that Dpb11 binds stably to CDK-phosphorylated RPA, the eukaryotic ssDNA binding protein, in the presence of branched DNA. A Dpb11 mutant specifically defective for DNA binding did not exhibit tight binding to RPA in the presence of DNA, suggesting that Dpb11-interaction with DNA may promote the recruitment of RPA to melted DNA. We then characterized a mutant of Dpb11 that is specifically defective in DNA binding in budding yeast cells. Expression of dpb11-m1,2,3,5,ΔC results in a substantial decrease in RPA recruitment to origins, suggesting that Dpb11 interaction with DNA may be required for RPA recruitment to origins. Expression of dpb11-m1,2,3,5,ΔC also results in diminished GINS interaction with Mcm2-7 during S phase, while Cdc45 interaction with Mcm2-7 is like wild-type. The reduced GINS interaction with Mcm2-7 may be an indirect consequence of diminished origin melting. We propose that the tight interaction between Dpb11, CDK-phosphorylated RPA, and branched-DNA may be required for the essential function of stabilizing melted origin DNA in vivo. We also propose an alternative model, wherein Dpb11-DNA interaction is required for some other function in DNA replication initiation, such as helicase activation. PMID:28467467

A novel motif in the yeast mitochondrial dynamin Dnm1 is essential for adaptor binding and membrane recruitment

PubMed Central

Bui, Huyen T.; Karren, Mary A.; Bhar, Debjani

2012-01-01

To initiate mitochondrial fission, dynamin-related proteins (DRPs) must bind specific adaptors on the outer mitochondrial membrane. The structural features underlying this interaction are poorly understood. Using yeast as a model, we show that the Insert B domain of the Dnm1 guanosine triphosphatase (a DRP) contains a novel motif required for association with the mitochondrial adaptor Mdv1. Mutation of this conserved motif specifically disrupted Dnm1–Mdv1 interactions, blocking Dnm1 recruitment and mitochondrial fission. Suppressor mutations in Mdv1 that restored Dnm1–Mdv1 interactions and fission identified potential protein-binding interfaces on the Mdv1 β-propeller domain. These results define the first known function for Insert B in DRP–adaptor interactions. Based on the variability of Insert B sequences and adaptor proteins, we propose that Insert B domains and mitochondrial adaptors have coevolved to meet the unique requirements for mitochondrial fission of different organisms. PMID:23148233

Physical Foundations of PTEN/Phosphoinositide Interaction

NASA Astrophysics Data System (ADS)

Gericke, Arne; Jiang, Zhiping; Redfern, Roberta E.; Kooijman, Edgar E.; Ross, Alonzo H.

2009-03-01

Phosphoinositides act as signaling molecules by recruiting critical effectors to specific subcellular membranes to regulate cell proliferation, apoptosis and cytoskeletal reorganization, which requires a tight regulation of phosphoinositide generation and turnover as well as a high degree of compartmentalization. PTEN is a phosphatase specific for the 3 position of the phosophoinositide ring that is deleted or mutated in many different disease states. PTEN association with membranes requires the interaction of its C2 domain with phosphatidylserine and the interaction of its N-terminal end with phosphatidylinositol-4,5-bisphophate (PI(4,5)P2). We have investigated PTEN/PI(4,5)P2 interaction and found that Lys13 is crucial for the observed binding. We also found that the presence of cholesterol enhances PTEN binding to mixed PI(4,5)P2/POPC vesicles. Fluorescence microscopy experiments utilizing GUVs yielded results consistent with enhanced phosphoinositide domain formation in the presence of cholesterol. These experiments were accompanied by zeta potential measurements and solid state MAS ^31P-NMR experiments aimed at investigating the ionization behavior of phosphoinositides.

Prion-specific Hsp40 function: The role of the auxilin homolog Swa2

PubMed Central

Oliver, Emily E.; Troisi, Elizabeth M.; Hines, Justin K.

2017-01-01

ABSTRACT Yeast prions are protein-based genetic elements that propagate through cell populations via cytosolic transfer from mother to daughter cell. Molecular chaperone proteins including Hsp70, the Hsp40/J-protein Sis1, and Hsp104 are required for continued prion propagation, however the specific requirements of chaperone proteins differ for various prions. We recently reported that Swa2, the yeast homolog of the mammalian protein auxilin, is specifically required for the propagation of the prion [URE3].1 [URE3] propagation requires both a functional J-domain and the tetratricopeptide repeat (TPR) domain of Swa2, but does not require Swa2 clathrin binding. We concluded that the TPR domain determines the specificity of the genetic interaction between Swa2 and [URE3], and that this domain likely interacts with one or more proteins with a C-terminal EEVD motif. Here we extend that analysis to incorporate additional data that supports this hypothesis. We also present new data eliminating Hsp104 as the relevant Swa2 binding partner and discuss our findings in the context of other recent work involving Hsp90. Based on these findings, we propose a new model for Swa2's involvement in [URE3] propagation in which Swa2 and Hsp90 mediate the formation of a multi-protein complex that increases the number of sites available for Hsp104 disaggregation. PMID:28574745

Usability of Interactive Computers in Exhibitions: Designing Knowledgeable Information for Visitors

ERIC Educational Resources Information Center

Bernier, Roxane

2003-01-01

This article investigates three types of content presentation (video documentary, computerized dictionary, and games) within interactive computer use at the Quebec Museum of Civilization. The visitors' viewpoint is particularly relevant for interface designing outcomes, since they argued that terminals require specific content display for…

Phosphorylation regulates the Star-PAP-PIPKIα interaction and directs specificity toward mRNA targets.

PubMed

Mohan, Nimmy; Sudheesh, A P; Francis, Nimmy; Anderson, Richard; Laishram, Rakesh S

2015-08-18

Star-PAP is a nuclear non-canonical poly(A) polymerase (PAP) that shows specificity toward mRNA targets. Star-PAP activity is stimulated by lipid messenger phosphatidyl inositol 4,5 bisphoshate (PI4,5P2) and is regulated by the associated Type I phosphatidylinositol-4-phosphate 5-kinase that synthesizes PI4,5P2 as well as protein kinases. These associated kinases act as coactivators of Star-PAP that regulates its activity and specificity toward mRNAs, yet the mechanism of control of these interactions are not defined. We identified a phosphorylated residue (serine 6, S6) on Star-PAP in the zinc finger region, the domain required for PIPKIα interaction. We show that S6 is phosphorylated by CKIα within the nucleus which is required for Star-PAP nuclear retention and interaction with PIPKIα. Unlike the CKIα mediated phosphorylation at the catalytic domain, Star-PAP S6 phosphorylation is insensitive to oxidative stress suggesting a signal mediated regulation of CKIα activity. S6 phosphorylation together with coactivator PIPKIα controlled select subset of Star-PAP target messages by regulating Star-PAP-mRNA association. Our results establish a novel role for phosphorylation in determining Star-PAP target mRNA specificity and regulation of 3'-end processing. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

Specifying Computer-Supported Collaboration Scripts

ERIC Educational Resources Information Center

Kobbe, Lars; Weinberger, Armin; Dillenbourg, Pierre; Harrer, Andreas; Hamalainen, Raija; Hakkinen, Paivi; Fischer, Frank

2007-01-01

Collaboration scripts facilitate social and cognitive processes of collaborative learning by shaping the way learners interact with each other. Computer-supported collaboration scripts generally suffer from the problem of being restrained to a specific learning platform. A standardization of collaboration scripts first requires a specification of…

A dermal HOX transcriptional program regulates site-specific epidermal fate

PubMed Central

Rinn, John L.; Wang, Jordon K.; Allen, Nancy; Brugmann, Samantha A.; Mikels, Amanda J.; Liu, Helen; Ridky, Todd W.; Stadler, H. Scott; Nusse, Roel; Helms, Jill A.; Chang, Howard Y.

2008-01-01

Reciprocal epithelial–mesenchymal interactions shape site-specific development of skin. Here we show that site-specific HOX expression in fibroblasts is cell-autonomous and epigenetically maintained. The distal-specific gene HOXA13 is continually required to maintain the distal-specific transcriptional program in adult fibroblasts, including expression of WNT5A, a morphogen required for distal development. The ability of distal fibroblasts to induce epidermal keratin 9, a distal-specific gene, is abrogated by depletion of HOXA13, but rescued by addition of WNT5A. Thus, maintenance of appropriate HOX transcriptional program in adult fibroblasts may serve as a source of positional memory to differentially pattern the epithelia during homeostasis and regeneration. PMID:18245445

A dermal HOX transcriptional program regulates site-specific epidermal fate.

PubMed

Rinn, John L; Wang, Jordon K; Allen, Nancy; Brugmann, Samantha A; Mikels, Amanda J; Liu, Helen; Ridky, Todd W; Stadler, H Scott; Nusse, Roel; Helms, Jill A; Chang, Howard Y

2008-02-01

Reciprocal epithelial-mesenchymal interactions shape site-specific development of skin. Here we show that site-specific HOX expression in fibroblasts is cell-autonomous and epigenetically maintained. The distal-specific gene HOXA13 is continually required to maintain the distal-specific transcriptional program in adult fibroblasts, including expression of WNT5A, a morphogen required for distal development. The ability of distal fibroblasts to induce epidermal keratin 9, a distal-specific gene, is abrogated by depletion of HOXA13, but rescued by addition of WNT5A. Thus, maintenance of appropriate HOX transcriptional program in adult fibroblasts may serve as a source of positional memory to differentially pattern the epithelia during homeostasis and regeneration.

Development and Testing of a Scale to Assess Interprofessional Education (IPE) Facilitation Skills

ERIC Educational Resources Information Center

Sargeant, Joan; Hill, Tanya; Breau, Lynn

2010-01-01

Introduction: Interprofessional education (IPE) is interactive and constructivist in nature and requires specific facilitation skills to engage participants in a unique body of content, interpersonal interaction, and learning from each other. This article describes the development and testing of a scale, the Interprofessional Facilitation Scale…

Deriving Heterospecific Self-Assembling Protein-Protein Interactions Using a Computational Interactome Screen.

PubMed

Crooks, Richard O; Baxter, Daniel; Panek, Anna S; Lubben, Anneke T; Mason, Jody M

2016-01-29

Interactions between naturally occurring proteins are highly specific, with protein-network imbalances associated with numerous diseases. For designed protein-protein interactions (PPIs), required specificity can be notoriously difficult to engineer. To accelerate this process, we have derived peptides that form heterospecific PPIs when combined. This is achieved using software that generates large virtual libraries of peptide sequences and searches within the resulting interactome for preferentially interacting peptides. To demonstrate feasibility, we have (i) generated 1536 peptide sequences based on the parallel dimeric coiled-coil motif and varied residues known to be important for stability and specificity, (ii) screened the 1,180,416 member interactome for predicted Tm values and (iii) used predicted Tm cutoff points to isolate eight peptides that form four heterospecific PPIs when combined. This required that all 32 hypothetical off-target interactions within the eight-peptide interactome be disfavoured and that the four desired interactions pair correctly. Lastly, we have verified the approach by characterising all 36 pairs within the interactome. In analysing the output, we hypothesised that several sequences are capable of adopting antiparallel orientations. We subsequently improved the software by removing sequences where doing so led to fully complementary electrostatic pairings. Our approach can be used to derive increasingly large and therefore complex sets of heterospecific PPIs with a wide range of potential downstream applications from disease modulation to the design of biomaterials and peptides in synthetic biology. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

Phytotechnology Technical and Regulatory Guidance and Decision Trees, Revised

DTIC Science & Technology

2009-02-01

upon planting . In other cases, it may require several seasons before the plant can interact with a contaminated zone at depth. Furthermore, it may...potential ecological exposures posed by the species planted need to be considered. The level of detail required is site specific and varies with the...to be assessed as well. However, the ecological exposure may not be directly from the consumption of the plant . Specifically, the U.S. Environmental

Apoptosis-linked Gene-2 (ALG-2)/Sec31 Interactions Regulate Endoplasmic Reticulum (ER)-to-Golgi Transport

PubMed Central

Helm, Jared R.; Bentley, Marvin; Thorsen, Kevin D.; Wang, Ting; Foltz, Lauren; Oorschot, Viola; Klumperman, Judith; Hay, Jesse C.

2014-01-01

Luminal calcium released from secretory organelles has been suggested to play a regulatory role in vesicle transport at several steps in the secretory pathway; however, its functional roles and effector pathways have not been elucidated. Here we demonstrate for the first time that specific luminal calcium depletion leads to a significant decrease in endoplasmic reticulum (ER)-to-Golgi transport rates in intact cells. Ultrastructural analysis revealed that luminal calcium depletion is accompanied by increased accumulation of intermediate compartment proteins in COPII buds and clusters of unfused COPII vesicles at ER exit sites. Furthermore, we present several lines of evidence suggesting that luminal calcium affected transport at least in part through calcium-dependent interactions between apoptosis-linked gene-2 (ALG-2) and the Sec31A proline-rich region: 1) targeted disruption of ALG-2/Sec31A interactions caused severe defects in ER-to-Golgi transport in intact cells; 2) effects of luminal calcium and ALG-2/Sec31A interactions on transport mutually required each other; and 3) Sec31A function in transport required luminal calcium. Morphological phenotypes of disrupted ALG-2/Sec31A interactions were characterized. We found that ALG-2/Sec31A interactions were not required for the localization of Sec31A to ER exit sites per se but appeared to acutely regulate the stability and trafficking of the cargo receptor p24 and the distribution of the vesicle tether protein p115. These results represent the first outline of a mechanism that connects luminal calcium to specific protein interactions regulating vesicle trafficking machinery. PMID:25006245

A Critical Role for the Nucleus Reuniens in Long-Term, But Not Short-Term Associative Recognition Memory Formation.

PubMed

Barker, Gareth R I; Warburton, Elizabeth Clea

2018-03-28

Recognition memory for single items requires the perirhinal cortex (PRH), whereas recognition of an item and its associated location requires a functional interaction among the PRH, hippocampus (HPC), and medial prefrontal cortex (mPFC). Although the precise mechanisms through which these interactions are effected are unknown, the nucleus reuniens (NRe) has bidirectional connections with each regions and thus may play a role in recognition memory. Here we investigated, in male rats, whether specific manipulations of NRe function affected performance of recognition memory for single items, object location, or object-in-place associations. Permanent lesions in the NRe significantly impaired long-term, but not short-term, object-in-place associative recognition memory, whereas single item recognition memory and object location memory were unaffected. Temporary inactivation of the NRe during distinct phases of the object-in-place task revealed its importance in both the encoding and retrieval stages of long-term associative recognition memory. Infusions of specific receptor antagonists showed that encoding was dependent on muscarinic and nicotinic cholinergic neurotransmission, whereas NMDA receptor neurotransmission was not required. Finally, we found that long-term object-in-place memory required protein synthesis within the NRe. These data reveal a specific role for the NRe in long-term associative recognition memory through its interactions with the HPC and mPFC, but not the PRH. The delay-dependent involvement of the NRe suggests that it is not a simple relay station between brain regions, but, rather, during high mnemonic demand, facilitates interactions between the mPFC and HPC, a process that requires both cholinergic neurotransmission and protein synthesis. SIGNIFICANCE STATEMENT Recognizing an object and its associated location, which is fundamental to our everyday memory, requires specific hippocampal-cortical interactions, potentially facilitated by the nucleus reuniens (NRe) of the thalamus. However, the role of the NRe itself in associative recognition memory is unknown. Here, we reveal the crucial role of the NRe in encoding and retrieval of long-term object-in-place memory, but not for remembrance of an individual object or individual location and such involvement is cholinergic receptor and protein synthesis dependent. This is the first demonstration that the NRe is a key node within an associative recognition memory network and is not just a simple relay for information within the network. Rather, we argue, the NRe actively modulates information processing during long-term associative memory formation. Copyright © 2018 the authors 0270-6474/18/383208-10$15.00/0.

Altered interactions within FY/AtCPSF complexes required for Arabidopsis FCA-mediated chromatin silencing

PubMed Central

Manzano, David; Marquardt, Sebastian; Jones, Alexandra M. E.; Bäurle, Isabel; Liu, Fuquan; Dean, Caroline

2009-01-01

The role of RNA metabolism in chromatin silencing is now widely recognized. We have studied the Arabidopsis RNA-binding protein FCA that down-regulates an endogenous floral repressor gene through a chromatin mechanism involving histone demethylase activity. This mechanism needs FCA to interact with an RNA 3′ processing/polyadenylation factor (FY/Pfs2p), but the subsequent events leading to chromatin changes are unknown. Here, we show that this FCA–FY interaction is required for general chromatin silencing roles where hairpin transgenes induce DNA methylation of an endogenous gene. We also show 2 conserved RNA processing factors, AtCPSF100 and AtCPSF160, but not FCA, are stably associated with FY in vivo and form a range of different-sized complexes. A hypomorphic fy allele producing a shorter protein, able to provide some FY functions but unable to interact with FCA, reduces abundance of some of the larger MW complexes. Suppressor mutants, which specifically disrupt the FY motif through which FCA interacts, also lacked these larger complexes. Our data support a model whereby FCA, perhaps after recognition of a specific RNA feature, transiently interacts with FY, an integral component of the canonical RNA 3′ processing machinery, changing the interactions of the different RNA processing components. These altered interactions would appear to be a necessary step in this RNA-mediated chromatin silencing. PMID:19439664

Altered interactions within FY/AtCPSF complexes required for Arabidopsis FCA-mediated chromatin silencing.

PubMed

Manzano, David; Marquardt, Sebastian; Jones, Alexandra M E; Bäurle, Isabel; Liu, Fuquan; Dean, Caroline

2009-05-26

The role of RNA metabolism in chromatin silencing is now widely recognized. We have studied the Arabidopsis RNA-binding protein FCA that down-regulates an endogenous floral repressor gene through a chromatin mechanism involving histone demethylase activity. This mechanism needs FCA to interact with an RNA 3' processing/polyadenylation factor (FY/Pfs2p), but the subsequent events leading to chromatin changes are unknown. Here, we show that this FCA-FY interaction is required for general chromatin silencing roles where hairpin transgenes induce DNA methylation of an endogenous gene. We also show 2 conserved RNA processing factors, AtCPSF100 and AtCPSF160, but not FCA, are stably associated with FY in vivo and form a range of different-sized complexes. A hypomorphic fy allele producing a shorter protein, able to provide some FY functions but unable to interact with FCA, reduces abundance of some of the larger MW complexes. Suppressor mutants, which specifically disrupt the FY motif through which FCA interacts, also lacked these larger complexes. Our data support a model whereby FCA, perhaps after recognition of a specific RNA feature, transiently interacts with FY, an integral component of the canonical RNA 3' processing machinery, changing the interactions of the different RNA processing components. These altered interactions would appear to be a necessary step in this RNA-mediated chromatin silencing.

Probing the human estrogen receptor-α binding requirements for phenolic mono- and di-hydroxyl compounds: A combined synthesis, binding and docking study

PubMed Central

McCullough, Christopher; Neumann, Terrence S.; Gone, Jayapal Reddy; He, Zhengjie; Herrild, Christian; Wondergem, Julie; Pandey, Rajesh K.; Donaldson, William A.; Sem, Daniel S.

2014-01-01

Various estrogen analogs were synthesized and tested for binding to human ERα using a fluorescence polarization displacement assay. Binding affinity and orientation were also predicted using docking calculations. Docking was able to accurately predict relative binding affinity and orientation for estradiol, but only if a tightly bound water molecule bridging Arg394/Glu353 is present. Di-hydroxyl compounds sometimes bind in two orientations, which are flipped in terms of relative positioning of their hydroxyl groups. Di-hydroxyl compounds were predicted to bind with their aliphatic hydroxyl group interacting with His524 in ERα. One nonsteroid-based dihdroxyl compound was 1000-fold specific for ERβ over ERα, and was also 25-fold specific for agonist ERβ versus antagonist activity. Docking predictions suggest this specificity may be due to interaction of the aliphatic hydroxyl with His475 in the agonist form of ERβ, versus with Thr299 in the antagonist form. But, the presence of this aliphatic hydroxyl is not required in all compounds, since mono-hydroxyl (phenolic) compounds bind ERα with high affinity, via hydroxyl hydrogen bonding interactions with the ERα Arg394/Glu353/water triad, and van der Waals interactions with the rest of the molecule. PMID:24315190

A Comparative Typology of Student and Institutional Expectations of Online Faculty

ERIC Educational Resources Information Center

Shaw, Melanie E.; Clowes, Meena C.; Burrus, Scott W. M.

2017-01-01

Online faculty must uphold institutional expectations for their performance. Typically, online institutions have specific guidelines for faculty-to-student interactions; yet, student expectations of faculty may not necessarily align with institutional requirements. This study included a typological analysis of institutional requirements for online…

HyCCAPP as a tool to characterize promoter DNA-protein interactions in Saccharomyces cerevisiae.

PubMed

Guillen-Ahlers, Hector; Rao, Prahlad K; Levenstein, Mark E; Kennedy-Darling, Julia; Perumalla, Danu S; Jadhav, Avinash Y L; Glenn, Jeremy P; Ludwig-Kubinski, Amy; Drigalenko, Eugene; Montoya, Maria J; Göring, Harald H; Anderson, Corianna D; Scalf, Mark; Gildersleeve, Heidi I S; Cole, Regina; Greene, Alexandra M; Oduro, Akua K; Lazarova, Katarina; Cesnik, Anthony J; Barfknecht, Jared; Cirillo, Lisa A; Gasch, Audrey P; Shortreed, Michael R; Smith, Lloyd M; Olivier, Michael

2016-06-01

Currently available methods for interrogating DNA-protein interactions at individual genomic loci have significant limitations, and make it difficult to work with unmodified cells or examine single-copy regions without specific antibodies. In this study, we describe a physiological application of the Hybridization Capture of Chromatin-Associated Proteins for Proteomics (HyCCAPP) methodology we have developed. Both novel and known locus-specific DNA-protein interactions were identified at the ENO2 and GAL1 promoter regions of Saccharomyces cerevisiae, and revealed subgroups of proteins present in significantly different levels at the loci in cells grown on glucose versus galactose as the carbon source. Results were validated using chromatin immunoprecipitation. Overall, our analysis demonstrates that HyCCAPP is an effective and flexible technology that does not require specific antibodies nor prior knowledge of locally occurring DNA-protein interactions and can now be used to identify changes in protein interactions at target regions in the genome in response to physiological challenges. Copyright © 2016 Elsevier Inc. All rights reserved.

Structure and Sequence Analyses of Clustered Protocadherins Reveal Antiparallel Interactions that Mediate Homophilic Specificity.

PubMed

Nicoludis, John M; Lau, Sze-Yi; Schärfe, Charlotta P I; Marks, Debora S; Weihofen, Wilhelm A; Gaudet, Rachelle

2015-11-03

Clustered protocadherin (Pcdh) proteins mediate dendritic self-avoidance in neurons via specific homophilic interactions in their extracellular cadherin (EC) domains. We determined crystal structures of EC1-EC3, containing the homophilic specificity-determining region, of two mouse clustered Pcdh isoforms (PcdhγA1 and PcdhγC3) to investigate the nature of the homophilic interaction. Within the crystal lattices, we observe antiparallel interfaces consistent with a role in trans cell-cell contact. Antiparallel dimerization is supported by evolutionary correlations. Two interfaces, located primarily on EC2-EC3, involve distinctive clustered Pcdh structure and sequence motifs, lack predicted glycosylation sites, and contain residues highly conserved in orthologs but not paralogs, pointing toward their biological significance as homophilic interaction interfaces. These two interfaces are similar yet distinct, reflecting a possible difference in interaction architecture between clustered Pcdh subfamilies. These structures initiate a molecular understanding of clustered Pcdh assemblies that are required to produce functional neuronal networks. Copyright © 2015 Elsevier Ltd. All rights reserved.

The evolution of meaning: spatio-temporal dynamics of visual object recognition.

PubMed

Clarke, Alex; Taylor, Kirsten I; Tyler, Lorraine K

2011-08-01

Research on the spatio-temporal dynamics of visual object recognition suggests a recurrent, interactive model whereby an initial feedforward sweep through the ventral stream to prefrontal cortex is followed by recurrent interactions. However, critical questions remain regarding the factors that mediate the degree of recurrent interactions necessary for meaningful object recognition. The novel prediction we test here is that recurrent interactivity is driven by increasing semantic integration demands as defined by the complexity of semantic information required by the task and driven by the stimuli. To test this prediction, we recorded magnetoencephalography data while participants named living and nonliving objects during two naming tasks. We found that the spatio-temporal dynamics of neural activity were modulated by the level of semantic integration required. Specifically, source reconstructed time courses and phase synchronization measures showed increased recurrent interactions as a function of semantic integration demands. These findings demonstrate that the cortical dynamics of object processing are modulated by the complexity of semantic information required from the visual input.

Non-covalent Interactions with SUMO and Ubiquitin Orchestrate Distinct Functions of the SLX4 Complex in Genome Maintenance

PubMed Central

Ouyang, Jian; Garner, Elizabeth; Hallet, Alexander; Nguyen, Hai Dang; Rickman, Kimberly A.; Gill, Grace; Smogorzewska, Agata; Zou, Lee

2014-01-01

SLX4, a coordinator of multiple DNA structure-specific endonucleases, is important for several DNA repair pathways. Non-covalent interactions of SLX4 with ubiquitin are required for localizing SLX4 to DNA-interstrand crosslinks (ICLs), yet how SLX4 is targeted to other functional contexts remains unclear. Here, we show that SLX4 binds SUMO-2/3 chains via SUMO-interacting motifs (SIMs). The SIMs of SLX4 are dispensable for ICL repair, but important for processing CPT-induced replication intermediates, suppressing fragile site instability, and localizing SLX4 to ALT telomeres. The localization of SLX4 to laser-induced DNA damage also requires the SIMs, as well as DNA-end resection, UBC9 and MDC1. Furthermore, the SUMO binding of SLX4 enhances its interaction with specific DNA-damage sensors or telomere-binding proteins, including RPA, MRE11-RAD50-NBS1 and TRF2. Thus, the interactions of SLX4 with SUMO and ubiquitin increase its affinity for factors recognizing different DNA lesions or telomeres, helping to direct the SLX4 complex in distinct functional contexts. PMID:25533185

A study of interactive control scheduling and economic assessment for robotic systems

NASA Technical Reports Server (NTRS)

1982-01-01

A class of interactive control systems is derived by generalizing interactive manipulator control systems. Tasks of interactive control systems can be represented as a network of a finite set of actions which have specific operational characteristics and specific resource requirements, and which are of limited duration. This has enabled the decomposition of the overall control algorithm simultaneously and asynchronously. The performance benefits of sensor referenced and computer-aided control of manipulators in a complex environment is evaluated. The first phase of the CURV arm control system software development and the basic features of the control algorithms and their software implementation are presented. An optimal solution for a production scheduling problem that will be easy to implement in practical situations is investigated.

Highly specific salt bridges govern bacteriophage P22 icosahedral capsid assembly: identification of the site in coat protein responsible for interaction with scaffolding protein.

PubMed

Cortines, Juliana R; Motwani, Tina; Vyas, Aashay A; Teschke, Carolyn M

2014-05-01

Icosahedral virus assembly requires a series of concerted and highly specific protein-protein interactions to produce a proper capsid. In bacteriophage P22, only coat protein (gp5) and scaffolding protein (gp8) are needed to assemble a procapsid-like particle, both in vivo and in vitro. In scaffolding protein's coat binding domain, residue R293 is required for procapsid assembly, while residue K296 is important but not essential. Here, we investigate the interaction of scaffolding protein with acidic residues in the N-arm of coat protein, since this interaction has been shown to be electrostatic. Through site-directed mutagenesis of genes 5 and 8, we show that changing coat protein N-arm residue 14 from aspartic acid to alanine causes a lethal phenotype. Coat protein residue D14 is shown by cross-linking to interact with scaffolding protein residue R293 and, thus, is intimately involved in proper procapsid assembly. To a lesser extent, coat protein N-arm residue E18 is also implicated in the interaction with scaffolding protein and is involved in capsid size determination, since a cysteine mutation at this site generated petite capsids. The final acidic residue in the N-arm that was tested, E15, is shown to only weakly interact with scaffolding protein's coat binding domain. This work supports growing evidence that surface charge density may be the driving force of virus capsid protein interactions. Bacteriophage P22 infects Salmonella enterica serovar Typhimurium and is a model for icosahedral viral capsid assembly. In this system, coat protein interacts with an internal scaffolding protein, triggering the assembly of an intermediate called a procapsid. Previously, we determined that there is a single amino acid in scaffolding protein required for P22 procapsid assembly, although others modulate affinity. Here, we identify partners in coat protein. We show experimentally that relatively weak interactions between coat and scaffolding proteins are capable of driving correctly shaped and sized procapsids and that the lack of these proper protein-protein interfaces leads to aberrant structures. The present work represents an important contribution supporting the hypothesis that virus capsid assembly is governed by seemingly simple interactions. The highly specific nature of the subunit interfaces suggests that these could be good targets for antivirals.

Long-Term Musical Group Interaction Has a Positive Influence on Empathy in Children

ERIC Educational Resources Information Center

Rabinowitch, Tal-Chen; Cross, Ian; Burnard, Pamela

2013-01-01

Musical group interaction (MGI) is a complex social setting requiring certain cognitive skills that may also elicit shared psychological states. We argue that many MGI-specific features may also be important for emotional empathy, the ability to experience another person's emotional state. We thus hypothesized that long-term repeated participation…

AgRISTARS: Yield model development/soil moisture. Interface control document

NASA Technical Reports Server (NTRS)

1980-01-01

The interactions and support functions required between the crop Yield Model Development (YMD) Project and Soil Moisture (SM) Project are defined. The requirements for YMD support of SM and vice-versa are outlined. Specific tasks in support of these interfaces are defined for development of support functions.

Field Guide for Designing Human Interaction with Intelligent Systems

NASA Technical Reports Server (NTRS)

Malin, Jane T.; Thronesbery, Carroll G.

1998-01-01

The characteristics of this Field Guide approach address the problems of designing innovative software to support user tasks. The requirements for novel software are difficult to specify a priori, because there is not sufficient understanding of how the users' tasks should be supported, and there are not obvious pre-existing design solutions. When the design team is in unfamiliar territory, care must be taken to avoid rushing into detailed design, requirements specification, or implementation of the wrong product. The challenge is to get the right design and requirements in an efficient, cost-effective manner. This document's purpose is to describe the methods we are using to design human interactions with intelligent systems which support Space Shuttle flight controllers in the Mission Control Center at NASA/Johnson Space Center. Although these software systems usually have some intelligent features, the design challenges arise primarily from the innovation needed in the software design. While these methods are tailored to our specific context, they should be extensible, and helpful to designers of human interaction with other types of automated systems. We review the unique features of this context so that you can determine how to apply these methods to your project Throughout this Field Guide, goals of the design methods are discussed. This should help designers understand how a specific method might need to be adapted to the project at hand.

Molecular interactions between chondroitin-dermatan sulfate and growth factors/receptors/matrix proteins.

PubMed

Mizumoto, Shuji; Yamada, Shuhei; Sugahara, Kazuyuki

2015-10-01

Recent functional studies on chondroitin sulfate-dermatan sulfate (CS-DS) demonstrated its indispensable roles in various biological events including brain development and cancer. CS-DS proteoglycans exert their physiological activity through interactions with specific proteins including growth factors, cell surface receptors, and matrix proteins. The characterization of these interactions is essential for regulating the biological functions of CS-DS proteoglycans. Although amino acid sequences on the bioactive proteins required for these interactions have already been elucidated, the specific saccharide sequences involved in the binding of CS-DS to target proteins have not yet been sufficiently identified. In this review, recent findings are described on the interaction between CS-DS and some proteins which are especially involved in the central nervous system and cancer development/metastasis. Copyright © 2015. Published by Elsevier Ltd.

MPQ-cytometry: a magnetism-based method for quantification of nanoparticle-cell interactions

NASA Astrophysics Data System (ADS)

Shipunova, V. O.; Nikitin, M. P.; Nikitin, P. I.; Deyev, S. M.

2016-06-01

Precise quantification of interactions between nanoparticles and living cells is among the imperative tasks for research in nanobiotechnology, nanotoxicology and biomedicine. To meet the challenge, a rapid method called MPQ-cytometry is developed, which measures the integral non-linear response produced by magnetically labeled nanoparticles in a cell sample with an original magnetic particle quantification (MPQ) technique. MPQ-cytometry provides a sensitivity limit 0.33 ng of nanoparticles and is devoid of a background signal present in many label-based assays. Each measurement takes only a few seconds, and no complicated sample preparation or data processing is required. The capabilities of the method have been demonstrated by quantification of interactions of iron oxide nanoparticles with eukaryotic cells. The total amount of targeted nanoparticles that specifically recognized the HER2/neu oncomarker on the human cancer cell surface was successfully measured, the specificity of interaction permitting the detection of HER2/neu positive cells in a cell mixture. Moreover, it has been shown that MPQ-cytometry analysis of a HER2/neu-specific iron oxide nanoparticle interaction with six cell lines of different tissue origins quantitatively reflects the HER2/neu status of the cells. High correlation of MPQ-cytometry data with those obtained by three other commonly used in molecular and cell biology methods supports consideration of this method as a prospective alternative for both quantifying cell-bound nanoparticles and estimating the expression level of cell surface antigens. The proposed method does not require expensive sophisticated equipment or highly skilled personnel and it can be easily applied for rapid diagnostics, especially under field conditions.Precise quantification of interactions between nanoparticles and living cells is among the imperative tasks for research in nanobiotechnology, nanotoxicology and biomedicine. To meet the challenge, a rapid method called MPQ-cytometry is developed, which measures the integral non-linear response produced by magnetically labeled nanoparticles in a cell sample with an original magnetic particle quantification (MPQ) technique. MPQ-cytometry provides a sensitivity limit 0.33 ng of nanoparticles and is devoid of a background signal present in many label-based assays. Each measurement takes only a few seconds, and no complicated sample preparation or data processing is required. The capabilities of the method have been demonstrated by quantification of interactions of iron oxide nanoparticles with eukaryotic cells. The total amount of targeted nanoparticles that specifically recognized the HER2/neu oncomarker on the human cancer cell surface was successfully measured, the specificity of interaction permitting the detection of HER2/neu positive cells in a cell mixture. Moreover, it has been shown that MPQ-cytometry analysis of a HER2/neu-specific iron oxide nanoparticle interaction with six cell lines of different tissue origins quantitatively reflects the HER2/neu status of the cells. High correlation of MPQ-cytometry data with those obtained by three other commonly used in molecular and cell biology methods supports consideration of this method as a prospective alternative for both quantifying cell-bound nanoparticles and estimating the expression level of cell surface antigens. The proposed method does not require expensive sophisticated equipment or highly skilled personnel and it can be easily applied for rapid diagnostics, especially under field conditions. Electronic supplementary information (ESI) available. See DOI: 10.1039/c6nr03507h

Quantifying domain-ligand affinities and specificities by high-throughput holdup assay

PubMed Central

Vincentelli, Renaud; Luck, Katja; Poirson, Juline; Polanowska, Jolanta; Abdat, Julie; Blémont, Marilyne; Turchetto, Jeremy; Iv, François; Ricquier, Kevin; Straub, Marie-Laure; Forster, Anne; Cassonnet, Patricia; Borg, Jean-Paul; Jacob, Yves; Masson, Murielle; Nominé, Yves; Reboul, Jérôme; Wolff, Nicolas; Charbonnier, Sebastian; Travé, Gilles

2015-01-01

Many protein interactions are mediated by small linear motifs interacting specifically with defined families of globular domains. Quantifying the specificity of a motif requires measuring and comparing its binding affinities to all its putative target domains. To this aim, we developed the high-throughput holdup assay, a chromatographic approach that can measure up to a thousand domain-motif equilibrium binding affinities per day. Extracts of overexpressed domains are incubated with peptide-coated resins and subjected to filtration. Binding affinities are deduced from microfluidic capillary electrophoresis of flow-throughs. After benchmarking the approach on 210 PDZ-peptide pairs with known affinities, we determined the affinities of two viral PDZ-binding motifs derived from Human Papillomavirus E6 oncoproteins for 209 PDZ domains covering 79% of the human PDZome. We obtained exquisite sequence-dependent binding profiles, describing quantitatively the PDZome recognition specificity of each motif. This approach, applicable to many categories of domain-ligand interactions, has a wide potential for quantifying the specificities of interactomes. PMID:26053890

Odorant binding protein 69a connects social interaction to modulation of social responsiveness in Drosophila.

PubMed

Bentzur, Assa; Shmueli, Anat; Omesi, Liora; Ryvkin, Julia; Knapp, Jon-Michael; Parnas, Moshe; Davis, Fred P; Shohat-Ophir, Galit

2018-04-01

Living in a social environment requires the ability to respond to specific social stimuli and to incorporate information obtained from prior interactions into future ones. One of the mechanisms that facilitates social interaction is pheromone-based communication. In Drosophila melanogaster, the male-specific pheromone cis-vaccenyl acetate (cVA) elicits different responses in male and female flies, and functions to modulate behavior in a context and experience-dependent manner. Although it is the most studied pheromone in flies, the mechanisms that determine the complexity of the response, its intensity and final output with respect to social context, sex and prior interaction, are still not well understood. Here we explored the functional link between social interaction and pheromone-based communication and discovered an odorant binding protein that links social interaction to sex specific changes in cVA related responses. Odorant binding protein 69a (Obp69a) is expressed in auxiliary cells and secreted into the olfactory sensilla. Its expression is inversely regulated in male and female flies by social interactions: cVA exposure reduces its levels in male flies and increases its levels in female flies. Increasing or decreasing Obp69a levels by genetic means establishes a functional link between Obp69a levels and the extent of male aggression and female receptivity. We show that activation of cVA-sensing neurons is sufficeint to regulate Obp69a levels in the absence of cVA, and requires active neurotransmission between the sensory neuron to the second order olfactory neuron. The cross-talk between sensory neurons and non-neuronal auxiliary cells at the olfactory sensilla, represents an additional component in the machinery that promotes behavioral plasticity to the same sensory stimuli in male and female flies.

Murine Polyomavirus Cell Surface Receptors Activate Distinct Signaling Pathways Required for Infection.

PubMed

O'Hara, Samantha D; Garcea, Robert L

2016-11-01

Virus binding to the cell surface triggers an array of host responses, including activation of specific signaling pathways that facilitate steps in virus entry. Using mouse polyomavirus (MuPyV), we identified host signaling pathways activated upon virus binding to mouse embryonic fibroblasts (MEFs). Pathways activated by MuPyV included the phosphatidylinositol 3-kinase (PI3K), FAK/SRC, and mitogen-activated protein kinase (MAPK) pathways. Gangliosides and α4-integrin are required receptors for MuPyV infection. MuPyV binding to both gangliosides and the α4-integrin receptors was required for activation of the PI3K pathway; however, either receptor interaction alone was sufficient for activation of the MAPK pathway. Using small-molecule inhibitors, we confirmed that the PI3K and FAK/SRC pathways were required for MuPyV infection, while the MAPK pathway was dispensable. Mechanistically, the PI3K pathway was required for MuPyV endocytosis, while the FAK/SRC pathway enabled trafficking of MuPyV along microtubules. Thus, MuPyV interactions with specific cell surface receptors facilitate activation of signaling pathways required for virus entry and trafficking. Understanding how different viruses manipulate cell signaling pathways through interactions with host receptors could lead to the identification of new therapeutic targets for viral infection. Virus binding to cell surface receptors initiates outside-in signaling that leads to virus endocytosis and subsequent virus trafficking. How different viruses manipulate cell signaling through interactions with host receptors remains unclear, and elucidation of the specific receptors and signaling pathways required for virus infection may lead to new therapeutic targets. In this study, we determined that gangliosides and α4-integrin mediate mouse polyomavirus (MuPyV) activation of host signaling pathways. Of these pathways, the PI3K and FAK/SRC pathways were required for MuPyV infection. Both the PI3K and FAK/SRC pathways have been implicated in human diseases, such as heart disease and cancer, and inhibitors directed against these pathways are currently being investigated as therapies. It is possible that these pathways play a role in human PyV infections and could be targeted to inhibit PyV infection in immunosuppressed patients. Copyright © 2016 O’Hara and Garcea.

Field-induced assembly of colloidal ellipsoids into well-defined microtubules

PubMed Central

Crassous, Jérôme J.; Mihut, Adriana M.; Wernersson, Erik; Pfleiderer, Patrick; Vermant, Jan; Linse, Per; Schurtenberger, Peter

2014-01-01

Current theoretical attempts to understand the reversible formation of stable microtubules and virus shells are generally based on shape-specific building blocks or monomers, where the local curvature of the resulting structure is explicitly built-in via the monomer geometry. Here we demonstrate that even simple ellipsoidal colloids can reversibly self-assemble into regular tubular structures when subjected to an alternating electric field. Supported by model calculations, we discuss the combined effects of anisotropic shape and field-induced dipolar interactions on the reversible formation of self-assembled structures. Our observations show that the formation of tubular structures through self-assembly requires much less geometrical and interaction specificity than previously thought, and advance our current understanding of the minimal requirements for self-assembly into regular virus-like structures. PMID:25409686

Social Networking Web Sites: Teaching Appropriate Social Competence to Students with Emotional and Behavioral Disorders

ERIC Educational Resources Information Center

Morgan, Joseph J.

2010-01-01

The Internet has opened a variety of different avenues for people to interact with each other. As new digital environments are developed, new sets of social skills are needed to appropriately interact. Students with emotional and behavioral disorders often have deficits in social competence and require specialized training in specific social…

An fMRI Study of the Social Competition in Healthy Subjects

ERIC Educational Resources Information Center

Polosan, M.; Baciu, M.; Cousin, E.; Perrone, M.; Pichat, C.; Bougerol, T.

2011-01-01

Social interaction requires the ability to infer another person's mental state (Theory of Mind, ToM) and also executive functions. This fMRI study aimed to identify the cerebral correlates activated by ToM during a specific social interaction, the human-human competition. In this framework, we tested a conflict resolution task (Stroop) adapted to…

Protein Kinase A Modulates Transforming Growth Factor-β Signaling through a Direct Interaction with Smad4 Protein*

PubMed Central

Yang, Huibin; Li, Gangyong; Wu, Jing-Jiang; Wang, Lidong; Uhler, Michael; Simeone, Diane M.

2013-01-01

Transforming growth factor β (TGFβ) signaling normally functions to regulate embryonic development and cellular homeostasis. It is increasingly recognized that TGFβ signaling is regulated by cross-talk with other signaling pathways. We previously reported that TGFβ activates protein kinase A (PKA) independent of cAMP through an interaction of an activated Smad3-Smad4 complex and the regulatory subunit of the PKA holoenzyme (PKA-R). Here we define the interaction domains of Smad4 and PKA-R and the functional consequences of this interaction. Using a series of Smad4 and PKA-R truncation mutants, we identified amino acids 290–300 of the Smad4 linker region as critical for the specific interaction of Smad4 and PKA-R. Co-immunoprecipitation assays showed that the B cAMP binding domain of PKA-R was sufficient for interaction with Smad4. Targeting of B domain regions conserved among all PKA-R isoforms and exposed on the molecular surface demonstrated that amino acids 281–285 and 320–329 were required for complex formation with Smad4. Interactions of these specific regions of Smad4 and PKA-R were necessary for TGFβ-mediated increases in PKA activity, CREB (cAMP-response element-binding protein) phosphorylation, induction of p21, and growth inhibition. Moreover, this Smad4-PKA interaction was required for TGFβ-induced epithelial mesenchymal transition, invasion of pancreatic tumor cells, and regulation of tumor growth in vivo. PMID:23362281

Teaching without Faculty: Policy Interactions and Their Effects on the Network of Teaching in German Higher Education

ERIC Educational Resources Information Center

Mitterle, Alexander; Würmann, Carsten; Bloch, Roland

2015-01-01

The impact of higher education reforms on teaching at faculty level in Germany has seldom been explored. Research on teaching at university so far centres on how to teach. Yet, before any (best) practice can take place, teaching requires a specific site where a specific teacher meets a specific number of students. To bring about teaching, teaching…

Declarative language design for interactive visualization.

PubMed

Heer, Jeffrey; Bostock, Michael

2010-01-01

We investigate the design of declarative, domain-specific languages for constructing interactive visualizations. By separating specification from execution, declarative languages can simplify development, enable unobtrusive optimization, and support retargeting across platforms. We describe the design of the Protovis specification language and its implementation within an object-oriented, statically-typed programming language (Java). We demonstrate how to support rich visualizations without requiring a toolkit-specific data model and extend Protovis to enable declarative specification of animated transitions. To support cross-platform deployment, we introduce rendering and event-handling infrastructures decoupled from the runtime platform, letting designers retarget visualization specifications (e.g., from desktop to mobile phone) with reduced effort. We also explore optimizations such as runtime compilation of visualization specifications, parallelized execution, and hardware-accelerated rendering. We present benchmark studies measuring the performance gains provided by these optimizations and compare performance to existing Java-based visualization tools, demonstrating scalability improvements exceeding an order of magnitude.

GIANT 2.0: genome-scale integrated analysis of gene networks in tissues.

PubMed

Wong, Aaron K; Krishnan, Arjun; Troyanskaya, Olga G

2018-05-25

GIANT2 (Genome-wide Integrated Analysis of gene Networks in Tissues) is an interactive web server that enables biomedical researchers to analyze their proteins and pathways of interest and generate hypotheses in the context of genome-scale functional maps of human tissues. The precise actions of genes are frequently dependent on their tissue context, yet direct assay of tissue-specific protein function and interactions remains infeasible in many normal human tissues and cell-types. With GIANT2, researchers can explore predicted tissue-specific functional roles of genes and reveal changes in those roles across tissues, all through interactive multi-network visualizations and analyses. Additionally, the NetWAS approach available through the server uses tissue-specific/cell-type networks predicted by GIANT2 to re-prioritize statistical associations from GWAS studies and identify disease-associated genes. GIANT2 predicts tissue-specific interactions by integrating diverse functional genomics data from now over 61 400 experiments for 283 diverse tissues and cell-types. GIANT2 does not require any registration or installation and is freely available for use at http://giant-v2.princeton.edu.

A history of studies that examine the interactions of Toxoplasma with its host cell: Emphasis on in vitro models.

PubMed

Boyle, Jon P; Radke, Jay R

2009-07-01

This review is a historical look at work carried out over the past 50 years examining interactions of Toxoplasma with the host cell and attempts to focus on some of the seminal experiments in the field. This early work formed the foundation for more recent studies aimed at identifying the host and parasite factors mediating key Toxoplasma-host cell interactions. We focus especially on those studies that were performed in vitro and provide discussions of the following general areas: (i) establishment of the parasitophorous vacuole, (ii) the requirement of specific host cell molecules for parasite replication, (iii) the scenarios under which the host cell can resist parasite replication and/or persistence, (iv) host species-specific and host strain-specific responses to Toxoplasma infection, and (v) Toxoplasma-induced immune modulation.

Demographic and habitat requirements for conservation of bull trout

Treesearch

Bruce E. Rieman; John D. Mclntyre

1993-01-01

Elements in bull trout biology, population dynamics, habitat, and biotic interactions important to conservation of the species are identified. Bull trout appear to have more specific habitat requirements than other salmonids, but no critical thresholds of acceptable habitat condition were found. Size, temporal variation, and spatial distribution are likely to influence...

Host–parasite fluctuating selection in the absence of specificity

PubMed Central

Ashby, Ben; White, Andy; Bowers, Roger; Buckling, Angus; Koskella, Britt

2017-01-01

Fluctuating selection driven by coevolution between hosts and parasites is important for the generation of host and parasite diversity across space and time. Theory has focused primarily on infection genetics, with highly specific ‘matching-allele’ frameworks more likely to generate fluctuating selection dynamics (FSD) than ‘gene-for-gene’ (generalist–specialist) frameworks. However, the environment, ecological feedbacks and life-history characteristics may all play a role in determining when FSD occurs. Here, we develop eco-evolutionary models with explicit ecological dynamics to explore the ecological, epidemiological and host life-history drivers of FSD. Our key result is to demonstrate for the first time, to our knowledge, that specificity between hosts and parasites is not required to generate FSD. Furthermore, highly specific host–parasite interactions produce unstable, less robust stochastic fluctuations in contrast to interactions that lack specificity altogether or those that vary from generalist to specialist, which produce predictable limit cycles. Given the ubiquity of ecological feedbacks and the variation in the nature of specificity in host–parasite interactions, our work emphasizes the underestimated potential for host–parasite coevolution to generate fluctuating selection. PMID:29093222

APOBEC3G Interacts with ssDNA by Two Modes: AFM Studies

NASA Astrophysics Data System (ADS)

Shlyakhtenko, Luda S.; Dutta, Samrat; Banga, Jaspreet; Li, Ming; Harris, Reuben S.; Lyubchenko, Yuri L.

2015-10-01

APOBEC3G (A3G) protein has antiviral activity against HIV and other pathogenic retroviruses. A3G has two domains: a catalytic C-terminal domain (CTD) that deaminates cytidine, and a N-terminal domain (NTD) that binds to ssDNA. Although abundant information exists about the biological activities of A3G protein, the interplay between sequence specific deaminase activity and A3G binding to ssDNA remains controversial. We used the topographic imaging and force spectroscopy modalities of Atomic Force Spectroscopy (AFM) to characterize the interaction of A3G protein with deaminase specific and nonspecific ssDNA substrates. AFM imaging demonstrated that A3G has elevated affinity for deaminase specific ssDNA than for nonspecific ssDNA. AFM force spectroscopy revealed two distinct binding modes by which A3G interacts with ssDNA. One mode requires sequence specificity, as demonstrated by stronger and more stable complexes with deaminase specific ssDNA than with nonspecific ssDNA. Overall these observations enforce prior studies suggesting that both domains of A3G contribute to the sequence specific binding of ssDNA.

APOBEC3G Interacts with ssDNA by Two Modes: AFM Studies.

PubMed

Shlyakhtenko, Luda S; Dutta, Samrat; Banga, Jaspreet; Li, Ming; Harris, Reuben S; Lyubchenko, Yuri L

2015-10-27

APOBEC3G (A3G) protein has antiviral activity against HIV and other pathogenic retroviruses. A3G has two domains: a catalytic C-terminal domain (CTD) that deaminates cytidine, and a N-terminal domain (NTD) that binds to ssDNA. Although abundant information exists about the biological activities of A3G protein, the interplay between sequence specific deaminase activity and A3G binding to ssDNA remains controversial. We used the topographic imaging and force spectroscopy modalities of Atomic Force Spectroscopy (AFM) to characterize the interaction of A3G protein with deaminase specific and nonspecific ssDNA substrates. AFM imaging demonstrated that A3G has elevated affinity for deaminase specific ssDNA than for nonspecific ssDNA. AFM force spectroscopy revealed two distinct binding modes by which A3G interacts with ssDNA. One mode requires sequence specificity, as demonstrated by stronger and more stable complexes with deaminase specific ssDNA than with nonspecific ssDNA. Overall these observations enforce prior studies suggesting that both domains of A3G contribute to the sequence specific binding of ssDNA.

Bacterial discrimination by Dictyostelid amoebae reveals the complexity of ancient interspecies interactions

PubMed Central

Nasser, Waleed; Santhanam, Balaji; Miranda, Edward Roshan; Parikh, Anup; Juneja, Kavina; Rot, Gregor; Dinh, Chris; Chen, Rui; Zupan, Blaz; Shaulsky, Gad; Kuspa, Adam

2014-01-01

Background Amoebae and bacteria interact within predator/prey and host/pathogen relationships, but the general response of amoeba to bacteria is not well understood. The amoeba Dictyostelium discoideum feeds on, and is colonized by diverse bacterial species including Gram-positive [Gram(+)] and Gram-negative [Gram(−)] bacteria, two major groups of bacteria that differ in structure and macromolecular composition. Results Transcriptional profiling of D. discoideum revealed sets of genes whose expression is enriched in amoebae interacting with different species of bacteria, including sets that appear specific to amoebae interacting with Gram(+), or with Gram(−) bacteria. In a genetic screen utilizing the growth of mutant amoebae on a variety of bacteria as a phenotypic readout, we identified amoebal genes that are only required for growth on Gram(+) bacteria, including one that encodes the cell surface protein gp130, as well as several genes that are only required for growth on Gram(−) bacteria including one that encodes a putative lysozyme, AlyL. These genes are required for parts of the transcriptional response of wild-type amoebae, and this allowed their classification into potential response pathways. Conclusions We have defined genes that are critical for amoebal survival during feeding on Gram(+), or Gram(−), bacteria which we propose form part of a regulatory network that allows D. discoideum to elicit specific cellular responses to different species of bacteria in order to optimize survival. PMID:23664307

A pollen-specific calmodulin-binding protein, NPG1, interacts with putative pectate lyases.

PubMed

Shin, Sung-Bong; Golovkin, Maxim; Reddy, Anireddy S N

2014-06-12

Previous genetic studies have revealed that a pollen-specific calmodulin-binding protein, No Pollen Germination 1 (NPG1), is required for pollen germination. However, its mode of action is unknown. Here we report direct interaction of NPG1 with pectate lyase-like proteins (PLLs). A truncated form of AtNPG1 lacking the N-terminal tetratricopeptide repeat 1 (TPR1) failed to interact with PLLs, suggesting that it is essential for NPG1 interaction with PLLs. Localization studies with AtNPG1 fused to a fluorescent reporter driven by its native promoter revealed its presence in the cytosol and cell wall of the pollen grain and the growing pollen tube of plasmolyzed pollen. Together, our data suggest that the function of NPG1 in regulating pollen germination is mediated through its interaction with PLLs, which may modify the pollen cell wall and regulate pollen tube emergence and growth.

Different modes of APC/C activation control growth and neuron-glia interaction in the developing Drosophila eye.

PubMed

Neuert, Helen; Yuva-Aydemir, Yeliz; Silies, Marion; Klämbt, Christian

2017-12-15

The development of the nervous system requires tight control of cell division, fate specification and migration. The anaphase-promoting complex/cyclosome (APC/C) is an E3 ubiquitin ligase that affects different steps of cell cycle progression, as well as having postmitotic functions in nervous system development. It can therefore link different developmental stages in one tissue. The two adaptor proteins, Fizzy/Cdc20 and Fizzy-related/Cdh1, confer APC/C substrate specificity. Here, we show that two distinct modes of APC/C function act during Drosophila eye development. Fizzy/Cdc20 controls the early growth of the eye disc anlage and the concomitant entry of glial cells onto the disc. In contrast, fzr/cdh1 acts during neuronal patterning and photoreceptor axon growth, and subsequently affects neuron-glia interaction. To further address the postmitotic role of Fzr/Cdh1 in controlling neuron-glia interaction, we identified a series of novel APC/C candidate substrates. Four of our candidate genes are required for fzr/cdh1 -dependent neuron-glia interaction, including the dynein light chain Dlc90F Taken together, our data show how different modes of APC/C activation can couple early growth and neuron-glia interaction during eye disc development. © 2017. Published by The Company of Biologists Ltd.

Factorization in large-scale many-body calculations

DOE PAGES

Johnson, Calvin W.; Ormand, W. Erich; Krastev, Plamen G.

2013-08-07

One approach for solving interacting many-fermion systems is the configuration-interaction method, also sometimes called the interacting shell model, where one finds eigenvalues of the Hamiltonian in a many-body basis of Slater determinants (antisymmetrized products of single-particle wavefunctions). The resulting Hamiltonian matrix is typically very sparse, but for large systems the nonzero matrix elements can nonetheless require terabytes or more of storage. An alternate algorithm, applicable to a broad class of systems with symmetry, in our case rotational invariance, is to exactly factorize both the basis and the interaction using additive/multiplicative quantum numbers; such an algorithm recreates the many-body matrix elementsmore » on the fly and can reduce the storage requirements by an order of magnitude or more. Here, we discuss factorization in general and introduce a novel, generalized factorization method, essentially a ‘double-factorization’ which speeds up basis generation and set-up of required arrays. Although we emphasize techniques, we also place factorization in the context of a specific (unpublished) configuration-interaction code, BIGSTICK, which runs both on serial and parallel machines, and discuss the savings in memory due to factorization.« less

Structures of the human Pals1 PDZ domain with and without ligand suggest gated access of Crb to the PDZ peptide-binding groove

PubMed Central

Ivanova, Marina E.; Fletcher, Georgina C.; O’Reilly, Nicola; Purkiss, Andrew G.; Thompson, Barry J.; McDonald, Neil Q.

2015-01-01

Many components of epithelial polarity protein complexes possess PDZ domains that are required for protein interaction and recruitment to the apical plasma membrane. Apical localization of the Crumbs (Crb) transmembrane protein requires a PDZ-mediated interaction with Pals1 (protein-associated with Lin7, Stardust, MPP5), a member of the p55 family of membrane-associated guanylate kinases (MAGUKs). This study describes the molecular interaction between the Crb carboxy-terminal motif (ERLI), which is required for Drosophila cell polarity, and the Pals1 PDZ domain using crystallography and fluorescence polarization. Only the last four Crb residues contribute to Pals1 PDZ-domain binding affinity, with specificity contributed by conserved charged interactions. Comparison of the Crb-bound Pals1 PDZ structure with an apo Pals1 structure reveals a key Phe side chain that gates access to the PDZ peptide-binding groove. Removal of this side chain enhances the binding affinity by more than fivefold, suggesting that access of Crb to Pals1 may be regulated by intradomain contacts or by protein–protein interaction. PMID:25760605

Receptor-Like Kinase RUPO Interacts with Potassium Transporters to Regulate Pollen Tube Growth and Integrity in Rice

PubMed Central

Liu, Lingtong; Zheng, Canhui; Kuang, Baijan; Wei, Liqin; Yan, Longfeng; Wang, Tai

2016-01-01

During sexual reproduction of flowering plants, the pollen tube grows fast and over a long distance within the pistil to deliver two sperms for double fertilization. Growing plant cells need to communicate constantly with external stimuli as well as monitor changes in surface tension of the cell wall and plasma membrane to coordinate these signals and internal growth machinery; however, the underlying mechanisms remain largely unknown. Here we show that the rice member of plant-specific receptor-like kinase CrRLK1Ls subfamily, Ruptured Pollen tube (RUPO), is specifically expressed in rice pollen. RUPO localizes to the apical plasma membrane and vesicle of pollen tubes and is required for male gamete transmission. K+ levels were greater in pollen of homozygous CRISPR-knockout lines than wild-type plants, and pollen tubes burst shortly after germination. We reveal the interaction of RUPO with high-affinity potassium transporters. Phosphorylation of RUPO established and dephosphorylation abolished the interaction. These results have revealed the receptor-like kinase as a regulator of high-affinity potassium transporters via phosphorylation-dependent interaction, and demonstrated a novel receptor-like kinase signaling pathway that mediates K+ homeostasis required for pollen tube growth and integrity. PMID:27447945

Dissection of specific binding of HIV-1 Gag to the 'packaging signal' in viral RNA.

PubMed

Comas-Garcia, Mauricio; Datta, Siddhartha Ak; Baker, Laura; Varma, Rajat; Gudla, Prabhakar R; Rein, Alan

2017-07-20

Selective packaging of HIV-1 genomic RNA (gRNA) requires the presence of a cis -acting RNA element called the 'packaging signal' (Ψ). However, the mechanism by which Ψ promotes selective packaging of the gRNA is not well understood. We used fluorescence correlation spectroscopy and quenching data to monitor the binding of recombinant HIV-1 Gag protein to Cy5-tagged 190-base RNAs. At physiological ionic strength, Gag binds with very similar, nanomolar affinities to both Ψ-containing and control RNAs. We challenged these interactions by adding excess competing tRNA; introducing mutations in Gag; or raising the ionic strength. These modifications all revealed high specificity for Ψ. This specificity is evidently obscured in physiological salt by non-specific, predominantly electrostatic interactions. This nonspecific activity was attenuated by mutations in the MA, CA, and NC domains, including CA mutations disrupting Gag-Gag interaction. We propose that gRNA is selectively packaged because binding to Ψ nucleates virion assembly with particular efficiency.

Revealing the Structural Complexity of Component Interactions of Topic-Specific PCK when Planning to Teach

NASA Astrophysics Data System (ADS)

Mavhunga, Elizabeth

2018-04-01

Teaching pedagogical content knowledge (PCK) at a topic-specific level requires clarity on the content-specific nature of the components employed, as well as the specific features that bring about the desirable depth in teacher explanations. Such understanding is often hazy; yet, it influences the nature of teacher tasks and learning opportunities afforded to pre-service teachers in a teaching program. The purpose of this study was twofold: firstly, to illuminate the emerging complexity when content-specific components of PCK interact when planning to teach a chemistry topic; and secondly, to identify the kinds of teacher tasks that promote the emergence of such complexity. Data collected were content representations (CoRes) in chemical equilibrium accompanied by expanded lesson outlines from 15 pre-service teachers in their final year of study towards a first degree in teaching (B Ed). The analysis involved extraction of episodes that exhibited component interaction by using a qualitative in-depth analysis method. The results revealed the structure in which the components of PCK in a topic interact among each other to be linear, interwoven, or a combination of the two. The interwoven interactions contained multiple components that connected explanations on different aspects of a concept, all working in a complementary manner. The most sophisticated component interactions emerged from teacher tasks on descriptions of a lesson sequence and a summary of a lesson. Recommendations in this study highlight core practices for making pedagogical transformation of topic content knowledge more accessible.

Autographa californica Nucleopolyhedrovirus AC141 (Exon0), a Potential E3 Ubiquitin Ligase, Interacts with Viral Ubiquitin and AC66 To Facilitate Nucleocapsid Egress.

PubMed

Biswas, Siddhartha; Willis, Leslie G; Fang, Minggang; Nie, Yingchao; Theilmann, David A

2018-02-01

During the infection cycle of Autographa californica multiple nucleopolyhedrovirus (AcMNPV), two forms of virions are produced, budded virus (BV) and occlusion-derived virus (ODV). Nucleocapsids that form BV have to egress from the nucleus, whereas nucleocapsids that form ODV remain inside the nucleus. The molecular mechanism that determines whether nucleocapsids remain inside or egress from the nucleus is unknown. AC141 (a predicted E3 ubiquitin ligase) and viral ubiquitin (vUbi) have both been shown to be required for efficient BV production. In this study, it was hypothesized that vUbi interacts with AC141, and in addition, that this interaction was required for BV production. Deletion of both ac141 and vubi restricted viral infection to a single cell, and BV production was completely eliminated. AC141 was ubiquitinated by either vUbi or cellular Ubi, and this interaction was required for optimal BV production. Nucleocapsids in BV, but not ODV, were shown to be specifically ubiquitinated by vUbi, including a 100-kDa protein, as well as high-molecular-weight conjugates. The viral ubiquitinated 100-kDa BV-specific nucleocapsid protein was identified as AC66, which is known to be required for BV production and was shown by coimmunoprecipitation and mass spectrometry to interact with AC141. Confocal microscopy also showed that AC141, AC66, and vUbi interact at the nuclear periphery. These results suggest that ubiquitination of nucleocapsid proteins by vUbi functions as a signal to determine if a nucleocapsid will egress from the nucleus and form BV or remain in the nucleus to form ODV. IMPORTANCE Baculoviruses produce two types of virions called occlusion-derived virus (ODV) and budded virus (BV). ODVs are required for oral infection, whereas BV enables the systemic spread of virus to all host tissues, which is critical for killing insects. One of the important steps for BV production is the export of nucleocapsids out of the nucleus. This study investigated the molecular mechanisms that enable the selection of nucleocapsids for nuclear export instead of being retained within the nucleus, where they would become ODV. Our data show that ubiquitination, a universal cellular process, specifically tags nucleocapsids of BV, but not those found in ODV, using a virus-encoded ubiquitin (vUbi). Therefore, ubiquitination may be the molecular signal that determines if a nucleocapsid is destined to form a BV, thus ensuring lethal infection of the host. © Crown copyright 2018.

Odor-Specific Habituation Arises from Interaction of Afferent Synaptic Adaptation and Intrinsic Synaptic Potentiation in Olfactory Cortex

ERIC Educational Resources Information Center

Linster, Christiane; Menon, Alka V.; Singh, Christopher Y.; Wilson, Donald A.

2009-01-01

Segmentation of target odorants from background odorants is a fundamental computational requirement for the olfactory system and is thought to be behaviorally mediated by olfactory habituation memory. Data from our laboratory have shown that odor-specific adaptation in piriform neurons, mediated at least partially by synaptic adaptation between…

Role of chirality in peptide-induced formation of cholesterol-rich domains

PubMed Central

2005-01-01

The chiral specificity of the interactions of peptides that induce the formation of cholesterol-rich domains has not been extensively investigated. Both the peptide and most lipids are chiral, so there is a possibility that interactions between peptide and lipid could require chiral recognition. On the other hand, in our models with small peptides, the extent of folding of the peptide to form a specific binding pocket is limited. We have determined that replacing cholesterol with its enantiomer, ent-cholesterol, alters the modulation of lipid organization by peptides. The phase-transition properties of SOPC (1-stearoyl-2-oleoylphosphatidylcholine):cholesterol [in a 6:4 ratio with 0.2 mol% PtdIns(4,5)P2] are not significantly altered when ent-cholesterol replaces cholesterol. However, in the presence of 10 mol% of a 19-amino-acid, N-terminally myristoylated fragment (myristoyl-GGKLSKKKKGYNVNDEKAK-amide) of the protein NAP-22 (neuronal axonal membrane protein), the lipid mixture containing cholesterol undergoes separation into cholesterol-rich and cholesterol-depleted domains. This does not occur when ent-cholesterol replaces cholesterol. In another example, when N-acetyl-Leu-Trp-Tyr-Ile-Lys-amide (N-acetyl-LWYIK-amide) is added to SOPC:cholesterol (7:3 ratio), there is a marked increase in the transition enthalpy of the phospholipid, indicating separation of a cholesterol-depleted domain of SOPC. This phenomenon completely disappears when ent-cholesterol replaces cholesterol. The all-D-isomer of N-acetyl-LWYIK-amide also induces the formation of cholesterol-rich domains with natural cholesterol, but does so to a lesser extent with ent-cholesterol. Thus specific peptide chirality is not required for interaction with cholesterol-containing membranes. However, a specific chirality of membrane lipids is required for peptide-induced formation of cholesterol-rich domains. PMID:15929726

Transforming Multidisciplinary Customer Requirements to Product Design Specifications

NASA Astrophysics Data System (ADS)

Ma, Xiao-Jie; Ding, Guo-Fu; Qin, Sheng-Feng; Li, Rong; Yan, Kai-Yin; Xiao, Shou-Ne; Yang, Guang-Wu

2017-09-01

With the increasing of complexity of complex mechatronic products, it is necessary to involve multidisciplinary design teams, thus, the traditional customer requirements modeling for a single discipline team becomes difficult to be applied in a multidisciplinary team and project since team members with various disciplinary backgrounds may have different interpretations of the customers' requirements. A new synthesized multidisciplinary customer requirements modeling method is provided for obtaining and describing the common understanding of customer requirements (CRs) and more importantly transferring them into a detailed and accurate product design specifications (PDS) to interact with different team members effectively. A case study of designing a high speed train verifies the rationality and feasibility of the proposed multidisciplinary requirement modeling method for complex mechatronic product development. This proposed research offersthe instruction to realize the customer-driven personalized customization of complex mechatronic product.

Integrating different perspectives on socialization theory and research: a domain-specific approach.

PubMed

Grusec, Joan E; Davidov, Maayan

2010-01-01

There are several different theoretical and research approaches to the study of socialization, characterized by frequently competing basic tenets and apparently contradictory evidence. As a way of integrating approaches and understanding discrepancies, it is proposed that socialization processes be viewed from a domain perspective, with each domain characterized by a particular form of social interaction between the object and agent of socialization and by specific socialization mechanisms and outcomes. It is argued that this approach requires researchers to identify the domain of social interaction they are investigating, to understand that phenotypically similar behaviors may belong to different domains, and to acknowledge that caregivers who are effective in one type of interaction may not be effective in another.

APID interactomes: providing proteome-based interactomes with controlled quality for multiple species and derived networks

PubMed Central

Alonso-López, Diego; Gutiérrez, Miguel A.; Lopes, Katia P.; Prieto, Carlos; Santamaría, Rodrigo; De Las Rivas, Javier

2016-01-01

APID (Agile Protein Interactomes DataServer) is an interactive web server that provides unified generation and delivery of protein interactomes mapped to their respective proteomes. This resource is a new, fully redesigned server that includes a comprehensive collection of protein interactomes for more than 400 organisms (25 of which include more than 500 interactions) produced by the integration of only experimentally validated protein–protein physical interactions. For each protein–protein interaction (PPI) the server includes currently reported information about its experimental validation to allow selection and filtering at different quality levels. As a whole, it provides easy access to the interactomes from specific species and includes a global uniform compendium of 90,379 distinct proteins and 678,441 singular interactions. APID integrates and unifies PPIs from major primary databases of molecular interactions, from other specific repositories and also from experimentally resolved 3D structures of protein complexes where more than two proteins were identified. For this purpose, a collection of 8,388 structures were analyzed to identify specific PPIs. APID also includes a new graph tool (based on Cytoscape.js) for visualization and interactive analyses of PPI networks. The server does not require registration and it is freely available for use at http://apid.dep.usal.es. PMID:27131791

Specificity of V1-V2 Orientation Networks in the Primate Visual Cortex

PubMed Central

Roe, Anna W.; Ts'o, Daniel Y.

2015-01-01

The computation of texture and shape involves integration of features of various orientations. Orientation networks within V1 tend to involve cells which share similar orientation selectivity. However, emergent properties in V2 require the integration of multiple orientations. We now show that, unlike interactions within V1, V1-V2 orientation interactions are much less synchronized and are not necessarily orientation dependent. We find V1-V2 orientation networks are of two types: a more tightly synchronized, orientation-preserving network and a less synchronized orientation-diverse network. We suggest that such diversity of V1-V2 interactions underlies the spatial and functional integration required for computation of higher order contour and shape in V2. PMID:26314798

EDITORIAL: SPECTROSCOPIC IMAGING

EPA Science Inventory

A foremost goal in biology is understanding the molecular basis of single cell behavior, as well as cell interactions that result in functioning tissues. Accomplishing this goal requires quantitative analysis of multiple, specific macromolecules (e.g. proteins, ligands and enzyme...

Specific binding of a HeLa cell nuclear protein to RNA sequences in the human immunodeficiency virus transactivating region.

PubMed Central

Gaynor, R; Soultanakis, E; Kuwabara, M; Garcia, J; Sigman, D S

1989-01-01

The transactivator protein, tat, encoded by the human immunodeficiency virus is a key regulator of viral transcription. Activation by the tat protein requires sequences downstream of the transcription initiation site called the transactivating region (TAR). RNA derived from the TAR is capable of forming a stable stem-loop structure and the maintenance of both the stem structure and the loop sequences located between +19 and +44 is required for complete in vivo activation by tat. Gel retardation assays with RNA from both wild-type and mutant TAR constructs generated in vitro with SP6 polymerase indicated specific binding of HeLa nuclear proteins to the TAR. To characterize this RNA-protein interaction, a method of chemical "imprinting" has been developed using photoactivated uranyl acetate as the nucleolytic agent. This reagent nicks RNA under physiological conditions at all four nucleotides in a reaction that is independent of sequence and secondary structure. Specific interaction of cellular proteins with TAR RNA could be detected by enhanced cleavages or imprints surrounding the loop region. Mutations that either disrupted stem base-pairing or extensively changed the primary sequence resulted in alterations in the cleavage pattern of the TAR RNA. Structural features of the TAR RNA stem-loop essential for tat activation are also required for specific binding of the HeLa cell nuclear protein. Images PMID:2544877

Dissociation of Paramyxovirus Interferon Evasion Activities: Universal and Virus-Specific Requirements for Conserved V Protein Amino Acids in MDA5 Interference ▿

PubMed Central

Ramachandran, Aparna; Horvath, Curt M.

2010-01-01

The V protein of the paramyxovirus subfamily Paramyxovirinae is an important virulence factor that can interfere with host innate immunity by inactivating the cytosolic pathogen recognition receptor MDA5. This interference is a result of a protein-protein interaction between the highly conserved carboxyl-terminal domain of the V protein and the helicase domain of MDA5. The V protein C-terminal domain (CTD) is an evolutionarily conserved 49- to 68-amino-acid region that coordinates two zinc atoms per protein chain. Site-directed mutagenesis of conserved residues in the V protein CTD has revealed both universal and virus-specific requirements for zinc coordination in MDA5 engagement and has also identified other conserved residues as critical for MDA5 interaction and interference. Mutation of these residues produces V proteins that are specifically defective for MDA5 interference and not impaired in targeting STAT1 for proteasomal degradation via the VDC ubiquitin ligase complex. Results demonstrate that mutation of conserved charged residues in the V proteins of Nipah virus, measles virus, and mumps virus also abolishes MDA5 interaction. These findings clearly define molecular determinants for MDA5 inhibition by the paramyxovirus V proteins. PMID:20719949

The Importance of Water for High Fidelity Information Processing and for Life

NASA Technical Reports Server (NTRS)

Hoehler, Tori M.; Pohorille, Andrew

2011-01-01

Is water an absolute prerequisite for life? Life depends on a variety of non-covalent interactions among molecules, the nature of which is determined as much by the solvent in which they occur as by the molecules themselves. Catalysis and information processing, two essential functions of life, require non-covalent molecular recognition with very high specificity. For example, to correctly reproduce a string consisting of 600,000 units of information (e.g ., 600 kilobases, equivalent to the genome of the smallest free living terrestrial organisms) with a 90% success rate requires specificity > 107 : 1 for the target molecule vs. incorrect alternatives. Such specificity requires (i) that the correct molecular association is energetically stabilized by at least 40 kJ/mol relative to alternatives, and (ii) that the system is able to sample among possible states (alternative molecular associations) rapidly enough to allow the system to fall under thermodynamic control and express the energetic stabilization. We argue that electrostatic interactions are required to confer the necessary energetic stabilization vs. a large library of molecular alternatives, and that a solvent with polarity and dielectric properties comparable to water is required for the system to sample among possible states and express thermodynamic control. Electrostatic associations can be made in non-polar solvents, but the resulting complexes are too stable to be "unmade" with sufficient frequency to confer thermodynamic control on the system. An electrostatic molecular complex representing 3 units of information (e.g., 3 base pairs) with specificity > 107 per unit has a stability in non-polar solvent comparable to that of a carbon-carbon bond at room temperature. These considerations suggest that water, or a solvent with properties very like water, is necessary to support high-fidelity information processing, and can therefore be considered a critical prerequisite for life.

The APP-Interacting Protein FE65 is Required for Hippocampus-Dependent Learning and Long-Term Potentiation

ERIC Educational Resources Information Center

Wang, Yan; Zhang, Ming; Moon, Changjong; Hu, Qubai; Wang, Baiping; Martin, George; Sun, Zhongsheng; Wang, Hongbing

2009-01-01

FE65 is expressed predominantly in the brain and interacts with the C-terminal domain of [beta]-amyloid precursor protein (APP). We examined hippocampus-dependent memory and in vivo long-term potentiation (LTP) at the CA1 synapses with isoform-specific FE65 knockout (p97FE65[superscript -/-]) mice. When examined using the Morris water maze,…

High-affinity DNA-binding Domains of Replication Protein A (RPA) Direct SMARCAL1-dependent Replication Fork Remodeling*

PubMed Central

Bhat, Kamakoti P.; Bétous, Rémy; Cortez, David

2015-01-01

SMARCAL1 catalyzes replication fork remodeling to maintain genome stability. It is recruited to replication forks via an interaction with replication protein A (RPA), the major ssDNA-binding protein in eukaryotic cells. In addition to directing its localization, RPA also activates SMARCAL1 on some fork substrates but inhibits it on others, thereby conferring substrate specificity to SMARCAL1 fork-remodeling reactions. We investigated the mechanism by which RPA regulates SMARCAL1. Our results indicate that although an interaction between SMARCAL1 and RPA is essential for SMARCAL1 activation, the location of the interacting surface on RPA is not. Counterintuitively, high-affinity DNA binding of RPA DNA-binding domain (DBD) A and DBD-B near the fork junction makes it easier for SMARCAL1 to remodel the fork, which requires removing RPA. We also found that RPA DBD-C and DBD-D are not required for SMARCAL1 regulation. Thus, the orientation of the high-affinity RPA DBDs at forks dictates SMARCAL1 substrate specificity. PMID:25552480

High-affinity DNA-binding domains of replication protein A (RPA) direct SMARCAL1-dependent replication fork remodeling.

PubMed

Bhat, Kamakoti P; Bétous, Rémy; Cortez, David

2015-02-13

SMARCAL1 catalyzes replication fork remodeling to maintain genome stability. It is recruited to replication forks via an interaction with replication protein A (RPA), the major ssDNA-binding protein in eukaryotic cells. In addition to directing its localization, RPA also activates SMARCAL1 on some fork substrates but inhibits it on others, thereby conferring substrate specificity to SMARCAL1 fork-remodeling reactions. We investigated the mechanism by which RPA regulates SMARCAL1. Our results indicate that although an interaction between SMARCAL1 and RPA is essential for SMARCAL1 activation, the location of the interacting surface on RPA is not. Counterintuitively, high-affinity DNA binding of RPA DNA-binding domain (DBD) A and DBD-B near the fork junction makes it easier for SMARCAL1 to remodel the fork, which requires removing RPA. We also found that RPA DBD-C and DBD-D are not required for SMARCAL1 regulation. Thus, the orientation of the high-affinity RPA DBDs at forks dictates SMARCAL1 substrate specificity. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Eukaryotic Elongation Factor 1A Interacts with the Upstream Pseudoknot Domain in the 3′ Untranslated Region of Tobacco Mosaic Virus RNA

PubMed Central

Zeenko, Vladimir V.; Ryabova, Lyubov A.; Spirin, Alexander S.; Rothnie, Helen M.; Hess, Daniel; Browning, Karen S.; Hohn, Thomas

2002-01-01

The genomic RNA of tobacco mosaic virus (TMV), like that of other positive-strand RNA viruses, acts as a template for both translation and replication. The highly structured 3′ untranslated region (UTR) of TMV RNAs plays an important role in both processes; it is not polyadenylated but ends with a tRNA-like structure (TLS) preceded by a conserved upstream pseudoknot domain (UPD). The TLS of tobamoviral RNAs can be specifically aminoacylated and, in this state, can interact with eukaryotic elongation factor 1A (eEF1A)/GTP with high affinity. Using a UV cross-linking assay, we detected another specific binding site for eEF1A/GTP, within the UPDs of TMV and crucifer-infecting tobamovirus (crTMV), that does not require aminoacylation. A mutational analysis revealed that UPD pseudoknot conformation and some conserved primary sequence elements are required for this interaction. Its possible role in the regulation of tobamovirus gene expression and replication is discussed. PMID:11991996

Interaction between repressor Opi1p and ER membrane protein Scs2p facilitates transit of phosphatidic acid from the ER to mitochondria and is essential for INO1 gene expression in the presence of choline

PubMed Central

Gaspar, Maria L.; Chang, Yu-Fang; Jesch, Stephen A.; Aregullin, Manuel; Henry, Susan A.

2017-01-01

In the yeast Saccharomyces cerevisiae, the Opi1p repressor controls the expression of INO1 via the Opi1p/Ino2p–Ino4p regulatory circuit. Inositol depletion favors Opi1p interaction with both Scs2p and phosphatidic acid at the endoplasmic reticulum (ER) membrane. Inositol supplementation, however, favors the translocation of Opi1p from the ER into the nucleus, where it interacts with the Ino2p–Ino4p complex, attenuating transcription of INO1. A strain devoid of Scs2p (scs2Δ) and a mutant, OPI1FFAT, lacking the ability to interact with Scs2p were utilized to examine the specific role(s) of the Opi1p–Scs2p interaction in the regulation of INO1 expression and overall lipid metabolism. Loss of the Opi1p–Scs2p interaction reduced INO1 expression and conferred inositol auxotrophy. Moreover, inositol depletion in strains lacking this interaction resulted in Opi1p being localized to sites of lipid droplet formation, coincident with increased synthesis of triacylglycerol. Supplementation of choline to inositol-depleted growth medium led to decreased TAG synthesis in all three strains. However, in strains lacking the Opi1p–Scs2p interaction, Opi1p remained in the nucleus, preventing expression of INO1. These data support the conclusion that a specific pool of phosphatidic acid, associated with lipid droplet formation in the perinuclear ER, is responsible for the initial rapid exit of Opi1p from the nucleus to the ER and is required for INO1 expression in the presence of choline. Moreover, the mitochondria-specific phospholipid, cardiolipin, was significantly reduced in both strains compromised for Opi1p–Scs2p interaction, indicating that this interaction is required for the transfer of phosphatidic acid from the ER to the mitochondria for cardiolipin synthesis. PMID:28924045

Interaction between repressor Opi1p and ER membrane protein Scs2p facilitates transit of phosphatidic acid from the ER to mitochondria and is essential for INO1 gene expression in the presence of choline.

PubMed

Gaspar, Maria L; Chang, Yu-Fang; Jesch, Stephen A; Aregullin, Manuel; Henry, Susan A

2017-11-10

In the yeast Saccharomyces cerevisiae , the Opi1p repressor controls the expression of INO1 via the Opi1p/Ino2p-Ino4p regulatory circuit. Inositol depletion favors Opi1p interaction with both Scs2p and phosphatidic acid at the endoplasmic reticulum (ER) membrane. Inositol supplementation, however, favors the translocation of Opi1p from the ER into the nucleus, where it interacts with the Ino2p-Ino4p complex, attenuating transcription of INO1 A strain devoid of Scs2p ( scs2 Δ) and a mutant, OPI1FFAT , lacking the ability to interact with Scs2p were utilized to examine the specific role(s) of the Opi1p-Scs2p interaction in the regulation of INO1 expression and overall lipid metabolism. Loss of the Opi1p-Scs2p interaction reduced INO1 expression and conferred inositol auxotrophy. Moreover, inositol depletion in strains lacking this interaction resulted in Opi1p being localized to sites of lipid droplet formation, coincident with increased synthesis of triacylglycerol. Supplementation of choline to inositol-depleted growth medium led to decreased TAG synthesis in all three strains. However, in strains lacking the Opi1p-Scs2p interaction, Opi1p remained in the nucleus, preventing expression of INO1 These data support the conclusion that a specific pool of phosphatidic acid, associated with lipid droplet formation in the perinuclear ER, is responsible for the initial rapid exit of Opi1p from the nucleus to the ER and is required for INO1 expression in the presence of choline. Moreover, the mitochondria-specific phospholipid, cardiolipin, was significantly reduced in both strains compromised for Opi1p-Scs2p interaction, indicating that this interaction is required for the transfer of phosphatidic acid from the ER to the mitochondria for cardiolipin synthesis. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

APOBEC3G Interacts with ssDNA by Two Modes: AFM Studies

PubMed Central

Shlyakhtenko, Luda S.; Dutta, Samrat; Banga, Jaspreet; Li, Ming; Harris, Reuben S.; Lyubchenko, Yuri L.

2015-01-01

APOBEC3G (A3G) protein has antiviral activity against HIV and other pathogenic retroviruses. A3G has two domains: a catalytic C-terminal domain (CTD) that deaminates cytidine, and a N-terminal domain (NTD) that binds to ssDNA. Although abundant information exists about the biological activities of A3G protein, the interplay between sequence specific deaminase activity and A3G binding to ssDNA remains controversial. We used the topographic imaging and force spectroscopy modalities of Atomic Force Spectroscopy (AFM) to characterize the interaction of A3G protein with deaminase specific and nonspecific ssDNA substrates. AFM imaging demonstrated that A3G has elevated affinity for deaminase specific ssDNA than for nonspecific ssDNA. AFM force spectroscopy revealed two distinct binding modes by which A3G interacts with ssDNA. One mode requires sequence specificity, as demonstrated by stronger and more stable complexes with deaminase specific ssDNA than with nonspecific ssDNA. Overall these observations enforce prior studies suggesting that both domains of A3G contribute to the sequence specific binding of ssDNA. PMID:26503602

A mammalian germ cell-specific RNA-binding protein interacts with ubiquitously expressed proteins involved in splice site selection

NASA Astrophysics Data System (ADS)

Elliott, David J.; Bourgeois, Cyril F.; Klink, Albrecht; Stévenin, James; Cooke, Howard J.

2000-05-01

RNA-binding motif (RBM) genes are found on all mammalian Y chromosomes and are implicated in spermatogenesis. Within human germ cells, RBM protein shows a similar nuclear distribution to components of the pre-mRNA splicing machinery. To address the function of RBM, we have used protein-protein interaction assays to test for possible physical interactions between these proteins. We find that RBM protein directly interacts with members of the SR family of splicing factors and, in addition, strongly interacts with itself. We have mapped the protein domains responsible for mediating these interactions and expressed the mouse RBM interaction region as a bacterial fusion protein. This fusion protein can pull-down several functionally active SR protein species from cell extracts. Depletion and add-back experiments indicate that these SR proteins are the only splicing factors bound by RBM which are required for the splicing of a panel of pre-mRNAs. Our results suggest that RBM protein is an evolutionarily conserved mammalian splicing regulator which operates as a germ cell-specific cofactor for more ubiquitously expressed pre-mRNA splicing activators.

Plant Virus–Insect Vector Interactions: Current and Potential Future Research Directions

PubMed Central

Dietzgen, Ralf G.; Mann, Krin S.; Johnson, Karyn N.

2016-01-01

Acquisition and transmission by an insect vector is central to the infection cycle of the majority of plant pathogenic viruses. Plant viruses can interact with their insect host in a variety of ways including both non-persistent and circulative transmission; in some cases, the latter involves virus replication in cells of the insect host. Replicating viruses can also elicit both innate and specific defense responses in the insect host. A consistent feature is that the interaction of the virus with its insect host/vector requires specific molecular interactions between virus and host, commonly via proteins. Understanding the interactions between plant viruses and their insect host can underpin approaches to protect plants from infection by interfering with virus uptake and transmission. Here, we provide a perspective focused on identifying novel approaches and research directions to facilitate control of plant viruses by better understanding and targeting virus–insect molecular interactions. We also draw parallels with molecular interactions in insect vectors of animal viruses, and consider technical advances for their control that may be more broadly applicable to plant virus vectors. PMID:27834855

Plant Virus-Insect Vector Interactions: Current and Potential Future Research Directions.

PubMed

Dietzgen, Ralf G; Mann, Krin S; Johnson, Karyn N

2016-11-09

Acquisition and transmission by an insect vector is central to the infection cycle of the majority of plant pathogenic viruses. Plant viruses can interact with their insect host in a variety of ways including both non-persistent and circulative transmission; in some cases, the latter involves virus replication in cells of the insect host. Replicating viruses can also elicit both innate and specific defense responses in the insect host. A consistent feature is that the interaction of the virus with its insect host/vector requires specific molecular interactions between virus and host, commonly via proteins. Understanding the interactions between plant viruses and their insect host can underpin approaches to protect plants from infection by interfering with virus uptake and transmission. Here, we provide a perspective focused on identifying novel approaches and research directions to facilitate control of plant viruses by better understanding and targeting virus-insect molecular interactions. We also draw parallels with molecular interactions in insect vectors of animal viruses, and consider technical advances for their control that may be more broadly applicable to plant virus vectors.

Quantitative analysis of small molecule-nucleic acid interactions with a biosensor surface and surface plasmon resonance detection.

PubMed

Liu, Yang; Wilson, W David

2010-01-01

Surface plasmon resonance (SPR) technology with biosensor surfaces has become a widely-used tool for the study of nucleic acid interactions without any labeling requirements. The method provides simultaneous kinetic and equilibrium characterization of the interactions of biomolecules as well as small molecule-biopolymer binding. SPR monitors molecular interactions in real time and provides significant advantages over optical or calorimetic methods for systems with strong binding coupled to small spectroscopic signals and/or reaction heats. A detailed and practical guide for nucleic acid interaction analysis using SPR-biosensor methods is presented. Details of the SPR technology and basic fundamentals are described with recommendations on the preparation of the SPR instrument, sensor chips, and samples, as well as extensive information on experimental design, quantitative and qualitative data analysis and presentation. A specific example of the interaction of a minor-groove-binding agent with DNA is evaluated by both kinetic and steady-state SPR methods to illustrate the technique. Since the molecules that bind cooperatively to specific DNA sequences are attractive for many applications, a cooperative small molecule-DNA interaction is also presented.

Different domains of the murine RNA polymerase I-specific termination factor mTTF-I serve distinct functions in transcription termination.

PubMed

Evers, R; Smid, A; Rudloff, U; Lottspeich, F; Grummt, I

1995-03-15

Termination of mouse ribosomal gene transcription by RNA polymerase I (Pol I) requires the specific interaction of a DNA binding protein, mTTF-I, with an 18 bp sequence element located downstream of the rRNA coding region. Here we describe the molecular cloning and functional characterization of the cDNA encoding this transcription termination factor. Recombinant mTTF-I binds specifically to the murine terminator elements and terminates Pol I transcription in a reconstituted in vitro system. Deletion analysis has defined a modular structure of mTTF-I comprising a dispensable N-terminal half, a large C-terminal DNA binding region and an internal domain which is required for transcription termination. Significantly, the C-terminal region of mTTF-I reveals striking homology to the DNA binding domains of the proto-oncogene c-Myb and the yeast transcription factor Reb1p. Site-directed mutagenesis of one of the tryptophan residues that is conserved in the homology region of c-Myb, Reb1p and mTTF-I abolishes specific DNA binding, a finding which underscores the functional relevance of these residues in DNA-protein interactions.

Different domains of the murine RNA polymerase I-specific termination factor mTTF-I serve distinct functions in transcription termination.

PubMed Central

Evers, R; Smid, A; Rudloff, U; Lottspeich, F; Grummt, I

1995-01-01

Termination of mouse ribosomal gene transcription by RNA polymerase I (Pol I) requires the specific interaction of a DNA binding protein, mTTF-I, with an 18 bp sequence element located downstream of the rRNA coding region. Here we describe the molecular cloning and functional characterization of the cDNA encoding this transcription termination factor. Recombinant mTTF-I binds specifically to the murine terminator elements and terminates Pol I transcription in a reconstituted in vitro system. Deletion analysis has defined a modular structure of mTTF-I comprising a dispensable N-terminal half, a large C-terminal DNA binding region and an internal domain which is required for transcription termination. Significantly, the C-terminal region of mTTF-I reveals striking homology to the DNA binding domains of the proto-oncogene c-Myb and the yeast transcription factor Reb1p. Site-directed mutagenesis of one of the tryptophan residues that is conserved in the homology region of c-Myb, Reb1p and mTTF-I abolishes specific DNA binding, a finding which underscores the functional relevance of these residues in DNA-protein interactions. Images PMID:7720715

21 CFR 201.56 - Requirements on content and format of labeling for human prescription drug and biological products.

Code of Federal Regulations, 2011 CFR

2011-04-01

... 21 Food and Drugs 4 2011-04-01 2011-04-01 false Requirements on content and format of labeling for human prescription drug and biological products. 201.56 Section 201.56 Food and Drugs FOOD AND DRUG... Contraindications Warnings and Precautions Adverse Reactions Drug Interactions Use in Specific Populations Full...

When action meets emotions: how facial displays of emotion influence goal-related behavior.

PubMed

Ferri, Francesca; Stoianov, Ivilin Peev; Gianelli, Claudia; D'Amico, Luigi; Borghi, Anna M; Gallese, Vittorio

2010-10-01

Many authors have proposed that facial expressions, by conveying emotional states of the person we are interacting with, influence the interaction behavior. We aimed at verifying how specific the effect is of the facial expressions of emotions of an individual (both their valence and relevance/specificity for the purpose of the action) with respect to how the action aimed at the same individual is executed. In addition, we investigated whether and how the effects of emotions on action execution are modulated by participants' empathic attitudes. We used a kinematic approach to analyze the simulation of feeding others, which consisted of recording the "feeding trajectory" by using a computer mouse. Actors could express different highly arousing emotions, namely happiness, disgust, anger, or a neutral expression. Response time was sensitive to the interaction between valence and relevance/specificity of emotion: disgust caused faster response. In addition, happiness induced slower feeding time and longer time to peak velocity, but only in blocks where it alternated with expressions of disgust. The kinematic profiles described how the effect of the specificity of the emotional context for feeding, namely a modulation of accuracy requirements, occurs. An early acceleration in kinematic relative-to-neutral feeding profiles occurred when actors expressed positive emotions (happiness) in blocks with specific-to-feeding negative emotions (disgust). On the other hand, the end-part of the action was slower when feeding happy with respect to neutral faces, confirming the increase of accuracy requirements and motor control. These kinematic effects were modulated by participants' empathic attitudes. In conclusion, the social dimension of emotions, that is, their ability to modulate others' action planning/execution, strictly depends on their relevance and specificity to the purpose of the action. This finding argues against a strict distinction between social and nonsocial emotions.

IE1 and hr facilitate the localization of Bombyx mori nucleopolyhedrovirus ORF8 to specific nuclear sites.

PubMed

Kang, WonKyung; Imai, Noriko; Kawasaki, Yu; Nagamine, Toshihiro; Matsumoto, Shogo

2005-11-01

The Bombyx mori nucleopolyhedrovirus (BmNPV) ORF8 protein has previously been reported to colocalize with IE1 to specific nuclear sites during infection. Transient expression of green fluorescent protein (GFP)-fused ORF8 showed the protein to have cytoplasmic localization, but following BmNPV infection the protein formed foci, suggesting that ORF8 requires some other viral factor(s) for this. Therefore, interacting factors were looked for using the yeast two-hybrid system and IE1 was identified. We mapped the interacting region of ORF8 using a yeast two-hybrid assay. An N-terminal region (residues 1-110) containing a predicted coiled-coil domain interacted with IE1, while a truncated N-terminal region (residues 1-78) that lacks this domain did not. In addition, a protein with a complete deletion of the N-terminal region failed to interact with IE1. These results suggest that the ORF8 N-terminal region containing the coiled-coil domain is required for the interaction with IE1. Next, whether IE1 plays a role in ORF8 localization was investigated. In the presence of IE1, GFP-ORF8 localized to the nucleus. In addition, cotransfection with a plasmid expressing IE1 and a plasmid containing the hr3 element resulted in nuclear foci formation. A GFP-fused ORF8 mutant protein containing the coiled-coil domain, previously shown to interact with IE1, also formed nuclear foci in the presence of IE1 and hr3. However, ORF8 mutant proteins that did not interact with IE1 failed to form nuclear foci. In contrast to wild-type IE1, focus formation was not observed for an IE1 mutant protein that was deficient in hr binding. These results suggest that IE1 and hr facilitate the localization of BmNPV ORF8 to specific nuclear sites.

T cell-B cell interactions in primary immunodeficiencies.

PubMed

Tangye, Stuart G; Deenick, Elissa K; Palendira, Umaimainthan; Ma, Cindy S

2012-02-01

Regulated interactions between cells of the immune system facilitate the generation of successful immune responses, thereby enabling efficient neutralization and clearance of pathogens and the establishment of both cell- and humoral-mediated immunological memory. The corollary of this is that impediments to efficient cell-cell interactions, normally necessary for differentiation and effector functions of immune cells, underly the clinical features and disease pathogenesis of primary immunodeficiencies. In affected individuals, these defects manifest as impaired long-term humoral immunity and susceptibility to infection by specific pathogens. In this review, we discuss the importance of, and requirements for, effective interactions between B cells and T cells during the formation of CD4(+) T follicular helper cells and the elicitation of cytotoxic function of virus-specific CD8(+) T cells, as well as how these processes are abrogated in primary immunodeficiencies due to loss-of-function mutations in defined genes. © 2012 New York Academy of Sciences.

CardioClassifier: disease- and gene-specific computational decision support for clinical genome interpretation.

PubMed

Whiffin, Nicola; Walsh, Roddy; Govind, Risha; Edwards, Matthew; Ahmad, Mian; Zhang, Xiaolei; Tayal, Upasana; Buchan, Rachel; Midwinter, William; Wilk, Alicja E; Najgebauer, Hanna; Francis, Catherine; Wilkinson, Sam; Monk, Thomas; Brett, Laura; O'Regan, Declan P; Prasad, Sanjay K; Morris-Rosendahl, Deborah J; Barton, Paul J R; Edwards, Elizabeth; Ware, James S; Cook, Stuart A

2018-01-25

PurposeInternationally adopted variant interpretation guidelines from the American College of Medical Genetics and Genomics (ACMG) are generic and require disease-specific refinement. Here we developed CardioClassifier (http://www.cardioclassifier.org), a semiautomated decision-support tool for inherited cardiac conditions (ICCs).MethodsCardioClassifier integrates data retrieved from multiple sources with user-input case-specific information, through an interactive interface, to support variant interpretation. Combining disease- and gene-specific knowledge with variant observations in large cohorts of cases and controls, we refined 14 computational ACMG criteria and created three ICC-specific rules.ResultsWe benchmarked CardioClassifier on 57 expertly curated variants and show full retrieval of all computational data, concordantly activating 87.3% of rules. A generic annotation tool identified fewer than half as many clinically actionable variants (64/219 vs. 156/219, Fisher's P = 1.1  ×  10 -18 ), with important false positives, illustrating the critical importance of disease and gene-specific annotations. CardioClassifier identified putatively disease-causing variants in 33.7% of 327 cardiomyopathy cases, comparable with leading ICC laboratories. Through addition of manually curated data, variants found in over 40% of cardiomyopathy cases are fully annotated, without requiring additional user-input data.ConclusionCardioClassifier is an ICC-specific decision-support tool that integrates expertly curated computational annotations with case-specific data to generate fast, reproducible, and interactive variant pathogenicity reports, according to best practice guidelines.GENETICS in MEDICINE advance online publication, 25 January 2018; doi:10.1038/gim.2017.258.

CCL22-specific Antibodies Reveal That Engagement of Two Distinct Binding Domains on CCL22 Is Required for CCR4-mediated Function.

PubMed

Santulli-Marotto, Sandra; Wheeler, John; Lacy, Eilyn R; Boakye, Ken; Luongo, Jennifer; Wu, Sheng-Jiun; Ryan, Mary

2015-12-01

CCL22 inactivation in vivo occurs by cleavage at the N-terminus; however, it is unclear whether this encompasses the entire site of CCR4 interaction. CCL17 also binds CCR4 and its function requires binding via two discrete binding sites. Using monoclonal antibodies (MAbs), we report that there are two separate sites on CCL22 that are required for CCR4-mediated function. The CCL22-specific antibodies bind with affinities of 632 ± 297 pM (MC2B7) and 308 ± 43 pM (MAB4391) and neither exhibited detectable binding to CCL17. Both antibodies are comparable in their ability to inhibit CCL22-mediated calcium mobilization; however, competition binding studies demonstrate that MC2B7 and MAB4391 bind to distinct epitopes on CCL22. Both antibodies inhibit function through CCR4, which is demonstrated by loss of β-arrestin recruitment in a reporter cell line. In both assays, blocking either site independently abolished CCL22 function, suggesting that concurrent engagement of both sites with CCR4 is necessary for function. This is the first demonstration that CCL22 has two distinct binding sites that are required for CCR4 function. These antibodies are valuable tools for better understanding the interaction and function of CCL22 and CCR4 and will potentially help further understanding of the differential outcomes of CCL17 and CCL22 interaction with CCR4.

Human DHEA sulfation requires direct interaction between PAPS synthase 2 and DHEA sulfotransferase SULT2A1.

PubMed

Mueller, Jonathan W; Idkowiak, Jan; Gesteira, Tarsis F; Vallet, Cecilia; Hardman, Rebecca; van den Boom, Johannes; Dhir, Vivek; Knauer, Shirley K; Rosta, Edina; Arlt, Wiebke

2018-06-22

The high-energy sulfate donor 3'-phosphoadenosine-5'-phosphosulfate (PAPS), generated by human PAPS synthase isoforms PAPSS1 and PAPSS2, is required for all human sulfation pathways. Sulfotransferase SULT2A1 uses PAPS for sulfation of the androgen precursor dehydroepiandrosterone (DHEA), thereby reducing downstream activation of DHEA to active androgens. Human PAPSS2 mutations manifest with undetectable DHEA sulfate, androgen excess, and metabolic disease, suggesting that ubiquitous PAPSS1 cannot compensate for deficient PAPSS2 in supporting DHEA sulfation. In knockdown studies in human adrenocortical NCI-H295R1 cells, we found that PAPSS2, but not PAPSS1, is required for efficient DHEA sulfation. Specific APS kinase activity, the rate-limiting step in PAPS biosynthesis, did not differ between PAPSS1 and PAPSS2. Co-expression of cytoplasmic SULT2A1 with a cytoplasmic PAPSS2 variant supported DHEA sulfation more efficiently than co-expression with nuclear PAPSS2 or nuclear/cytosolic PAPSS1. Proximity ligation assays revealed protein-protein interactions between SULT2A1 and PAPSS2 and, to a lesser extent, PAPSS1. Molecular docking studies showed a putative binding site for SULT2A1 within the PAPSS2 APS kinase domain. Energy-dependent scoring of docking solutions identified the interaction as specific for the PAPSS2 and SULT2A1 isoforms. These findings elucidate the mechanistic basis for the selective requirement for PAPSS2 in human DHEA sulfation. © 2018 Mueller et al.

Rule-based modeling: a computational approach for studying biomolecular site dynamics in cell signaling systems

PubMed Central

Chylek, Lily A.; Harris, Leonard A.; Tung, Chang-Shung; Faeder, James R.; Lopez, Carlos F.

2013-01-01

Rule-based modeling was developed to address the limitations of traditional approaches for modeling chemical kinetics in cell signaling systems. These systems consist of multiple interacting biomolecules (e.g., proteins), which themselves consist of multiple parts (e.g., domains, linear motifs, and sites of phosphorylation). Consequently, biomolecules that mediate information processing generally have the potential to interact in multiple ways, with the number of possible complexes and post-translational modification states tending to grow exponentially with the number of binary interactions considered. As a result, only large reaction networks capture all possible consequences of the molecular interactions that occur in a cell signaling system, which is problematic because traditional modeling approaches for chemical kinetics (e.g., ordinary differential equations) require explicit network specification. This problem is circumvented through representation of interactions in terms of local rules. With this approach, network specification is implicit and model specification is concise. Concise representation results in a coarse graining of chemical kinetics, which is introduced because all reactions implied by a rule inherit the rate law associated with that rule. Coarse graining can be appropriate if interactions are modular, and the coarseness of a model can be adjusted as needed. Rules can be specified using specialized model-specification languages, and recently developed tools designed for specification of rule-based models allow one to leverage powerful software engineering capabilities. A rule-based model comprises a set of rules, which can be processed by general-purpose simulation and analysis tools to achieve different objectives (e.g., to perform either a deterministic or stochastic simulation). PMID:24123887

Mapping the distribution of specific antibody interaction forces on individual red blood cells

NASA Astrophysics Data System (ADS)

Yeow, Natasha; Tabor, Rico F.; Garnier, Gil

2017-02-01

Current blood typing methods rely on the agglutination of red blood cells (RBCs) to macroscopically indicate a positive result. An indirect agglutination mechanism is required when blood typing with IgG forms of antibodies. To date, the interaction forces between anti-IgG and IgG antibodies have been poorly quantified, and blood group related antigens have never been quantified with the atomic force microscope (AFM). Instead, the total intensity resulting from fluorescent-tagged antibodies adsorbed on RBC has been measured to calculate an average antigen density on a series of RBCs. In this study we mapped specific antibody interaction forces on the RBC surface. AFM cantilever tips functionalized with anti-IgG were used to probe RBCs incubated with specific IgG antibodies. This work provides unique insight into antibody-antigen interactions in their native cell-bound location, and crucially, on a per-cell basis rather than an ensemble average set of properties. Force profiles obtained from the AFM directly provide not only the anti-IgG - IgG antibody interaction force, but also the spatial distribution and density of antigens over a single cell. This new understanding might be translated into the development of very selective and quantitative interactions that underpin the action of drugs in the treatment of frontier illnesses.

DASMI: exchanging, annotating and assessing molecular interaction data.

PubMed

Blankenburg, Hagen; Finn, Robert D; Prlić, Andreas; Jenkinson, Andrew M; Ramírez, Fidel; Emig, Dorothea; Schelhorn, Sven-Eric; Büch, Joachim; Lengauer, Thomas; Albrecht, Mario

2009-05-15

Ever increasing amounts of biological interaction data are being accumulated worldwide, but they are currently not readily accessible to the biologist at a single site. New techniques are required for retrieving, sharing and presenting data spread over the Internet. We introduce the DASMI system for the dynamic exchange, annotation and assessment of molecular interaction data. DASMI is based on the widely used Distributed Annotation System (DAS) and consists of a data exchange specification, web servers for providing the interaction data and clients for data integration and visualization. The decentralized architecture of DASMI affords the online retrieval of the most recent data from distributed sources and databases. DASMI can also be extended easily by adding new data sources and clients. We describe all DASMI components and demonstrate their use for protein and domain interactions. The DASMI tools are available at http://www.dasmi.de/ and http://ipfam.sanger.ac.uk/graph. The DAS registry and the DAS 1.53E specification is found at http://www.dasregistry.org/.

Gaussian Processes for Data-Efficient Learning in Robotics and Control.

PubMed

Deisenroth, Marc Peter; Fox, Dieter; Rasmussen, Carl Edward

2015-02-01

Autonomous learning has been a promising direction in control and robotics for more than a decade since data-driven learning allows to reduce the amount of engineering knowledge, which is otherwise required. However, autonomous reinforcement learning (RL) approaches typically require many interactions with the system to learn controllers, which is a practical limitation in real systems, such as robots, where many interactions can be impractical and time consuming. To address this problem, current learning approaches typically require task-specific knowledge in form of expert demonstrations, realistic simulators, pre-shaped policies, or specific knowledge about the underlying dynamics. In this paper, we follow a different approach and speed up learning by extracting more information from data. In particular, we learn a probabilistic, non-parametric Gaussian process transition model of the system. By explicitly incorporating model uncertainty into long-term planning and controller learning our approach reduces the effects of model errors, a key problem in model-based learning. Compared to state-of-the art RL our model-based policy search method achieves an unprecedented speed of learning. We demonstrate its applicability to autonomous learning in real robot and control tasks.

15 CFR 27.102 - Definitions.

Code of Federal Regulations, 2011 CFR

2011-01-01

... specific responsibility for regulating as a research activity, (for example, Investigational New Drug requirements administered by the Food and Drug Administration). It does not include research activities which...) conducting research obtains (1) Data through intervention or interaction with the individual, or (2...

28 CFR 46.102 - Definitions.

Code of Federal Regulations, 2011 CFR

2011-07-01

... specific responsibility for regulating as a research activity, (for example, Investigational New Drug requirements administered by the Food and Drug Administration). It does not include research activities which...) conducting research obtains (1) Data through intervention or interaction with the individual, or (2...

16 CFR 1028.102 - Definitions.

Code of Federal Regulations, 2011 CFR

2011-01-01

... specific responsibility for regulating as a research activity, (for example, Investigational New Drug requirements administered by the Food and Drug Administration). It does not include research activities which...) conducting research obtains: (1) Data through intervention or interaction with the individual, or (2...

28 CFR 46.102 - Definitions.

Code of Federal Regulations, 2010 CFR

2010-07-01

... specific responsibility for regulating as a research activity, (for example, Investigational New Drug requirements administered by the Food and Drug Administration). It does not include research activities which...) conducting research obtains (1) Data through intervention or interaction with the individual, or (2...

14 CFR 1230.102 - Definitions.

Code of Federal Regulations, 2010 CFR

2010-01-01

... specific responsibility for regulating as a research activity, (for example, Investigational New Drug requirements administered by the Food and Drug Administration). It does not include research activities which...) conducting research obtains (1) Data through intervention or interaction with the individual, or (2...

14 CFR 1230.102 - Definitions.

Code of Federal Regulations, 2011 CFR

2011-01-01

... specific responsibility for regulating as a research activity, (for example, Investigational New Drug requirements administered by the Food and Drug Administration). It does not include research activities which...) conducting research obtains (1) Data through intervention or interaction with the individual, or (2...

40 CFR 26.102 - Definitions.

Code of Federal Regulations, 2011 CFR

2011-07-01

... specific responsibility for regulating as a research activity, (for example, Investigational New Drug requirements administered by the Food and Drug Administration). It does not include research activities which...) conducting research obtains (1) Data through intervention or interaction with the individual, or (2...

15 CFR 27.102 - Definitions.

Code of Federal Regulations, 2010 CFR

2010-01-01

... specific responsibility for regulating as a research activity, (for example, Investigational New Drug requirements administered by the Food and Drug Administration). It does not include research activities which...) conducting research obtains (1) Data through intervention or interaction with the individual, or (2...

34 CFR 97.102 - Definitions.

Code of Federal Regulations, 2011 CFR

2011-07-01

... specific responsibility for regulating as a research activity, (for example, Investigational New Drug requirements administered by the Food and Drug Administration). It does not include research activities which...) conducting research obtains (1) Data through intervention or interaction with the individual, or (2...

40 CFR 26.102 - Definitions.

Code of Federal Regulations, 2010 CFR

2010-07-01

... specific responsibility for regulating as a research activity, (for example, Investigational New Drug requirements administered by the Food and Drug Administration). It does not include research activities which...) conducting research obtains (1) Data through intervention or interaction with the individual, or (2...

34 CFR 97.102 - Definitions.

Code of Federal Regulations, 2010 CFR

2010-07-01

... specific responsibility for regulating as a research activity, (for example, Investigational New Drug requirements administered by the Food and Drug Administration). It does not include research activities which...) conducting research obtains (1) Data through intervention or interaction with the individual, or (2...

Birth/death process model

NASA Technical Reports Server (NTRS)

Solloway, C. B.; Wakeland, W.

1976-01-01

First-order Markov model developed on digital computer for population with specific characteristics. System is user interactive, self-documenting, and does not require user to have complete understanding of underlying model details. Contains thorough error-checking algorithms on input and default capabilities.

Aeronautical Systems Division (AFSC) Program Management Resource Document.

DTIC Science & Technology

1981-09-30

assure that the detail desing adeuately satisfies the requirements contained in the Part I Development Specifications and to allow the PM to formally...disciplines. He becomes a source of integrated information concerning a particular program and an interaction point for coordinating the di- verse...consider cost, schedule, and technical factors not only individually, but also their interaction on each other (5 60). The fact that a program has

Social Experience Is Sufficient to Modulate Sleep Need of Drosophila without Increasing Wakefulness.

PubMed

Lone, Shahnaz Rahman; Potdar, Sheetal; Srivastava, Manishi; Sharma, Vijay Kumar

2016-01-01

Organisms quickly learn about their surroundings and display synaptic plasticity which is thought to be critical for their survival. For example, fruit flies Drosophila melanogaster exposed to highly enriched social environment are found to show increased synaptic connections and a corresponding increase in sleep. Here we asked if social environment comprising a pair of same-sex individuals could enhance sleep in the participating individuals. To study this, we maintained individuals of D. melanogaster in same-sex pairs for a period of 1 to 4 days, and after separation, monitored sleep of the previously socialized and solitary individuals under similar conditions. Males maintained in pairs for 3 or more days were found to sleep significantly more during daytime and showed a tendency to fall asleep sooner as compared to solitary controls (both measures together are henceforth referred to as "sleep-enhancement"). This sleep phenotype is not strain-specific as it is observed in males from three different "wild type" strains of D. melanogaster. Previous studies on social interaction mediated sleep-enhancement presumed 'waking experience' during the interaction to be the primary underlying cause; however, we found sleep-enhancement to occur without any significant increase in wakefulness. Furthermore, while sleep-enhancement due to group-wise social interaction requires Pigment Dispersing Factor (PDF) positive neurons; PDF positive and CRYPTOCHROME (CRY) positive circadian clock neurons and the core circadian clock genes are not required for sleep-enhancement to occur when males interact in pairs. Pair-wise social interaction mediated sleep-enhancement requires dopamine and olfactory signaling, while visual and gustatory signaling systems seem to be dispensable. These results suggest that socialization alone (without any change in wakefulness) is sufficient to cause sleep-enhancement in fruit fly D. melanogaster males, and that its neuronal control is context-specific.

Real-Time Interactive Facilities Associated With A 3-D Medical Workstation

NASA Astrophysics Data System (ADS)

Goldwasser, S. M.; Reynolds, R. A.; Talton, D.; Walsh, E.

1986-06-01

Biomedical workstations of the future will incorporate three-dimensional interactive capabilities which provide real-time response to most common operator requests. Such systems will find application in many areas of medicine including clinical diagnosis, surgical and radiation therapy planning, biomedical research based on functional imaging, and medical education. This paper considers the requirements of these future systems in terms of image quality, performance, and the interactive environment, and examines the relationship of workstation capabilities to specific medical applications. We describe a prototype physician's workstation that we have designed and built to meet many of these requirements (using conventional graphics technology in conjunction with a custom real-time 3-D processor), and give an account of the remaining issues and challenges that future designers of such systems will have to address.

Ames interactive molecular model building system - A 3-D computer modelling system applied to the study of the origin of life

NASA Technical Reports Server (NTRS)

Coeckelenbergh, Y.; Macelroy, R. D.; Rein, R.

1978-01-01

The investigation of specific interactions among biological molecules must take into consideration the stereochemistry of the structures. Thus, models of the molecules are essential for describing the spatial organization of potentially interacting groups, and estimations of conformation are required for a description of spatial organization. Both the function of visualizing molecules, and that of estimating conformation through calculations of energy, are part of the molecular modeling system described in the present paper. The potential uses of the system in investigating some aspects of the origin of life rest on the assumption that translation of conformation from genetic elements to catalytic elements would have been required for the development of the first replicating systems subject to the process of biological evolution.

A Traveling Wave Ion Mobility Spectrometry (TWIMS) Study of the Robo1-Heparan Sulfate Interaction

NASA Astrophysics Data System (ADS)

Zhao, Yuejie; Yang, Jeong Yeh; Thieker, David F.; Xu, Yongmei; Zong, Chengli; Boons, Geert-Jan; Liu, Jian; Woods, Robert J.; Moremen, Kelley W.; Amster, I. Jonathan

2018-03-01

Roundabout 1 (Robo1) interacts with its receptor Slit to regulate axon guidance, axon branching, and dendritic development in the nervous system and to regulate morphogenesis and many cell functions in the nonneuronal tissues. This interaction is known to be critically regulated by heparan sulfate (HS). Previous studies suggest that HS is required to promote the binding of Robo1 to Slit to form the minimal signaling complex, but the molecular details and the structural requirements of HS for this interaction are still unclear. Here, we describe the application of traveling wave ion mobility spectrometry (TWIMS) to study the conformational details of the Robo1-HS interaction. The results suggest that Robo1 exists in two conformations that differ by their compactness and capability to interact with HS. The results also suggest that the highly flexible interdomain hinge region connecting the Ig1 and Ig2 domains of Robo1 plays an important functional role in promoting the Robo1-Slit interaction. Moreover, variations in the sulfation pattern and size of HS were found to affect its binding affinity and selectivity to interact with different conformations of Robo1. Both MS measurements and CIU experiments show that the Robo1-HS interaction requires the presence of a specific size and pattern of modification of HS. Furthermore, the effect of N-glycosylation on the conformation of Robo1 and its binding modes with HS is reported. [Figure not available: see fulltext.

A Sequence in the loop domain of hepatitis C virus E2 protein identified in silico as crucial for the selective binding to human CD81

PubMed Central

Chang, Chun-Chun; Hsu, Hao-Jen; Yen, Jui-Hung; Lo, Shih-Yen

2017-01-01

Hepatitis C virus (HCV) is a species-specific pathogenic virus that infects only humans and chimpanzees. Previous studies have indicated that interactions between the HCV E2 protein and CD81 on host cells are required for HCV infection. To determine the crucial factors for species-specific interactions at the molecular level, this study employed in silico molecular docking involving molecular dynamic simulations of the binding of HCV E2 onto human and rat CD81s. In vitro experiments including surface plasmon resonance measurements and cellular binding assays were applied for simple validations of the in silico results. The in silico studies identified two binding regions on the HCV E2 loop domain, namely E2-site1 and E2-site2, as being crucial for the interactions with CD81s, with the E2-site2 as the determinant factor for human-specific binding. Free energy calculations indicated that the E2/CD81 binding process might follow a two-step model involving (i) the electrostatic interaction-driven initial binding of human-specific E2-site2, followed by (ii) changes in the E2 orientation to facilitate the hydrophobic and van der Waals interaction-driven binding of E2-site1. The sequence of the human-specific, stronger-binding E2-site2 could serve as a candidate template for the future development of HCV-inhibiting peptide drugs. PMID:28481946

Gia/Mthl5 is an aorta specific GPCR required for Drosophila heart tube morphology and normal pericardial cell positioning.

PubMed

Patel, Meghna V; Zhu, Jun-Yi; Jiang, Zhiping; Richman, Adam; VanBerkum, Mark F A; Han, Zhe

2016-06-01

G-protein signaling is known to be required for cell-cell contacts during the development of the Drosophila dorsal vessel. However, the identity of the G protein-coupled receptor (GPCR) that regulates this signaling pathway activity is unknown. Here we describe the identification of a novel cardiac specific GPCR, called Gia, for "GPCR in aorta". Gia is the only heart-specific GPCR identified in Drosophila to date and it is specifically expressed in cardioblasts that fuse at the dorsal midline to become the aorta. Gia is the only Drosophila gene so far identified for which expression is entirely restricted to cells of the aorta. Deletion of Gia led to a broken-hearted phenotype, characterized by pericardial cells dissociated from cardioblasts and abnormal distribution of cell junction proteins. Both phenotypes were similar to those observed in mutants of the heterotrimeric cardiac G proteins. Lack of Gia also led to defects in the alignment and fusion of cardioblasts in the aorta. Gia forms a protein complex with G-αo47A, the alpha subunit of the heterotrimeric cardiac G proteins and interacts genetically with G-αo47A during cardiac morphogenesis. Our study identified Gia as an essential aorta-specific GPCR that functions upstream of cardiac heterotrimeric G proteins and is required for morphological integrity of the aorta during heart tube formation. These studies lead to a redefinition of the bro phenotype, to encompass morphological integrity of the heart tube as well as cardioblast-pericardial cell spatial interactions. Copyright © 2016 Elsevier Inc. All rights reserved.

Selective layer-free blood serum ionogram based on ion-specific interactions with a nanotransistor

NASA Astrophysics Data System (ADS)

Sivakumarasamy, R.; Hartkamp, R.; Siboulet, B.; Dufrêche, J.-F.; Nishiguchi, K.; Fujiwara, A.; Clément, N.

2018-05-01

Despite being ubiquitous in the fields of chemistry and biology, the ion-specific effects of electrolytes pose major challenges for researchers. A lack of understanding about ion-specific surface interactions has hampered the development and application of materials for (bio-)chemical sensor applications. Here, we show that scaling a silicon nanotransistor sensor down to 25 nm provides a unique opportunity to understand and exploit ion-specific surface interactions, yielding a surface that is highly sensitive to cations and inert to pH. The unprecedented sensitivity of these devices to Na+ and divalent ions can be attributed to an overscreening effect via molecular dynamics. The surface potential of multi-ion solutions is well described by the sum of the electrochemical potentials of each cation, enabling selective measurements of a target ion concentration without requiring a selective organic layer. We use these features to construct a blood serum ionogram for Na+, K+, Ca2+ and Mg2+, in an important step towards the development of a versatile, durable and mobile chemical or blood diagnostic tool.

Direct interaction of the Golgi V-ATPase a-subunit isoform with PI(4)P drives localization of Golgi V-ATPases in yeast.

PubMed

Banerjee, Subhrajit; Kane, Patricia M

2017-09-15

Luminal pH and phosphoinositide content are fundamental features of organelle identity. Vacuolar H + -ATPases (V-ATPases) drive organelle acidification in all eukaryotes, and membrane-bound a-subunit isoforms of the V-ATPase are implicated in organelle-specific targeting and regulation. Earlier work demonstrated that the endolysosomal lipid PI(3,5)P 2 activates V-ATPases containing the vacuolar a-subunit isoform in Saccharomyces cerevisiae Here we demonstrate that PI(4)P, the predominant Golgi phosphatidylinositol (PI) species, directly interacts with the cytosolic amino terminal (NT) domain of the yeast Golgi V-ATPase a-isoform Stv1. Lysine-84 of Stv1NT is essential for interaction with PI(4)P in vitro and in vivo, and interaction with PI(4)P is required for efficient localization of Stv1-containing V-ATPases. The cytosolic NT domain of the human V-ATPase a2 isoform specifically interacts with PI(4)P in vitro, consistent with its Golgi localization and function. We propose that NT domains of V o a-subunit isoforms interact specifically with PI lipids in their organelles of residence. These interactions can transmit organelle-specific targeting or regulation information to V-ATPases. © 2017 Banerjee and Kane. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

Planetary mission requirements, technology and design considerations for a solar electric propulsion stage

NASA Technical Reports Server (NTRS)

Cork, M. J.; Hastrup, R. C.; Menard, W. A.; Olson, R. N.

1979-01-01

High energy planetary missions such as comet rendezvous, Saturn orbiter and asteroid rendezvous require development of a Solar Electric Propulsion Stage (SEPS) for augmentation of the Shuttle-IUS. Performance and functional requirements placed on the SEPS are presented. These requirements will be used in evolution of the SEPS design, which must be highly interactive with both the spacecraft and the mission design. Previous design studies have identified critical SEPS technology areas and some specific design solutions which are also presented in the paper.

Governing & Foundation Boards: Fostering Successful Relationships.

ERIC Educational Resources Information Center

Legon, Richard D.

1996-01-01

Public colleges and universities increasingly raise private funds through institutional foundations. A healthy relationship between governing and foundation boards requires open communication, regular interaction, and shared information. The fulcrum of this relationship is the college's president, who has specific responsibilities toward each…

An Exquisitely Specific PDZ/Target Recognition Revealed by the Structure of INAD PDZ3 in Complex with TRP Channel Tail.

PubMed

Ye, Fei; Liu, Wei; Shang, Yuan; Zhang, Mingjie

2016-03-01

The vast majority of PDZ domains are known to bind to a few C-terminal tail residues of target proteins with modest binding affinities and specificities. Such promiscuous PDZ/target interactions are not compatible with highly specific physiological functions of PDZ domain proteins and their targets. Here, we report an unexpected PDZ/target binding occurring between the scaffold protein inactivation no afterpotential D (INAD) and transient receptor potential (TRP) channel in Drosophila photoreceptors. The C-terminal 15 residues of TRP are required for the specific interaction with INAD PDZ3. The INAD PDZ3/TRP peptide complex structure reveals that only the extreme C-terminal Leu of TRP binds to the canonical αB/βB groove of INAD PDZ3. The rest of the TRP peptide, by forming a β hairpin structure, binds to a surface away from the αB/βB groove of PDZ3 and contributes to the majority of the binding energy. Thus, the INAD PDZ3/TRP channel interaction is exquisitely specific and represents a new mode of PDZ/target recognitions. Copyright © 2016 Elsevier Ltd. All rights reserved.

Caenorhabditis elegans fibroblast growth factor receptor signaling can occur independently of the multi-substrate adaptor FRS2.

PubMed

Lo, Te-Wen; Bennett, Daniel C; Goodman, S Jay; Stern, Michael J

2010-06-01

The components of receptor tyrosine kinase signaling complexes help to define the specificity of the effects of their activation. The Caenorhabditis elegans fibroblast growth factor receptor (FGFR), EGL-15, regulates a number of processes, including sex myoblast (SM) migration guidance and fluid homeostasis, both of which require a Grb2/Sos/Ras cassette of signaling components. Here we show that SEM-5/Grb2 can bind directly to EGL-15 to mediate SM chemoattraction. A yeast two-hybrid screen identified SEM-5 as able to interact with the carboxy-terminal domain (CTD) of EGL-15, a domain that is specifically required for SM chemoattraction. This interaction requires the SEM-5 SH2-binding motifs present in the CTD (Y(1009) and Y(1087)), and these sites are required for the CTD role of EGL-15 in SM chemoattraction. SEM-5, but not the SEM-5 binding sites located in the CTD, is required for the fluid homeostasis function of EGL-15, indicating that SEM-5 can link to EGL-15 through an alternative mechanism. The multi-substrate adaptor protein FRS2 serves to link vertebrate FGFRs to Grb2. In C. elegans, an FRS2-like gene, rog-1, functions upstream of a Ras/MAPK pathway for oocyte maturation but is not required for EGL-15 function. Thus, unlike the vertebrate FGFRs, which require the multi-substrate adaptor FRS2 to recruit Grb2, EGL-15 can recruit SEM-5/Grb2 directly.

Structure of the Human FANCL RING-Ube2T Complex Reveals Determinants of Cognate E3-E2 Selection

PubMed Central

Hodson, Charlotte; Purkiss, Andrew; Miles, Jennifer Anne; Walden, Helen

2014-01-01

Summary The combination of an E2 ubiquitin-conjugating enzyme with an E3 ubiquitin-ligase is essential for ubiquitin modification of a substrate. Moreover, the pairing dictates both the substrate choice and the modification type. The molecular details of generic E3-E2 interactions are well established. Nevertheless, the determinants of selective, specific E3-E2 recognition are not understood. There are ∼40 E2s and ∼600 E3s giving rise to a possible ∼24,000 E3-E2 pairs. Using the Fanconi Anemia pathway exclusive E3-E2 pair, FANCL-Ube2T, we report the atomic structure of the FANCL RING-Ube2T complex, revealing a specific and extensive network of additional electrostatic and hydrophobic interactions. Furthermore, we show that these specific interactions are required for selection of Ube2T over other E2s by FANCL. PMID:24389026

Specialized rules of gene transcription in male germ cells: the CREM paradigm.

PubMed

Monaco, Lucia; Kotaja, Noora; Fienga, Giulia; Hogeveen, Kevin; Kolthur, Ullas S; Kimmins, Sarah; Brancorsini, Stefano; Macho, Betina; Sassone-Corsi, Paolo

2004-12-01

Specialized transcription complexes that coordinate the differentiation programme of spermatogenesis have been found in germ cells, which display specific differences in the components of the general transcription machinery. The TATA-binding protein family and its associated cofactors, for example, show upregulated expression in testis. In this physiological context, transcriptional control mediated by the activator cAMP response element modulator (CREM) represents an established paradigm. Somatic cell activation by CREM requires its phosphorylation at a unique regulatory site (Ser117) and subsequent interaction with the ubiquitous coactivator CREB-binding protein. In testis, CREM transcriptional activity is controlled through interaction with a tissue-specific partner, activator of CREM in the testis (ACT), which confers a powerful, phosphorylation-independent activation capacity. The function of ACT was found to be regulated by the testis-specific kinesin KIF17b. Here we discuss some aspects of the testis-specific transcription machinery, whose function is essential for the process of spermatogenesis.

Editing of EIA coded, numerically controlled, machine tool tapes

NASA Technical Reports Server (NTRS)

Weiner, J. M.

1975-01-01

Editing of numerically controlled (N/C) machine tool tapes (8-level paper tape) using an interactive graphic display processor is described. A rapid technique required for correcting production errors in N/C tapes was developed using the interactive text editor on the IMLAC PDS-ID graphic display system and two special programs resident on disk. The correction technique and special programs for processing N/C tapes coded to EIA specifications are discussed.

Web-based interactive visualization in a Grid-enabled neuroimaging application using HTML5.

PubMed

Siewert, René; Specovius, Svenja; Wu, Jie; Krefting, Dagmar

2012-01-01

Interactive visualization and correction of intermediate results are required in many medical image analysis pipelines. To allow certain interaction in the remote execution of compute- and data-intensive applications, new features of HTML5 are used. They allow for transparent integration of user interaction into Grid- or Cloud-enabled scientific workflows. Both 2D and 3D visualization and data manipulation can be performed through a scientific gateway without the need to install specific software or web browser plugins. The possibilities of web-based visualization are presented along the FreeSurfer-pipeline, a popular compute- and data-intensive software tool for quantitative neuroimaging.

Learning of embodied interaction dynamics with recurrent neural networks: some exploratory experiments.

PubMed

Oubbati, Mohamed; Kord, Bahram; Koprinkova-Hristova, Petia; Palm, Günther

2014-04-01

The new tendency of artificial intelligence suggests that intelligence must be seen as a result of the interaction between brains, bodies and environments. This view implies that designing sophisticated behaviour requires a primary focus on how agents are functionally coupled to their environments. Under this perspective, we present early results with the application of reservoir computing as an efficient tool to understand how behaviour emerges from interaction. Specifically, we present reservoir computing models, that are inspired by imitation learning designs, to extract the essential components of behaviour that results from agent-environment interaction dynamics. Experimental results using a mobile robot are reported to validate the learning architectures.

Learning of embodied interaction dynamics with recurrent neural networks: some exploratory experiments

NASA Astrophysics Data System (ADS)

Oubbati, Mohamed; Kord, Bahram; Koprinkova-Hristova, Petia; Palm, Günther

2014-04-01

The new tendency of artificial intelligence suggests that intelligence must be seen as a result of the interaction between brains, bodies and environments. This view implies that designing sophisticated behaviour requires a primary focus on how agents are functionally coupled to their environments. Under this perspective, we present early results with the application of reservoir computing as an efficient tool to understand how behaviour emerges from interaction. Specifically, we present reservoir computing models, that are inspired by imitation learning designs, to extract the essential components of behaviour that results from agent-environment interaction dynamics. Experimental results using a mobile robot are reported to validate the learning architectures.

Binding affinities of vascular endothelial growth factor (VEGF) for heparin-derived oligosaccharides

PubMed Central

Zhao, Wenjing; McCallum, Scott A.; Xiao, Zhongping; Zhang, Fuming; Linhardt, Robert J.

2011-01-01

Heparin and heparan sulphate (HS) exert their wide range of biological activities by interacting with extracellular protein ligands. Among these important protein ligands are various angiogenic growth factors and cytokines. HS-binding to vascular endothelial growth factor (VEGF) regulates multiple aspects of vascular development and function through its specific interaction with HS. Many studies have focused on HS-derived or HS-mimicking structures for the characterization of VEGF165 interaction with HS. Using a heparinase 1-prepared small library of heparin-derived oligosaccharides ranging from hexasaccharide to octadecasaccharide, we systematically investigated the heparin-specific structural features required for VEGF binding. We report the apparent affinities for the association between the heparin-derived oligosaccharides with both VEGF165 and VEGF55, a peptide construct encompassing exclusively the heparin-binding domain of VEGF165. An octasaccharide was the minimum size of oligosaccharide within the library to efficiently bind to both forms of VEGF and that a tetradecasaccharide displayed an effective binding affinity to VEGF165 comparable to unfractionated heparin. The range of relative apparent binding affinities among VEGF and the panel of heparin-derived oligosaccharides demonstrate that VEGF binding affinity likely depends on the specific structural features of these oligosaccharides including their degree of sulphation and sugar ring stereochemistry and conformation. Notably, the unique 3-O-sulpho group found within the specific antithrombin binding site of heparin is not required for VEGF165 binding. These findings afford new insight into the inherent kinetics and affinities for VEGF association with heparin and heparin-derived oligosaccharides with key residue specific modifications and may potentially benefit the future design of oligosaccharide-based anti-angiogenesis drugs. PMID:21658003

Secondary structure encodes a cooperative tertiary folding funnel in the Azoarcus ribozyme

PubMed Central

Mustoe, Anthony M.; Al-Hashimi, Hashim M.; Brooks, Charles L.

2016-01-01

A requirement for specific RNA folding is that the free-energy landscape discriminate against non-native folds. While tertiary interactions are critical for stabilizing the native fold, they are relatively non-specific, suggesting additional mechanisms contribute to tertiary folding specificity. In this study, we use coarse-grained molecular dynamics simulations to explore how secondary structure shapes the tertiary free-energy landscape of the Azoarcus ribozyme. We show that steric and connectivity constraints posed by secondary structure strongly limit the accessible conformational space of the ribozyme, and that these so-called topological constraints in turn pose strong free-energy penalties on forming different tertiary contacts. Notably, native A-minor and base-triple interactions form with low conformational free energy, while non-native tetraloop/tetraloop–receptor interactions are penalized by high conformational free energies. Topological constraints also give rise to strong cooperativity between distal tertiary interactions, quantitatively matching prior experimental measurements. The specificity of the folding landscape is further enhanced as tertiary contacts place additional constraints on the conformational space, progressively funneling the molecule to the native state. These results indicate that secondary structure assists the ribozyme in navigating the otherwise rugged tertiary folding landscape, and further emphasize topological constraints as a key force in RNA folding. PMID:26481360

Commercial technologies from the SP-100 program

NASA Astrophysics Data System (ADS)

Truscello, Vincent C.; Fujita, Toshio; Mondt, Jack F.

1995-01-01

For more than a decade, the Jet Propulsion Labortory (JPL) and Los Alamos National Laboratory (LANL) have managed a multi-agency funded effort to develop a space reactor power system. This SP-100 Program has developed technologies required for space power systems that can be implemented in the industrial and commercial sectors to improve our competitiveness in the global economy. Initial steps taken to transfer this technology from the laboratories to industrial and commercial entities within the United States include: (1) identifying specific technologies having commercial potential; (2) distributing information describing the identified technologies and interacting with interested commercial and industrial entities to develop application-specific details and requirements; and (3) providing a technological data base that leads to transfer of technology or the forming of teaming arrangements to accomplish the transfer by tailoring the technology to meet application-specific requirements. SP-100 technologies having commercial potential encompass fabrication processes, devices, and components. Examples are a process for bonding refractory metals to graphite, a device to sense the position of an actuator and a component to enable rotating machines to operate without supplying lubrication ( a self-lubricating ball bearing). Shortly after the National Aeronautics and Space Administration (NASA) Regional Technology Transfer Centers widely disseminated information covering SP-100 technologies, over one hundred expressions of interest were received. These early responses indicate that there is a large potential benefit in transferring SP-100 technology. Interactions with industrial and commercial entities have identified a substantial need for creating teaming arrangements involving the interested entity and personnel from laboratories and their contractors, who have the knowledge and ability to tailor the technology to meet application-specific requirements.

21 CFR 201.57 - Specific requirements on content and format of labeling for human prescription drug and...

Code of Federal Regulations, 2013 CFR

2013-04-01

... comparative rates of occurrence cannot be reliably determined (e.g., adverse reactions were observed only in... in vivo study designs or results (e.g., drug interaction studies), may be included in this section if...

21 CFR 201.57 - Specific requirements on content and format of labeling for human prescription drug and...

Code of Federal Regulations, 2012 CFR

2012-04-01

... comparative rates of occurrence cannot be reliably determined (e.g., adverse reactions were observed only in... in vivo study designs or results (e.g., drug interaction studies), may be included in this section if...

21 CFR 201.57 - Specific requirements on content and format of labeling for human prescription drug and...

Code of Federal Regulations, 2014 CFR

2014-04-01

... comparative rates of occurrence cannot be reliably determined (e.g., adverse reactions were observed only in... in vivo study designs or results (e.g., drug interaction studies), may be included in this section if...

Genetic Interactions Between the Meiosis-Specific Cohesin Components, STAG3, REC8, and RAD21L.

PubMed

Ward, Ayobami; Hopkins, Jessica; Mckay, Matthew; Murray, Steve; Jordan, Philip W

2016-06-01

Cohesin is an essential structural component of chromosomes that ensures accurate chromosome segregation during mitosis and meiosis. Previous studies have shown that there are cohesin complexes specific to meiosis, required to mediate homologous chromosome pairing, synapsis, recombination, and segregation. Meiosis-specific cohesin complexes consist of two structural maintenance of chromosomes proteins (SMC1α/SMC1β and SMC3), an α-kleisin protein (RAD21, RAD21L, or REC8), and a stromal antigen protein (STAG1, 2, or 3). STAG3 is exclusively expressed during meiosis, and is the predominant STAG protein component of cohesin complexes in primary spermatocytes from mouse, interacting directly with each α-kleisin subunit. REC8 and RAD21L are also meiosis-specific cohesin components. Stag3 mutant spermatocytes arrest in early prophase ("zygotene-like" stage), displaying failed homolog synapsis and persistent DNA damage, as a result of unstable loading of cohesin onto the chromosome axes. Interestingly, Rec8, Rad21L double mutants resulted in an earlier "leptotene-like" arrest, accompanied by complete absence of STAG3 loading. To assess genetic interactions between STAG3 and α-kleisin subunits RAD21L and REC8, our lab generated Stag3, Rad21L, and Stag3, Rec8 double knockout mice, and compared them to the Rec8, Rad21L double mutant. These double mutants are phenotypically distinct from one another, and more severe than each single knockout mutant with regards to chromosome axis formation, cohesin loading, and sister chromatid cohesion. The Stag3, Rad21L, and Stag3, Rec8 double mutants both progress further into prophase I than the Rec8, Rad21L double mutant. Our genetic analysis demonstrates that cohesins containing STAG3 and REC8 are the main complex required for centromeric cohesion, and RAD21L cohesins are required for normal clustering of pericentromeric heterochromatin. Furthermore, the STAG3/REC8 and STAG3/RAD21L cohesins are the primary cohesins required for axis formation. Copyright © 2016 Ward et al.

ROTAS: a rotamer-dependent, atomic statistical potential for assessment and prediction of protein structures.

PubMed

Park, Jungkap; Saitou, Kazuhiro

2014-09-18

Multibody potentials accounting for cooperative effects of molecular interactions have shown better accuracy than typical pairwise potentials. The main challenge in the development of such potentials is to find relevant structural features that characterize the tightly folded proteins. Also, the side-chains of residues adopt several specific, staggered conformations, known as rotamers within protein structures. Different molecular conformations result in different dipole moments and induce charge reorientations. However, until now modeling of the rotameric state of residues had not been incorporated into the development of multibody potentials for modeling non-bonded interactions in protein structures. In this study, we develop a new multibody statistical potential which can account for the influence of rotameric states on the specificity of atomic interactions. In this potential, named "rotamer-dependent atomic statistical potential" (ROTAS), the interaction between two atoms is specified by not only the distance and relative orientation but also by two state parameters concerning the rotameric state of the residues to which the interacting atoms belong. It was clearly found that the rotameric state is correlated to the specificity of atomic interactions. Such rotamer-dependencies are not limited to specific type or certain range of interactions. The performance of ROTAS was tested using 13 sets of decoys and was compared to those of existing atomic-level statistical potentials which incorporate orientation-dependent energy terms. The results show that ROTAS performs better than other competing potentials not only in native structure recognition, but also in best model selection and correlation coefficients between energy and model quality. A new multibody statistical potential, ROTAS accounting for the influence of rotameric states on the specificity of atomic interactions was developed and tested on decoy sets. The results show that ROTAS has improved ability to recognize native structure from decoy models compared to other potentials. The effectiveness of ROTAS may provide insightful information for the development of many applications which require accurate side-chain modeling such as protein design, mutation analysis, and docking simulation.

Glycoprotein interactions in paramyxovirus fusion

PubMed Central

Iorio, Ronald M; Melanson, Vanessa R; Mahon, Paul J

2009-01-01

The Paramyxoviridae are enveloped, negative-stranded RNA viruses, some of which recognize sialic acid-containing receptors, while others recognize specific proteinaceous receptors. The major cytopathic effect of paramyxovirus infection is membrane fusion-induced syncytium formation. Paramyxoviruses are unusual in that the receptor-binding and fusion-promoting activities reside on two different spike structures, the attachment and fusion glycoproteins, respectively. For most paramyxoviruses, this distribution of functions requires a mechanism by which the two processes can be linked for the promotion of fusion. This is accomplished by a virus-specific interaction between the two proteins. An increasing body of evidence supports the notion that members of this family of viruses utilize this glycoprotein interaction in different ways in order to mediate the regulation of the fusion protein activation, depending on the type of receptor utilized by the virus. PMID:20161127

The Intrinsically Disordered Regions of the Drosophila melanogaster Hox Protein Ultrabithorax Select Interacting Proteins Based on Partner Topology

PubMed Central

Hsiao, Hao-Ching; Gonzalez, Kim L.; Catanese, Daniel J.; Jordy, Kristopher E.; Matthews, Kathleen S.; Bondos, Sarah E.

2014-01-01

Interactions between structured proteins require a complementary topology and surface chemistry to form sufficient contacts for stable binding. However, approximately one third of protein interactions are estimated to involve intrinsically disordered regions of proteins. The dynamic nature of disordered regions before and, in some cases, after binding calls into question the role of partner topology in forming protein interactions. To understand how intrinsically disordered proteins identify the correct interacting partner proteins, we evaluated interactions formed by the Drosophila melanogaster Hox transcription factor Ultrabithorax (Ubx), which contains both structured and disordered regions. Ubx binding proteins are enriched in specific folds: 23 of its 39 partners include one of 7 folds, out of the 1195 folds recognized by SCOP. For the proteins harboring the two most populated folds, DNA-RNA binding 3-helical bundles and α-α superhelices, the regions of the partner proteins that exhibit these preferred folds are sufficient for Ubx binding. Three disorder-containing regions in Ubx are required to bind these partners. These regions are either alternatively spliced or multiply phosphorylated, providing a mechanism for cellular processes to regulate Ubx-partner interactions. Indeed, partner topology correlates with the ability of individual partner proteins to bind Ubx spliceoforms. Partners bind different disordered regions within Ubx to varying extents, creating the potential for competition between partners and cooperative binding by partners. The ability of partners to bind regions of Ubx that activate transcription and regulate DNA binding provides a mechanism for partners to modulate transcription regulation by Ubx, and suggests that one role of disorder in Ubx is to coordinate multiple molecular functions in response to tissue-specific cues. PMID:25286318

Studying Protein-Protein Interactions by Biotin AP-Tagged Pulldown and LTQ-Orbitrap Mass Spectrometry.

PubMed

Xie, Zhongqiu; Jia, Yuemeng; Li, Hui

2017-01-01

The study of protein-protein interactions represents a key aspect of biological research. Identifying unknown protein binding partners using mass spectrometry (MS)-based proteomics has evolved into an indispensable strategy in drug discovery. The classic approach of immunoprecipitation with specific antibodies against the proteins of interest has limitations, such as the need for immunoprecipitation-qualified antibody. The biotin AP-tag pull-down system has the advantage of high specificity, ease of use, and no requirement for antibody. It is based on the high specificity, high affinity interaction between biotin and streptavidin. After pulldown, in-gel tryptic digestion and tandem mass spectrometry (MS/MS) analysis of sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) protein bands can be performed. In this work, we provide protocols that can be used for the identification of proteins that interact with FOXM1, a protein that has recently emerged as a potential biomarker and drug target in oncotherapy, as an example. We focus on the pull-down procedure and assess the efficacy of the pulldown with known FOXM1 interactors such as β-catenin. We use a high performance LTQ Orbitrap MSn system that combines rapid LTQ ion trap data acquisition with high mass accuracy Orbitrap analysis to identify the interacting proteins.

Inositol Pentakisphosphate Isomers Bind PH Domains with Varying Specificity and Inhibit Phosphoinositide Interactions

DOE Office of Scientific and Technical Information (OSTI.GOV)

S Jackson; S Al-Saigh; C Schultz

2011-12-31

PH domains represent one of the most common domains in the human proteome. These domains are recognized as important mediators of protein-phosphoinositide and protein-protein interactions. Phosphoinositides are lipid components of the membrane that function as signaling molecules by targeting proteins to their sites of action. Phosphoinositide based signaling pathways govern a diverse range of important cellular processes including membrane remodeling, differentiation, proliferation and survival. Myo-Inositol phosphates are soluble signaling molecules that are structurally similar to the head groups of phosphoinositides. These molecules have been proposed to function, at least in part, by regulating PH domain-phosphoinositide interactions. Given the structural similaritymore » of inositol phosphates we were interested in examining the specificity of PH domains towards the family of myo-inositol pentakisphosphate isomers. In work reported here we demonstrate that the C-terminal PH domain of pleckstrin possesses the specificity required to discriminate between different myo-inositol pentakisphosphate isomers. The structural basis for this specificity was determined using high-resolution crystal structures. Moreover, we show that while the PH domain of Grp1 does not possess this high degree of specificity, the PH domain of protein kinase B does. These results demonstrate that some PH domains possess enough specificity to discriminate between myo-inositol pentakisphosphate isomers allowing for these molecules to differentially regulate interactions with phosphoinositides. Furthermore, this work contributes to the growing body of evidence supporting myo-inositol phosphates as regulators of important PH domain-phosphoinositide interactions. Finally, in addition to expanding our knowledge of cellular signaling, these results provide a basis for developing tools to probe biological pathway.« less

The interplay of non-specific binding, target-mediated clearance and FcRn interactions on the pharmacokinetics of humanized antibodies

PubMed Central

Datta-Mannan, Amita; Lu, Jirong; Witcher, Derrick R; Leung, Donmienne; Tang, Ying; Wroblewski, Victor J

2015-01-01

The application of protein engineering technologies toward successfully improving antibody pharmacokinetics has been challenging due to the multiplicity of biochemical factors that influence monoclonal antibody (mAb) disposition in vivo. Physiological factors including interactions with the neonatal Fc receptor (FcRn) and specific antigen binding properties of mAbs, along with biophysical properties of the mAbs themselves play a critical role. It has become evident that applying an integrated approach to understand the relative contribution of these factors is critical to rationally guide and apply engineering strategies to optimize mAb pharmacokinetics. The study presented here evaluated the influence of unintended non-specific interactions on the disposition of mAbs whose clearance rates are governed predominantly by either non-specific (FcRn) or target-mediated processes. The pharmacokinetics of 8 mAbs representing a diverse range of these properties was evaluated in cynomolgus monkeys. Results revealed complementarity-determining region (CDR) charge patch engineering to decrease charge-related non-specific binding can have a significant impact on improving the clearance. In contrast, the influence of enhanced in vitro FcRn binding was mixed, and related to both the strength of charge interaction and the general mechanism predominant in governing the clearance of the particular mAb. Overall, improved pharmacokinetics through enhanced FcRn interactions were apparent for a CDR charge-patch normalized mAb which was affected by non-specific clearance. The findings in this report are an important demonstration that mAb pharmacokinetics requires optimization on a case-by-case basis to improve the design of molecules with increased therapeutic application. PMID:26337808

40 CFR 1042.205 - Application requirements.

Code of Federal Regulations, 2010 CFR

2010-07-01

...'s specifications and other basic parameters of the engine's design and emission controls. List the fuel type on which your engines are designed to operate (for example, ultra low-sulfur diesel fuel... or modulate the same parameter. Describe whether the strategies interact in a comparative or additive...

40 CFR 1042.205 - Application requirements.

Code of Federal Regulations, 2014 CFR

2014-07-01

...'s specifications and other basic parameters of the engine's design and emission controls. List the fuel type on which your engines are designed to operate (for example, ultra low-sulfur diesel fuel... or modulate the same parameter. Describe whether the strategies interact in a comparative or additive...

40 CFR 1042.205 - Application requirements.

Code of Federal Regulations, 2011 CFR

2011-07-01

...'s specifications and other basic parameters of the engine's design and emission controls. List the fuel type on which your engines are designed to operate (for example, ultra low-sulfur diesel fuel... or modulate the same parameter. Describe whether the strategies interact in a comparative or additive...

40 CFR 1042.205 - Application requirements.

Code of Federal Regulations, 2012 CFR

2012-07-01

...'s specifications and other basic parameters of the engine's design and emission controls. List the fuel type on which your engines are designed to operate (for example, ultra low-sulfur diesel fuel... or modulate the same parameter. Describe whether the strategies interact in a comparative or additive...

40 CFR 1042.205 - Application requirements.

Code of Federal Regulations, 2013 CFR

2013-07-01

...'s specifications and other basic parameters of the engine's design and emission controls. List the fuel type on which your engines are designed to operate (for example, ultra low-sulfur diesel fuel... or modulate the same parameter. Describe whether the strategies interact in a comparative or additive...

Observe, Explain, Connect

ERIC Educational Resources Information Center

Vardell, Sylvia M.; Wong, Janet S.

2014-01-01

Like science, poetry often involves a high level of abstraction in language and ideas, requiring specific critical-thinking skills and promoting interaction. Casteel and Isom (1994) acknowledge, "The literacy processes are the means by which science content is learned because content information is rooted in written and oral language"…

Apprentices' Learning of Occupationally Informed Practical Judgment

ERIC Educational Resources Information Center

Chan, Selena

2015-01-01

Learning to become trade workers requires developing the ability to make practical workplace-based judgments, often centred around difficult to articulate trade "know-how" or tacit knowledge. Apprentices learn discipline specific ways of doing, thinking, feeling and being from experts, peers and through interactions with occupational…

Neuropilin-1 interacts with the second branchial arch microenvironment to mediate chick neural crest cell dynamics

PubMed Central

McLennan, Rebecca; Kulesa, Paul M.

2011-01-01

Cranial neural crest cells (NCCs) require neuropilin signaling to reach and invade the branchial arches. Here, we use an in vivo chick model to investigate whether the neuropilin-1 knockdown phenotype is specific to the second branchial arch (ba2), changes in NCC behaviors and phenotypic consequences, and whether neuropilins work together to facilitate entry into and invasion of ba2. We find that cranial NCCs with reduced neuropilin-1 expression displayed shorter protrusions and decreased cell body and nuclear length-to-width ratios characteristic of a loss in polarity and motility, after specific interaction with ba2. Directed NCC migration was rescued by transplantation of transfected cells into rhombomere 4 of younger hosts. Lastly, reduction of neuropilin-2 expression by shRNA either solely or with reduction of neuropilin-1 expression did not lead to a stronger head phenotype. Thus, NCCs, independent of rhombomere origin, require neuropilin-1, but not neuropilin-2 to maintain polarity and directed migration into ba2. PMID:20503363

Business Process Management

NASA Astrophysics Data System (ADS)

Hantry, Francois; Papazoglou, Mike; van den Heuvel, Willem-Jan; Haque, Rafique; Whelan, Eoin; Carroll, Noel; Karastoyanova, Dimka; Leymann, Frank; Nikolaou, Christos; Lammersdorf, Winfried; Hacid, Mohand-Said

Business process management is one of the core drivers of business innovation and is based on strategic technology and capable of creating and successfully executing end-to-end business processes. The trend will be to move from relatively stable, organization-specific applications to more dynamic, high-value ones where business process interactions and trends are examined closely to understand more accurately an application's requirements. Such collaborative, complex end-to-end service interactions give rise to the concept of Service Networks (SNs).

Gain in Transcriptional Activity by Primate-specific Coevolution of Melanoma Antigen-A11 and Its Interaction Site in Androgen Receptor*

PubMed Central

Liu, Qiang; Su, Shifeng; Blackwelder, Amanda J.; Minges, John T.; Wilson, Elizabeth M.

2011-01-01

Male sex development and growth occur in response to high affinity androgen binding to the androgen receptor (AR). In contrast to complete amino acid sequence conservation in the AR DNA and ligand binding domains among mammals, a primate-specific difference in the AR NH2-terminal region that regulates the NH2- and carboxyl-terminal (N/C) interaction enables direct binding to melanoma antigen-A11 (MAGE-11), an AR coregulator that is also primate-specific. Human, mouse, and rat AR share the same NH2-terminal 23FQNLF27 sequence that mediates the androgen-dependent N/C interaction. However, the mouse and rat AR FXXLF motif is flanked by Ala33 that evolved to Val33 in primates. Human AR Val33 was required to interact directly with MAGE-11 and for the inhibitory effect of the AR N/C interaction on activation function 2 that was relieved by MAGE-11. The functional importance of MAGE-11 was indicated by decreased human AR regulation of an androgen-dependent endogenous gene using lentivirus short hairpin RNAs and by the greater transcriptional strength of human compared with mouse AR. MAGE-11 increased progesterone and glucocorticoid receptor activity independently of binding an FXXLF motif by interacting with p300 and p160 coactivators. We conclude that the coevolution of the AR NH2-terminal sequence and MAGE-11 expression among primates provides increased regulatory control over activation domain dominance. Primate-specific expression of MAGE-11 results in greater steroid receptor transcriptional activity through direct interactions with the human AR FXXLF motif region and indirectly through steroid receptor-associated p300 and p160 coactivators. PMID:21730049

ChIP-chip.

PubMed

Kim, Tae Hoon; Dekker, Job

2018-05-01

ChIP-chip can be used to analyze protein-DNA interactions in a region-wide and genome-wide manner. DNA microarrays contain PCR products or oligonucleotide probes that are designed to represent genomic sequences. Identification of genomic sites that interact with a specific protein is based on competitive hybridization of the ChIP-enriched DNA and the input DNA to DNA microarrays. The ChIP-chip protocol can be divided into two main sections: Amplification of ChIP DNA and hybridization of ChIP DNA to arrays. A large amount of DNA is required to hybridize to DNA arrays, and hybridization to a set of multiple commercial arrays that represent the entire human genome requires two rounds of PCR amplifications. The relative hybridization intensity of ChIP DNA and that of the input DNA is used to determine whether the probe sequence is a potential site of protein-DNA interaction. Resolution of actual genomic sites bound by the protein is dependent on the size of the chromatin and on the genomic distance between the probes on the array. As with expression profiling using gene chips, ChIP-chip experiments require multiple replicates for reliable statistical measure of protein-DNA interactions. © 2018 Cold Spring Harbor Laboratory Press.

A tale of two sequences: microRNA-target chimeric reads.

PubMed

Broughton, James P; Pasquinelli, Amy E

2016-04-04

In animals, a functional interaction between a microRNA (miRNA) and its target RNA requires only partial base pairing. The limited number of base pair interactions required for miRNA targeting provides miRNAs with broad regulatory potential and also makes target prediction challenging. Computational approaches to target prediction have focused on identifying miRNA target sites based on known sequence features that are important for canonical targeting and may miss non-canonical targets. Current state-of-the-art experimental approaches, such as CLIP-seq (cross-linking immunoprecipitation with sequencing), PAR-CLIP (photoactivatable-ribonucleoside-enhanced CLIP), and iCLIP (individual-nucleotide resolution CLIP), require inference of which miRNA is bound at each site. Recently, the development of methods to ligate miRNAs to their target RNAs during the preparation of sequencing libraries has provided a new tool for the identification of miRNA target sites. The chimeric, or hybrid, miRNA-target reads that are produced by these methods unambiguously identify the miRNA bound at a specific target site. The information provided by these chimeric reads has revealed extensive non-canonical interactions between miRNAs and their target mRNAs, and identified many novel interactions between miRNAs and noncoding RNAs.

Challenges and dreams: physics of weak interactions essential to life

PubMed Central

Chien, Peter; Gierasch, Lila M.

2014-01-01

Biological systems display stunning capacities to self-organize. Moreover, their subcellular architectures are dynamic and responsive to changing needs and conditions. Key to these properties are manifold weak “quinary” interactions that have evolved to create specific spatial networks of macromolecules. These specific arrangements of molecules enable signals to be propagated over distances much greater than molecular dimensions, create phase separations that define functional regions in cells, and amplify cellular responses to changes in their environments. A major challenge is to develop biochemical tools and physical models to describe the panoply of weak interactions operating in cells. We also need better approaches to measure the biases in the spatial distributions of cellular macromolecules that result from the integrated action of multiple weak interactions. Partnerships between cell biologists, biochemists, and physicists are required to deploy these methods. Together these approaches will help us realize the dream of understanding the biological “glue” that sustains life at a molecular and cellular level. PMID:25368424

Spacelab user interaction

NASA Technical Reports Server (NTRS)

1975-01-01

The results of the third and final phase of a study undertaken to define means of optimizing the Spacelab experiment data system by interactively manipulating the flow of data were presented. A number of payload applicable interactive techniques and an integrated interaction system for each of two possible payloads are described. These interaction systems have been functionally defined and are accompanied with block diagrams, hardware specifications, software sizing and speed requirements, operational procedures and cost/benefits analysis data for both onboard and ground based system elements. It is shown that accrued benefits are attributable to a reduction in data processing costs obtained by, generally, a considerable reduction in the quantity of data that might otherwise be generated without interaction. One other additional anticipated benefit includes the increased scientific value obtained by the quicker return of all useful data.

FY is an RNA 3' end-processing factor that interacts with FCA to control the Arabidopsis floral transition.

PubMed

Simpson, Gordon G; Dijkwel, Paul P; Quesada, Victor; Henderson, Ian; Dean, Caroline

2003-06-13

The nuclear RNA binding protein, FCA, promotes Arabidopsis reproductive development. FCA contains a WW protein interaction domain that is essential for FCA function. We have identified FY as a protein partner for this domain. FY belongs to a highly conserved group of eukaryotic proteins represented in Saccharomyces cerevisiae by the RNA 3' end-processing factor, Pfs2p. FY regulates RNA 3' end processing in Arabidopsis as evidenced through its role in FCA regulation. FCA expression is autoregulated through the use of different polyadenylation sites within the FCA pre-mRNA, and the FCA/FY interaction is required for efficient selection of the promoter-proximal polyadenylation site. The FCA/FY interaction is also required for the downregulation of the floral repressor FLC. We propose that FCA controls 3' end formation of specific transcripts and that in higher eukaryotes, proteins homologous to FY may have evolved as sites of association for regulators of RNA 3' end processing.

Quantitative Investigation of Protein-Nucleic Acid Interactions by Biosensor Surface Plasmon Resonance.

PubMed

Wang, Shuo; Poon, Gregory M K; Wilson, W David

2015-01-01

Biosensor-surface plasmon resonance (SPR) technology has emerged as a powerful label-free approach for the study of nucleic acid interactions in real time. The method provides simultaneous equilibrium and kinetic characterization for biomolecular interactions with low sample requirements and without the need for external probes. A detailed and practical guide for protein-DNA interaction analyses using biosensor-SPR methods is presented. Details of SPR technology and basic fundamentals are described with recommendations on the preparation of the SPR instrument, sensor chips and samples, experimental design, quantitative and qualitative data analyses and presentation. A specific example of the interaction of a transcription factor with DNA is provided with results evaluated by both kinetic and steady-state SPR methods.

New levels of language processing complexity and organization revealed by granger causation.

PubMed

Gow, David W; Caplan, David N

2012-01-01

Granger causation analysis of high spatiotemporal resolution reconstructions of brain activation offers a new window on the dynamic interactions between brain areas that support language processing. Premised on the observation that causes both precede and uniquely predict their effects, this approach provides an intuitive, model-free means of identifying directed causal interactions in the brain. It requires the analysis of all non-redundant potentially interacting signals, and has shown that even "early" processes such as speech perception involve interactions of many areas in a strikingly large network that extends well beyond traditional left hemisphere perisylvian cortex that play out over hundreds of milliseconds. In this paper we describe this technique and review several general findings that reframe the way we think about language processing and brain function in general. These include the extent and complexity of language processing networks, the central role of interactive processing dynamics, the role of processing hubs where the input from many distinct brain regions are integrated, and the degree to which task requirements and stimulus properties influence processing dynamics and inform our understanding of "language-specific" localized processes.

A convenient method for synthesis of glyconanoparticles for colorimetric measuring carbohydrate-protein interactions

PubMed Central

Chuang, Yen-Jun; Zhou, Xichun; Pan, Zhengwei; Turchi, Craig

2009-01-01

Carbohydrate functionalized nanoparticles, i.e., the glyconanoparticles, have wide application ranging from studies of carbohydrate-protein interactions, in vivo cell imaging, biolabeling, etc. Currently reported methods for preparation of glyconanoaprticles require multi-step modifications of carbohydrates moieties to conjugate to nanoparticle surface. However, the required synthetic manipulations are difficult and time consuming. We report herewith a simple and versatile method for preparing glyconanoparticles. This method is based on the utilization of clean and convenient microwave irradiation energy for one-step, site-specific conjugation of unmodified carbohydrates onto hydrazide-functionalized Au nanoparticles. A colorimetric assay that utilizes the ensemble of gold glyconanoparticles and Concanavalin A (ConA) was also presented. This feasible assay system was developed to analyze multivalent interactions and to determine the dissociation constant (Kd) for five kind of Au glyconanoparticles with lectin. Surface plasmon changes of the Au glyconanparticles as a function of lectin-carbohydrate interactions were measured and the dissociation constants were determined based on non-linear curve fitting. The strength of the interaction of carbohydrates with ConA was found to be as follows: Maltose > Mannose > Glucose > Lactose > MAN5. PMID:19698698

Two-colored fluorescence correlation spectroscopy screening for LC3-P62 interaction inhibitors.

PubMed

Tsuganezawa, Keiko; Shinohara, Yoshiyasu; Ogawa, Naoko; Tsuboi, Shun; Okada, Norihisa; Mori, Masumi; Yokoyama, Shigeyuki; Noda, Nobuo N; Inagaki, Fuyuhiko; Ohsumi, Yoshinori; Tanaka, Akiko

2013-10-01

The fluorescence correlation spectroscopy (FCS)-based competitive binding assay to screen for protein-protein interaction inhibitors is a highly sensitive method as compared with the fluorescent polarization assay used conventionally. However, the FCS assay identifies many false-positive compounds, which requires specifically designed orthogonal screenings. A two-colored application of the FCS-based screening was newly developed, and inhibitors of a protein-protein interaction, involving selective autophagy, were selected. We focused on the interaction of LC3 with the adaptor protein p62, because the interaction is crucial to degrade the specific target proteins recruited by p62. First, about 10,000 compounds were subjected to the FCS-based competitive assay using a TAMRA-labeled p62-derived probe, and 29 hit compounds were selected. Next, the obtained hits were evaluated by the second FCS assay, using an Alexa647-labeled p62-derived probe to remove the false-positive compounds, and six hit compounds inhibited the interaction. Finally, we tested all 29 compounds by surface plasmon resonance-based competitive binding assay to evaluate their inhibition of the LC3-p62 interaction and selected two inhibitors with IC50 values less than 2 µM. The two-colored FCS-based screening was shown to be effective to screen for protein-protein interaction inhibitors.

Facilitating interpersonal interaction and learning online: linking theory and practice.

PubMed

Sargeant, Joan; Curran, Vernon; Allen, Michael; Jarvis-Selinger, Sandra; Ho, Kendall

2006-01-01

An earlier study of physicians' perceptions of interactive online learning showed that these were shaped both by program design and quality and the quality and quantity of interpersonal interaction. We explore instructor roles in enhancing online learning through interpersonal interaction and the learning theories that inform these. This was a qualitative study using focus groups and interviews. Using purposive sampling, 50 physicians were recruited based on their experience with interactive online CME and face-to-face CME. Qualitative thematic and interpretive analysis was used. Two facilitation roles appeared key: creating a comfortable learning environment and enhancing the educational value of electronic discussions. Comfort developed gradually, and specific interventions like facilitating introductions and sharing experiences in a friendly, informative manner were helpful. As in facilitating effective small-group learning, instructors' thoughtful use of techniques that facilitated constructive interaction based on learner's needs and practice demands contributed to the educational value of interpersonal interactions. Facilitators require enhanced skills to engage learners in meaningful interaction and to overcome the transactional distance of online learning. The use of learning theories, including behavioral, cognitive, social, humanistic, and constructivist, can strengthen the educational design and facilitation of online programs. Preparation for online facilitation should include instruction in the roles and techniques required and the theories that inform them.

The RSF1 histone-remodelling factor facilitates DNA double-strand break repair by recruiting centromeric and Fanconi Anaemia proteins.

PubMed

Pessina, Fabio; Lowndes, Noel F

2014-05-01

ATM is a central regulator of the cellular responses to DNA double-strand breaks (DSBs). Here we identify a biochemical interaction between ATM and RSF1 and we characterise the role of RSF1 in this response. The ATM-RSF1 interaction is dependent upon both DSBs and ATM kinase activity. Together with SNF2H/SMARCA5, RSF1 forms the RSF chromatin-remodelling complex. Although RSF1 is specific to the RSF complex, SNF2H/SMARCA5 is a catalytic subunit of several other chromatin-remodelling complexes. Although not required for checkpoint signalling, RSF1 is required for efficient repair of DSBs via both end-joining and homology-directed repair. Specifically, the ATM-dependent recruitment to sites of DSBs of the histone fold proteins CENPS/MHF1 and CENPX/MHF2, previously identified at centromeres, is RSF1-dependent. In turn these proteins recruit and regulate the mono-ubiquitination of the Fanconi Anaemia proteins FANCD2 and FANCI. We propose that by depositing CENPS/MHF1 and CENPX/MHF2, the RSF complex either directly or indirectly contributes to the reorganisation of chromatin around DSBs that is required for efficient DNA repair.

Cell cycle-regulated PLEIADE/AtMAP65-3 links membrane and microtubule dynamics during plant cytokinesis.

PubMed

Steiner, Alexander; Rybak, Katarzyna; Altmann, Melina; McFarlane, Heather E; Klaeger, Susan; Nguyen, Ngoc; Facher, Eva; Ivakov, Alexander; Wanner, Gerhard; Kuster, Bernhard; Persson, Staffan; Braun, Pascal; Hauser, Marie-Theres; Assaad, Farhah F

2016-11-01

Cytokinesis, the partitioning of the cytoplasm following nuclear division, requires extensive coordination between cell cycle cues, membrane trafficking and microtubule dynamics. Plant cytokinesis occurs within a transient membrane compartment known as the cell plate, to which vesicles are delivered by a plant-specific microtubule array, the phragmoplast. While membrane proteins required for cytokinesis are known, how these are coordinated with microtubule dynamics and regulated by cell cycle cues remains unclear. Here, we document physical and genetic interactions between Transport Protein Particle II (TRAPPII) tethering factors and microtubule-associated proteins of the PLEIADE/AtMAP65 family. These interactions do not specifically affect the recruitment of either TRAPPII or MAP65 proteins to the cell plate or midzone. Rather, and based on single versus double mutant phenotypes, it appears that they are required to coordinate cytokinesis with the nuclear division cycle. As MAP65 family members are known to be targets of cell cycle-regulated kinases, our results provide a conceptual framework for how membrane and microtubule dynamics may be coordinated with each other and with the nuclear cycle during plant cytokinesis. © 2016 The Authors The Plant Journal © 2016 John Wiley & Sons Ltd.

Mapping the physical network of cellular interactions.

PubMed

Boisset, Jean-Charles; Vivié, Judith; Grün, Dominic; Muraro, Mauro J; Lyubimova, Anna; van Oudenaarden, Alexander

2018-05-21

A cell's function is influenced by the environment, or niche, in which it resides. Studies of niches usually require assumptions about the cell types present, which impedes the discovery of new cell types or interactions. Here we describe ProximID, an approach for building a cellular network based on physical cell interaction and single-cell mRNA sequencing, and show that it can be used to discover new preferential cellular interactions without prior knowledge of component cell types. ProximID found specific interactions between megakaryocytes and mature neutrophils and between plasma cells and myeloblasts and/or promyelocytes (precursors of neutrophils) in mouse bone marrow, and it identified a Tac1 + enteroendocrine cell-Lgr5 + stem cell interaction in small intestine crypts. This strategy can be used to discover new niches or preferential interactions in a variety of organs.

Physical and Chemical Toeholds for Exoplanet Bioastronomy

NASA Technical Reports Server (NTRS)

Hoehler, Tori

2013-01-01

If a search for exoplanet life were mounted today, the likely focus would be to detect oxygen (or ozone) in the atmosphere of a water-bearing rocky planet orbiting roughly 1AU from a G-type star. This appropriately conservative and practical default is necessary in large part because biological input on the question of where and how to look for life has progressed little beyond a purely empirical reliance on the example of terrestrial biology. However, fundamental physical and chemical considerations may impose significant yet universal constraints on biological potential. The liquid water + oxygen paradigm will be considered as an example, with a focus on the question, is liquid water a prerequisite for life? . Life requires a solvent to mediate interactions among biological molecules. A key class of these interactions is molecular recognition with high specificity, which is essential for high fidelity catalysis and (especially) information processing. For example, to correctly reproduce a string consisting of 600,000 units of information (e.g., 600 kilobases, equivalent to the genome of the smallest free living terrestrial organisms) with a 90% success rate requires specificity greater than 10(exp 7):1 for the target molecule vs. incorrect alternatives. Such specificity requires (i) that the correct molecular association is energetically stabilized by at least 40 kJ/mol relative to alternatives, and (ii) that the system is able to sample among possible states (alternative molecular associations) rapidly enough to allow the system to fall under thermodynamic control and express the energetic stabilization. We argue that electrostatic interactions are required to confer the necessary energetic stabilization vs. a large library of molecular alternatives, and that a solvent with polarity and dielectric properties comparable to water is required for the system to sample among possible states and express thermodynamic control. Electrostatic associations can be made in non-polar solvents, but the resulting complexes are too stable to be "unmade" with sufficient frequency to confer thermodynamic control on the system. Such considerations suggest that water, or a solvent with properties very like water, is necessary to support high-fidelity information processing a feature that must be common to all biology and can therefore be considered a critical prerequisite for life.

The RNA Export Factor, Nxt1, Is Required for Tissue Specific Transcriptional Regulation

PubMed Central

Jiang, Jianqiao; White-Cooper, Helen

2013-01-01

The highly conserved, Nxf/Nxt (TAP/p15) RNA nuclear export pathway is important for export of most mRNAs from the nucleus, by interacting with mRNAs and promoting their passage through nuclear pores. Nxt1 is essential for viability; using a partial loss of function allele, we reveal a role for this gene in tissue specific transcription. We show that many Drosophila melanogaster testis-specific mRNAs require Nxt1 for their accumulation. The transcripts that require Nxt1 also depend on a testis-specific transcription complex, tMAC. We show that loss of Nxt1 leads to reduced transcription of tMAC targets. A reporter transcript from a tMAC-dependent promoter is under-expressed in Nxt1 mutants, however the same transcript accumulates in mutants if driven by a tMAC-independent promoter. Thus, in Drosophila primary spermatocytes, the transcription factor used to activate expression of a transcript, rather than the RNA sequence itself or the core transcription machinery, determines whether this expression requires Nxt1. We additionally find that transcripts from intron-less genes are more sensitive to loss of Nxt1 function than those from intron-containing genes and propose a mechanism in which transcript processing feeds back to increase activity of a tissue specific transcription complex. PMID:23754955

The stylar 120 kDa glycoprotein is required for S-specific pollen rejection in Nicotiana.

PubMed

Hancock, C Nathan; Kent, Lia; McClure, Bruce A

2005-09-01

S-RNase participates in at least three mechanisms of pollen rejection. It functions in S-specific pollen rejection (self-incompatibility) and in at least two distinct interspecific mechanisms of pollen rejection in Nicotiana. S-specific pollen rejection and rejection of pollen from Nicotiana plumbaginifolia also require additional stylar proteins. Transmitting-tract-specific (TTS) protein, 120 kDa glycoprotein (120K) and pistil extensin-like protein III (PELP III) are stylar glycoproteins that bind S-RNase in vitro and are also known to interact with pollen. Here we tested whether these glycoproteins have a direct role in pollen rejection. 120K shows the most polymorphism in size between Nicotiana species. Larger 120K-like proteins are often correlated with S-specific pollen rejection. Sequencing results suggest that the polymorphism primarily reflects differences in glycosylation, although indels also occur in the predicted polypeptides. Using RNA interference (RNAi), we suppressed expression of 120K to determine if it is required for S-specific pollen rejection. Transgenic SC N. plumbaginifolia x SI Nicotiana alata (S105S105 or SC10SC10) hybrids with no detectable 120K were unable to perform S-specific pollen rejection. Thus, 120K has a direct role in S-specific pollen rejection. However, suppression of 120K had no effect on rejection of N. plumbaginifolia pollen. In contrast, suppression of HT-B, a factor previously implicated in S-specific pollen rejection, disrupts rejection of N. plumbaginifolia pollen. Thus, S-specific pollen rejection and rejection of N. plumbaginifolia pollen are mechanistically distinct, because they require different non-S-RNase factors.

T7 RNA polymerase non-specifically transcribes and induces disassembly of DNA nanostructures

PubMed Central

Schaffter, Samuel W; Green, Leopold N; Schneider, Joanna; Subramanian, Hari K K; Schulman, Rebecca

2018-01-01

Abstract The use of proteins that bind and catalyze reactions with DNA alongside DNA nanostructures has broadened the functionality of DNA devices. DNA binding proteins have been used to specifically pattern and tune structural properties of DNA nanostructures and polymerases have been employed to directly and indirectly drive structural changes in DNA structures and devices. Despite these advances, undesired and poorly understood interactions between DNA nanostructures and proteins that bind DNA continue to negatively affect the performance and stability of DNA devices used in conjunction with enzymes. A better understanding of these undesired interactions will enable the construction of robust DNA nanostructure-enzyme hybrid systems. Here, we investigate the undesired disassembly of DNA nanotubes in the presence of viral RNA polymerases (RNAPs) under conditions used for in vitro transcription. We show that nanotubes and individual nanotube monomers (tiles) are non-specifically transcribed by T7 RNAP, and that RNA transcripts produced during non-specific transcription disassemble the nanotubes. Disassembly requires a single-stranded overhang on the nanotube tiles where transcripts can bind and initiate disassembly through strand displacement, suggesting that single-stranded domains on other DNA nanostructures could cause unexpected interactions in the presence of viral RNA polymerases. PMID:29718412

T7 RNA polymerase non-specifically transcribes and induces disassembly of DNA nanostructures.

PubMed

Schaffter, Samuel W; Green, Leopold N; Schneider, Joanna; Subramanian, Hari K K; Schulman, Rebecca; Franco, Elisa

2018-06-01

The use of proteins that bind and catalyze reactions with DNA alongside DNA nanostructures has broadened the functionality of DNA devices. DNA binding proteins have been used to specifically pattern and tune structural properties of DNA nanostructures and polymerases have been employed to directly and indirectly drive structural changes in DNA structures and devices. Despite these advances, undesired and poorly understood interactions between DNA nanostructures and proteins that bind DNA continue to negatively affect the performance and stability of DNA devices used in conjunction with enzymes. A better understanding of these undesired interactions will enable the construction of robust DNA nanostructure-enzyme hybrid systems. Here, we investigate the undesired disassembly of DNA nanotubes in the presence of viral RNA polymerases (RNAPs) under conditions used for in vitro transcription. We show that nanotubes and individual nanotube monomers (tiles) are non-specifically transcribed by T7 RNAP, and that RNA transcripts produced during non-specific transcription disassemble the nanotubes. Disassembly requires a single-stranded overhang on the nanotube tiles where transcripts can bind and initiate disassembly through strand displacement, suggesting that single-stranded domains on other DNA nanostructures could cause unexpected interactions in the presence of viral RNA polymerases.

Double-stranded RNA-binding protein 4 is required for resistance signaling against viral and bacterial pathogens.

PubMed

Zhu, Shifeng; Jeong, Rae-Dong; Lim, Gah-Hyun; Yu, Keshun; Wang, Caixia; Chandra-Shekara, A C; Navarre, Duroy; Klessig, Daniel F; Kachroo, Aardra; Kachroo, Pradeep

2013-09-26

Plant viruses often encode suppressors of host RNA silencing machinery, which occasionally function as avirulence factors that are recognized by host resistance (R) proteins. For example, the Arabidopsis R protein, hypersensitive response to TCV (HRT), recognizes the turnip crinkle virus (TCV) coat protein (CP). HRT-mediated resistance requires the RNA-silencing component double-stranded RNA-binding protein 4 (DRB4) even though it neither is associated with the accumulation of TCV-specific small RNA nor requires the RNA silencing suppressor function of CP. HRT interacts with the cytosolic fraction of DRB4. Interestingly, TCV infection both increases the cytosolic DRB4 pool and inhibits the HRT-DRB4 interaction. The virulent R8A CP derivative, which induces a subset of HRT-derived responses, also disrupts this interaction. The differential localization of DRB4 in the presence of wild-type and R8A CP implies the importance of subcellular compartmentalization of DRB4. The requirement of DRB4 in resistance to bacterial infection suggests a universal role in R-mediated defense signaling. Copyright © 2013 The Authors. Published by Elsevier Inc. All rights reserved.

Molecular mechanism of ERK dephosphorylation by striatal-enriched protein tyrosine phosphatase (STEP)

PubMed Central

Li, Hui; Li, Kang-shuai; Su, Jing; Chen, Lai-Zhong; Xu, Yun-Fei; Wang, Hong-Mei; Gong, Zheng; Cui, Guo-Ying; Yu, Xiao; Wang, Kai; Yao, Wei; Xin, Tao; Li, Min-Yong; Xiao, Kun-Hong; An, Xiao-fei; Huo, Yuqing; Xu, Zhi-gang; Sun, Jin-Peng; Pang, Qi

2013-01-01

Striatal-enriched tyrosine phosphatase (STEP) is an important regulator of neuronal synaptic plasticity, and its abnormal level or activity contributes to cognitive disorders. One crucial downstream effector and direct substrate of STEP is extracellular signal-regulated protein kinase (ERK), which has important functions in spine stabilisation and action potential transmission. The inhibition of STEP activity toward phospho-ERK has the potential to treat neuronal diseases, but the detailed mechanism underlying the dephosphorylation of phospho-ERK by STEP is not known. Therefore, we examined STEP activity toward pNPP, phospho-tyrosine-containing peptides, and the full-length phospho-ERK protein using STEP mutants with different structural features. STEP was found to be a highly efficient ERK tyrosine phosphatase that required both its N-terminal regulatory region and key residues in its active site. Specifically, both KIM and KIS of STEP were required for ERK interaction. In addition to the N-terminal KIS region, S245, hydrophobic residues L249/L251, and basic residues R242/R243 located in the KIM region were important in controlling STEP activity toward phospho-ERK. Further kinetic experiments revealed subtle structural differences between STEP and HePTP that affected the interactions of their KIMs with ERK. Moreover, STEP recognised specific positions of a phospho-ERK peptide sequence through its active site, and the contact of STEP F311 with phospho-ERK V205 and T207 were crucial interactions. Taken together, our results not only provide the information for interactions between ERK and STEP, but will also help in the development of specific strategies to target STEP-ERK recognition, which could serve as a potential therapy for neurological disorders. PMID:24117863

Organic Ion Transporters and Statin Drug Interactions.

PubMed

Kellick, Kenneth

2017-11-25

Statin drug-drug interactions (DDIs) are both troublesome to patients as well as costly to medical resources. The ability to predict and avoid these events could lead to improved outcomes as well as patient satisfaction. This review will explore efforts to better understand and predict these interactions specifically related to one drug transport system, the organic anion-transporting polypeptides (OATPs) specifically OATP1B1 and OATP1B3. Since the publication of the discovery of OATPs, there have been various pharmacokinetic models that have been proposed to explain the variation in pharmacokinetic and clinical effects related to the OATPs. The effects in transport activity appear to be partially related to the individual polymorphisms studied. Drug-drug interactions can occur when other drugs compete for the metabolic site on the OATPs. Various medications are identified as substrates and/or inhibitors of the OATPs, thereby complicating the ability to fully predict the impact on levels and effects. All of the models reviewed claim successes but show limited clinical utility. There are specific populations that have been identified, predominately various Asian descendants that require lower doses of statins to avoid adverse events. The concept of attributing these actions to the OATPs has been explored, but current models cannot accurately predict statin blood levels or elimination constants. The current research only points to the differences in the human genome and the single-nucleotide polymorphisms that exist between us. Based upon the currently available studies, there is beginning to be a glimmer in the understanding how different populations respond to statin transport and elimination. Additionally and unfortunately, there are other enzymes to be studied to better predict patient differences. Clearly, there has been much work completed, yet many more questions require answering to better understand these transport proteins.

Measurements of weak interactions between truncated substrates and a hammerhead ribozyme by competitive kinetic analyses: implications for the design of new and efficient ribozymes with high sequence specificity

PubMed Central

Kasai, Yasuhiro; Shizuku, Hideki; Takagi, Yasuomi; Warashina, Masaki; Taira, Kazunari

2002-01-01

Exploitation of ribozymes in a practical setting requires high catalytic activity and strong specificity. The hammerhead ribozyme R32 has considerable potential in this regard since it has very high catalytic activity. In this study, we have examined how R32 recognizes and cleaves a specific substrate, focusing on the mechanism behind the specificity. Comparing rates of cleavage of a substrate in a mixture that included the correct substrate and various substrates with point mutations, we found that R32 cleaved the correct substrate specifically and at a high rate. To clarify the source of this strong specificity, we quantified the weak interactions between R32 and various truncated substrates, using truncated substrates as competitive inhibitors since they were not readily cleaved during kinetic measurements of cleavage of the correct substrate, S11. We found that the strong specificity of the cleavage reaction was due to a closed form of R32 with a hairpin structure. The self-complementary structure within R32 enabled the ribozyme to discriminate between the correct substrate and a mismatched substrate. Since this hairpin motif did not increase the Km (it did not inhibit the binding interaction) or decrease the kcat (it did not decrease the cleavage rate), this kind of hairpin structure might be useful for the design of new ribozymes with strong specificity and high activity. PMID:12034825

Switching between simple cognitive tasks: the interaction of top-down and bottom-up factors

NASA Technical Reports Server (NTRS)

Ruthruff, E.; Remington, R. W.; Johnston, J. C.

2001-01-01

How do top-down factors (e.g., task expectancy) and bottom-up factors (e.g., task recency) interact to produce an overall level of task readiness? This question was addressed by factorially manipulating task expectancy and task repetition in a task-switching paradigm. The effects of expectancy and repetition on response time tended to interact underadditively, but only because the traditional binary task-repetition variable lumps together all switch trials, ignoring variation in task lag. When the task-recency variable was scaled continuously, all 4 experiments instead showed additivity between expectancy and recency. The results indicated that expectancy and recency influence different stages of mental processing. One specific possibility (the configuration-execution model) is that task expectancy affects the time required to configure upcoming central operations, whereas task recency affects the time required to actually execute those central operations.

A flexible flight display research system using a ground-based interactive graphics terminal

NASA Technical Reports Server (NTRS)

Hatfield, J. J.; Elkins, H. C.; Batson, V. M.; Poole, W. L.

1975-01-01

Requirements and research areas for the air transportation system of the 1980 to 1990's were reviewed briefly to establish the need for a flexible flight display generation research tool. Specific display capabilities required by aeronautical researchers are listed and a conceptual system for providing these capabilities is described. The conceptual system uses a ground-based interactive graphics terminal driven by real-time radar and telemetry data to generate dynamic, experimental flight displays. These displays are scan converted to television format, processed, and transmitted to the cockpits of evaluation aircraft. The attendant advantages of a Flight Display Research System (FDRS) designed to employ this concept are presented. The detailed implementation of an FDRS is described. The basic characteristics of the interactive graphics terminal and supporting display electronic subsystems are presented and the resulting system capability is summarized. Finally, the system status and utilization are reviewed.

Interactive Web Graphs with Fewer Restrictions

NASA Technical Reports Server (NTRS)

Fiedler, James

2012-01-01

There is growing popularity for interactive, statistical web graphs and programs to generate them. However, it seems that these programs tend to be somewhat restricted in which web browsers and statistical software are supported. For example, the software might use SVG (e.g., Protovis, gridSVG) or HTML canvas, both of which exclude most versions of Internet Explorer, or the software might be made specifically for R (gridSVG, CRanvas), thus excluding users of other stats software. There are more general tools (d3, Rapha lJS) which are compatible with most browsers, but using one of these to make statistical graphs requires more coding than is probably desired, and requires learning a new tool. This talk will present a method for making interactive web graphs, which, by design, attempts to support as many browsers and as many statistical programs as possible, while also aiming to be relatively easy to use and relatively easy to extend.

Mediator-dependent Nuclear Receptor Functions

PubMed Central

Chen, Wei; Roeder, Robert

2011-01-01

As gene-specific transcription factors, nuclear hormone receptors are broadly involved in many important biological processes. Their function on target genes requires the stepwise assembly of different coactivator complexes that facilitate chromatin remodeling and subsequent preinitiation complex (PIC) formation and function. Mediator has proved to be a crucial, and general, nuclear receptor-interacting coactivator, with demonstrated functions in transcription steps ranging from chromatin remodeling to subsequent PIC formation and function. Here we discuss (i) our current understanding of pathways that nuclear receptors and other interacting cofactors employ to recruit Mediator to target gene enhancers and promoters, including conditional requirements for the strong NR-Mediator interactions mediated by the NR AF2 domain and the MED1 LXXLLL motifs and (ii) mechanisms by which Mediator acts to transmit signals from enhancer-bound nuclear receptors to the general transcription machinery at core promoters to effect PIC formation and function. PMID:21854863

Students' experiences with interactivity and learning in a high school physics multimedia distance learning course

NASA Astrophysics Data System (ADS)

Villarreal-Stewart, Irene

The purpose guiding this research has been to learn about and describe the phenomena of interactivity from the learners' perspectives and to learn which of the interactivity affordances and practices were actually used by students and why in the process of learning physics using an interactive multimedia distance learning course system. The bigger purpose behind learning about and describing interactivity has been to gain knowledge and perspective for its instructional design to benefit the learner, the school as curriculum implementer, and instructional media designers to create better products. Qualitative methodology in the interpretivist tradition was used, that is, in-depth interviews and on-site observations, to gain understanding of interactivity from the learners' perspective and to gain understanding of the student learning context impacting and shaping the students' interactivity experiences. NVivo was used to sort, organize and index data. All data were read on three levels: literally, interpretively, and reflexively; and were read comparatively to other perspectives to get descriptions and interpretations that were holistic to the implementation and had potential insight to improve practice for instructional designers, teachers, administrators, specifically to improve the learning experience for students. Site-Specific Findings: Students watched videos, resisted using phone and e-mail, and worked math problems to demonstrate learning, which resulted in very little interactivity, virtually no dialogue about physics, no physical activity, one-way communication, multifaceted dissatisfaction, student need for teacher involvement in the learning enterprise, student appreciation for interactivity, and expressed desire for a real, live teacher. I also found that some students did experience the system as interactive, did experience learner control and self-directed learning, and despite dissatisfaction, liked and appreciated the course. Wider Applications: Interactivity, a design element, requires scaffolding and nurturing in implementation. The variable and changing context of implementation suggests the requirement for its consideration in design work. The study suggests that during implementation the integrity of design as a whole and flexibility within the design are important continuing considerations. Recommendations. (1) implementation supervision by the school district, (2) use of a language and activity-based theory of learning and teaching and (3) dialogic inquiry (Wells, 1999) to continue learning about interactivity.

Animal-specific C-terminal domain links myeloblastosis oncoprotein (Myb) to an ancient repressor complex

PubMed Central

Andrejka, Laura; Wen, Hong; Ashton, Jonathan; Grant, Megan; Iori, Kevin; Wang, Amy; Manak, J. Robert; Lipsick, Joseph S.

2011-01-01

Members of the Myb oncoprotein and E2F-Rb tumor suppressor protein families are present within the same highly conserved multiprotein transcriptional repressor complex, named either as Myb and synthetic multivuval class B (Myb-MuvB) or as Drosophila Rb E2F and Myb-interacting proteins (dREAM). We now report that the animal-specific C terminus of Drosophila Myb but not the more highly conserved N-terminal DNA-binding domain is necessary and sufficient for (i) adult viability, (ii) proper localization to chromosomes in vivo, (iii) regulation of gene expression in vivo, and (iv) interaction with the highly conserved core of the MuvB/dREAM transcriptional repressor complex. In addition, we have identified a conserved peptide motif that is required for this interaction. Our results imply that an ancient function of Myb in regulating G2/M genes in both plants and animals appears to have been transferred from the DNA-binding domain to the animal-specific C-terminal domain. Increased expression of B-MYB/MYBL2, the human ortholog of Drosophila Myb, correlates with poor prognosis in human patients with breast cancer. Therefore, our results imply that the specific interaction of the C terminus of Myb with the MuvB/dREAM core complex may provide an attractive target for the development of cancer therapeutics. PMID:21969598

Defining Gas Turbine Engine Performance Requirements for the Large Civil TiltRotor (LCTR2)

NASA Technical Reports Server (NTRS)

Snyder, Christopher A.

2013-01-01

Defining specific engine requirements is a critical part of identifying technologies and operational models for potential future rotary wing vehicles. NASA's Fundamental Aeronautics Program, Subsonic Rotary Wing Project has identified the Large Civil TiltRotor (LCTR) as the configuration to best meet technology goals. This notional vehicle concept has evolved with more clearly defined mission and operational requirements to the LCTR-iteration 2 (LCTR2). This paper reports on efforts to further review and refine the LCTR2 analyses to ascertain specific engine requirements and propulsion sizing criteria. The baseline mission and other design or operational requirements are reviewed. Analysis tools are described to help understand their interactions and underlying assumptions. Various design and operational conditions are presented and explained for their contribution to defining operational and engine requirements. These identified engine requirements are discussed to suggest which are most critical to the engine sizing and operation. The most-critical engine requirements are compared to in-house NASA engine simulations to try to ascertain which operational requirements define engine requirements versus points within the available engine operational capability. Finally, results are summarized with suggestions for future efforts to improve analysis capabilities, and better define and refine mission and operational requirements.

Exosites in the substrate specificity of blood coagulation reactions.

PubMed

Bock, P E; Panizzi, P; Verhamme, I M A

2007-07-01

The specificity of blood coagulation proteinases for substrate, inhibitor, and effector recognition is mediated by exosites on the surfaces of the catalytic domains, physically separated from the catalytic site. Some thrombin ligands bind specifically to either exosite I or II, while others engage both exosites. The involvement of different, overlapping constellations of exosite residues enables binding of structurally diverse ligands. The flexibility of the thrombin structure is central to the mechanism of complex formation and the specificity of exosite interactions. Encounter complex formation is driven by electrostatic ligand-exosite interactions, followed by conformational rearrangement to a stable complex. Exosites on some zymogens are in low affinity proexosite states and are expressed concomitant with catalytic site activation. The requirement for exosite expression controls the specificity of assembly of catalytic complexes on the coagulation pathway, such as the membrane-bound factor Xa*factor Va (prothrombinase) complex, and prevents premature assembly. Substrate recognition by prothrombinase involves a two-step mechanism with initial docking of prothrombin to exosites, followed by a conformational change to engage the FXa catalytic site. Prothrombin and its activation intermediates bind prothrombinase in two alternative conformations determined by the zymogen to proteinase transition that are hypothesized to involve prothrombin (pro)exosite I interactions with FVa, which underpin the sequential activation pathway. The role of exosites as the major source of substrate specificity has stimulated development of exosite-targeted anticoagulants for treatment of thrombosis.

Ligand requirements for involvement of PKCε in synergistic analgesic interactions between spinal μ and δ opioid receptors.

PubMed

Schuster, D J; Metcalf, M D; Kitto, K F; Messing, R O; Fairbanks, C A; Wilcox, G L

2015-01-01

We recently found that PKCε was required for spinal analgesic synergy between two GPCRs, δ opioid receptors and α2 A adrenoceptors, co-located in the same cellular subpopulation. We sought to determine if co-delivery of μ and δ opioid receptor agonists would similarly result in synergy requiring PKCε. Combinations of μ and δ opioid receptor agonists were co-administered intrathecally by direct lumbar puncture to PKCε-wild-type (PKCε-WT) and -knockout (PKCε-KO) mice. Antinociception was assessed using the hot-water tail-flick assay. Drug interactions were evaluated by isobolographic analysis. All agonists produced comparable antinociception in both PKCε-WT and PKCε-KO mice. Of 19 agonist combinations that produced analgesic synergy, only 3 required PKCε for a synergistic interaction. In these three combinations, one of the agonists was morphine, although not all combinations involving morphine required PKCε. Morphine + deltorphin II and morphine + deltorphin I required PKCε for synergy, whereas a similar combination, morphine + deltorphin, did not. Additionally, morphine + oxymorphindole required PKCε for synergy, whereas a similar combination, morphine + oxycodindole, did not. We discovered biased agonism for a specific signalling pathway at the level of spinally co-delivered opioid agonists. As the bias is only revealed by an appropriate ligand combination and cannot be accounted for by a single drug, it is likely that the receptors these agonists act on are interacting with each other. Our results support the existence of μ and δ opioid receptor heteromers at the spinal level in vivo. This article is part of a themed section on Opioids: New Pathways to Functional Selectivity. To view the other articles in this section visit http://dx.doi.org/10.1111/bph.2015.172.issue-2. © 2014 The British Pharmacological Society.

Structural mechanism underlying capsaicin binding and activation of TRPV1 ion channel

PubMed Central

Cheng, Wei; Yang, Wei; Yu, Peilin; Song, Zhenzhen; Yarov-Yarovoy, Vladimir; Zheng, Jie

2015-01-01

Capsaicin bestows spiciness by activating TRPV1 channel with exquisite potency and selectivity. Capsaicin-bound channel structure was previously resolved by cryo-EM at 4.2-to-4.5 Å resolution, however important details required for mechanistic understandings are unavailable: capsaicin was registered as a small electron density, reflecting neither its chemical structure nor specific ligand-channel interactions. We obtained the missing atomic-level details by iterative computation, which were confirmed by systematic site-specific functional tests. We observed that the bound capsaicin takes “tail-up, head-down” configurations. The vanillyl and amide groups form specific interactions to anchor its bound position, while the aliphatic tail may sample a range of conformations, making it invisible in cryo-EM images. Capsaicin stabilizes the open state by “pull-and-contact” interactions between the vanillyl group and the S4-S5 linker. Our study provided a structural mechanism for the agonistic function of capsaicin and its analogs, and demonstrated an effective approach to obtain atomic level information from cryo-EM structures. PMID:26053297

The Public Trustee: Ostrich, Mule or Owl?

ERIC Educational Resources Information Center

Pittenger, John C.

The role of state college trustees and their interaction with state government are considered. It is suggested that the trustee should be concerned about what is best for postsecondary education in the state generally as well as for the specific institution. The future trends indicate declining college enrollments, which require planning…

False Belief and Language Comprehension in Cantonese-Speaking Children

ERIC Educational Resources Information Center

Cheung, Him

2006-01-01

The current research compared two accounts of the relation between language and false belief in children, namely that (a) language is generally related to false belief because both require secondary representation in a social-interactional context and that (b) specific language structures that explicitly code meta representation contribute…

Kpna7 interacts with egg-specific nuclear factors in rainbow trout (Oncorhynchus mykiss)

USDA-ARS?s Scientific Manuscript database

Nuclear proteins are required for initiation of transcription in early embryos before embryonic genome activation. The regulation of transportation of nuclear proteins is mediated by transport factors known as importins (karyopherins). Kpna7 is a newly discovered member of the importin a family, whi...

Virus-induced gene silencing of WRKY53 and an inducible phenylalanine ammonia-lyase in wheat reduces aphid resistance

USDA-ARS?s Scientific Manuscript database

Although several wheat genes differentially expressed during the Russian wheat aphid resistance response have recently been identified, their requirement for and specific role in resistance remain unclear. Progress in wheat-aphid interaction research is hampered by inadequate collections of mutant g...

Is Speech Learning "Gated" by the Social Brain?

ERIC Educational Resources Information Center

Kuhl, Patricia K.

2007-01-01

I advance the hypothesis that the earliest phases of language acquisition--the developmental transition from an initial universal state of language processing to one that is language-specific--requires social interaction. Relating human language learning to a broader set of neurobiological cases of communicative development, I argue that the…

Mars Observer: Phase 0 safety review data package

NASA Technical Reports Server (NTRS)

1986-01-01

The Mars Observer Program has as its primary objectives a study of the geochemistry, atmospheric dynamics, atmosphere/surface interactions, seasonal variations, and magnetic field characteristics of Mars. The Mars Observer Spacecraft, safety critical spacecraft subsystems, ground support equipment, ground operations scenario, requirements matrix, and equipment specifications are described.

Development of a 13 kW Hall Thruster Propulsion System Performance Model for AEPS

NASA Technical Reports Server (NTRS)

Stanley, Steven; Allen, May; Goodfellow, Keith; Chew, Gilbert; Rapetti, Ryan; Tofil, Todd; Herman, Dan; Jackson, Jerry; Myers, Roger

2017-01-01

The Advanced Electric Propulsion System (AEPS) program will develop a flight 13kW Hall thruster propulsion system based on NASA's HERMeS thruster. The AEPS system includes the Hall Thruster, the Power Processing Unit (PPU) and the Xenon Flow Controller (XFC). These three primary components must operate together to ensure that the system generates the required combinations of thrust and specific impulse at the required system efficiencies for the desired system lifetime. At the highest level, the AEPS system will be integrated into the spacecraft and will receive power, propellant, and commands from the spacecraft. Power and propellant flow rates will be determined by the throttle set points commanded by the spacecraft. Within the system, the major control loop is between the mass flow rate and thruster current, with time-dependencies required to handle all expected transients, and additional, much slower interactions between the thruster and cathode temperatures, flow controller and PPU. The internal system interactions generally occur on shorter timescales than the spacecraft interactions, though certain failure modes may require rapid responses from the spacecraft. The AEPS system performance model is designed to account for all these interactions in a way that allows evaluation of the sensitivity of the system to expected changes over the planned mission as well as to assess the impacts of normal component and assembly variability during the production phase of the program. This effort describes the plan for the system performance model development, correlation to NASA test data, and how the model will be used to evaluate the critical internal and external interactions. The results will ensure the component requirements do not unnecessarily drive the system cost or overly constrain the development program. Finally, the model will be available to quickly troubleshoot any future unforeseen development challenges.

Choline Metabolites: Gene by Diet Interactions

PubMed Central

Smallwood, Tangi; Allayee, Hooman; Bennett, Brian J.

2015-01-01

Purpose of review This review highlights recent advances in our understanding of the interactions between genetic polymorphisms in genes that metabolize choline and the dietary requirements of choline and how these interactions relate to human health and disease. Recent findings The importance of choline as an essential nutrient has been well established but our appreciation of the interaction between our underlying genetic architecture and dietary choline requirements is only beginning. It has been shown in both human and animal studies that choline deficiencies contribute to diseases such as non-alcoholic fatty liver disease and various neurodegenerative diseases. An adequate supply of dietary choline is important for optimum development, highlighted by the increased maternal requirements during fetal development and in breast-fed infants. We discuss recent studies investigating variants in PEMT and MTHFR1 that are associated with a variety of birth defects. In addition to genetic interactions, we discuss several recent studies that uncover changes in fetal global methylation patterns in response to maternal dietary choline intake that result in changes in gene expression in the offspring. In contrast to the developmental role of adequate choline, there is now an appreciation of the role choline has in cardiovascular disease through the gut microbiota-mediated metabolite trimethylamine N-oxide. This pathway highlights some of our understanding of how the microbiome affects nutrient processing and bioavailability. Finally, in order to better characterize the genetic architecture regulating choline requirements, we discuss recent results focused on identifying polymorphisms that regulate choline and its derivative products. Summary Here we discuss recent studies that have advanced our understanding of how specific alleles in key choline metabolism genes are related to dietary choline requirements and human disease. PMID:26655287

FACT complex is required for DNA demethylation at heterochromatin during reproduction in Arabidopsis.

PubMed

Frost, Jennifer M; Kim, M Yvonne; Park, Guen Tae; Hsieh, Ping-Hung; Nakamura, Miyuki; Lin, Samuel J H; Yoo, Hyunjin; Choi, Jaemyung; Ikeda, Yoko; Kinoshita, Tetsu; Choi, Yeonhee; Zilberman, Daniel; Fischer, Robert L

2018-05-15

The DEMETER (DME) DNA glycosylase catalyzes genome-wide DNA demethylation and is required for endosperm genomic imprinting and embryo viability. Targets of DME-mediated DNA demethylation reside in small, euchromatic, AT-rich transposons and at the boundaries of large transposons, but how DME interacts with these diverse chromatin states is unknown. The STRUCTURE SPECIFIC RECOGNITION PROTEIN 1 (SSRP1) subunit of the chromatin remodeler FACT (facilitates chromatin transactions), was previously shown to be involved in the DME-dependent regulation of genomic imprinting in Arabidopsis endosperm. Therefore, to investigate the interaction between DME and chromatin, we focused on the activity of the two FACT subunits, SSRP1 and SUPPRESSOR of TY16 (SPT16), during reproduction in Arabidopsis We found that FACT colocalizes with nuclear DME in vivo, and that DME has two classes of target sites, the first being euchromatic and accessible to DME, but the second, representing over half of DME targets, requiring the action of FACT for DME-mediated DNA demethylation genome-wide. Our results show that the FACT-dependent DME targets are GC-rich heterochromatin domains with high nucleosome occupancy enriched with H3K9me2 and H3K27me1. Further, we demonstrate that heterochromatin-associated linker histone H1 specifically mediates the requirement for FACT at a subset of DME-target loci. Overall, our results demonstrate that FACT is required for DME targeting by facilitating its access to heterochromatin. Copyright © 2018 the Author(s). Published by PNAS.

A 'user friendly' geographic information system in a color interactive digital image processing system environment

NASA Technical Reports Server (NTRS)

Campbell, W. J.; Goldberg, M.

1982-01-01

NASA's Eastern Regional Remote Sensing Applications Center (ERRSAC) has recognized the need to accommodate spatial analysis techniques in its remote sensing technology transfer program. A computerized Geographic Information System to incorporate remotely sensed data, specifically Landsat, with other relevant data was considered a realistic approach to address a given resource problem. Questions arose concerning the selection of a suitable available software system to demonstrate, train, and undertake demonstration projects with ERRSAC's user community. The very specific requirements for such a system are discussed. The solution found involved the addition of geographic information processing functions to the Interactive Digital Image Manipulation System (IDIMS). Details regarding the functions of the new integrated system are examined along with the characteristics of the software.

Structures of the human Pals1 PDZ domain with and without ligand suggest gated access of Crb to the PDZ peptide-binding groove

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ivanova, Marina E.; Fletcher, Georgina C.; O’Reilly, Nicola

2015-03-01

This study characterizes the interaction between the carboxy-terminal (ERLI) motif of the essential polarity protein Crb and the Pals1/Stardust PDZ-domain protein. Structures of human Pals1 PDZ with and without a Crb peptide are described, explaining the highly conserved nature of the ERLI motif and revealing a sterically blocked peptide-binding groove in the absence of ligand. Many components of epithelial polarity protein complexes possess PDZ domains that are required for protein interaction and recruitment to the apical plasma membrane. Apical localization of the Crumbs (Crb) transmembrane protein requires a PDZ-mediated interaction with Pals1 (protein-associated with Lin7, Stardust, MPP5), a member ofmore » the p55 family of membrane-associated guanylate kinases (MAGUKs). This study describes the molecular interaction between the Crb carboxy-terminal motif (ERLI), which is required for Drosophila cell polarity, and the Pals1 PDZ domain using crystallography and fluorescence polarization. Only the last four Crb residues contribute to Pals1 PDZ-domain binding affinity, with specificity contributed by conserved charged interactions. Comparison of the Crb-bound Pals1 PDZ structure with an apo Pals1 structure reveals a key Phe side chain that gates access to the PDZ peptide-binding groove. Removal of this side chain enhances the binding affinity by more than fivefold, suggesting that access of Crb to Pals1 may be regulated by intradomain contacts or by protein–protein interaction.« less

Multidisciplinary model-based-engineering for laser weapon systems: recent progress

NASA Astrophysics Data System (ADS)

Coy, Steve; Panthaki, Malcolm

2013-09-01

We are working to develop a comprehensive, integrated software framework and toolset to support model-based engineering (MBE) of laser weapons systems. MBE has been identified by the Office of the Director, Defense Science and Engineering as one of four potentially "game-changing" technologies that could bring about revolutionary advances across the entire DoD research and development and procurement cycle. To be effective, however, MBE requires robust underlying modeling and simulation technologies capable of modeling all the pertinent systems, subsystems, components, effects, and interactions at any level of fidelity that may be required in order to support crucial design decisions at any point in the system development lifecycle. Very often the greatest technical challenges are posed by systems involving interactions that cut across two or more distinct scientific or engineering domains; even in cases where there are excellent tools available for modeling each individual domain, generally none of these domain-specific tools can be used to model the cross-domain interactions. In the case of laser weapons systems R&D these tools need to be able to support modeling of systems involving combined interactions among structures, thermal, and optical effects, including both ray optics and wave optics, controls, atmospheric effects, target interaction, computational fluid dynamics, and spatiotemporal interactions between lasing light and the laser gain medium. To address this problem we are working to extend Comet™, to add the addition modeling and simulation capabilities required for this particular application area. In this paper we will describe our progress to date.

Methods for the In Vitro Characterization of Nanomedicines—Biological Component Interaction

PubMed Central

Fornaguera, Cristina; Solans, Conxita

2017-01-01

The design of colloidal nanosystems intended for biomedical applications, specifically in the field of personalized medicine, has increased notably in the last years. Consequently, a variety of characterization techniques devoted to studying nanomedicine interactions with proteins and cells have been developed, since a deep characterization of nanosystems is required before starting preclinical and clinical studies. In this context, this review aims to summarize the main techniques used to assess the interaction of nanomedicines with biological systems, highlighting their advantages and disadvantages. Testing designed nanomaterials with these techniques is required in order to have more information about their behavior on a physiological environment. Moreover, techniques used to study the interaction of nanomedicines with proteins, such as albumin and fibrinogen, are summarized. These interactions are not desired, since they usually are the first signal to the body for the activation of the immune system, which leads to the clearance of the exogenous components. On the other hand, techniques for studying the cell toxicity of nanosystems are also summarized, since this information is required before starting preclinical steps. The translation of knowledge from novel designed nanosystems at a research laboratory scale to real human therapies is usually a limiting or even a final point due to the lack of systematic studies regarding these two aspects: nanoparticle interaction with biological components and nanoparticle cytotoxicity. In conclusion, this review will be a useful support for those scientists aiming to develop nanosystems for nanomedicine purposes. PMID:28134833

Simulation of blood flow in deformable vessels using subject-specific geometry and spatially varying wall properties

PubMed Central

Xiong, Guanglei; Figueroa, C. Alberto; Xiao, Nan; Taylor, Charles A.

2011-01-01

SUMMARY Simulation of blood flow using image-based models and computational fluid dynamics has found widespread application to quantifying hemodynamic factors relevant to the initiation and progression of cardiovascular diseases and for planning interventions. Methods for creating subject-specific geometric models from medical imaging data have improved substantially in the last decade but for many problems, still require significant user interaction. In addition, while fluid–structure interaction methods are being employed to model blood flow and vessel wall dynamics, tissue properties are often assumed to be uniform. In this paper, we propose a novel workflow for simulating blood flow using subject-specific geometry and spatially varying wall properties. The geometric model construction is based on 3D segmentation and geometric processing. Variable wall properties are assigned to the model based on combining centerline-based and surface-based methods. We finally demonstrate these new methods using an idealized cylindrical model and two subject-specific vascular models with thoracic and cerebral aneurysms. PMID:21765984

Hedgehog signaling and laminin play unique and synergistic roles in muscle development.

PubMed

Peterson, Matthew T; Henry, Clarissa A

2010-03-01

Hedgehog (Hh) signaling and laminin-111, a basement membrane protein, are required for early muscle development. Hh signaling specifies different populations of muscle fibers and laminin-111 is critical for early muscle morphogenesis. However, additional requirements for Hh signaling and laminin during later phases of muscle development are not known. Furthermore, interactions between Hh signaling and laminin in this context are unknown. We used laminin gamma1 mutant zebrafish and cyclopamine to block Hh signal transduction separately and in combination to investigate their functions and interactions. We found that both Hh signaling and laminin are required for normal myosin chain expression. In addition, Hh signaling and laminin act synergistically during fast-twitch fiber elongation: fast muscle cells do not elongate in embryos deficient for both Hh signaling and laminin. Finally, we present evidence that suggests that Hh signaling is indirectly required via slow fiber specification for recovery of fast fiber elongation in laminin gamma1 mutant embryos. Copyright (c) 2010 Wiley-Liss, Inc.

Space Station Freedom solar array panels plasma interaction test facility

NASA Technical Reports Server (NTRS)

Martin, Donald F.; Mellott, Kenneth D.

1989-01-01

The Space Station Freedom Power System will make extensive use of photovoltaic (PV) power generation. The phase 1 power system consists of two PV power modules each capable of delivering 37.5 KW of conditioned power to the user. Each PV module consists of two solar arrays. Each solar array is made up of two solar blankets. Each solar blanket contains 82 PV panels. The PV power modules provide a 160 V nominal operating voltage. Previous research has shown that there are electrical interactions between a plasma environment and a photovoltaic power source. The interactions take two forms: parasitic current loss (occurs when the currect produced by the PV panel leaves at a high potential point and travels through the plasma to a lower potential point, effectively shorting that portion of the PV panel); and arcing (occurs when the PV panel electrically discharges into the plasma). The PV solar array panel plasma interaction test was conceived to evaluate the effects of these interactions on the Space Station Freedom type PV panels as well as to conduct further research. The test article consists of two active solar array panels in series. Each panel consists of two hundred 8 cm x 8 cm silicon solar cells. The test requirements dictated specifications in the following areas: plasma environment/plasma sheath; outgassing; thermal requirements; solar simulation; and data collection requirements.

Distinct Mechanisms of Recognizing Endosomal Sorting Complex Required for Transport III (ESCRT-III) Protein IST1 by Different Microtubule Interacting and Trafficking (MIT) Domains*

PubMed Central

Guo, Emily Z.; Xu, Zhaohui

2015-01-01

The endosomal sorting complex required for transport (ESCRT) machinery is responsible for membrane remodeling in a number of biological processes including multivesicular body biogenesis, cytokinesis, and enveloped virus budding. In mammalian cells, efficient abscission during cytokinesis requires proper function of the ESCRT-III protein IST1, which binds to the microtubule interacting and trafficking (MIT) domains of VPS4, LIP5, and Spartin via its C-terminal MIT-interacting motif (MIM). Here, we studied the molecular interactions between IST1 and the three MIT domain-containing proteins to understand the structural basis that governs pairwise MIT-MIM interaction. Crystal structures of the three molecular complexes revealed that IST1 binds to the MIT domains of VPS4, LIP5, and Spartin using two different mechanisms (MIM1 mode versus MIM3 mode). Structural comparison revealed that structural features in both MIT and MIM contribute to determine the specific binding mechanism. Within the IST1 MIM sequence, two phenylalanine residues were shown to be important in discriminating MIM1 versus MIM3 binding. These observations enabled us to deduce a preliminary binding code, which we applied to provide CHMP2A, a protein that normally only binds the MIT domain in the MIM1 mode, the additional ability to bind the MIT domain of Spartin in the MIM3 mode. PMID:25657007

Array-based sensing using nanoparticles: an alternative approach for cancer diagnostics.

PubMed

Le, Ngoc D B; Yazdani, Mahdieh; Rotello, Vincent M

2014-07-01

Array-based sensing using nanoparticles (NPs) provides an attractive alternative to specific biomarker-focused strategies for cancer diagnosis. The physical and chemical properties of NPs provide both the recognition and transduction capabilities required for biosensing. Array-based sensors utilize a combined response from the interactions between sensors and analytes to generate a distinct pattern (fingerprint) for each analyte. These interactions can be the result of either the combination of multiple specific biomarker recognition (specific binding) or multiple selective binding responses, known as chemical nose sensing. The versatility of the latter array-based sensing using NPs can facilitate the development of new personalized diagnostic methodologies in cancer diagnostics, a necessary evolution in the current healthcare system to better provide personalized treatments. This review will describe the basic principle of array-based sensors, along with providing examples of both invasive and noninvasive samples used in cancer diagnosis.

ATP-independent reversal of a membrane protein aggregate by a chloroplast SRP

PubMed Central

Jaru-Ampornpan, Peera; Shen, Kuang; Lam, Vinh Q.; Ali, Mona; Doniach, Sebastian; Jia, Tony Z.; Shan, Shu-ou

2010-01-01

Membrane proteins impose enormous challenges to cellular protein homeostasis during their post-translational targeting, and require chaperones to keep them soluble and translocation-competent. Here we show that a novel targeting factor in the chloroplast Signal Recognition Particle (cpSRP), cpSRP43, is a highly specific molecular chaperone that efficiently reverses the aggregation of its substrate proteins. In contrast to AAA+-chaperones, cpSRP43 utilizes specific binding interactions with its substrate to mediate its disaggregase activity. This ‘disaggregase’ capability can allow targeting machineries to more effectively capture their protein substrates, and emphasizes a close connection between protein folding and trafficking processes. Moreover, cpSRP43 provides the first example of an ATP-independent disaggregase, and demonstrates that efficient reversal of protein aggregation can be attained by specific binding interactions between a chaperone and its substrate. PMID:20424608

Formal verification of a fault tolerant clock synchronization algorithm

NASA Technical Reports Server (NTRS)

Rushby, John; Vonhenke, Frieder

1989-01-01

A formal specification and mechanically assisted verification of the interactive convergence clock synchronization algorithm of Lamport and Melliar-Smith is described. Several technical flaws in the analysis given by Lamport and Melliar-Smith were discovered, even though their presentation is unusally precise and detailed. It seems that these flaws were not detected by informal peer scrutiny. The flaws are discussed and a revised presentation of the analysis is given that not only corrects the flaws but is also more precise and easier to follow. Some of the corrections to the flaws require slight modifications to the original assumptions underlying the algorithm and to the constraints on its parameters, and thus change the external specifications of the algorithm. The formal analysis of the interactive convergence clock synchronization algorithm was performed using the Enhanced Hierarchical Development Methodology (EHDM) formal specification and verification environment. This application of EHDM provides a demonstration of some of the capabilities of the system.

Management of optics. [for HEAO-2 X ray telescope

NASA Technical Reports Server (NTRS)

Kirchner, T. E.; Russell, M.

1981-01-01

American Science and Engineering, Inc., designed the large X-ray optic for the HEAO-2 X-ray Telescope. The key element in this project was the High Resolution Mirror Assembly (HRMA), subcontracting the fabrication of the optical surfaces and their assembly and alignment. The roles and organization of the key participants in the creation of HRMA are defined, and the degree of interaction between the groups is described. Management of this effort was extremely complex because of the intricate weaving of responsibilities, and AS&E, as HEAO-2 Program managers, needed to be well versed in the scientific objectives, the technical requirements, the program requirements, and the subcontract management. Understanding these factors was essential for implementing both technical and management controls, such as schedule and budget constraints, in-process control, residence requirements, and scientist review and feedback. Despite unforeseen technical problems and interaction differences, the HEAO-2 was built on schedule and to specification.

Drug-nutrient interactions in the intensive care unit: literature review and current recommendations.

PubMed

Heldt, Tatiane; Loss, Sergio Henrique

2013-01-01

To describe the interactions between drugs and nutrients and their frequency in the intensive care unit and to assess the professional team's awareness regarding this subject. The keywords "drug interactions" and "nutrition therapy" were searched in the PubMed (specifically MeSH) electronic database. The studies were systematically reviewed for descriptions of the types of interactions between drugs and nutrients, including their frequency and consequences. Sixty-seven articles were found. Among these, 20 articles were appropriate for the methodology adopted and accomplished the objectives of the study. Of these 20 articles, 14 articles described interactions between drugs and enteral nutrition, three described interactions between drugs and parenteral nutrition, and three described the importance and care required to avoid such interactions. The literature about drug and nutrient interactions is limited and suggests the inability of health care teams to recognize the potential for these interactions. Possibly, the elaboration of a protocol to evaluate drug-nutrient interactions will increase the safety and efficacy of therapeutics.

Drug-nutrient interactions in the intensive care unit: literature review and current recommendations

PubMed Central

Heldt, Tatiane; Loss, Sergio Henrique

2013-01-01

Objective To describe the interactions between drugs and nutrients and their frequency in the intensive care unit and to assess the professional team's awareness regarding this subject. Methods The keywords "drug interactions" and "nutrition therapy" were searched in the PubMed (specifically MeSH) electronic database. The studies were systematically reviewed for descriptions of the types of interactions between drugs and nutrients, including their frequency and consequences. Results Sixty-seven articles were found. Among these, 20 articles were appropriate for the methodology adopted and accomplished the objectives of the study. Of these 20 articles, 14 articles described interactions between drugs and enteral nutrition, three described interactions between drugs and parenteral nutrition, and three described the importance and care required to avoid such interactions. Conclusions The literature about drug and nutrient interactions is limited and suggests the inability of health care teams to recognize the potential for these interactions. Possibly, the elaboration of a protocol to evaluate drug-nutrient interactions will increase the safety and efficacy of therapeutics. PMID:23917982

Adenovirus E1A functions as a cofactor for retinoic acid receptor beta (RAR beta) through direct interaction with RAR beta.

PubMed

Folkers, G E; van der Saag, P T

1995-11-01

Transcription regulation by DNA-bound activators is thought to be mediated by a direct interaction between these proteins and TATA-binding protein (TBP), TFIIB, or TBP-associated factors, although occasionally cofactors or adapters are required. For ligand-induced activation by the retinoic acid receptor-retinoid X receptor (RAR-RXR) heterodimer, the RAR beta 2 promoter is dependent on the presence of E1A or E1A-like activity, since this promoter is activated by retinoic acid only in cells expressing such proteins. The mechanism underlying this E1A requirement is largely unknown. We now show that direct interaction between RAR and E1A is a requirement for retinoic acid-induced RAR beta 2 activation. The activity of the hormone-dependent activation function 2 (AF-2) of RAR beta is upregulated by E1A, and an interaction between this region and E1A was observed, but not with AF-1 or AF-2 of RXR alpha. This interaction is dependent on conserved region III (CRIII), the 13S mRNA-specific region of E1A. Deletion analysis within this region indicated that the complete CRIII is needed for activation. The putative zinc finger region is crucial, probably as a consequence of interaction with TBP. Furthermore, the region surrounding amino acid 178, partially overlapping with the TBP binding region, is involved in both binding to and activation by AF-2. We propose that E1A functions as a cofactor by interacting with both TBP and RAR, thereby stabilizing the preinitiation complex.

Determining the Specificity of Cascade Binding, Interference, and Primed Adaptation In Vivo in the Escherichia coli Type I-E CRISPR-Cas System

PubMed Central

Cooper, Lauren A.; Stringer, Anne M.

2018-01-01

ABSTRACT In clustered regularly interspaced short palindromic repeat (CRISPR)-Cas (CRISPR-associated) immunity systems, short CRISPR RNAs (crRNAs) are bound by Cas proteins, and these complexes target invading nucleic acid molecules for degradation in a process known as interference. In type I CRISPR-Cas systems, the Cas protein complex that binds DNA is known as Cascade. Association of Cascade with target DNA can also lead to acquisition of new immunity elements in a process known as primed adaptation. Here, we assess the specificity determinants for Cascade-DNA interaction, interference, and primed adaptation in vivo, for the type I-E system of Escherichia coli. Remarkably, as few as 5 bp of crRNA-DNA are sufficient for association of Cascade with a DNA target. Consequently, a single crRNA promotes Cascade association with numerous off-target sites, and the endogenous E. coli crRNAs direct Cascade binding to >100 chromosomal sites. In contrast to the low specificity of Cascade-DNA interactions, >18 bp are required for both interference and primed adaptation. Hence, Cascade binding to suboptimal, off-target sites is inert. Our data support a model in which the initial Cascade association with DNA targets requires only limited sequence complementarity at the crRNA 5′ end whereas recruitment and/or activation of the Cas3 nuclease, a prerequisite for interference and primed adaptation, requires extensive base pairing. PMID:29666291

Interactions between HIV-1 Gag and Viral RNA Genome Enhance Virion Assembly.

PubMed

Dilley, Kari A; Nikolaitchik, Olga A; Galli, Andrea; Burdick, Ryan C; Levine, Louis; Li, Kelvin; Rein, Alan; Pathak, Vinay K; Hu, Wei-Shau

2017-08-15

Most HIV-1 virions contain two copies of full-length viral RNA, indicating that genome packaging is efficient and tightly regulated. However, the structural protein Gag is the only component required for the assembly of noninfectious viruslike particles, and the viral RNA is dispensable in this process. The mechanism that allows HIV-1 to achieve such high efficiency of genome packaging when a packageable viral RNA is not required for virus assembly is currently unknown. In this report, we examined the role of HIV-1 RNA in virus assembly and found that packageable HIV-1 RNA enhances particle production when Gag is expressed at levels similar to those in cells containing one provirus. However, such enhancement is diminished when Gag is overexpressed, suggesting that the effects of viral RNA can be replaced by increased Gag concentration in cells. We also showed that the specific interactions between Gag and viral RNA are required for the enhancement of particle production. Taken together, these studies are consistent with our previous hypothesis that specific dimeric viral RNA-Gag interactions are the nucleation event of infectious virion assembly, ensuring that one RNA dimer is packaged into each nascent virion. These studies shed light on the mechanism by which HIV-1 achieves efficient genome packaging during virus assembly. IMPORTANCE Retrovirus assembly is a well-choreographed event, during which many viral and cellular components come together to generate infectious virions. The viral RNA genome carries the genetic information to new host cells, providing instructions to generate new virions, and therefore is essential for virion infectivity. In this report, we show that the specific interaction of the viral RNA genome with the structural protein Gag facilitates virion assembly and particle production. These findings resolve the conundrum that HIV-1 RNA is selectively packaged into virions with high efficiency despite being dispensable for virion assembly. Understanding the mechanism used by HIV-1 to ensure genome packaging provides significant insights into viral assembly and replication. Copyright © 2017 American Society for Microbiology.

Characterization of Type Three Secretion System Translocator Interactions with Phospholipid Membranes.

PubMed

Adam, Philip R; Barta, Michael L; Dickenson, Nicholas E

2017-01-01

In vitro characterization of type III secretion system (T3SS) translocator proteins has proven challenging due to complex purification schemes and their hydrophobic nature that often requires detergents to provide protein solubility and stability. Here, we provide experimental details for several techniques that overcome these hurdles, allowing for the direct characterization of the Shigella translocator protein IpaB with respect to phospholipid membrane interaction. The techniques specifically discussed in this chapter include membrane interaction/liposome flotation, liposome sensitive fluorescence quenching, and protein-mediated liposome disruption assays. These assays have provided valuable insight into the role of IpaB in T3SS-mediated phospholipid membrane interactions by Shigella and should readily extend to other members of this important class of proteins.

Dynamics of the Ternary Complex Formed by c-Myc Interactor JPO2, Transcriptional Co-activator LEDGF/p75, and Chromatin*

PubMed Central

Hendrix, Jelle; van Heertum, Bart; Vanstreels, Els; Daelemans, Dirk; De Rijck, Jan

2014-01-01

Lens epithelium-derived growth factor (LEDGF/p75) is a transcriptional co-activator involved in targeting human immunodeficiency virus (HIV) integration and the development of MLL fusion-mediated acute leukemia. A previous study revealed that LEDGF/p75 dynamically scans the chromatin, and upon interaction with HIV-1 integrase, their complex is locked on chromatin. At present, it is not known whether LEDGF/p75-mediated chromatin locking is typical for interacting proteins. Here, we employed continuous photobleaching and fluorescence correlation and cross-correlation spectroscopy to investigate in vivo chromatin binding of JPO2, a LEDGF/p75- and c-Myc-interacting protein involved in transcriptional regulation. In the absence of LEDGF/p75, JPO2 performs chromatin scanning inherent to transcription factors. However, whereas the dynamics of JPO2 chromatin binding are decelerated upon interaction with LEDGF/p75, very strong locking of their complex onto chromatin is absent. Similar results were obtained with the domesticated transposase PogZ, another cellular interaction partner of LEDGF/p75. We furthermore show that diffusive JPO2 can oligomerize; that JPO2 and LEDGF/p75 interact directly and specifically in vivo through the specific interaction domain of JPO2 and the C-terminal domain of LEDGF/p75, comprising the integrase-binding domain; and that modulation of JPO2 dynamics requires a functional PWWP domain in LEDGF/p75. Our results suggest that the dynamics of the LEDGF/p75-chromatin interaction depend on the specific partner and that strong chromatin locking is not a property of all LEDGF/p75-binding proteins. PMID:24634210

Developmental Stage of Parasites Influences the Structure of Fish-Parasite Networks

PubMed Central

Bellay, Sybelle; de Oliveira, Edson Fontes; Almeida-Neto, Mário; Lima Junior, Dilermando Pereira; Takemoto, Ricardo Massato; Luque, José Luis

2013-01-01

Specialized interactions tend to be more common in systems that require strong reciprocal adaptation between species, such as those observed between parasites and hosts. Parasites exhibit a high diversity of species and life history strategies, presenting host specificity which increases the complexity of these antagonistic systems. However, most studies are limited to the description of interactions between a few parasite and host species, which restricts our understanding of these systems as a whole. We investigated the effect of the developmental stage of the parasite on the structure of 30 metazoan fish-parasite networks, with an emphasis on the specificity of the interactions, connectance and modularity. We assessed the functional role of each species in modular networks and its interactions within and among the modules according to the developmental stage (larval and adult) and taxonomic group of the parasites. We observed that most parasite and host species perform a few interactions but that parasites at the larval stage tended to be generalists, increasing the network connectivity within and among modules. The parasite groups did not differ among each other in the number of interactions within and among the modules when considering only species at the larval stage. However, the same groups of adult individuals differed from each other in their interaction patterns, which were related to variations in the degree of host specificity at this stage. Our results show that the interaction pattern of fishes with parasites, such as acanthocephalans, cestodes, digeneans and nematodes, is more closely associated with their developmental stage than their phylogenetic history. This finding corroborates the hypothesis that the life history of parasites results in adaptations that cross phylogenetic boundaries. PMID:24124506

Developmental stage of parasites influences the structure of fish-parasite networks.

PubMed

Bellay, Sybelle; de Oliveira, Edson Fontes; Almeida-Neto, Mário; Lima Junior, Dilermando Pereira; Takemoto, Ricardo Massato; Luque, José Luis

2013-01-01

Specialized interactions tend to be more common in systems that require strong reciprocal adaptation between species, such as those observed between parasites and hosts. Parasites exhibit a high diversity of species and life history strategies, presenting host specificity which increases the complexity of these antagonistic systems. However, most studies are limited to the description of interactions between a few parasite and host species, which restricts our understanding of these systems as a whole. We investigated the effect of the developmental stage of the parasite on the structure of 30 metazoan fish-parasite networks, with an emphasis on the specificity of the interactions, connectance and modularity. We assessed the functional role of each species in modular networks and its interactions within and among the modules according to the developmental stage (larval and adult) and taxonomic group of the parasites. We observed that most parasite and host species perform a few interactions but that parasites at the larval stage tended to be generalists, increasing the network connectivity within and among modules. The parasite groups did not differ among each other in the number of interactions within and among the modules when considering only species at the larval stage. However, the same groups of adult individuals differed from each other in their interaction patterns, which were related to variations in the degree of host specificity at this stage. Our results show that the interaction pattern of fishes with parasites, such as acanthocephalans, cestodes, digeneans and nematodes, is more closely associated with their developmental stage than their phylogenetic history. This finding corroborates the hypothesis that the life history of parasites results in adaptations that cross phylogenetic boundaries.

AKAP18:PKA-RIIα structure reveals crucial anchor points for recognition of regulatory subunits of PKA

PubMed Central

Götz, Frank; Roske, Yvette; Schulz, Maike Svenja; Autenrieth, Karolin; Bertinetti, Daniela; Faelber, Katja; Zühlke, Kerstin; Kreuchwig, Annika; Kennedy, Eileen J.; Krause, Gerd; Daumke, Oliver; Herberg, Friedrich W.; Heinemann, Udo; Klussmann, Enno

2016-01-01

A-kinase anchoring proteins (AKAPs) interact with the dimerization/docking (D/D) domains of regulatory subunits of the ubiquitous protein kinase A (PKA). AKAPs tether PKA to defined cellular compartments establishing distinct pools to increase the specificity of PKA signalling. Here, we elucidated the structure of an extended PKA-binding domain of AKAP18β bound to the D/D domain of the regulatory RIIα subunits of PKA. We identified three hydrophilic anchor points in AKAP18β outside the core PKA-binding domain, which mediate contacts with the D/D domain. Such anchor points are conserved within AKAPs that bind regulatory RII subunits of PKA. We derived a different set of anchor points in AKAPs binding regulatory RI subunits of PKA. In vitro and cell-based experiments confirm the relevance of these sites for the interaction of RII subunits with AKAP18 and of RI subunits with the RI-specific smAKAP. Thus we report a novel mechanism governing interactions of AKAPs with PKA. The sequence specificity of each AKAP around the anchor points and the requirement of these points for the tight binding of PKA allow the development of selective inhibitors to unequivocally ascribe cellular functions to the AKAP18-PKA and other AKAP-PKA interactions. PMID:27102985

Hepatitis-C-virus-like internal ribosome entry sites displace eIF3 to gain access to the 40S subunit

NASA Astrophysics Data System (ADS)

Hashem, Yaser; Des Georges, Amedee; Dhote, Vidya; Langlois, Robert; Liao, Hstau Y.; Grassucci, Robert A.; Pestova, Tatyana V.; Hellen, Christopher U. T.; Frank, Joachim

2013-11-01

Hepatitis C virus (HCV) and classical swine fever virus (CSFV) messenger RNAs contain related (HCV-like) internal ribosome entry sites (IRESs) that promote 5'-end independent initiation of translation, requiring only a subset of the eukaryotic initiation factors (eIFs) needed for canonical initiation on cellular mRNAs. Initiation on HCV-like IRESs relies on their specific interaction with the 40S subunit, which places the initiation codon into the P site, where it directly base-pairs with eIF2-bound initiator methionyl transfer RNA to form a 48S initiation complex. However, all HCV-like IRESs also specifically interact with eIF3 (refs 2, 5, 6, 7, 9, 10, 11, 12), but the role of this interaction in IRES-mediated initiation has remained unknown. During canonical initiation, eIF3 binds to the 40S subunit as a component of the 43S pre-initiation complex, and comparison of the ribosomal positions of eIF3 and the HCV IRES revealed that they overlap, so that their rearrangement would be required for formation of ribosomal complexes containing both components. Here we present a cryo-electron microscopy reconstruction of a 40S ribosomal complex containing eIF3 and the CSFV IRES. Remarkably, although the position and interactions of the CSFV IRES with the 40S subunit in this complex are similar to those of the HCV IRES in the 40S-IRES binary complex, eIF3 is completely displaced from its ribosomal position in the 43S complex, and instead interacts through its ribosome-binding surface exclusively with the apical region of domain III of the IRES. Our results suggest a role for the specific interaction of HCV-like IRESs with eIF3 in preventing ribosomal association of eIF3, which could serve two purposes: relieving the competition between the IRES and eIF3 for a common binding site on the 40S subunit, and reducing formation of 43S complexes, thereby favouring translation of viral mRNAs.

HCV-like IRESs displace eIF3 to gain access to the 40S subunit

PubMed Central

Hashem, Yaser; des Georges, Amedee; Dhote, Vidya; Langlois, Robert; Liao, Hstau Y.; Grassucci, Robert A.; Pestova, Tatyana V.; Hellen, Christopher U.T.; Frank, Joachim

2014-01-01

Hepatitis C virus (HCV) and Classical swine fever virus (CSFV) mRNAs contain related (HCV-like) internal ribosome entry sites (IRESs) that promote 5’-end independent initiation of translation, requiring only a subset of the eukaryotic initiation factors (eIFs) needed for canonical initiation on cellular mRNAs1. Initiation on HCV-like IRESs relies on their specific interaction with the 40S subunit2–8, which places the initiation codon into the P site, where it directly base-pairs with eIF2-bound Met-tRNAiMet to form a 48S initiation complex. However, all HCV-like IRESs also specifically interact with eIF32,5–7,9–12, but the role of this interaction in IRES-mediated initiation has remained unknown. During canonical initiation, eIF3 binds to the 40S subunit as a component of the 43S pre-initiation complex, and comparison of the ribosomal positions of eIF313 and the HCV IRES8 revealed that they overlap, so that their rearrangement would be required for formation of ribosomal complexes containing both components13. Here, we present a cryo-electron microscopy reconstruction of a 40S ribosomal complex containing eIF3 and the CSFV IRES. Strikingly, although the position and interactions of the CSFV IRES with the 40S subunit in this complex are similar to those of the HCV IRES in the 40S/IRES binary complex8, eIF3 is completely displaced from its ribosomal position in the 43S complex, and instead interacts through its ribosome-binding surface exclusively with the apical region of domain III of the IRES. Our results suggest a role for the specific interaction of HCV-like IRESs with eIF3 in preventing ribosomal association of eIF3, which could serve two purposes: relieving the competition between the IRES and eIF3 for a common binding site on the 40S subunit, and reducing formation of 43S complexes, thereby favoring translation of viral mRNAs. PMID:24185006

Obscurin Targets Ankyrin-B and Protein Phosphatase 2A to the Cardiac M-line*

PubMed Central

Cunha, Shane R.; Mohler, Peter J.

2008-01-01

Ankyrin-B targets ion channels and transporters in excitable cells. Dysfunction in ankyrin-B-based pathways results in defects in cardiac physiology. Despite a wealth of knowledge regarding the role of ankyrin-B for cardiac function, little is known regarding the mechanisms underlying ankyrin-B regulation. Moreover, the pathways underlying ankyrin-B targeting in heart are unclear. We report that alternative splicing regulates ankyrin-B localization and function in cardiomyocytes. Specifically, we identify a novel exon (exon 43′) in the ankyrin-B regulatory domain that mediates interaction with the Rho-GEF obscurin. Ankyrin-B transcripts harboring exon 43′ represent the primary cardiac isoform in human and mouse. We demonstrate that ankyrin-B and obscurin are co-localized at the M-line of myocytes and co-immunoprecipitate from heart. We define the structural requirements for ankyrin-B/obscurin interaction to two motifs in the ankyrin-B regulatory domain and demonstrate that both are critical for obscurin/ankyrin-B interaction. In addition, we demonstrate that interaction with obscurin is required for ankyrin-B M-line targeting. Specifically, both obscurin-binding motifs are required for the M-line targeting of a GFP-ankyrin-B regulatory domain. Moreover, this construct acts as a dominant-negative by competing with endogenous ankyrin-B for obscurin-binding at the M-line, thus providing a powerful new tool to evaluate the function of obscurin/ankyrin-B interactions. With this new tool, we demonstrate that the obscurin/ankyrin-B interaction is critical for recruitment of PP2A to the cardiac M-line. Together, these data provide the first evidence for the molecular basis of ankyrin-B and PP2A targeting and function at the cardiac M-line. Finally, we report that ankyrin-B R1788W is localized adjacent to the ankyrin-B obscurin-binding motif and increases binding activity for obscurin. In summary, our new findings demonstrate that ANK2 is subject to alternative splicing that gives rise to unique polypeptides with diverse roles in cardiac function. PMID:18782775

Mapping regions of Epstein-Barr virus (EBV) glycoprotein B (gB) important for fusion function with gH/gL

DOE Office of Scientific and Technical Information (OSTI.GOV)

Plate, Aileen E.; Reimer, Jessica J.; Jardetzky, Theodore S.

Glycoproteins gB and gH/gL are required for entry of Epstein-Barr virus (EBV) into cells, but the role of each glycoprotein and how they function together to mediate fusion is unclear. Analysis of the functional homology of gB from the closely related primate gammaherpesvirus, rhesus lymphocryptovirus (Rh-LCV), showed that EBV gB could not complement Rh gB due to a species-specific dependence between gB and gL. To map domains of gB required for this interaction, we constructed a panel of EBV/Rh gB chimeric proteins. Analysis showed that insertion of Rh gB from residues 456 to 807 restored fusion function of EBV gBmore » with Rh gH/gL, suggesting this region of gB is important for interaction with gH/gL. Split YFP bimolecular complementation (BiFC) provided evidence of an interaction between EBV gB and gH/gL. Together, our results suggest the importance of a gB-gH/gL interaction in EBV-mediated fusion with B cells requiring the region of EBV gB from 456 to 807.« less

Specificity and autoregulation of Notch binding by tandem WW domains in suppressor of Deltex.

PubMed

Jennings, Martin D; Blankley, Richard T; Baron, Martin; Golovanov, Alexander P; Avis, Johanna M

2007-09-28

WW domains target proline-tyrosine (PY) motifs and frequently function as tandem pairs. When studied in isolation, single WW domains are notably promiscuous and regulatory mechanisms are undoubtedly required to ensure selective interactions. Here, we show that the fourth WW domain (WW4) of Suppressor of Deltex, a modular Nedd4-like protein that down-regulates the Notch receptor, is the primary mediator of a direct interaction with a Notch-PY motif. A natural Trp to Phe substitution in WW4 reduces its affinity for general PY sequences and enhances selective interaction with the Notch-PY motif via compensatory specificity-determining interactions with PY-flanking residues. When WW4 is paired with WW3, domain-domain association, impeding proper folding, competes with Notch-PY binding to WW4. This novel mode of autoinhibition is relieved by binding of another ligand to WW3. Such cooperativity may facilitate the transient regulatory interactions observed in vivo between Su(dx) and Notch in the endocytic pathway. The highly conserved tandem arrangement of WW domains in Nedd4 proteins, and similar arrangements in more diverse proteins, suggests domain-domain communication may be integral to regulation of their associated cellular activities.

Flavivirus Replication Complex Assembly Revealed by DNAJC14 Functional Mapping

PubMed Central

Yi, Zhigang; Yuan, Zhenghong; Rice, Charles M.

2012-01-01

DNAJC14 is an Hsp40 family member that broadly modulates flavivirus replication. The mechanism by which DNAJC14 stoichiometrically participates in flavivirus replication complex (RC) formation is unknown; both reduced and elevated levels result in replication inhibition. Using yellow fever virus (YFV), we demonstrate that DNAJC14 redistributes and clusters with YFV nonstructural proteins via a transmembrane domain and a newly identified membrane-binding domain (MBD), which both mediate targeting to detergent-resistant membranes. Furthermore, the RC and DNAJC14 reside as part of a protein interaction network that remains after 1% Triton solubilization. Mutagenesis studies demonstrate that entry into this protein interaction network requires the DNAJC14 C-terminal self-interaction domain. Fusion of the DNAJC14 MBD and self-interaction domain with another Hsp40 family protein is sufficient to confer YFV-inhibitory activity. Our findings support a novel model of DNAJC14 action that includes specific membrane targeting of both DNAJC14 and YFV replication proteins, the formation of protein interactions, and a microdomain-specific chaperone event leading to RC formation. This process alters the properties of the RC membrane and results in the formation of a protein scaffold that maintains the RC. PMID:22915803

A high-confidence interaction map identifies SIRT1 as a mediator of acetylation of USP22 and the SAGA coactivator complex.

PubMed

Armour, Sean M; Bennett, Eric J; Braun, Craig R; Zhang, Xiao-Yong; McMahon, Steven B; Gygi, Steven P; Harper, J Wade; Sinclair, David A

2013-04-01

Although many functions and targets have been attributed to the histone and protein deacetylase SIRT1, a comprehensive analysis of SIRT1 binding proteins yielding a high-confidence interaction map has not been established. Using a comparative statistical analysis of binding partners, we have assembled a high-confidence SIRT1 interactome. Employing this method, we identified the deubiquitinating enzyme ubiquitin-specific protease 22 (USP22), a component of the deubiquitinating module (DUBm) of the SAGA transcriptional coactivating complex, as a SIRT1-interacting partner. We found that this interaction is highly specific, requires the ZnF-UBP domain of USP22, and is disrupted by the inactivating H363Y mutation within SIRT1. Moreover, we show that USP22 is acetylated on multiple lysine residues and that alteration of a single lysine (K129) within the ZnF-UBP domain is sufficient to alter interaction of the DUBm with the core SAGA complex. Furthermore, USP22-mediated recruitment of SIRT1 activity promotes the deacetylation of individual SAGA complex components. Our results indicate an important role of SIRT1-mediated deacetylation in regulating the formation of DUBm subcomplexes within the larger SAGA complex.

Selective redox regulation of cytokine receptor signaling by extracellular thioredoxin-1

PubMed Central

Schwertassek, Ulla; Balmer, Yves; Gutscher, Marcus; Weingarten, Lars; Preuss, Marc; Engelhard, Johanna; Winkler, Monique; Dick, Tobias P

2007-01-01

The thiol-disulfide oxidoreductase thioredoxin-1 (Trx1) is known to be secreted by leukocytes and to exhibit cytokine-like properties. Extracellular effects of Trx1 require a functional active site, suggesting a redox-based mechanism of action. However, specific cell surface proteins and pathways coupling extracellular Trx1 redox activity to cellular responses have not been identified so far. Using a mechanism-based kinetic trapping technique to identify disulfide exchange interactions on the intact surface of living lymphocytes, we found that Trx1 catalytically interacts with a single principal target protein. This target protein was identified as the tumor necrosis factor receptor superfamily member 8 (TNFRSF8/CD30). We demonstrate that the redox interaction is highly specific for both Trx1 and CD30 and that the redox state of CD30 determines its ability to engage the cognate ligand and transduce signals. Furthermore, we confirm that Trx1 affects CD30-dependent changes in lymphocyte effector function. Thus, we conclude that receptor–ligand signaling interactions can be selectively regulated by an extracellular redox catalyst. PMID:17557078

L1CAM/Neuroglian controls the axon–axon interactions establishing layered and lobular mushroom body architecture

PubMed Central

Siegenthaler, Dominique; Enneking, Eva-Maria; Moreno, Eliza

2015-01-01

The establishment of neuronal circuits depends on the guidance of axons both along and in between axonal populations of different identity; however, the molecular principles controlling axon–axon interactions in vivo remain largely elusive. We demonstrate that the Drosophila melanogaster L1CAM homologue Neuroglian mediates adhesion between functionally distinct mushroom body axon populations to enforce and control appropriate projections into distinct axonal layers and lobes essential for olfactory learning and memory. We addressed the regulatory mechanisms controlling homophilic Neuroglian-mediated cell adhesion by analyzing targeted mutations of extra- and intracellular Neuroglian domains in combination with cell type–specific rescue assays in vivo. We demonstrate independent and cooperative domain requirements: intercalating growth depends on homophilic adhesion mediated by extracellular Ig domains. For functional cluster formation, intracellular Ankyrin2 association is sufficient on one side of the trans-axonal complex whereas Moesin association is likely required simultaneously in both interacting axonal populations. Together, our results provide novel mechanistic insights into cell adhesion molecule–mediated axon–axon interactions that enable precise assembly of complex neuronal circuits. PMID:25825519

L1CAM/Neuroglian controls the axon-axon interactions establishing layered and lobular mushroom body architecture.

PubMed

Siegenthaler, Dominique; Enneking, Eva-Maria; Moreno, Eliza; Pielage, Jan

2015-03-30

The establishment of neuronal circuits depends on the guidance of axons both along and in between axonal populations of different identity; however, the molecular principles controlling axon-axon interactions in vivo remain largely elusive. We demonstrate that the Drosophila melanogaster L1CAM homologue Neuroglian mediates adhesion between functionally distinct mushroom body axon populations to enforce and control appropriate projections into distinct axonal layers and lobes essential for olfactory learning and memory. We addressed the regulatory mechanisms controlling homophilic Neuroglian-mediated cell adhesion by analyzing targeted mutations of extra- and intracellular Neuroglian domains in combination with cell type-specific rescue assays in vivo. We demonstrate independent and cooperative domain requirements: intercalating growth depends on homophilic adhesion mediated by extracellular Ig domains. For functional cluster formation, intracellular Ankyrin2 association is sufficient on one side of the trans-axonal complex whereas Moesin association is likely required simultaneously in both interacting axonal populations. Together, our results provide novel mechanistic insights into cell adhesion molecule-mediated axon-axon interactions that enable precise assembly of complex neuronal circuits. © 2015 Siegenthaler et al.

Gene environment interaction studies in depression and suicidal behavior: An update.

PubMed

Mandelli, Laura; Serretti, Alessandro

2013-12-01

Increasing evidence supports the involvement of both heritable and environmental risk factors in major depression (MD) and suicidal behavior (SB). Studies investigating gene-environment interaction (G × E) may be useful for elucidating the role of biological mechanisms in the risk for mental disorders. In the present paper, we review the literature regarding the interaction between genes modulating brain functions and stressful life events in the etiology of MD and SB and discuss their potential added benefit compared to genetic studies only. Within the context of G × E investigation, thus far, only a few reliable results have been obtained, although some genes have consistently shown interactive effects with environmental risk in MD and, to a lesser extent, in SB. Further investigation is required to disentangle the direct and mediated effects that are common or specific to MD and SB. Since traditional G × E studies overall suffer from important methodological limitations, further effort is required to develop novel methodological strategies with an interdisciplinary approach. Copyright © 2013 Elsevier Ltd. All rights reserved.

A race-specific interaction between vitamin K status and statin use during warfarin therapy initiation

USDA-ARS?s Scientific Manuscript database

Vitamin K (VK) is required for the post-translational modification of several clotting factors. Warfarin is a vitamin K antagonist and anticoagulant. The most common dietary and circulating form of VK is phylloquinone (PK). PK is lipid soluble, carried by triglyceride-rich lipoproteins, and shares a...

Learning Soft Skills at Work: An Interview with Annalee Luhman

ERIC Educational Resources Information Center

Davis, Barbara D.; Muir, Clive

2004-01-01

Soft skills are attitudes and behaviors displayed in interactions among individuals that affect the outcomes of such encounters. These differ from hard skills, which are the technical knowledge and abilities required to perform specific job-related tasks more formally stated in job descriptions. In the past, it was felt that managers and employees…

Conditions of Educational Environment for the Development of Teenagers' Moral Relations

ERIC Educational Resources Information Center

Khuziakhmetov, Anvar N.; Shafikova, Gulnaz R.; Kapranova, Vera A.

2015-01-01

The relevance of the study is conditioned by the fact that social relations require from a school graduator not only the subject specific competences, but also the ability to build interpersonal interaction based on moral norms. Therefore, this article is aimed to reveal the conditions of the school educational environment organization promoting…

In Search of a New Paradigm for Teaching English as an International Language

ERIC Educational Resources Information Center

Canagarajah, Suresh

2014-01-01

In the context of more diverse communicative practices and social relations in globalization, scholars are increasingly defining English as constituting socially constructed situational norms in specific contexts of interaction, and not a homogeneous language or even discrete varieties of English. This shift requires treating pragmatics and not…

Team-Based Learning in Anatomy: An Efficient, Effective, and Economical Strategy

ERIC Educational Resources Information Center

Vasan, Nagaswami S.; DeFouw, David O.; Compton, Scott

2011-01-01

Team-based learning (TBL) strategy is being adopted in medical education to implement interactive small group learning. We have modified classical TBL to fit our curricular needs and approach. Anatomy lectures were replaced with TBL that required preparation of assigned content specific discussion topics (in the text referred as "discussion…

Teaching through Interactive Pictures: Computer, Video and Human Realities.

ERIC Educational Resources Information Center

Strauss, Andre

A technique designed to make the classroom use of videotapes for second language teaching for specific purposes more efficient is described. The technique begins with a classroom review of basic vocabulary and structures, which is then tested with a computer exercise. A second stage requires discussion and memorization of vocabulary and phrases…

Intercultural Training for Business Expatriates: Cultural-Specific Training Needs of US Business Expatriates on Assignment to Taiwan

ERIC Educational Resources Information Center

Chien, Tien-Chen; McLean, Gary N.

2004-01-01

US companies' expansion across political boundaries requires personnel to work in foreign facilities with vastly different local cultures. Intercultural training facilitates adjustment to the foreign environment and improved interaction with host country nationals. This study assessed US expatriates and their Taiwanese colleagues and identified…

Can Being Scared Cause Tummy Aches? Naive Theories, Ambiguous Evidence, and Preschoolers' Causal Inferences

ERIC Educational Resources Information Center

Schulz, Laura E.; Bonawitz, Elizabeth Baraff; Griffiths, Thomas L.

2007-01-01

Causal learning requires integrating constraints provided by domain-specific theories with domain-general statistical learning. In order to investigate the interaction between these factors, the authors presented preschoolers with stories pitting their existing theories against statistical evidence. Each child heard 2 stories in which 2 candidate…

Digital Technology in Teaching International Business: Is a Tradeoff between Richness and Reach Required?

ERIC Educational Resources Information Center

Wymbs, Cliff; Kijne, Hugo

2003-01-01

This analysis extends the traditional marketing tradeoffs between richness (depth of knowledge) and reach (geographic area coverage) to the emerging technology-mediated education industry, and then specifically evaluates their effect on the teaching of international business. It asserts that interactive learning, particularly as it applies to team…

A three-tiered approach for linking pharmacokinetic ...

EPA Pesticide Factsheets

The power of the adverse outcome pathway (AOP) framework arises from its utilization of pathway-based data to describe the initial interaction of a chemical with a molecular target (molecular initiating event; (MIE), followed by a progression through a series of key events that lead to an adverse outcome relevant for regulatory purposes. The AOP itself is not chemical specific, thus providing the biological context necessary for interpreting high throughput (HT) toxicity screening results. Application of the AOP framework and HT predictions in ecological and human health risk assessment, however, requires the consideration of chemical-specific properties that influence external exposure doses and target tissue doses. To address this requirement, a three-tiered approach was developed to provide a workflow for connecting biology-based AOPs to biochemical-based pharmacokinetic properties (absorption, distribution, metabolism, excretion; ADME), and then to chemical/human activity-based exposure pathways. This approach included: (1) The power of the adverse outcome pathway (AOP) framework arisesfrom its utilization of pathway-based data to describe the initial interaction of a chemical with a molecular target (molecular initiating event; (MIE), followed by a progression through a series of key events that lead to an adverse outcome relevant for regulatory purposes. The AOP itself is not chemical specific, thus providing the biological context necessary for interpreti

Advanced aeroservoelastic stabilization techniques for hypersonic flight vehicles

NASA Technical Reports Server (NTRS)

Chan, Samuel Y.; Cheng, Peter Y.; Myers, Thomas T.; Klyde, David H.; Magdaleno, Raymond E.; Mcruer, Duane T.

1992-01-01

Advanced high performance vehicles, including Single-Stage-To-Orbit (SSTO) hypersonic flight vehicles, that are statically unstable, require higher bandwidth flight control systems to compensate for the instability resulting in interactions between the flight control system, the engine/propulsion dynamics, and the low frequency structural modes. Military specifications, such as MIL-F-9490D and MIL-F-87242, tend to limit treatment of structural modes to conventional gain stabilization techniques. The conventional gain stabilization techniques, however, introduce low frequency effective time delays which can be troublesome from a flying qualities standpoint. These time delays can be alleviated by appropriate blending of gain and phase stabilization techniques (referred to as Hybrid Phase Stabilization or HPS) for the low frequency structural modes. The potential of using HPS for compensating structural mode interaction was previously explored. It was shown that effective time delay was significantly reduced with the use of HPS; however, the HPS design was seen to have greater residual response than a conventional gain stablized design. Additional work performed to advance and refine the HPS design procedure, to further develop residual response metrics as a basis for alternative structural stability specifications, and to develop strategies for validating HPS design and specification concepts in manned simulation is presented. Stabilization design sensitivity to structural uncertainties and aircraft-centered requirements are also assessed.

The RSF1 Histone-Remodelling Factor Facilitates DNA Double-Strand Break Repair by Recruiting Centromeric and Fanconi Anaemia Proteins

PubMed Central

Pessina, Fabio; Lowndes, Noel F.

2014-01-01

ATM is a central regulator of the cellular responses to DNA double-strand breaks (DSBs). Here we identify a biochemical interaction between ATM and RSF1 and we characterise the role of RSF1 in this response. The ATM–RSF1 interaction is dependent upon both DSBs and ATM kinase activity. Together with SNF2H/SMARCA5, RSF1 forms the RSF chromatin-remodelling complex. Although RSF1 is specific to the RSF complex, SNF2H/SMARCA5 is a catalytic subunit of several other chromatin-remodelling complexes. Although not required for checkpoint signalling, RSF1 is required for efficient repair of DSBs via both end-joining and homology-directed repair. Specifically, the ATM-dependent recruitment to sites of DSBs of the histone fold proteins CENPS/MHF1 and CENPX/MHF2, previously identified at centromeres, is RSF1-dependent. In turn these proteins recruit and regulate the mono-ubiquitination of the Fanconi Anaemia proteins FANCD2 and FANCI. We propose that by depositing CENPS/MHF1 and CENPX/MHF2, the RSF complex either directly or indirectly contributes to the reorganisation of chromatin around DSBs that is required for efficient DNA repair. PMID:24800743

Spacecraft Environmental Interactions Technology 1983

DTIC Science & Technology

1985-01-01

sr. 144 S+ +. + ’ J9 • "=• _Z7 `16~ r’r f,"::,irC 14 0 ral Irl -r -4: 3J’ r. .4 1 .. i. . Figure 3. - Isocontour maps as in figiire 1 for the average ...Geophysics Laboratory ....... . . . . . . . . . .. 125 AVERAGE AND WORST-CASE SPECIFICATIONS OF PRECIPITATING AURORAL ELECTRON ENVIRONMENT David A...force now, In volume S we d( ’elop a road- map of how a technology program would co-tinue to develop the technology base required for the specific

Affinity adsorption of cells to surfaces and strategies for cell detachment.

PubMed

Hubble, John

2007-01-01

The use of bio-specific interactions for the separation and recovery of bio-molecules is now widely established and in many cases the technique has successfully crossed the divide between bench and process scale operation. Although the major specificity advantage of affinity-based separations also applies to systems intended for cell fractionation, developments in this area have been slower. Many of the problems encountered result from attempts to take techniques developed for molecular systems and, with only minor modification to the conditions used, apply them for the separation of cells. This approach tends to ignore or at least trivialise the problems, which arise from the heterogeneous nature of a cell suspension and the multivalent nature of the cell/surface interaction. To develop viable separation processes on a larger scale, effective contacting strategies are required in separators that also allow detachment or recovery protocols that overcome the enhanced binding strength generated by multivalent interactions. The effects of interaction valency on interaction strength needs to be assessed and approaches developed to allow effective detachment and recovery of adsorbed cells without compromising cell viability. This article considers the influence of operating conditions on cell attachment and the extent to which multivalent interactions determine the strength of cell binding and subsequent detachment.

Global investigation of protein-protein interactions in yeast Saccharomyces cerevisiae using re-occurring short polypeptide sequences.

PubMed

Pitre, S; North, C; Alamgir, M; Jessulat, M; Chan, A; Luo, X; Green, J R; Dumontier, M; Dehne, F; Golshani, A

2008-08-01

Protein-protein interaction (PPI) maps provide insight into cellular biology and have received considerable attention in the post-genomic era. While large-scale experimental approaches have generated large collections of experimentally determined PPIs, technical limitations preclude certain PPIs from detection. Recently, we demonstrated that yeast PPIs can be computationally predicted using re-occurring short polypeptide sequences between known interacting protein pairs. However, the computational requirements and low specificity made this method unsuitable for large-scale investigations. Here, we report an improved approach, which exhibits a specificity of approximately 99.95% and executes 16,000 times faster. Importantly, we report the first all-to-all sequence-based computational screen of PPIs in yeast, Saccharomyces cerevisiae in which we identify 29,589 high confidence interactions of approximately 2 x 10(7) possible pairs. Of these, 14,438 PPIs have not been previously reported and may represent novel interactions. In particular, these results reveal a richer set of membrane protein interactions, not readily amenable to experimental investigations. From the novel PPIs, a novel putative protein complex comprised largely of membrane proteins was revealed. In addition, two novel gene functions were predicted and experimentally confirmed to affect the efficiency of non-homologous end-joining, providing further support for the usefulness of the identified PPIs in biological investigations.

Nanophotonic detection of freely interacting molecules on a single influenza virus

NASA Astrophysics Data System (ADS)

Kang, Pilgyu; Schein, Perry; Serey, Xavier; O'Dell, Dakota; Erickson, David

2015-07-01

Biomolecular interactions, such as antibody-antigen binding, are fundamental to many biological processes. At present, most techniques for analyzing these interactions require immobilizing one or both of the interacting molecules on an assay plate or a sensor surface. This is convenient experimentally but can constrain the natural binding affinity and capacity of the molecules, resulting in data that can deviate from the natural free-solution behavior. Here we demonstrate a label-free method for analyzing free-solution interactions between a single influenza virus and specific antibodies at the single particle level using near-field optical trapping and light-scattering techniques. We determine the number of specific antibodies binding to an optically trapped influenza virus by analyzing the change of the Brownian fluctuations of the virus. We develop an analytical model that determines the increased size of the virus resulting from antibodies binding to the virus membrane with uncertainty of ±1-2 nm. We present stoichiometric results of 26 ± 4 (6.8 ± 1.1 attogram) anti-influenza antibodies binding to an H1N1 influenza virus. Our technique can be applied to a wide range of molecular interactions because the nanophotonic tweezer can handle molecules from tens to thousands of nanometers in diameter.

Smart Swarms of Bacteria-Inspired Agents with Performance Adaptable Interactions

PubMed Central

Shklarsh, Adi; Ariel, Gil; Schneidman, Elad; Ben-Jacob, Eshel

2011-01-01

Collective navigation and swarming have been studied in animal groups, such as fish schools, bird flocks, bacteria, and slime molds. Computer modeling has shown that collective behavior of simple agents can result from simple interactions between the agents, which include short range repulsion, intermediate range alignment, and long range attraction. Here we study collective navigation of bacteria-inspired smart agents in complex terrains, with adaptive interactions that depend on performance. More specifically, each agent adjusts its interactions with the other agents according to its local environment – by decreasing the peers' influence while navigating in a beneficial direction, and increasing it otherwise. We show that inclusion of such performance dependent adaptable interactions significantly improves the collective swarming performance, leading to highly efficient navigation, especially in complex terrains. Notably, to afford such adaptable interactions, each modeled agent requires only simple computational capabilities with short-term memory, which can easily be implemented in simple swarming robots. PMID:21980274

Smart swarms of bacteria-inspired agents with performance adaptable interactions.

PubMed

Shklarsh, Adi; Ariel, Gil; Schneidman, Elad; Ben-Jacob, Eshel

2011-09-01

Collective navigation and swarming have been studied in animal groups, such as fish schools, bird flocks, bacteria, and slime molds. Computer modeling has shown that collective behavior of simple agents can result from simple interactions between the agents, which include short range repulsion, intermediate range alignment, and long range attraction. Here we study collective navigation of bacteria-inspired smart agents in complex terrains, with adaptive interactions that depend on performance. More specifically, each agent adjusts its interactions with the other agents according to its local environment--by decreasing the peers' influence while navigating in a beneficial direction, and increasing it otherwise. We show that inclusion of such performance dependent adaptable interactions significantly improves the collective swarming performance, leading to highly efficient navigation, especially in complex terrains. Notably, to afford such adaptable interactions, each modeled agent requires only simple computational capabilities with short-term memory, which can easily be implemented in simple swarming robots.

Integrating distributed multimedia systems and interactive television networks

NASA Astrophysics Data System (ADS)

Shvartsman, Alex A.

1996-01-01

Recent advances in networks, storage and video delivery systems are about to make commercial deployment of interactive multimedia services over digital television networks a reality. The emerging components individually have the potential to satisfy the technical requirements in the near future. However, no single vendor is offering a complete end-to-end commercially-deployable and scalable interactive multimedia applications systems over digital/analog television systems. Integrating a large set of maturing sub-assemblies and interactive multimedia applications is a major task in deploying such systems. Here we deal with integration issues, requirements and trade-offs in building delivery platforms and applications for interactive television services. Such integration efforts must overcome lack of standards, and deal with unpredictable development cycles and quality problems of leading- edge technology. There are also the conflicting goals of optimizing systems for video delivery while enabling highly interactive distributed applications. It is becoming possible to deliver continuous video streams from specific sources, but it is difficult and expensive to provide the ability to rapidly switch among multiple sources of video and data. Finally, there is the ever- present challenge of integrating and deploying expensive systems whose scalability and extensibility is limited, while ensuring some resiliency in the face of inevitable changes. This proceedings version of the paper is an extended abstract.

Interaction of COP1 and UVR8 regulates UV-B-induced photomorphogenesis and stress acclimation in Arabidopsis

PubMed Central

Favory, Jean-Jacques; Stec, Agnieszka; Gruber, Henriette; Rizzini, Luca; Oravecz, Attila; Funk, Markus; Albert, Andreas; Cloix, Catherine; Jenkins, Gareth I; Oakeley, Edward J; Seidlitz, Harald K; Nagy, Ferenc; Ulm, Roman

2009-01-01

The ultraviolet-B (UV-B) portion of the solar radiation functions as an environmental signal for which plants have evolved specific and sensitive UV-B perception systems. The UV-B-specific UV RESPONSE LOCUS 8 (UVR8) and the multifunctional E3 ubiquitin ligase CONSTITUTIVELY PHOTOMORPHOGENIC 1 (COP1) are key regulators of the UV-B response. We show here that uvr8-null mutants are deficient in UV-B-induced photomorphogenesis and hypersensitive to UV-B stress, whereas overexpression of UVR8 results in enhanced UV-B photomorphogenesis, acclimation and tolerance to UV-B stress. By using sun simulators, we provide evidence at the physiological level that UV-B acclimation mediated by the UV-B-specific photoregulatory pathway is indeed required for survival in sunlight. At the molecular level, we demonstrate that the wild type but not the mutant UVR8 and COP1 proteins directly interact in a UV-B-dependent, rapid manner in planta. These data collectively suggest that UV-B-specific interaction of COP1 and UVR8 in the nucleus is a very early step in signalling and responsible for the plant's coordinated response to UV-B ensuring UV-B acclimation and protection in the natural environment. PMID:19165148

A minimally invasive optical trapping system to understand cellular interactions at onset of an immune response

PubMed Central

Glass, David G.; McAlinden, Niall; Millington, Owain R.

2017-01-01

T-cells and antigen presenting cells are an essential part of the adaptive immune response system and how they interact is crucial in how the body effectively fights infection or responds to vaccines. Much of the experimental work studying interaction forces between cells has looked at the average properties of bulk samples of cells or applied microscopy to image the dynamic contact between these cells. In this paper we present a novel optical trapping technique for interrogating the force of this interaction and measuring relative interaction forces at the single-cell level. A triple-spot optical trap is used to directly manipulate the cells of interest without introducing foreign bodies such as beads to the system. The optical trap is used to directly control the initiation of cell-cell contact and, subsequently to terminate the interaction at a defined time point. The laser beam power required to separate immune cell pairs is determined and correlates with the force applied by the optical trap. As proof of concept, the antigen-specific increase in interaction force between a dendritic cell and a specific T-cell is demonstrated. Furthermore, it is demonstrated that this interaction force is completely abrogated when T-cell signalling is blocked. As a result the potential of using optical trapping to interrogate cellular interactions at the single cell level without the need to introduce foreign bodies such as beads is clearly demonstrated. PMID:29220398

Spacelab data analysis and interactive control study

NASA Technical Reports Server (NTRS)

Tarbell, T. D.; Drake, J. F.

1980-01-01

The study consisted of two main tasks, a series of interviews of Spacelab users and a survey of data processing and display equipment. Findings from the user interviews on questions of interactive control, downlink data formats, and Spacelab computer software development are presented. Equipment for quick look processing and display of scientific data in the Spacelab Payload Operations Control Center (POCC) was surveyed. Results of this survey effort are discussed in detail, along with recommendations for NASA development of several specific display systems which meet common requirements of many Spacelab experiments.

Dmc1 Functions in a Saccharomyces Cerevisiae Meiotic Pathway That Is Largely Independent of the Rad51 Pathway

PubMed Central

Dresser, M. E.; Ewing, D. J.; Conrad, M. N.; Dominguez, A. M.; Barstead, R.; Jiang, H.; Kodadek, T.

1997-01-01

Meiotic recombination in the yeast Saccharomyces cerevisiae requires two similar recA-like proteins, Dmc1p and Rad51p. A screen for dominant meiotic mutants provided DMC1-G126D, a dominant allele mutated in the conserved ATP-binding site (specifically, the A-loop motif) that confers a null phenotype. A recessive null allele, dmc1-K69E, was isolated as an intragenic suppressor of DMC1-G126D. Dmc1-K69Ep, unlike Dmc1p, does not interact homotypically in a two-hybrid assay, although it does interact with other fusion proteins identified by two-hybrid screen with Dmc1p. Dmc1p, unlike Rad51p, does not interact in the two-hybrid assay with Rad52p or Rad54p. However, Dmc1p does interact with Tid1p, a Rad54p homologue, with Tid4p, a Rad16p homologue, and with other fusion proteins that do not interact with Rad51p, suggesting that Dmc1p and Rad51p function in separate, though possibly overlapping, recombinational repair complexes. Epistasis analysis suggests that DMC1 and RAD51 function in separate pathways responsible for meiotic recombination. Taken together, our results are consistent with a requirement for DMC1 for meiosis-specific entry of DNA double-strand break ends into chromatin. Interestingly, the pattern on CHEF gels of chromosome fragments that result from meiotic DNA double-strand break formation is different in DMC1 mutant strains from that seen in rad50S strains. PMID:9335591

Context-specific requirements of functional domains of the Spectraplakin Short stop in vivo.

PubMed

Bottenberg, Wolfgang; Sanchez-Soriano, Natalia; Alves-Silva, Juliana; Hahn, Ines; Mende, Michael; Prokop, Andreas

2009-07-01

Spectraplakins are large multifunctional cytoskeletal interacting molecules implicated in various processes, including gastrulation, wound healing, skin blistering and neuronal degeneration. It has been speculated that the various functional domains and regions found in Spectraplakins are used in context-specific manners, a model which would provide a crucial explanation for the multifunctional nature of Spectraplakins. Here we tested this possibility by studying domain requirements of the Drosophila Spectraplakin Short stop (Shot) in three different cellular contexts in vivo: (1) neuronal growth, which requires dynamic actin-microtubule interaction; (2) formation and maintenance of tendon cells, which depends on highly stabilised arrays of actin filaments and microtubules, and (3) compartmentalisation in neurons, which is likely to involve cortical F-actin networks. Using these cellular contexts for rescue experiments with Shot deletion constructs in shot mutant background, a number of differential domain requirements were uncovered. First, binding of Shot to F-actin through the first Calponin domain is essential in neuronal contexts but dispensable in tendon cells. This finding is supported by our analyses of shot(kakP2) mutant embryos, which produce only endogenous isoforms lacking the first Calponin domain. Thus, our data demonstrate a functional relevance for these isoforms in vivo. Second, we provide the first functional role for the Plakin domain of Shot, which has a strong requirement for compartmentalisation in neurons and axonal growth, demonstrating that Plakin domains of long Spectraplakin isoforms are of functional relevance. Like the Calponin domain, also the Plakin domain is dispensable in tendon cells, and the currently assumed role of Shot as a linker of microtubules to the tendon cell surface may have to be reconsidered. Third, we demonstrate a function of Shot as an actin-microtubule linker in dendritic growth, thus shedding new light into principal growth mechanisms of this neurite type. Taken together, our data clearly support the view that Spectraplakins function in tissue-specific modes in vivo, and even domains believed to be crucial for Spectraplakin function can be dispensable in specific contexts.

H3K9me2/3 Binding of the MBT Domain Protein LIN-61 Is Essential for Caenorhabditis elegans Vulva Development

PubMed Central

Koester-Eiserfunke, Nora; Fischle, Wolfgang

2011-01-01

MBT domain proteins are involved in developmental processes and tumorigenesis. In vitro binding and mutagenesis studies have shown that individual MBT domains within clustered MBT repeat regions bind mono- and dimethylated histone lysine residues with little to no sequence specificity but discriminate against the tri- and unmethylated states. However, the exact function of promiscuous histone methyl-lysine binding in the biology of MBT domain proteins has not been elucidated. Here, we show that the Caenorhabditis elegans four MBT domain protein LIN-61, in contrast to other MBT repeat factors, specifically interacts with histone H3 when methylated on lysine 9, displaying a strong preference for di- and trimethylated states (H3K9me2/3). Although the fourth MBT repeat is implicated in this interaction, H3K9me2/3 binding minimally requires MBT repeats two to four. Further, mutagenesis of residues conserved with other methyl-lysine binding MBT regions in the fourth MBT repeat does not abolish interaction, implicating a distinct binding mode. In vivo, H3K9me2/3 interaction of LIN-61 is required for C. elegans vulva development within the synMuvB pathway. Mutant LIN-61 proteins deficient in H3K9me2/3 binding fail to rescue lin-61 synMuvB function. Also, previously identified point mutant synMuvB alleles are deficient in H3K9me2/3 interaction although these target residues that are outside of the fourth MBT repeat. Interestingly, lin-61 genetically interacts with two other synMuvB genes, hpl-2, an HP1 homologous H3K9me2/3 binding factor, and met-2, a SETDB1 homologous H3K9 methyl transferase (H3K9MT), in determining C. elegans vulva development and fertility. Besides identifying the first sequence specific and di-/trimethylation binding MBT domain protein, our studies imply complex multi-domain regulation of ligand interaction of MBT domains. Our results also introduce a mechanistic link between LIN-61 function and biology, and they establish interplay of the H3K9me2/3 binding proteins, LIN-61 and HPL-2, as well as the H3K9MT MET-2 in distinct developmental pathways. PMID:21437264

Academic-industry partnerships in addressing nutrition--[infection-immunity-inflammation] interactions.

PubMed

Uauy, Ricardo

2007-10-01

The interaction between nutrition and infection is a key determinant of human health. Traditionally the interaction has centered on the role of nutrients in defining host defenses and the impact of infection in defining nutritional needs and status. Over the past decades the interaction has expanded its scope to encompass the role of specific nutrients in defining acquired immune function, in the modulation of inflammatory processes and on the virulence of the infectious agent itself. More recently the role of micronutrients and fatty acids on the response of cells and tissues to hypoxic and toxic damage has been recognized suggesting a fourth dimension to the interaction. The list of nutrients affecting infection, immunity, inflammation and cell injury has expanded from traditional protein-energy supply to several vitamins, multiple minerals and more recently specific lipid components of the diet. The promise of nutrition in the defense against infection, inflammation and tissue injury has spawned a thriving pharma-nutritional supplement industry and the development of novel foods that require appropriate evaluation of efficacy, safety and effectiveness relative to costs. Academics need to aware of the ethics and the pitfalls in the interaction with industry; conversely industry has to define its role in the process of bringing new knowledge to useful products. The process needs to be interactive, transparent and clearly place public interest above all other considerations.

Modeling of Tool-Tissue Interactions for Computer-Based Surgical Simulation: A Literature Review

PubMed Central

Misra, Sarthak; Ramesh, K. T.; Okamura, Allison M.

2009-01-01

Surgical simulators present a safe and potentially effective method for surgical training, and can also be used in robot-assisted surgery for pre- and intra-operative planning. Accurate modeling of the interaction between surgical instruments and organs has been recognized as a key requirement in the development of high-fidelity surgical simulators. Researchers have attempted to model tool-tissue interactions in a wide variety of ways, which can be broadly classified as (1) linear elasticity-based, (2) nonlinear (hyperelastic) elasticity-based finite element (FE) methods, and (3) other techniques that not based on FE methods or continuum mechanics. Realistic modeling of organ deformation requires populating the model with real tissue data (which are difficult to acquire in vivo) and simulating organ response in real time (which is computationally expensive). Further, it is challenging to account for connective tissue supporting the organ, friction, and topological changes resulting from tool-tissue interactions during invasive surgical procedures. Overcoming such obstacles will not only help us to model tool-tissue interactions in real time, but also enable realistic force feedback to the user during surgical simulation. This review paper classifies the existing research on tool-tissue interactions for surgical simulators specifically based on the modeling techniques employed and the kind of surgical operation being simulated, in order to inform and motivate future research on improved tool-tissue interaction models. PMID:20119508

Study of multi-megawatt technology needs for photovoltaic space power systems, volume 2

NASA Technical Reports Server (NTRS)

Peterson, D. M.; Pleasant, R. L.

1981-01-01

Possible missions requiring multimegawatt photovoltaic space power systems in the 1990's time frame and power system technology needs associated with these missions are examined. Four specific task areas were considered: (1) missions requiring power in the 1-10 megawatt average power region; (2) alternative power systems and component technologies; (3) technology goals and sensitivity trades and analyses; and (4) technology recommendations. Specific concepts for photovoltaic power approaches considered were: planar arrays, concentrating arrays, hybrid systems using Rankine engines, thermophotovoltaic approaches; all with various photovoltaic cell component technologies. Various AC/DC power management approaches, and battery, fuel cell, and flywheel energy storage concepts are evaluated. Interactions with the electrical ion engine injection and stationkeeping system are also considered.

Isolation and characterization of a J domain protein that interacts with ARC1 from ornamental kale (Brassica oleracea var. acephala).

PubMed

Lan, Xingguo; Yang, Jia; Cao, Mingming; Wang, Yanhong; Kawabata, Saneyuki; Li, Yuhua

2015-05-01

A novel J domain protein, JDP1, was isolated from ornamental kale. The C-terminus of JDP1 specifically interacted with ARC1, which has a conserved role in self-incompatibility signaling. Armadillo (ARM)-repeat containing 1 (ARC1) plays a conserved role in self-incompatibility signaling across the Brassicaceae and functions downstream of the S-locus receptor kinase. Here, we identified a J domain protein 1 (JDP1) that interacts with ARC1 using a yeast two-hybrid screen against a stigma cDNA library from ornamental kale (Brassica oleracea var. acephala). JDP1, a 38.4-kDa protein with 344 amino acids, is a member of the Hsp40 family. Fragment JDP1(57-344), originally isolated from a yeast two-hybrid cDNA library, interacted specifically with ARC1 in yeast two-hybrid assays. The N-terminus of JDP1 (JDP1(1-68)) contains a J domain, and the C-terminus of JDP1 (JDP1(69-344)) contains an X domain of unknown function. However, JDP1(69-344) was required and sufficient for interaction with ARC1 in yeast two-hybrid assays and in vitro binding assays. Moreover, JDP1(69-344) regulated the trafficking of ARC1 from the cytoplasm to the plasma membrane by interacting with ARC1 in Arabidopsis mesophyll protoplasts. Finally, Tyr(8) in the JDP1 N-terminal region was identified to be the specific site for regulating the interaction between JDP1 and BoARC1 in yeast two-hybrid assays. Possible roles of JDP1 as an interactor with ARC1 in Brassica are discussed.

V and V of Lexical, Syntactic and Semantic Properties for Interactive Systems Through Model Checking of Formal Description of Dialog

NASA Technical Reports Server (NTRS)

Brat, Guillaume P.; Martinie, Celia; Palanque, Philippe

2013-01-01

During early phases of the development of an interactive system, future system properties are identified (through interaction with end users in the brainstorming and prototyping phase of the application, or by other stakehold-ers) imposing requirements on the final system. They can be specific to the application under development or generic to all applications such as usability principles. Instances of specific properties include visibility of the aircraft altitude, speed… in the cockpit and the continuous possibility of disengaging the autopilot in whatever state the aircraft is. Instances of generic properties include availability of undo (for undoable functions) and availability of a progression bar for functions lasting more than four seconds. While behavioral models of interactive systems using formal description techniques provide complete and unambiguous descriptions of states and state changes, it does not provide explicit representation of the absence or presence of properties. Assessing that the system that has been built is the right system remains a challenge usually met through extensive use and acceptance tests. By the explicit representation of properties and the availability of tools to support checking these properties, it becomes possible to provide developers with means for systematic exploration of the behavioral models and assessment of the presence or absence of these properties. This paper proposes the synergistic use two tools for checking both generic and specific properties of interactive applications: Petshop and Java PathFinder. Petshop is dedicated to the description of interactive system behavior. Java PathFinder is dedicated to the runtime verification of Java applications and as an extension dedicated to User Interfaces. This approach is exemplified on a safety critical application in the area of interactive cockpits for large civil aircrafts.

Repression by PRDM13 is critical for generating precision in neuronal identity

PubMed Central

Kollipara, Rahul K; Ma, Zhenzhong; Borromeo, Mark D; Chang, Joshua C

2017-01-01

The mechanisms that activate some genes while silencing others are critical to ensure precision in lineage specification as multipotent progenitors become restricted in cell fate. During neurodevelopment, these mechanisms are required to generate the diversity of neuronal subtypes found in the nervous system. Here we report interactions between basic helix-loop-helix (bHLH) transcriptional activators and the transcriptional repressor PRDM13 that are critical for specifying dorsal spinal cord neurons. PRDM13 inhibits gene expression programs for excitatory neuronal lineages in the dorsal neural tube. Strikingly, PRDM13 also ensures a battery of ventral neural tube specification genes such as Olig1, Olig2 and Prdm12 are excluded dorsally. PRDM13 does this via recruitment to chromatin by multiple neural bHLH factors to restrict gene expression in specific neuronal lineages. Together these findings highlight the function of PRDM13 in repressing the activity of bHLH transcriptional activators that together are required to achieve precise neuronal specification during mouse development. PMID:28850031

Implementation of meiosis prophase I programme requires a conserved retinoid-independent stabilizer of meiotic transcripts

PubMed Central

Abby, Emilie; Tourpin, Sophie; Ribeiro, Jonathan; Daniel, Katrin; Messiaen, Sébastien; Moison, Delphine; Guerquin, Justine; Gaillard, Jean-Charles; Armengaud, Jean; Langa, Francina; Toth, Attila; Martini, Emmanuelle; Livera, Gabriel

2016-01-01

Sexual reproduction is crucially dependent on meiosis, a conserved, specialized cell division programme that is essential for the production of haploid gametes. Here we demonstrate that fertility and the implementation of the meiotic programme require a previously uncharacterized meiosis-specific protein, MEIOC. Meioc invalidation in mice induces early and pleiotropic meiotic defects in males and females. MEIOC prevents meiotic transcript degradation and interacts with an RNA helicase that binds numerous meiotic mRNAs. Our results indicate that proper engagement into meiosis necessitates the specific stabilization of meiotic transcripts, a previously little-appreciated feature in mammals. Remarkably, the upregulation of MEIOC at the onset of meiosis does not require retinoic acid and STRA8 signalling. Thus, we propose that the complete induction of the meiotic programme requires both retinoic acid-dependent and -independent mechanisms. The latter process involving post-transcriptional regulation likely represents an ancestral mechanism, given that MEIOC homologues are conserved throughout multicellular animals. PMID:26742488

Discovery of Cellular Proteins Required for the Early Steps of HCV Infection Using Integrative Genomics

PubMed Central

Yang, Jae-Seong; Kwon, Oh Sung; Kim, Sanguk; Jang, Sung Key

2013-01-01

Successful viral infection requires intimate communication between virus and host cell, a process that absolutely requires various host proteins. However, current efforts to discover novel host proteins as therapeutic targets for viral infection are difficult. Here, we developed an integrative-genomics approach to predict human genes involved in the early steps of hepatitis C virus (HCV) infection. By integrating HCV and human protein associations, co-expression data, and tight junction-tetraspanin web specific networks, we identified host proteins required for the early steps in HCV infection. Moreover, we validated the roles of newly identified proteins in HCV infection by knocking down their expression using small interfering RNAs. Specifically, a novel host factor CD63 was shown to directly interact with HCV E2 protein. We further demonstrated that an antibody against CD63 blocked HCV infection, indicating that CD63 may serve as a new therapeutic target for HCV-related diseases. The candidate gene list provides a source for identification of new therapeutic targets. PMID:23593195

A haptic interface for virtual simulation of endoscopic surgery.

PubMed

Rosenberg, L B; Stredney, D

1996-01-01

Virtual reality can be described as a convincingly realistic and naturally interactive simulation in which the user is given a first person illusion of being immersed within a computer generated environment While virtual reality systems offer great potential to reduce the cost and increase the quality of medical training, many technical challenges must be overcome before such simulation platforms offer effective alternatives to more traditional training means. A primary challenge in developing effective virtual reality systems is designing the human interface hardware which allows rich sensory information to be presented to users in natural ways. When simulating a given manual procedure, task specific human interface requirements dictate task specific human interface hardware. The following paper explores the design of human interface hardware that satisfies the task specific requirements of virtual reality simulation of Endoscopic surgical procedures. Design parameters were derived through direct cadaver studies and interviews with surgeons. Final hardware design is presented.

Semantic technologies in a decision support system

NASA Astrophysics Data System (ADS)

Wasielewska, K.; Ganzha, M.; Paprzycki, M.; Bǎdicǎ, C.; Ivanovic, M.; Lirkov, I.

2015-10-01

The aim of our work is to design a decision support system based on ontological representation of domain(s) and semantic technologies. Specifically, we consider the case when Grid / Cloud user describes his/her requirements regarding a "resource" as a class expression from an ontology, while the instances of (the same) ontology represent available resources. The goal is to help the user to find the best option with respect to his/her requirements, while remembering that user's knowledge may be "limited." In this context, we discuss multiple approaches based on semantic data processing, which involve different "forms" of user interaction with the system. Specifically, we consider: (a) ontological matchmaking based on SPARQL queries and class expression, (b) graph-based semantic closeness of instances representing user requirements (constructed from the class expression) and available resources, and (c) multicriterial analysis based on the AHP method, which utilizes expert domain knowledge (also ontologically represented).

Supporting secure programming in web applications through interactive static analysis.

PubMed

Zhu, Jun; Xie, Jing; Lipford, Heather Richter; Chu, Bill

2014-07-01

Many security incidents are caused by software developers' failure to adhere to secure programming practices. Static analysis tools have been used to detect software vulnerabilities. However, their wide usage by developers is limited by the special training required to write rules customized to application-specific logic. Our approach is interactive static analysis, to integrate static analysis into Integrated Development Environment (IDE) and provide in-situ secure programming support to help developers prevent vulnerabilities during code construction. No additional training is required nor are there any assumptions on ways programs are built. Our work is motivated in part by the observation that many vulnerabilities are introduced due to failure to practice secure programming by knowledgeable developers. We implemented a prototype interactive static analysis tool as a plug-in for Java in Eclipse. Our technical evaluation of our prototype detected multiple zero-day vulnerabilities in a large open source project. Our evaluations also suggest that false positives may be limited to a very small class of use cases.

Supporting secure programming in web applications through interactive static analysis

PubMed Central

Zhu, Jun; Xie, Jing; Lipford, Heather Richter; Chu, Bill

2013-01-01

Many security incidents are caused by software developers’ failure to adhere to secure programming practices. Static analysis tools have been used to detect software vulnerabilities. However, their wide usage by developers is limited by the special training required to write rules customized to application-specific logic. Our approach is interactive static analysis, to integrate static analysis into Integrated Development Environment (IDE) and provide in-situ secure programming support to help developers prevent vulnerabilities during code construction. No additional training is required nor are there any assumptions on ways programs are built. Our work is motivated in part by the observation that many vulnerabilities are introduced due to failure to practice secure programming by knowledgeable developers. We implemented a prototype interactive static analysis tool as a plug-in for Java in Eclipse. Our technical evaluation of our prototype detected multiple zero-day vulnerabilities in a large open source project. Our evaluations also suggest that false positives may be limited to a very small class of use cases. PMID:25685513

Issues in conducting epidemiologic research among elders: lessons from the MOBILIZE Boston Study.

PubMed

Samelson, Elizabeth J; Kelsey, Jennifer L; Kiel, Douglas P; Roman, Anthony M; Cupples, L Adrienne; Freeman, Marcie B; Jones, Richard N; Hannan, Marian T; Leveille, Suzanne G; Gagnon, Margaret M; Lipsitz, Lewis A

2008-12-15

Conducting research in elderly populations is important, but challenging. In this paper, the authors describe specific challenges that have arisen and solutions that have been used in carrying out The MOBILIZE Boston Study, a community-based, prospective cohort study in Massachusetts focusing on falls among 765 participants aged 70 years or older enrolled during 2005-2007. To recruit older individuals, face-to-face interactions are more effective than less personal approaches. Use of a board of community leaders facilitated community acceptance of the research. Establishing eligibility for potential participants required several interactions, so resources must be anticipated in advance. Assuring a safe and warm environment for elderly participants and offering a positive experience are a vital priority. Adequate funding, planning, and monitoring are required to provide transportation and a fully accessible environment in which to conduct study procedures as well as to select personnel highly skilled in interacting with elders. It is hoped that this paper will encourage and inform future epidemiologic research in this important segment of the population.

Joining the dots - protein-RNA interactions mediating local mRNA translation in neurons.

PubMed

Gallagher, Christopher; Ramos, Andres

2018-06-01

Establishing and maintaining the complex network of connections required for neuronal communication requires the transport and in situ translation of large groups of mRNAs to create local proteomes. In this Review, we discuss the regulation of local mRNA translation in neurons and the RNA-binding proteins that recognise RNA zipcode elements and connect the mRNAs to the cellular transport networks, as well as regulate their translation control. However, mRNA recognition by the regulatory proteins is mediated by the combinatorial action of multiple RNA-binding domains. This increases the specificity and affinity of the interaction, while allowing the protein to recognise a diverse set of targets and mediate a range of mechanisms for translational regulation. The structural and molecular understanding of the interactions can be used together with novel microscopy and transcriptome-wide data to build a mechanistic framework for the regulation of local mRNA translation. © 2018 Federation of European Biochemical Societies.

Combining ANOVA-PCA with POCHEMON to analyse micro-organism development in a polymicrobial environment.

PubMed

Geurts, Brigitte P; Neerincx, Anne H; Bertrand, Samuel; Leemans, Manja A A P; Postma, Geert J; Wolfender, Jean-Luc; Cristescu, Simona M; Buydens, Lutgarde M C; Jansen, Jeroen J

2017-04-22

Revealing the biochemistry associated to micro-organismal interspecies interactions is highly relevant for many purposes. Each pathogen has a characteristic metabolic fingerprint that allows identification based on their unique multivariate biochemistry. When pathogen species come into mutual contact, their co-culture will display a chemistry that may be attributed both to mixing of the characteristic chemistries of the mono-cultures and to competition between the pathogens. Therefore, investigating pathogen development in a polymicrobial environment requires dedicated chemometric methods to untangle and focus upon these sources of variation. The multivariate data analysis method Projected Orthogonalised Chemical Encounter Monitoring (POCHEMON) is dedicated to highlight metabolites characteristic for the interaction of two micro-organisms in co-culture. However, this approach is currently limited to a single time-point, while development of polymicrobial interactions may be highly dynamic. A well-known multivariate implementation of Analysis of Variance (ANOVA) uses Principal Component Analysis (ANOVA-PCA). This allows the overall dynamics to be separated from the pathogen-specific chemistry to analyse the contributions of both aspects separately. For this reason, we propose to integrate ANOVA-PCA with the POCHEMON approach to disentangle the pathogen dynamics and the specific biochemistry in interspecies interactions. Two complementary case studies show great potential for both liquid and gas chromatography - mass spectrometry to reveal novel information on chemistry specific to interspecies interaction during pathogen development. Copyright © 2017 The Author(s). Published by Elsevier B.V. All rights reserved.

Peptide ligands specific to the oxidized form of escherichia coli thioredoxin.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Scholle, M. D.; Banach, B. S.; Hamdan, S. M.

Thioredoxin (Trx) is a highly conserved redox protein involved in several essential cellular processes. In this study, our goal was to isolate peptide ligands to Escherichia coli Trx that mimic protein-protein interactions, specifically the T7 polymerase-Trx interaction. To do this, we subjected Trx to affinity selection against a panel of linear and cysteine-constrained peptides using M13 phage display. A novel cyclized conserved peptide sequence, with a motif of C(D/N/S/T/G)D(S/T)-hydrophobic-C-X-hydrophobic-P, was isolated to Trx. These peptides bound specifically to the E. coli Trx when compared to the human and spirulina homologs. An alanine substitution of the active site cysteines (CGPC) resultedmore » in a significant loss of peptide binding affinity to the Cys-32 mutant. The peptides were also characterized in the context of Trx's role as a processivity factor of the T7 DNA polymerase (gp5). As the interaction between gp5 and Trx normally takes place under reducing conditions, which might interfere with the conformation of the disulfide-bridged peptides, we made use of a 22 residue deletion mutant of gp5 in the thioredoxin binding domain (gp5{Delta}22) that bypassed the requirements of reducing conditions to interact with Trx. A competition study revealed that the peptide selectively inhibits the interaction of gp5{Delta}22 with Trx, under oxidizing conditions, with an IC50 of {approx} 10 {micro}M.« less

An Amoebal Grazer of Cyanobacteria Requires Cobalamin Produced by Heterotrophic Bacteria.

PubMed

Ma, Amy T; Beld, Joris; Brahamsha, Bianca

2017-05-15

Amoebae are unicellular eukaryotes that consume microbial prey through phagocytosis, playing a role in shaping microbial food webs. Many amoebal species can be cultivated axenically in rich media or monoxenically with a single bacterial prey species. Here, we characterize heterolobosean amoeba LPG3, a recent natural isolate, which is unable to grow on unicellular cyanobacteria, its primary food source, in the absence of a heterotrophic bacterium, a Pseudomonas species coisolate. To investigate the molecular basis of this requirement for heterotrophic bacteria, we performed a screen using the defined nonredundant transposon library of Vibrio cholerae , which implicated genes in corrinoid uptake and biosynthesis. Furthermore, cobalamin synthase deletion mutations in V. cholerae and the Pseudomonas species coisolate do not support the growth of amoeba LPG3 on cyanobacteria. While cyanobacteria are robust producers of a corrinoid variant called pseudocobalamin, this variant does not support the growth of amoeba LPG3. Instead, we show that it requires cobalamin that is produced by the Pseudomonas species coisolate. The diversity of eukaryotes utilizing corrinoids is poorly understood, and this amoebal corrinoid auxotroph serves as a model for examining predator-prey interactions and micronutrient transfer in bacterivores underpinning microbial food webs. IMPORTANCE Cyanobacteria are important primary producers in aquatic environments, where they are grazed upon by a variety of phagotrophic protists and, hence, have an impact on nutrient flux at the base of microbial food webs. Here, we characterize amoebal isolate LPG3, which consumes cyanobacteria as its primary food source but also requires heterotrophic bacteria as a source of corrinoid vitamins. Amoeba LPG3 specifically requires the corrinoid variant produced by heterotrophic bacteria and cannot grow on cyanobacteria alone, as they produce a different corrinoid variant. This same corrinoid specificity is also exhibited by other eukaryotes, including humans and algae. This amoebal model system allows us to dissect predator-prey interactions to uncover factors that may shape microbial food webs while also providing insight into corrinoid specificity in eukaryotes. Copyright © 2017 American Society for Microbiology.

An Amoebal Grazer of Cyanobacteria Requires Cobalamin Produced by Heterotrophic Bacteria

PubMed Central

Beld, Joris; Brahamsha, Bianca

2017-01-01

ABSTRACT Amoebae are unicellular eukaryotes that consume microbial prey through phagocytosis, playing a role in shaping microbial food webs. Many amoebal species can be cultivated axenically in rich media or monoxenically with a single bacterial prey species. Here, we characterize heterolobosean amoeba LPG3, a recent natural isolate, which is unable to grow on unicellular cyanobacteria, its primary food source, in the absence of a heterotrophic bacterium, a Pseudomonas species coisolate. To investigate the molecular basis of this requirement for heterotrophic bacteria, we performed a screen using the defined nonredundant transposon library of Vibrio cholerae, which implicated genes in corrinoid uptake and biosynthesis. Furthermore, cobalamin synthase deletion mutations in V. cholerae and the Pseudomonas species coisolate do not support the growth of amoeba LPG3 on cyanobacteria. While cyanobacteria are robust producers of a corrinoid variant called pseudocobalamin, this variant does not support the growth of amoeba LPG3. Instead, we show that it requires cobalamin that is produced by the Pseudomonas species coisolate. The diversity of eukaryotes utilizing corrinoids is poorly understood, and this amoebal corrinoid auxotroph serves as a model for examining predator-prey interactions and micronutrient transfer in bacterivores underpinning microbial food webs. IMPORTANCE Cyanobacteria are important primary producers in aquatic environments, where they are grazed upon by a variety of phagotrophic protists and, hence, have an impact on nutrient flux at the base of microbial food webs. Here, we characterize amoebal isolate LPG3, which consumes cyanobacteria as its primary food source but also requires heterotrophic bacteria as a source of corrinoid vitamins. Amoeba LPG3 specifically requires the corrinoid variant produced by heterotrophic bacteria and cannot grow on cyanobacteria alone, as they produce a different corrinoid variant. This same corrinoid specificity is also exhibited by other eukaryotes, including humans and algae. This amoebal model system allows us to dissect predator-prey interactions to uncover factors that may shape microbial food webs while also providing insight into corrinoid specificity in eukaryotes. PMID:28283521

Distinct mechanisms of recognizing endosomal sorting complex required for transport III (ESCRT-III) protein IST1 by different microtubule interacting and trafficking (MIT) domains.

PubMed

Guo, Emily Z; Xu, Zhaohui

2015-03-27

The endosomal sorting complex required for transport (ESCRT) machinery is responsible for membrane remodeling in a number of biological processes including multivesicular body biogenesis, cytokinesis, and enveloped virus budding. In mammalian cells, efficient abscission during cytokinesis requires proper function of the ESCRT-III protein IST1, which binds to the microtubule interacting and trafficking (MIT) domains of VPS4, LIP5, and Spartin via its C-terminal MIT-interacting motif (MIM). Here, we studied the molecular interactions between IST1 and the three MIT domain-containing proteins to understand the structural basis that governs pairwise MIT-MIM interaction. Crystal structures of the three molecular complexes revealed that IST1 binds to the MIT domains of VPS4, LIP5, and Spartin using two different mechanisms (MIM1 mode versus MIM3 mode). Structural comparison revealed that structural features in both MIT and MIM contribute to determine the specific binding mechanism. Within the IST1 MIM sequence, two phenylalanine residues were shown to be important in discriminating MIM1 versus MIM3 binding. These observations enabled us to deduce a preliminary binding code, which we applied to provide CHMP2A, a protein that normally only binds the MIT domain in the MIM1 mode, the additional ability to bind the MIT domain of Spartin in the MIM3 mode. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Distinct Mechanisms of Recognizing Endosomal Sorting Complex Required for Transport III (ESCRT-III) Protein IST1 by Different Microtubule Interacting and Trafficking (MIT) Domains

DOE PAGES

Guo, Emily Z.; Xu, Zhaohui

2015-02-05

The endosomal sorting complex required for transport (ESCRT) machinery is responsible for membrane remodeling in a number of biological processes including multivesicular body biogenesis, cytokinesis, and enveloped virus budding. In mammalian cells, efficient abscission during cytokinesis requires proper function of the ESCRT-III protein IST1, which binds to the microtubule interacting and trafficking (MIT) domains of VPS4, LIP5, and Spartin via its C-terminal MIT-interacting motif (MIM). In this paper, we studied the molecular interactions between IST1 and the three MIT domain-containing proteins to understand the structural basis that governs pairwise MIT-MIM interaction. Crystal structures of the three molecular complexes revealed thatmore » IST1 binds to the MIT domains of VPS4, LIP5, and Spartin using two different mechanisms (MIM1 mode versus MIM3 mode). Structural comparison revealed that structural features in both MIT and MIM contribute to determine the specific binding mechanism. Within the IST1 MIM sequence, two phenylalanine residues were shown to be important in discriminating MIM1 versus MIM3 binding. Finally, these observations enabled us to deduce a preliminary binding code, which we applied to provide CHMP2A, a protein that normally only binds the MIT domain in the MIM1 mode, the additional ability to bind the MIT domain of Spartin in the MIM3 mode.« less

Cross- and Co-Packaging of Retroviral RNAs and Their Consequences

PubMed Central

Ali, Lizna M.; Rizvi, Tahir A.; Mustafa, Farah

2016-01-01

Retroviruses belong to the family Retroviridae and are ribonucleoprotein (RNP) particles that contain a dimeric RNA genome. Retroviral particle assembly is a complex process, and how the virus is able to recognize and specifically capture the genomic RNA (gRNA) among millions of other cellular and spliced retroviral RNAs has been the subject of extensive investigation over the last two decades. The specificity towards RNA packaging requires higher order interactions of the retroviral gRNA with the structural Gag proteins. Moreover, several retroviruses have been shown to have the ability to cross-/co-package gRNA from other retroviruses, despite little sequence homology. This review will compare the determinants of gRNA encapsidation among different retroviruses, followed by an examination of our current understanding of the interaction between diverse viral genomes and heterologous proteins, leading to their cross-/co-packaging. Retroviruses are well-known serious animal and human pathogens, and such a cross-/co-packaging phenomenon could result in the generation of novel viral variants with unknown pathogenic potential. At the same time, however, an enhanced understanding of the molecular mechanisms involved in these specific interactions makes retroviruses an attractive target for anti-viral drugs, vaccines, and vectors for human gene therapy. PMID:27727192

Cross- and Co-Packaging of Retroviral RNAs and Their Consequences.

PubMed

Ali, Lizna M; Rizvi, Tahir A; Mustafa, Farah

2016-10-11

Retroviruses belong to the family Retroviridae and are ribonucleoprotein (RNP) particles that contain a dimeric RNA genome. Retroviral particle assembly is a complex process, and how the virus is able to recognize and specifically capture the genomic RNA (gRNA) among millions of other cellular and spliced retroviral RNAs has been the subject of extensive investigation over the last two decades. The specificity towards RNA packaging requires higher order interactions of the retroviral gRNA with the structural Gag proteins. Moreover, several retroviruses have been shown to have the ability to cross-/co-package gRNA from other retroviruses, despite little sequence homology. This review will compare the determinants of gRNA encapsidation among different retroviruses, followed by an examination of our current understanding of the interaction between diverse viral genomes and heterologous proteins, leading to their cross-/co-packaging. Retroviruses are well-known serious animal and human pathogens, and such a cross-/co-packaging phenomenon could result in the generation of novel viral variants with unknown pathogenic potential. At the same time, however, an enhanced understanding of the molecular mechanisms involved in these specific interactions makes retroviruses an attractive target for anti-viral drugs, vaccines, and vectors for human gene therapy.

Superglue from bacteria: unbreakable bridges for protein nanotechnology.

PubMed

Veggiani, Gianluca; Zakeri, Bijan; Howarth, Mark

2014-10-01

Biotechnology is often limited by weak interactions. We suggest that an ideal interaction between proteins would be covalent, specific, require addition of only a peptide tag to the protein of interest, and form under a wide range of conditions. Here we summarize peptide tags that are able to form spontaneous amide bonds, based on harnessing reactions of adhesion proteins from the bacterium Streptococcus pyogenes. These include the irreversible peptide-protein interaction of SpyTag with SpyCatcher, as well as irreversible peptide-peptide interactions via SpyLigase. We describe existing applications, including polymerization to enhance cancer cell capture, assembly of living biomaterial, access to diverse protein shapes, and improved enzyme resilience. We also indicate future opportunities for resisting biological force and extending the scope of protein nanotechnology. Copyright © 2014 Elsevier Ltd. All rights reserved.

ISS Mini AERCam Radio Frequency (RF) Coverage Analysis Using iCAT Development Tool

NASA Technical Reports Server (NTRS)

Bolen, Steve; Vazquez, Luis; Sham, Catherine; Fredrickson, Steven; Fink, Patrick; Cox, Jan; Phan, Chau; Panneton, Robert

2003-01-01

The long-term goals of the National Aeronautics and Space Administration's (NASA's) Human Exploration and Development of Space (HEDS) enterprise may require the development of autonomous free-flier (FF) robotic devices to operate within the vicinity of low-Earth orbiting spacecraft to supplement human extravehicular activities (EVAs) in space. Future missions could require external visual inspection of the spacecraft that would be difficult, or dangerous, for humans to perform. Under some circumstance, it may be necessary to employ an un-tethered communications link between the FF and the users. The interactive coverage analysis tool (ICAT) is a software tool that has been developed to perform critical analysis of the communications link performance for a FF operating in the vicinity of the International Space Station (ISS) external environment. The tool allows users to interactively change multiple parameters of the communications link parameters to efficiently perform systems engineering trades on network performance. These trades can be directly translated into design and requirements specifications. This tool significantly reduces the development time in determining a communications network topology by allowing multiple parameters to be changed, and the results of link coverage to be statistically characterized and plotted interactively.

The Use of Arabidopsis to Study Interactions between Parasitic Angiosperms and Their Plant Hosts

PubMed Central

Goldwasser, Y.; Westwood, J. H.; Yoder, J. I.

2002-01-01

Parasitic plants invade host plants in order to rob them of water, minerals and nutrients. The consequences to the infected hosts can be debilitating and some of the world's most pernicious agricultural weeds are parasitic. Parasitic genera of the Scrophulariaceae and Orobanchaceae directly invade roots of neighboring plants via underground structures called haustoria. The mechanisms by which these parasites identify and associate with host plants present unsurpassed opportunities for studying chemical signaling in plant-plant interactions. Seeds of some parasites require specific host factors for efficient germination, thereby insuring the availability of an appropriate host root prior to germination. A second set of signal molecules is required to induce haustorium development and the beginning of heterotrophy. Later stages in parasitism also require the presence of host factors, although these have not yet been well characterized. Arabidopsis is being used as a model host plant to identify genetic loci associated with stimulating parasite germination, haustorium development, and parasite support. Arabidopsis is also being employed to explore how host plants respond to parasite attack. Current methodologies and recent findings in Arabidopsis – parasitic plant interactions will be discussed. PMID:22303205

PDZ Domain-containing 1 (PDZK1) Protein Regulates Phospholipase C-β3 (PLC-β3)-specific Activation of Somatostatin by Forming a Ternary Complex with PLC-β3 and Somatostatin Receptors*

PubMed Central

Kim, Jung Kuk; Kwon, Ohman; Kim, Jinho; Kim, Eung-Kyun; Park, Hye Kyung; Lee, Ji Eun; Kim, Kyung Lock; Choi, Jung Woong; Lim, Seyoung; Seok, Heon; Lee-Kwon, Whaseon; Choi, Jang Hyun; Kang, Byoung Heon; Kim, Sanguk; Ryu, Sung Ho; Suh, Pann-Ghill

2012-01-01

Phospholipase C-β (PLC-β) is a key molecule in G protein-coupled receptor (GPCR)-mediated signaling. Many studies have shown that the four PLC-β subtypes have different physiological functions despite their similar structures. Because the PLC-β subtypes possess different PDZ-binding motifs, they have the potential to interact with different PDZ proteins. In this study, we identified PDZ domain-containing 1 (PDZK1) as a PDZ protein that specifically interacts with PLC-β3. To elucidate the functional roles of PDZK1, we next screened for potential interacting proteins of PDZK1 and identified the somatostatin receptors (SSTRs) as another protein that interacts with PDZK1. Through these interactions, PDZK1 assembles as a ternary complex with PLC-β3 and SSTRs. Interestingly, the expression of PDZK1 and PLC-β3, but not PLC-β1, markedly potentiated SST-induced PLC activation. However, disruption of the ternary complex inhibited SST-induced PLC activation, which suggests that PDZK1-mediated complex formation is required for the specific activation of PLC-β3 by SST. Consistent with this observation, the knockdown of PDZK1 or PLC-β3, but not that of PLC-β1, significantly inhibited SST-induced intracellular Ca2+ mobilization, which further attenuated subsequent ERK1/2 phosphorylation. Taken together, our results strongly suggest that the formation of a complex between SSTRs, PDZK1, and PLC-β3 is essential for the specific activation of PLC-β3 and the subsequent physiologic responses by SST. PMID:22528496

Hermes (Rbpms) is a Critical Component of RNP Complexes that Sequester Germline RNAs during Oogenesis.

PubMed

Aguero, Tristan; Zhou, Yi; Kloc, Malgorzata; Chang, Patrick; Houliston, Evelyn; King, Mary Lou

2016-03-01

The germ cell lineage in Xenopus is specified by the inheritance of germ plasm that assembles within the mitochondrial cloud or Balbiani body in stage I oocytes. Specific RNAs, such as nanos1 , localize to the germ plasm. nanos1 has the essential germline function of blocking somatic gene expression and thus preventing Primordial Germ Cell (PGC) loss and sterility. Hermes/Rbpms protein and nanos RNA co-localize within germinal granules, diagnostic electron dense particles found within the germ plasm. Previous work indicates that nanos accumulates within the germ plasm through a diffusion/entrapment mechanism. Here we show that Hermes/Rbpms interacts with nanos through sequence specific RNA localization signals found in the nanos -3'UTR. Importantly, Hermes/Rbpms specifically binds nanos , but not Vg1 RNA in the nucleus of stage I oocytes. In vitro binding data show that Hermes/Rbpms requires additional factors that are present in stage I oocytes in order to bind nanos1 . One such factor may be hnRNP I, identified in a yeast-2-hybrid screen as directly interacting with Hermes/Rbpms. We suggest that Hermes/Rbpms functions as part of a RNP complex in the nucleus that facilitates selection of germline RNAs for germ plasm localization. We propose that Hermes/Rbpms is required for nanos RNA to form within the germinal granules and in this way, participates in the germline specific translational repression and sequestration of nanos RNA .

Hermes (Rbpms) is a Critical Component of RNP Complexes that Sequester Germline RNAs during Oogenesis

PubMed Central

Aguero, Tristan; Zhou, Yi; Kloc, Malgorzata; Chang, Patrick; Houliston, Evelyn; King, Mary Lou

2016-01-01

The germ cell lineage in Xenopus is specified by the inheritance of germ plasm that assembles within the mitochondrial cloud or Balbiani body in stage I oocytes. Specific RNAs, such as nanos1, localize to the germ plasm. nanos1 has the essential germline function of blocking somatic gene expression and thus preventing Primordial Germ Cell (PGC) loss and sterility. Hermes/Rbpms protein and nanos RNA co-localize within germinal granules, diagnostic electron dense particles found within the germ plasm. Previous work indicates that nanos accumulates within the germ plasm through a diffusion/entrapment mechanism. Here we show that Hermes/Rbpms interacts with nanos through sequence specific RNA localization signals found in the nanos-3′UTR. Importantly, Hermes/Rbpms specifically binds nanos, but not Vg1 RNA in the nucleus of stage I oocytes. In vitro binding data show that Hermes/Rbpms requires additional factors that are present in stage I oocytes in order to bind nanos1. One such factor may be hnRNP I, identified in a yeast-2-hybrid screen as directly interacting with Hermes/Rbpms. We suggest that Hermes/Rbpms functions as part of a RNP complex in the nucleus that facilitates selection of germline RNAs for germ plasm localization. We propose that Hermes/Rbpms is required for nanos RNA to form within the germinal granules and in this way, participates in the germline specific translational repression and sequestration of nanos RNA. PMID:26998427

Type III restriction-modification enzymes: a historical perspective.

PubMed

Rao, Desirazu N; Dryden, David T F; Bheemanaik, Shivakumara

2014-01-01

Restriction endonucleases interact with DNA at specific sites leading to cleavage of DNA. Bacterial DNA is protected from restriction endonuclease cleavage by modifying the DNA using a DNA methyltransferase. Based on their molecular structure, sequence recognition, cleavage position and cofactor requirements, restriction-modification (R-M) systems are classified into four groups. Type III R-M enzymes need to interact with two separate unmethylated DNA sequences in inversely repeated head-to-head orientations for efficient cleavage to occur at a defined location (25-27 bp downstream of one of the recognition sites). Like the Type I R-M enzymes, Type III R-M enzymes possess a sequence-specific ATPase activity for DNA cleavage. ATP hydrolysis is required for the long-distance communication between the sites before cleavage. Different models, based on 1D diffusion and/or 3D-DNA looping, exist to explain how the long-distance interaction between the two recognition sites takes place. Type III R-M systems are found in most sequenced bacteria. Genome sequencing of many pathogenic bacteria also shows the presence of a number of phase-variable Type III R-M systems, which play a role in virulence. A growing number of these enzymes are being subjected to biochemical and genetic studies, which, when combined with ongoing structural analyses, promise to provide details for mechanisms of DNA recognition and catalysis.

Functional cortical network in alpha band correlates with social bargaining.

PubMed

Billeke, Pablo; Zamorano, Francisco; Chavez, Mario; Cosmelli, Diego; Aboitiz, Francisco

2014-01-01

Solving demanding tasks requires fast and flexible coordination among different brain areas. Everyday examples of this are the social dilemmas in which goals tend to clash, requiring one to weigh alternative courses of action in limited time. In spite of this fact, there are few studies that directly address the dynamics of flexible brain network integration during social interaction. To study the preceding, we carried out EEG recordings while subjects played a repeated version of the Ultimatum Game in both human (social) and computer (non-social) conditions. We found phase synchrony (inter-site-phase-clustering) modulation in alpha band that was specific to the human condition and independent of power modulation. The strength and patterns of the inter-site-phase-clustering of the cortical networks were also modulated, and these modulations were mainly in frontal and parietal regions. Moreover, changes in the individuals' alpha network structure correlated with the risk of the offers made only in social conditions. This correlation was independent of changes in power and inter-site-phase-clustering strength. Our results indicate that, when subjects believe they are participating in a social interaction, a specific modulation of functional cortical networks in alpha band takes place, suggesting that phase synchrony of alpha oscillations could serve as a mechanism by which different brain areas flexibly interact in order to adapt ongoing behavior in socially demanding contexts.

Functional Cortical Network in Alpha Band Correlates with Social Bargaining

PubMed Central

Billeke, Pablo; Zamorano, Francisco; Chavez, Mario; Cosmelli, Diego; Aboitiz, Francisco

2014-01-01

Solving demanding tasks requires fast and flexible coordination among different brain areas. Everyday examples of this are the social dilemmas in which goals tend to clash, requiring one to weigh alternative courses of action in limited time. In spite of this fact, there are few studies that directly address the dynamics of flexible brain network integration during social interaction. To study the preceding, we carried out EEG recordings while subjects played a repeated version of the Ultimatum Game in both human (social) and computer (non-social) conditions. We found phase synchrony (inter-site-phase-clustering) modulation in alpha band that was specific to the human condition and independent of power modulation. The strength and patterns of the inter-site-phase-clustering of the cortical networks were also modulated, and these modulations were mainly in frontal and parietal regions. Moreover, changes in the individuals’ alpha network structure correlated with the risk of the offers made only in social conditions. This correlation was independent of changes in power and inter-site-phase-clustering strength. Our results indicate that, when subjects believe they are participating in a social interaction, a specific modulation of functional cortical networks in alpha band takes place, suggesting that phase synchrony of alpha oscillations could serve as a mechanism by which different brain areas flexibly interact in order to adapt ongoing behavior in socially demanding contexts. PMID:25286240

Abelson Interactor 1 (Abi1) and Its Interaction with Wiskott-Aldrich Syndrome Protein (Wasp) Are Critical for Proper Eye Formation in Xenopus Embryos*

PubMed Central

Singh, Arvinder; Winterbottom, Emily F.; Ji, Yon Ju; Hwang, Yoo-Seok; Daar, Ira O.

2013-01-01

Abl interactor 1 (Abi1) is a scaffold protein that plays a central role in the regulation of actin cytoskeleton dynamics as a constituent of several key protein complexes, and homozygous loss of this protein leads to embryonic lethality in mice. Because this scaffold protein has been shown in cultured cells to be a critical component of pathways controlling cell migration and actin regulation at cell-cell contacts, we were interested to investigate the in vivo role of Abi1 in morphogenesis during the development of Xenopus embryos. Using morpholino-mediated translation inhibition, we demonstrate that knockdown of Abi1 in the whole embryo, or specifically in eye field progenitor cells, leads to disruption of eye morphogenesis. Moreover, signaling through the Src homology 3 domain of Abi1 is critical for proper movement of retinal progenitor cells into the eye field and their appropriate differentiation, and this process is dependent upon an interaction with the nucleation-promoting factor Wasp (Wiskott-Aldrich syndrome protein). Collectively, our data demonstrate that the Abi1 scaffold protein is an essential regulator of cell movement processes required for normal eye development in Xenopus embryos and specifically requires an Src homology 3 domain-dependent interaction with Wasp to regulate this complex morphogenetic process. PMID:23558677

The Impact of a Simulation and Problem-Based Learning Design Project on Student Learning and Teamwork Skills. CSE Technical Report.

ERIC Educational Resources Information Center

Chung, Gregory K. W. K.

This study examined a civil engineering capstone course that embedded a sophisticated simulation-based task within instruction. Students (n=28) were required to conduct a hazardous waste site investigation using simulation software designed specifically for the course (Interactive Site Investigation Software) (ISIS). The software simulated…

The National Shipbuilding Research Program. Proceedings of the REAPS Technical Symposium held June 15-16, 1976 Atlanta, Georgia

DTIC Science & Technology

1976-06-01

the imformation needed to perform the design funct ions, but will also provide the user with the details required to interact with other individuals...several calls of basic drafting (DRA) macros. The deck heights are constant andincluded in the coding. Al - identifies the specific frame to be retrived

What Teachers Should Say and How They Should Say It

ERIC Educational Resources Information Center

Conderman, Greg; Johnston-Rodriguez, Sarah; Hartman, Paula; Kemp, Drew

2010-01-01

Many teachers have discovered that their positions require them to communicate effectively with families, team members, and other colleagues. In fact, teachers today indicate that much of their day is spent navigating adult-to-adult interactions for which they feel ill-prepared. Specifically, teachers may not know what to say or how to react when…

On pseudosupersymmetric oscillators and reality of relativistic energies for vector mesons

NASA Technical Reports Server (NTRS)

Beckers, Jules; Debergh, Nathalie

1995-01-01

Specific oscillators - hereafter called pseudosupersymmetric oscillators - appear as interesting nonrelativistic concepts in connection with the study of relativistic vector mesons interacting with an external constant magnetic field when the real character of the energy eigenvalues is required as expected. A new pseudosupersymmetric quantum mechanics can then be developed and the corresponding pseudosupersymmetries can be pointed out.

Immune Tolerance in Multiple Sclerosis

PubMed Central

Goverman, Joan M.

2011-01-01

Summary Multiple sclerosis is believed to be mediated by T cells specific for myelin antigens that circulate harmlessly in the periphery of healthy individuals until they are erroneously by an environmental stimulus. Upon activation, the T cells enter the central nervous system and orchestrate an immune response against myelin. To understand the initial steps in the pathogenesis of multiple sclerosis, it is important to identify the mechanisms that maintain T-cell tolerance to myelin antigens and to understand how some myelin-specific T cells escape tolerance and what conditions lead to their activation. Central tolerance strongly shapes the peripheral repertoire of myelin-specific T cells, as most myelin-specific T cells are eliminated by clonal deletion in the thymus. Self-reactive T cells that escape central tolerance are generally capable only of low-avidity interactions with antigen-presenting cells. Despite the low avidity of these interactions, peripheral tolerance mechanisms are required to prevent spontaneous autoimmunity. Multiple peripheral tolerance mechanisms for myelin-specific T cells have been indentified, the most important of which appears to be regulatory T cells. While most studies have focused on CD4+ myelin-specific T cells, interesting differences in tolerance mechanisms and the conditions that abrogate these mechanisms have recently been described for CD8+ myelin-specific T cells. PMID:21488900

Abasic pivot substitution harnesses target specificity of RNA interference

PubMed Central

Lee, Hye-Sook; Seok, Heeyoung; Lee, Dong Ha; Ham, Juyoung; Lee, Wooje; Youm, Emilia Moonkyung; Yoo, Jin Seon; Lee, Yong-Seung; Jang, Eun-Sook; Chi, Sung Wook

2015-01-01

Gene silencing via RNA interference inadvertently represses hundreds of off-target transcripts. Because small interfering RNAs (siRNAs) can function as microRNAs, avoiding miRNA-like off-target repression is a major challenge. Functional miRNA–target interactions are known to pre-require transitional nucleation, base pairs from position 2 to the pivot (position 6). Here, by substituting nucleotide in pivot with abasic spacers, which prevent base pairing and alleviate steric hindrance, we eliminate miRNA-like off-target repression while preserving on-target activity at ∼80–100%. Specifically, miR-124 containing dSpacer pivot substitution (6pi) loses seed-mediated transcriptome-wide target interactions, repression activity and biological function, whereas other conventional modifications are ineffective. Application of 6pi allows PCSK9 siRNA to efficiently lower plasma cholesterol concentration in vivo, and abolish potentially deleterious off-target phenotypes. The smallest spacer, C3, also shows the same improvement in target specificity. Abasic pivot substitution serves as a general means to harness the specificity of siRNA experiments and therapeutic applications. PMID:26679372

Elucidating nitric oxide synthase domain interactions by molecular dynamics.

PubMed

Hollingsworth, Scott A; Holden, Jeffrey K; Li, Huiying; Poulos, Thomas L

2016-02-01

Nitric oxide synthase (NOS) is a multidomain enzyme that catalyzes the production of nitric oxide (NO) by oxidizing L-Arg to NO and L-citrulline. NO production requires multiple interdomain electron transfer steps between the flavin mononucleotide (FMN) and heme domain. Specifically, NADPH-derived electrons are transferred to the heme-containing oxygenase domain via the flavin adenine dinucleotide (FAD) and FMN containing reductase domains. While crystal structures are available for both the reductase and oxygenase domains of NOS, to date there is no atomic level structural information on domain interactions required for the final FMN-to-heme electron transfer step. Here, we evaluate a model of this final electron transfer step for the heme-FMN-calmodulin NOS complex based on the recent biophysical studies using a 105-ns molecular dynamics trajectory. The resulting equilibrated complex structure is very stable and provides a detailed prediction of interdomain contacts required for stabilizing the NOS output state. The resulting equilibrated complex model agrees well with previous experimental work and provides a detailed working model of the final NOS electron transfer step required for NO biosynthesis. © 2015 The Protein Society.

Molecular biomimetics: GEPI-based biological routes to technology.

PubMed

Tamerler, Candan; Khatayevich, Dmitriy; Gungormus, Mustafa; Kacar, Turgay; Oren, E Emre; Hnilova, Marketa; Sarikaya, Mehmet

2010-01-01

In nature, the viability of biological systems is sustained via specific interactions among the tens of thousands of proteins, the major building blocks of organisms from the simplest single-celled to the most complex multicellular species. Biomolecule-material interaction is accomplished with molecular specificity and efficiency leading to the formation of controlled structures and functions at all scales of dimensional hierarchy. Through evolution, Mother Nature developed molecular recognition by successive cycles of mutation and selection. Molecular specificity of probe-target interactions, e.g., ligand-receptor, antigen-antibody, is always based on specific peptide molecular recognition. Using biology as a guide, we can now understand, engineer, and control peptide-material interactions and exploit them as a new design tool for novel materials and systems. We adapted the protocols of combinatorially designed peptide libraries, via both cell surface or phage display methods; using these we select short peptides with specificity to a variety of practical materials. These genetically engineered peptides for inorganics (GEPI) are then studied experimentally to establish their binding kinetics and surface stability. The bound peptide structure and conformations are interrogated both experimentally and via modeling, and self-assembly characteristics are tested via atomic force microscopy. We further engineer the peptide binding and assembly characteristics using a computational biomimetics approach where bioinformatics based peptide-sequence similarity analysis is developed to design higher generation function-specific peptides. The molecular biomimetic approach opens up new avenues for the design and utilization of multifunctional molecular systems in a wide-range of applications from tissue engineering, disease diagnostics, and therapeutics to various areas of nanotechnology where integration is required among inorganic, organic and biological materials. Here, we describe lessons from biology with examples of protein-mediated functional biological materials, explain how novel peptides can be designed with specific affinity to inorganic solids using evolutionary engineering approaches, give examples of their potential utilizations in technology and medicine, and, finally, provide a summary of challenges and future prospects. (c) 2010 Wiley Periodicals, Inc.

Plasmacytoid Dendritic Cells Require Direct Infection To Sustain the Pulmonary Influenza A Virus-Specific CD8 T Cell Response

PubMed Central

Hemann, Emily A.; Sjaastad, Louisa E.; Langlois, Ryan A.

2015-01-01

ABSTRACT Following influenza A virus (IAV) infection, development of a robust IAV-specific CD8 T cell response is required for clearance of primary infection and enhances memory protection. Following IAV infection, plasmacytoid dendritic cells (pDC) or CD8α+ DC regulate pulmonary effector CD8 T cell responses within the lung. Without this DC-T cell interaction, insufficient effector CD8 T cells are maintained in the lungs, leading to enhanced morbidity and mortality. Previous studies have demonstrated that pDC are capable of classical presentation or cross-presentation of IAV antigens and could potentially regulate IAV-specific CD8 T cell responses through either mechanism. Our results demonstrate that pDC from the lungs of donor mice infected with an IAV that is not able to replicate in hematopoietic cells (142t-IAV), unlike donor pDC isolated from the lungs of control infected mice, are not able to rescue the host IAV-specific CD8 T cell response from apoptosis. This indicates that pDC must utilize the direct presentation pathway for this rescue. This inability of pDC from 142t-IAV donors to rescue the IAV-specific CD8 T cell response is not due to differences in the overall ability of 142t-IAV to replicate within the lungs or generate defective viral genomes or to differences in levels of costimulatory molecules required for this interaction. We further demonstrate that bypassing the antigen presentation pathway by coating the 142t-IAV pDC with IAV peptide epitopes restores their ability to rescue the IAV-specific CD8 T cell response. IMPORTANCE IAV continues to be a global health burden, infecting 5 to 20% of the global population annually. Continued investigation into the mechanisms that mediate protective immune responses against IAV is important to improving current vaccination and antiviral strategies antagonistic toward IAV. Our findings presented herein demonstrate a key requirement for pDC promotion of effector CD8 T cell survival: that rather than utilizing cross-presentation, pDC must be infected and utilize the endogenous pathway for presentation of antigens to CD8 T cells during in vivo IAV infections. This suggests that targeting presentation via the endogenous pathway in pDC could be important for the development of unique antiviral cellular therapies. PMID:26719269

A RET-ER81-NRG1 Signaling Pathway Drives the Development of Pacinian Corpuscles.

PubMed

Fleming, Michael S; Li, Jian J; Ramos, Daniel; Li, Tong; Talmage, David A; Abe, Shin-Ichi; Arber, Silvia; Luo, Wenqin

2016-10-05

Axon-Schwann cell interactions are crucial for the development, function, and repair of the peripheral nervous system, but mechanisms underlying communication between axons and nonmyelinating Schwann cells are unclear. Here, we show that ER81 is functionally required in a subset of mouse RET + mechanosensory neurons for formation of Pacinian corpuscles, which are composed of a single myelinated axon and multiple layers of nonmyelinating Schwann cells, and Ret is required for the maintenance of Er81 expression. Interestingly, Er81 mutants have normal myelination but exhibit deficient interactions between axons and corpuscle-forming nonmyelinating Schwann cells. Finally, ablating Neuregulin-1 (Nrg1) in mechanosensory neurons results in no Pacinian corpuscles, and an Nrg1 isoform not required for communication with myelinating Schwann cells is specifically decreased in Er81-null somatosensory neurons. Collectively, our results suggest that a RET-ER81-NRG1 signaling pathway promotes axon communication with nonmyelinating Schwann cells, and that neurons use distinct mechanisms to interact with different types of Schwann cells. Communication between neurons and Schwann cells is critical for development, normal function, and regeneration of the peripheral nervous system. Despite many studies about axonal communication with myelinating Schwann cells, mostly via a specific isoform of Neuregulin1, the molecular nature of axonal communication with nonmyelinating Schwann cells is poorly understood. Here, we described a RET-ER81-Neuregulin1 signaling pathway in neurons innervating Pacinian corpuscle somatosensory end organs, which is essential for communication between the innervating axon and the end organ nonmyelinating Schwann cells. We also showed that this signaling pathway uses isoforms of Neuregulin1 that are not involved in myelination, providing evidence that neurons use different isoforms of Neuregulin1 to interact with different types of Schwann cells. Copyright © 2016 the authors 0270-6474/16/3610337-19$15.00/0.

Systematic discovery of Xist RNA binding proteins

PubMed Central

Chu, Ci; Zhang, Qiangfeng Cliff; da Rocha, Simão Teixeira; Flynn, Ryan A.; Bharadwaj, Maheetha; Calabrese, J. Mauro; Magnuson, Terry; Heard, Edith; Chang, Howard Y.

2015-01-01

Summary Noncoding RNAs (ncRNAs) function with associated proteins to effect complex structural and regulatory outcomes. To reveal the composition and dynamics of specific noncoding RNA- protein complexes (RNPs) in vivo, we developed comprehensive identification of RNA-binding proteins by mass spectrometry (ChIRP-MS). ChIRP-MS analysis of four ncRNAs captures key protein interactors, including a U1-specific link to the 3′ RNA processing machinery. Xist, an essential lncRNA for X-chromosome inactivation (XCI), interacts with 81 proteins from chromatin modification, nuclear matrix, and RNA remodeling pathways. The Xist RNA-protein particle assembles in two steps coupled with the transition from pluripotency to differentiation. Specific interactors include HnrnpK that participates in Xist-mediated gene silencing and histone modifications, but not Xist localization and Drosophila Split ends homolog Spen that interacts via the A-repeat domain of Xist and is required for gene silencing. Thus, Xist lncRNA engages with proteins in a modular and developmentally controlled manner to coordinate chromatin spreading and silencing. PMID:25843628

A FYVE zinc finger domain protein specifically links mRNA transport to endosome trafficking.

PubMed

Pohlmann, Thomas; Baumann, Sebastian; Haag, Carl; Albrecht, Mario; Feldbrügge, Michael

2015-05-18

An emerging theme in cellular logistics is the close connection between mRNA and membrane trafficking. A prominent example is the microtubule-dependent transport of mRNAs and associated ribosomes on endosomes. This coordinated process is crucial for correct septin filamentation and efficient growth of polarised cells, such as fungal hyphae. Despite detailed knowledge on the key RNA-binding protein and the molecular motors involved, it is unclear how mRNAs are connected to membranes during transport. Here, we identify a novel factor containing a FYVE zinc finger domain for interaction with endosomal lipids and a new PAM2-like domain required for interaction with the MLLE domain of the key RNA-binding protein. Consistently, loss of this FYVE domain protein leads to specific defects in mRNA, ribosome, and septin transport without affecting general functions of endosomes or their movement. Hence, this is the first endosomal component specific for mRNP trafficking uncovering a new mechanism to couple mRNPs to endosomes.

Who Is Influencing Whom? Latino Parent–Child Request Interactions and Product Purchases in Food Retail Environments

PubMed Central

Castro, Iana A.; Calderon, Joanna; Ayala, Guadalupe X.

2017-01-01

This study examines Latino parent–child interactions about foods and beverages requested in food retail environments in San Diego, CA. It seeks to extend our understanding of parent–child request interactions and purchases by studying how the number of product request interactions and purchases differ based on four factors that have been understudied in previous parent–child interaction research: parent gender, child gender, product healthfulness, and who initiated the request interaction (parent or child). By unobtrusively observing Latino parent–child dyads for the duration of a brief shopping trip, we found that parent and child gender are related to the number of request interactions initiated by parents and children. For gender-specific child-initiated request interactions, sons initiated more request interactions with fathers while daughters initiated more request interactions with mothers. Most request interactions were for products that were categorized as calorie dense, and a higher percentage of these products were purchased as a result of parent-initiated (vs. child-initiated) request interactions. The results provide important considerations for practitioners and researchers working on improving nutrition and reducing obesity. Assumptions about who is influencing whom in food store request interactions are challenged, requiring more research. PMID:29081718

rigor mortis encodes a novel nuclear receptor interacting protein required for ecdysone signaling during Drosophila larval development.

PubMed

Gates, Julie; Lam, Geanette; Ortiz, José A; Losson, Régine; Thummel, Carl S

2004-01-01

Pulses of the steroid hormone ecdysone trigger the major developmental transitions in Drosophila, including molting and puparium formation. The ecdysone signal is transduced by the EcR/USP nuclear receptor heterodimer that binds to specific response elements in the genome and directly regulates target gene transcription. We describe a novel nuclear receptor interacting protein encoded by rigor mortis (rig) that is required for ecdysone responses during larval development. rig mutants display defects in molting, delayed larval development, larval lethality, duplicated mouth parts, and defects in puparium formation--phenotypes that resemble those seen in EcR, usp, E75A and betaFTZ-F1 mutants. Although the expression of these nuclear receptor genes is essentially normal in rig mutant larvae, the ecdysone-triggered switch in E74 isoform expression is defective. rig encodes a protein with multiple WD-40 repeats and an LXXLL motif, sequences that act as specific protein-protein interaction domains. Consistent with the presence of these elements and the lethal phenotypes of rig mutants, Rig protein interacts with several Drosophila nuclear receptors in GST pull-down experiments, including EcR, USP, DHR3, SVP and betaFTZ-F1. The ligand binding domain of betaFTZ-F1 is sufficient for this interaction, which can occur in an AF-2-independent manner. Antibody stains reveal that Rig protein is present in the brain and imaginal discs of second and third instar larvae, where it is restricted to the cytoplasm. In larval salivary gland and midgut cells, however, Rig shuttles between the cytoplasm and nucleus in a spatially and temporally regulated manner, at times that correlate with the major lethal phase of rig mutants and major switches in ecdysone-regulated gene expression. Taken together, these data indicate that rig exerts essential functions during larval development through gene-specific effects on ecdysone-regulated transcription, most likely as a cofactor for one or more nuclear receptors. Furthermore, the dynamic intracellular redistribution of Rig protein suggests that it may act to refine spatial and temporal responses to ecdysone during development.

Cataloging the biomedical world of pain through semi-automated curation of molecular interactions

PubMed Central

Jamieson, Daniel G.; Roberts, Phoebe M.; Robertson, David L.; Sidders, Ben; Nenadic, Goran

2013-01-01

The vast collection of biomedical literature and its continued expansion has presented a number of challenges to researchers who require structured findings to stay abreast of and analyze molecular mechanisms relevant to their domain of interest. By structuring literature content into topic-specific machine-readable databases, the aggregate data from multiple articles can be used to infer trends that can be compared and contrasted with similar findings from topic-independent resources. Our study presents a generalized procedure for semi-automatically creating a custom topic-specific molecular interaction database through the use of text mining to assist manual curation. We apply the procedure to capture molecular events that underlie ‘pain’, a complex phenomenon with a large societal burden and unmet medical need. We describe how existing text mining solutions are used to build a pain-specific corpus, extract molecular events from it, add context to the extracted events and assess their relevance. The pain-specific corpus contains 765 692 documents from Medline and PubMed Central, from which we extracted 356 499 unique normalized molecular events, with 261 438 single protein events and 93 271 molecular interactions supplied by BioContext. Event chains are annotated with negation, speculation, anatomy, Gene Ontology terms, mutations, pain and disease relevance, which collectively provide detailed insight into how that event chain is associated with pain. The extracted relations are visualized in a wiki platform (wiki-pain.org) that enables efficient manual curation and exploration of the molecular mechanisms that underlie pain. Curation of 1500 grouped event chains ranked by pain relevance revealed 613 accurately extracted unique molecular interactions that in the future can be used to study the underlying mechanisms involved in pain. Our approach demonstrates that combining existing text mining tools with domain-specific terms and wiki-based visualization can facilitate rapid curation of molecular interactions to create a custom database. Database URL: ••• PMID:23707966

Cataloging the biomedical world of pain through semi-automated curation of molecular interactions.

PubMed

Jamieson, Daniel G; Roberts, Phoebe M; Robertson, David L; Sidders, Ben; Nenadic, Goran

2013-01-01

The vast collection of biomedical literature and its continued expansion has presented a number of challenges to researchers who require structured findings to stay abreast of and analyze molecular mechanisms relevant to their domain of interest. By structuring literature content into topic-specific machine-readable databases, the aggregate data from multiple articles can be used to infer trends that can be compared and contrasted with similar findings from topic-independent resources. Our study presents a generalized procedure for semi-automatically creating a custom topic-specific molecular interaction database through the use of text mining to assist manual curation. We apply the procedure to capture molecular events that underlie 'pain', a complex phenomenon with a large societal burden and unmet medical need. We describe how existing text mining solutions are used to build a pain-specific corpus, extract molecular events from it, add context to the extracted events and assess their relevance. The pain-specific corpus contains 765 692 documents from Medline and PubMed Central, from which we extracted 356 499 unique normalized molecular events, with 261 438 single protein events and 93 271 molecular interactions supplied by BioContext. Event chains are annotated with negation, speculation, anatomy, Gene Ontology terms, mutations, pain and disease relevance, which collectively provide detailed insight into how that event chain is associated with pain. The extracted relations are visualized in a wiki platform (wiki-pain.org) that enables efficient manual curation and exploration of the molecular mechanisms that underlie pain. Curation of 1500 grouped event chains ranked by pain relevance revealed 613 accurately extracted unique molecular interactions that in the future can be used to study the underlying mechanisms involved in pain. Our approach demonstrates that combining existing text mining tools with domain-specific terms and wiki-based visualization can facilitate rapid curation of molecular interactions to create a custom database. Database URL: •••

A High-Confidence Interaction Map Identifies SIRT1 as a Mediator of Acetylation of USP22 and the SAGA Coactivator Complex

PubMed Central

Armour, Sean M.; Bennett, Eric J.; Braun, Craig R.; Zhang, Xiao-Yong; McMahon, Steven B.; Gygi, Steven P.; Harper, J. Wade

2013-01-01

Although many functions and targets have been attributed to the histone and protein deacetylase SIRT1, a comprehensive analysis of SIRT1 binding proteins yielding a high-confidence interaction map has not been established. Using a comparative statistical analysis of binding partners, we have assembled a high-confidence SIRT1 interactome. Employing this method, we identified the deubiquitinating enzyme ubiquitin-specific protease 22 (USP22), a component of the deubiquitinating module (DUBm) of the SAGA transcriptional coactivating complex, as a SIRT1-interacting partner. We found that this interaction is highly specific, requires the ZnF-UBP domain of USP22, and is disrupted by the inactivating H363Y mutation within SIRT1. Moreover, we show that USP22 is acetylated on multiple lysine residues and that alteration of a single lysine (K129) within the ZnF-UBP domain is sufficient to alter interaction of the DUBm with the core SAGA complex. Furthermore, USP22-mediated recruitment of SIRT1 activity promotes the deacetylation of individual SAGA complex components. Our results indicate an important role of SIRT1-mediated deacetylation in regulating the formation of DUBm subcomplexes within the larger SAGA complex. PMID:23382074

AFD: an application for bi-molecular interaction using axial frequency distribution.

PubMed

Raza, Saad; Azam, Syed Sikander

2018-03-06

Conformational flexibility and generalized structural features are responsible for specific phenomena existing in biological pathways. With advancements in computational chemistry, novel approaches and new methods are required to compare the dynamic nature of biomolecules, which are crucial not only to address dynamic functional relationships but also to gain detailed insights into the disturbance and positional fluctuation responsible for functional shifts. Keeping this in mind, axial frequency distribution (AFD) has been developed, designed, and implemented. AFD can profoundly represent distribution and density of ligand atom around a particular atom or set of atoms. It enabled us to obtain an explanation of local movements and rotations, which are not significantly highlighted by any other structural and dynamical parameters. AFD can be implemented on biological models representing ligand and protein interactions. It shows a comprehensive view of the binding pattern of ligand by exploring the distribution of atoms relative to the x-y plane of the system. By taking a relative centroid on protein or ligand, molecular interactions like hydrogen bonds, van der Waals, polar or ionic interaction can be analyzed to cater the ligand movement, stabilization or flexibility with respect to the protein. The AFD graph resulted in the residual depiction of bi-molecular interaction in gradient form which can yield specific information depending upon the system of interest.

A new reference implementation of the PSICQUIC web service.

PubMed

del-Toro, Noemi; Dumousseau, Marine; Orchard, Sandra; Jimenez, Rafael C; Galeota, Eugenia; Launay, Guillaume; Goll, Johannes; Breuer, Karin; Ono, Keiichiro; Salwinski, Lukasz; Hermjakob, Henning

2013-07-01

The Proteomics Standard Initiative Common QUery InterfaCe (PSICQUIC) specification was created by the Human Proteome Organization Proteomics Standards Initiative (HUPO-PSI) to enable computational access to molecular-interaction data resources by means of a standard Web Service and query language. Currently providing >150 million binary interaction evidences from 28 servers globally, the PSICQUIC interface allows the concurrent search of multiple molecular-interaction information resources using a single query. Here, we present an extension of the PSICQUIC specification (version 1.3), which has been released to be compliant with the enhanced standards in molecular interactions. The new release also includes a new reference implementation of the PSICQUIC server available to the data providers. It offers augmented web service capabilities and improves the user experience. PSICQUIC has been running for almost 5 years, with a user base growing from only 4 data providers to 28 (April 2013) allowing access to 151 310 109 binary interactions. The power of this web service is shown in PSICQUIC View web application, an example of how to simultaneously query, browse and download results from the different PSICQUIC servers. This application is free and open to all users with no login requirement (http://www.ebi.ac.uk/Tools/webservices/psicquic/view/main.xhtml).

Interactions Between the Diatom Thalassiosira pseudonana and Heterotrophic Bacterium Pelagibacter ubique Examined in Co-Culture

NASA Astrophysics Data System (ADS)

Moore, E.; Giovannoni, S. J.; Halsey, K.

2016-02-01

Recent studies employing network theory have focused attention on the importance of interactions between taxa in microbial plankton community ecology. In this study, the centric diatom Thalassiosira pseudonana and the heterotrophic bacterium Pelagibacter ubique HTCC1062, both coastal organisms, were cultured independently and together in co-culture to investigate phytoplankton-bacteria interactions at the molecular level. A basis for predicting synergistic interactions between these plankton taxa can be found in published descriptions of specific vitamin, nutrient, and carbon compound requirements for their growth, but no experiments have been done that measure their physiological and molecular responses to co-culturing. When grown in co-culture with T. pseudonana, HTCC1062 grew slightly, but significantly faster than in monoculture. The generation time of HTCC1062 grown in co-culture was consistently 2.5 to 6.3 hours faster than when grown in monoculture (p-value <0.05). There was no significant difference in the growth rate of T. pseudonana between co- and monoculture. These results indicate that HTCC1062 is benefiting from the presence of the diatom by an unknown mechanism. Messenger RNA sequencing is being utilized to examine transcriptional responses that may provide insight into specific interactions that contribute to the observed shift in Pelagibacter growth rates.

Target-specific NMR detection of protein-ligand interactions with antibody-relayed 15N-group selective STD.

PubMed

Hetényi, Anasztázia; Hegedűs, Zsófia; Fajka-Boja, Roberta; Monostori, Éva; Kövér, Katalin E; Martinek, Tamás A

2016-12-01

Fragment-based drug design has been successfully applied to challenging targets where the detection of the weak protein-ligand interactions is a key element. 1 H saturation transfer difference (STD) NMR spectroscopy is a powerful technique for this work but it requires pure homogeneous proteins as targets. Monoclonal antibody (mAb)-relayed 15 N-GS STD spectroscopy has been developed to resolve the problem of protein mixtures and impure proteins. A 15 N-labelled target-specific mAb is selectively irradiated and the saturation is relayed through the target to the ligand. Tests on the anti-Gal-1 mAb/Gal-1/lactose system showed that the approach is experimentally feasible in a reasonable time frame. This method allows detection and identification of binding molecules directly from a protein mixture in a multicomponent system.

Through its F-BAR and RhoGAP domains, Rgd1p acts in different polarized growth processes in budding yeast

PubMed Central

Lefebvre, Fabien; Prouzet-Mauléon, Valérie; Vieillemard, Aurélie; Thoraval, Didier; Crouzet, Marc

2009-01-01

Protein domain architecture can be used to construct supramolecular structures, to carry out specific functions and to mediate signaling in prokaryotic and eukaryotic cells. The Rgd1p protein of budding yeast contains two domains with different functions in the cell: the F-BAR and RhoGAP domains. The F-BAR domain has been shown to interact with membrane phospholipids and is thought to induce or sense membrane curvature. The RhoGAP domain activates the GTP hydrolysis of two Rho GTPases, thereby regulating different cellular pathways. Specific molecular interactions with the F-BAR and RhoGAP domains, cell signaling and interplay between these domains may allow the Rgd1p protein to act in several different biological processes, all of which are required for polarized growth in yeast. PMID:19704907

Regulatory T cells in the control of host-microorganism interactions (*).

PubMed

Belkaid, Yasmine; Tarbell, Kristin

2009-01-01

Each microenvironment requires a specific set of regulatory elements that are finely and constantly tuned to maintain local homeostasis. Various populations of regulatory T cells contribute to the maintenance of this equilibrium and establishment of controlled immune responses. In particular, regulatory T cells limit the magnitude of effector responses, which may result in failure to adequately control infection. However, regulatory T cells also help limit collateral tissue damage caused by vigorous antimicrobial immune responses against pathogenic microbes as well as commensals. In this review, we describe various situations in which the balance between regulatory T cells and effector immune functions influence the outcome of host-microorganism coexistence and discuss current hypotheses and points of polemic associated with the origin, target, and antigen specificity of both endogenous and induced regulatory T cells during these interactions.

High-Throughput Live-Cell Microscopy Analysis of Association Between Chromosome Domains and the Nucleolus in S. cerevisiae.

PubMed

Wang, Renjie; Normand, Christophe; Gadal, Olivier

2016-01-01

Spatial organization of the genome has important impacts on all aspects of chromosome biology, including transcription, replication, and DNA repair. Frequent interactions of some chromosome domains with specific nuclear compartments, such as the nucleolus, are now well documented using genome-scale methods. However, direct measurement of distance and interaction frequency between loci requires microscopic observation of specific genomic domains and the nucleolus, followed by image analysis to allow quantification. The fluorescent repressor operator system (FROS) is an invaluable method to fluorescently tag DNA sequences and investigate chromosome position and dynamics in living cells. This chapter describes a combination of methods to define motion and region of confinement of a locus relative to the nucleolus in cell's nucleus, from fluorescence acquisition to automated image analysis using two dedicated pipelines.

Eukaryotic DNA Ligases: Structural and Functional Insights

PubMed Central

Ellenberger, Tom; Tomkinson, Alan E.

2010-01-01

DNA ligases are required for DNA replication, repair, and recombination. In eukaryotes, there are three families of ATP-dependent DNA ligases. Members of the DNA ligase I and IV families are found in all eukaryotes, whereas DNA ligase III family members are restricted to vertebrates. These enzymes share a common catalytic region comprising a DNA-binding domain, a nucleotidyltransferase (NTase) domain, and an oligonucleotide/oligosaccharide binding (OB)-fold domain. The catalytic region encircles nicked DNA with each of the domains contacting the DNA duplex. The unique segments adjacent to the catalytic region of eukaryotic DNA ligases are involved in specific protein-protein interactions with a growing number of DNA replication and repair proteins. These interactions determine the specific cellular functions of the DNA ligase isozymes. In mammals, defects in DNA ligation have been linked with an increased incidence of cancer and neurodegeneration. PMID:18518823

A Protein Linkage Map of the P2 Nonstructural Proteins of Poliovirus

PubMed Central

Cuconati, Andrea; Xiang, Wenkai; Lahser, Frederick; Pfister, Thomas; Wimmer, Eckard

1998-01-01

The yeast two-hybrid system was used to catalog all detectable interactions among the P2 nonstructural cleavage products of poliovirus type 1 (Mahoney). Evidence has been obtained for specific associations among 2Apro, 2BC, 2C, and 2B. Specifically, 2Apro can interact with itself and 2BC and its cleavage products (2B and 2C) interact in all possible combinations, with the exception of 2C/2C. Detected interactions were confirmed in vitro by a glutathione S-transferase pulldown assay, which allowed us to detect 2C/2C association. trans-dominant-negative mutants of 2B (K. Johnson and P. J. Sarnow, J. Virol. 65:4341–4349, 1991) were examined and were found to retain interaction with wild-type 2B, perhaps reflecting a need for 2B multimerization in viral RNA replication. The multimerization of 2B was examined further by screening a mutagenized library for 2B variants that have lost the ability to bind wild-type 2B. The screen identified two nonconservative missense mutations within a central hydrophobic region, as well as truncations and frameshifts that implicate the C terminus in homointeraction. Introduction of the missense mutations into the genome of the virus conferred a quasi-infectious phenotype, an observation strongly suggesting that the 2B/2B interaction is required for replication of the viral genome. PMID:9445030

Cyclophilin B binding to platelets supports calcium-dependent adhesion to collagen.

PubMed

Allain, F; Durieux, S; Denys, A; Carpentier, M; Spik, G

1999-08-01

We have recently reported that cyclophilin B (CyPB), a secreted cyclosporine-binding protein, could bind to T lymphocytes through interactions with two types of binding sites. The first ones, referred to as type I, involve interactions with the conserved domain of CyPB and promote the endocytosis of surface-bound ligand, while the second type of binding sites, termed type II, are represented by glycosaminoglycans (GAG). Here, we further investigated the interactions of CyPB with blood cell populations. In addition to lymphocytes, CyPB was found to interact mainly with platelets. The binding is specific, with a dissociation constant (kd) of 9 +/- 3 nmol/L and the number of sites estimated at 960 +/- 60 per cell. Platelet glycosaminoglycans are not required for the interactions, but the binding is dramatically reduced by active cyclosporine derivatives. We then analyzed the biologic effects of CyPB and found a significant increase in platelet adhesion to collagen. Concurrently, CyPB initiates a transmembranous influx of Ca(2+) and induces the phosphorylation of the P-20 light chains of myosin. Taken together, the present results demonstrate for the first time that extracellular CyPB specifically interacts with platelets through a functional receptor related to the lymphocyte type I binding sites and might act by regulating the activity of a receptor-operated membrane Ca(2+) channel.

Patient portal readiness among postpartum patients in a safety net setting.

PubMed

Wieland, Daryl; Gibeau, Anne; Dewey, Caitlin; Roshto, Melanie; Frankel, Hilary

2017-07-05

Maternity patients interact with the healthcare system over an approximately ten-month interval, requiring multiple visits, acquiring pregnancy-specific education, and sharing health information among providers. Many features of a web-based patient portal could help pregnant women manage their interactions with the healthcare system; however, it is unclear whether pregnant women in safety-net settings have the resources, skills or interest required for portal adoption. In this study of postpartum patients in a safety net hospital, we aimed to: (1) determine if patients have the technical resources and skills to access a portal, (2) gain insight into their interest in health information, and (3) identify the perceived utility of portal features and potential barriers to adoption. We developed a structured questionnaire to collect demographics from postpartum patients and measure use of technology and the internet, self-reported literacy, interest in health information, awareness of portal functions, and perceived barriers to use. The questionnaire was administered in person to women in an inpatient setting. Of the 100 participants surveyed, 95% reported routine internet use and 56% used it to search for health information. Most participants had never heard of a patient portal, yet 92% believed that the portal functions were important. The two most appealing functions were to check results and manage appointments. Most participants in this study have the required resources such as a device and familiarity with the internet to access a patient portal including an interest in interacting with a healthcare institution via electronic means. Pregnancy is a critical episode of care where active engagement with the healthcare system can influence outcomes. Healthcare systems and portal developers should consider ways to tailor a portal to address the specific health needs of a maternity population including those in a safety net setting.

Profilin is required for viral morphogenesis, syncytium formation, and cell-specific stress fiber induction by respiratory syncytial virus

PubMed Central

Bitko, Vira; Oldenburg, Anja; Garmon, Nicolle E; Barik, Sailen

2003-01-01

Background Actin is required for the gene expression and morphogenesis of respiratory syncytial virus (RSV), a clinically important Pneumovirus of the Paramyxoviridae family. In HEp-2 cells, RSV infection also induces actin stress fibers, which may be important in the immunopathology of the RSV disease. Profilin, a major regulator of actin polymerization, stimulates viral transcription in vitro. Thus, we tested the role of profilin in RSV growth and RSV-actin interactions in cultured cells (ex vivo). Results We tested three cell lines: HEp-2 (human), A549 (human), and L2 (rat). In all three, RSV grew well and produced fused cells (syncytium), and two RSV proteins, namely, the phosphoprotein P and the nucleocapsid protein N, associated with profilin. In contrast, induction of actin stress fibers by RSV occurred in HEp-2 and L2 cells, but not in A549. Knockdown of profilin by RNA interference had a small effect on viral macromolecule synthesis but strongly inhibited maturation of progeny virions, cell fusion, and induction of stress fibers. Conclusions Profilin plays a cardinal role in RSV-mediated cell fusion and viral maturation. In contrast, interaction of profilin with the viral transcriptional proteins P and N may only nominally activate viral RNA-dependent RNA polymerase. Stress fiber formation is a cell-specific response to infection, requiring profilin and perhaps other signaling molecules that are absent in certain cell lines. Stress fibers per se play no role in RSV replication in cell culture. Clearly, the cellular architecture controls multiple steps of host-RSV interaction, some of which are regulated by profilin. PMID:12740026

Alterations in the Proteome of the Euprymna scolopes Light Organ in Response to Symbiotic Vibrio fischeri

PubMed Central

Doino Lemus, Judith; McFall-Ngai, Margaret J.

2000-01-01

During the onset of the cooperative association between the Hawaiian sepiolid squid Euprymna scolopes and the marine luminous bacterium Vibrio fischeri, the anatomy and morphology of the host's symbiotic organ undergo dramatic changes that require interaction with the bacteria. This morphogenetic process involves an array of tissues, including those in direct contact with, as well as those remote from, the symbiotic bacteria. The bacteria induce the developmental program soon after colonization of the organ, although complete morphogenesis requires 96 h. In this study, to determine critical time points, we examined the biochemistry underlying bacterium-induced host development using two-dimensional polyacrylamide gel electrophoresis. Specifically, V. fischeri-induced changes in the soluble proteome of the symbiotic organ during the first 96 h of symbiosis were identified by comparing the protein profiles of symbiont-colonized and uncolonized organs. Both symbiosis-related changes and age-related changes were analyzed to determine what proportion of the differences in the proteomes was the result of specific responses to interaction with bacteria. Although no differences were detected over the first 24 h, numerous symbiosis-related changes became apparent at 48 and 96 h and were more abundant than age-related changes. In addition, many age-related protein changes occurred 48 h sooner in symbiotic animals, suggesting that the interaction of squid tissue with V. fischeri cells accelerates certain developmental processes of the symbiotic organ. These data suggest that V. fischeri-induced modifications in host tissues that occur in the first 24 h of the symbiosis are independent of marked alterations in the patterns of abundant proteins but that the full 4-day morphogenetic program requires significant alteration of the host soluble proteome. PMID:10966433

Alterations in the proteome of the Euprymna scolopes light organ in response to symbiotic Vibrio fischeri.

PubMed

Doino Lemus, J; McFall-Ngai, M J

2000-09-01

During the onset of the cooperative association between the Hawaiian sepiolid squid Euprymna scolopes and the marine luminous bacterium Vibrio fischeri, the anatomy and morphology of the host's symbiotic organ undergo dramatic changes that require interaction with the bacteria. This morphogenetic process involves an array of tissues, including those in direct contact with, as well as those remote from, the symbiotic bacteria. The bacteria induce the developmental program soon after colonization of the organ, although complete morphogenesis requires 96 h. In this study, to determine critical time points, we examined the biochemistry underlying bacterium-induced host development using two-dimensional polyacrylamide gel electrophoresis. Specifically, V. fischeri-induced changes in the soluble proteome of the symbiotic organ during the first 96 h of symbiosis were identified by comparing the protein profiles of symbiont-colonized and uncolonized organs. Both symbiosis-related changes and age-related changes were analyzed to determine what proportion of the differences in the proteomes was the result of specific responses to interaction with bacteria. Although no differences were detected over the first 24 h, numerous symbiosis-related changes became apparent at 48 and 96 h and were more abundant than age-related changes. In addition, many age-related protein changes occurred 48 h sooner in symbiotic animals, suggesting that the interaction of squid tissue with V. fischeri cells accelerates certain developmental processes of the symbiotic organ. These data suggest that V. fischeri-induced modifications in host tissues that occur in the first 24 h of the symbiosis are independent of marked alterations in the patterns of abundant proteins but that the full 4-day morphogenetic program requires significant alteration of the host soluble proteome.

Physical interaction of the activator protein-1 factors c-Fos and c-Jun with Cbfa1 for collagenase-3 promoter activation

NASA Technical Reports Server (NTRS)

D'Alonzo, Richard C.; Selvamurugan, Nagarajan; Karsenty, Gerard; Partridge, Nicola C.

2002-01-01

Previously, we determined that the activator protein-1 (AP-1)-binding site and the runt domain (RD)-binding site and their binding proteins, c-Fos.c-Jun and Cbfa, regulate the collagenase-3 promoter in parathyroid hormone-treated and differentiating osteoblasts. Here we show that Cbfa1 and c-Fos.c-Jun appear to cooperatively bind the RD- and AP-1-binding sites and form ternary structures in vitro. Both in vitro and in vivo co-immunoprecipitation and yeast two-hybrid studies further demonstrate interaction between Cbfa1 with c-Fos and c-Jun in the absence of phosphorylation and without binding to DNA. Additionally, only the runt domain of Cbfa1 was required for interaction with c-Jun and c-Fos. In mammalian cells, overexpression of Cbfa1 enhanced c-Jun activation of AP-1-binding site promoter activity, demonstrating functional interaction. Finally, insertion of base pairs that disrupted the helical phasing between the AP-1- and RD-binding sites also inhibited collagenase-3 promoter activation. Thus, we provide direct evidence that Cbfa1 and c-Fos.c-Jun physically interact and cooperatively bind the AP-1- and RD-binding sites in the collagenase-3 promoter. Moreover, the AP-1- and RD-binding sites appear to be organized in a specific required helical arrangement that facilitates transcription factor interaction and enables promoter activation.

Interactions between late acting proteins required for peptidoglycan synthesis during sporulation

PubMed Central

Fay, Allison; Meyer, Pablo; Dworkin, Jonathan

2010-01-01

The requirement of peptidoglycan synthesis for growth complicates the analysis of interactions between proteins involved in this pathway. In particular, the later steps that involve membrane-linked substrates have proven largely recalcitrant to in vivo analysis. Here we have taken advantage of the peptidoglycan synthesis that occurs during sporulation in Bacillus subtilis to examine the interactions between SpoVE, a non-essential, sporulation-specific homolog of the well-conserved and essential SEDS proteins, and SpoVD, a non-essential class B penicillin binding protein (PBP). We found that localization of SpoVD is dependent on SpoVE and that SpoVD protects SpoVE from in vivo proteolysis. Co-immunoprecipitations and Fluorescence Resonance Energy Transfer experiments indicated that SpoVE and SpoVD interact and co-affinity purification in E. coli demonstrated that this interaction is direct. Finally, we generated a functional protein consisting of a SpoVE-SpoVD fusion and found that a loss-of-function point mutation in either part of the fusion resulted in a loss of function of the entire fusion that was not complemented by a wild type protein. Thus, SpoVE has a direct and functional interaction with SpoVD and this conclusion will facilitate understanding the essential function SpoVE and related SEDS proteins such as FtsW and RodA play in bacterial growth and division. PMID:20417640

Peripheral stator of the yeast V-ATPase: stoichiometry and specificity of interaction between the EG complex and subunits C and H.

PubMed

Féthière, James; Venzke, David; Madden, Dean R; Böttcher, Bettina

2005-12-06

V-ATPases are multisubunit membrane protein complexes that use the energy provided by ATP hydrolysis to generate a proton gradient across various intracellular and plasma membranes. In doing so, they maintain an acidic pH in the lumen of intracellular organelles and acidify extracellular milieu to support specific cellular functions. V-ATPases are structurally similar to the F1F0-ATP synthase, with an intrinsic membrane domain (V0) and an extrinsic peripheral domain (V1) joined by several connecting elements. To gain a clear functional understanding of the catalytic mechanism, and of the stability requirements for regulatory processes in the enzyme, a clear topology of the enzyme has to be established. In particular, the composition and arrangement of the peripheral stator subunits must be firmly settled, as these play specific roles in catalysis and regulation. We have designed a strategy allowing us to coexpress different combinations of these subunits to delineate specific interactions. In this study, we report the interaction between the peripheral stator EG complex and subunits C and H of the V-ATPase from the yeast Saccharomyces cerevisae. A combination of analytical gel filtration, native gel electrophoresis, and ultracentrifugation analysis allowed us to ascertain the homogeneity and molar mass of the purified EGC complex as well as of the EG complex, supporting the formation of 1:1(:1) stoichiometric complexes. The EGC complex can be formed in vitro by combining equimolar amounts of subunit C and the EG subcomplex and results most likely from the initial interaction between subunits E and C.

A new analytical method for characterizing nonlinear visual processes with stimuli of arbitrary distribution: Theory and applications.

PubMed

Hayashi, Ryusuke; Watanabe, Osamu; Yokoyama, Hiroki; Nishida, Shin'ya

2017-06-01

Characterization of the functional relationship between sensory inputs and neuronal or observers' perceptual responses is one of the fundamental goals of systems neuroscience and psychophysics. Conventional methods, such as reverse correlation and spike-triggered data analyses are limited in their ability to resolve complex and inherently nonlinear neuronal/perceptual processes because these methods require input stimuli to be Gaussian with a zero mean. Recent studies have shown that analyses based on a generalized linear model (GLM) do not require such specific input characteristics and have advantages over conventional methods. GLM, however, relies on iterative optimization algorithms and its calculation costs become very expensive when estimating the nonlinear parameters of a large-scale system using large volumes of data. In this paper, we introduce a new analytical method for identifying a nonlinear system without relying on iterative calculations and yet also not requiring any specific stimulus distribution. We demonstrate the results of numerical simulations, showing that our noniterative method is as accurate as GLM in estimating nonlinear parameters in many cases and outperforms conventional, spike-triggered data analyses. As an example of the application of our method to actual psychophysical data, we investigated how different spatiotemporal frequency channels interact in assessments of motion direction. The nonlinear interaction estimated by our method was consistent with findings from previous vision studies and supports the validity of our method for nonlinear system identification.

A Novel RNA Polymerase I Transcription Initiation Factor, TIF-IE, Commits rRNA Genes by Interaction with TIF-IB, Not by DNA Binding

PubMed Central

Al-Khouri, Anna Maria; Paule, Marvin R.

2002-01-01

In the small, free-living amoeba Acanthamoeba castellanii, rRNA transcription requires, in addition to RNA polymerase I, a single DNA-binding factor, transcription initiation factor IB (TIF-IB). TIF-IB is a multimeric protein that contains TATA-binding protein (TBP) and four TBP-associated factors that are specific for polymerase I transcription. TIF-IB is required for accurate and promoter-specific initiation of rRNA transcription, recruiting and positioning the polymerase on the start site by protein-protein interaction. In A. castellanii, partially purified TIF-IB can form a persistent complex with the ribosomal DNA (rDNA) promoter while homogeneous TIF-IB cannot. An additional factor, TIF-IE, is required along with homogeneous TIF-IB for the formation of a stable complex on the rDNA core promoter. We show that TIF-IE by itself, however, does not bind to the rDNA promoter and thus differs in its mechanism from the upstream binding factor and upstream activating factor, which carry out similar complex-stabilizing functions in vertebrates and yeast, respectively. In addition to its presence in impure TIF-IB, TIF-IE is found in highly purified fractions of polymerase I, with which it associates. Renaturation of polypeptides excised from sodium dodecyl sulfate-polyacrylamide gels showed that a 141-kDa polypeptide possesses all the known activities of TIF-IE. PMID:11784852

A novel RNA polymerase I transcription initiation factor, TIF-IE, commits rRNA genes by interaction with TIF-IB, not by DNA binding.

PubMed

Al-Khouri, Anna Maria; Paule, Marvin R

2002-02-01

In the small, free-living amoeba Acanthamoeba castellanii, rRNA transcription requires, in addition to RNA polymerase I, a single DNA-binding factor, transcription initiation factor IB (TIF-IB). TIF-IB is a multimeric protein that contains TATA-binding protein (TBP) and four TBP-associated factors that are specific for polymerase I transcription. TIF-IB is required for accurate and promoter-specific initiation of rRNA transcription, recruiting and positioning the polymerase on the start site by protein-protein interaction. In A. castellanii, partially purified TIF-IB can form a persistent complex with the ribosomal DNA (rDNA) promoter while homogeneous TIF-IB cannot. An additional factor, TIF-IE, is required along with homogeneous TIF-IB for the formation of a stable complex on the rDNA core promoter. We show that TIF-IE by itself, however, does not bind to the rDNA promoter and thus differs in its mechanism from the upstream binding factor and upstream activating factor, which carry out similar complex-stabilizing functions in vertebrates and yeast, respectively. In addition to its presence in impure TIF-IB, TIF-IE is found in highly purified fractions of polymerase I, with which it associates. Renaturation of polypeptides excised from sodium dodecyl sulfate-polyacrylamide gels showed that a 141-kDa polypeptide possesses all the known activities of TIF-IE.

Vaccine development: From concept to early clinical testing.

PubMed

Cunningham, Anthony L; Garçon, Nathalie; Leo, Oberdan; Friedland, Leonard R; Strugnell, Richard; Laupèze, Béatrice; Doherty, Mark; Stern, Peter

2016-12-20

In the 21st century, an array of microbiological and molecular allow antigens for new vaccines to be specifically identified, designed, produced and delivered with the aim of optimising the induction of a protective immune response against a well-defined immunogen. New knowledge about the functioning of the immune system and host pathogen interactions has stimulated the rational design of vaccines. The design toolbox includes vaccines made from whole pathogens, protein subunits, polysaccharides, pathogen-like particles, use of viral/bacterial vectors, plus adjuvants and conjugation technology to increase and broaden the immune response. Processes such as recombinant DNA technology can simplify the complexity of manufacturing and facilitate consistent production of large quantities of antigen. Any new vaccine development is greatly enhanced by, and requires integration of information concerning: 1. Pathogen life-cycle & epidemiology. Knowledge of pathogen structure, route of entry, interaction with cellular receptors, subsequent replication sites and disease-causing mechanisms are all important to identify antigens suitable for disease prevention. The demographics of infection, specific risk groups and age-specific infection rates determine which population to immunise, and at what age. 2. Immune control & escape. Interactions between the host and pathogen are explored, with determination of the relative importance of antibodies, T-cells of different types and innate immunity, immune escape strategies during infection, and possible immune correlates of protection. This information guides identification and selection of antigen and the specific immune response required for protection. 3. Antigen selection & vaccine formulation. The selected antigen is formulated to remain suitably immunogenic and stable over time, induce an immune response that is likely to be protective, plus be amenable to eventual scale-up to commercial production. 4. Vaccine preclinical & clinical testing. The candidate vaccine must be tested for immunogenicity, safety and efficacy in preclinical and appropriately designed clinical trials. This review considers these processes using examples of differing pathogenic challenges, including human papillomavirus, malaria, and ebola. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.

Validating the disruption of proliferating cell nuclear antigen interactions in the development of targeted cancer therapeutics.

PubMed

Smith, Shanna J; Hickey, Robert J; Malkas, Linda H

2016-01-01

Human DNA replication and repair is a highly coordinated process involving the specifically timed actions of numerous proteins and enzymes. Many of these proteins require interaction with proliferating cell nuclear antigen (PCNA) for activation within the process. The interdomain connector loop (IDCL) of PCNA provides a docking site for many of those proteins, suggesting that this region is critically important in the regulation of cellular function. Previous work in this laboratory has demonstrated that a peptide mimicking a specific region of the IDCL (caPeptide) has the ability to disrupt key protein-protein interactions between PCNA and its binding partners, thereby inhibiting DNA replication within the cells. In this study, we confirm the ability of the caPeptide to disrupt DNA replication function using both intact cell and in vitro DNA replication assays. Further, we were able to demonstrate that treatment with caPeptide results in a decrease of polymerase δ activity that correlates with the observed decrease in DNA replication. We have also successfully developed a surface plasmon resonance (SPR) assay to validate the disruption of the PCNA-pol δ interaction with caPeptide.

Knowledge is power: how conceptual knowledge transforms visual cognition.

PubMed

Collins, Jessica A; Olson, Ingrid R

2014-08-01

In this review, we synthesize the existing literature demonstrating the dynamic interplay between conceptual knowledge and visual perceptual processing. We consider two theoretical frameworks that demonstrate interactions between processes and brain areas traditionally considered perceptual or conceptual. Specifically, we discuss categorical perception, in which visual objects are represented according to category membership, and highlight studies showing that category knowledge can penetrate early stages of visual analysis. We next discuss the embodied account of conceptual knowledge, which holds that concepts are instantiated in the same neural regions required for specific types of perception and action, and discuss the limitations of this framework. We additionally consider studies showing that gaining abstract semantic knowledge about objects and faces leads to behavioral and electrophysiological changes that are indicative of more efficient stimulus processing. Finally, we consider the role that perceiver goals and motivation may play in shaping the interaction between conceptual and perceptual processing. We hope to demonstrate how pervasive such interactions between motivation, conceptual knowledge, and perceptual processing are in our understanding of the visual environment, and to demonstrate the need for future research aimed at understanding how such interactions arise in the brain.

Physical Studies of P450–P450 Interactions: Predicting Quaternary Structures of P450 Complexes in Membranes from Their X-ray Crystal Structures

PubMed Central

Reed, James R.; Backes, Wayne L.

2017-01-01

Cytochrome P450 enzymes, which catalyze oxygenation reactions of both exogenous and endogenous chemicals, are membrane bound proteins that require interaction with their redox partners in order to function. Those responsible for drug and foreign compound metabolism are localized primarily in the endoplasmic reticulum of liver, lung, intestine, and other tissues. More recently, the potential for P450 enzymes to exist as supramolecular complexes has been shown by the demonstration of both homomeric and heteromeric complexes. The P450 units in these complexes are heterogeneous with respect to their distribution and function, and the interaction of different P450s can influence P450-specific metabolism. The goal of this review is to examine the evidence supporting the existence of physical complexes among P450 enzymes. Additionally, the review examines the crystal lattices of different P450 enzymes derived from X-ray diffraction data to make assumptions regarding possible quaternary structures in membranes and in turn, to predict how the quaternary structures could influence metabolism and explain the functional effects of specific P450–P450 interactions. PMID:28194112

Knowledge is Power: How Conceptual Knowledge Transforms Visual Cognition

PubMed Central

Collins, Jessica A.; Olson, Ingrid R.

2014-01-01

In this review we synthesize the existing literature demonstrating the dynamic interplay between conceptual knowledge and visual perceptual processing. We consider two theoretical frameworks demonstrating interactions between processes and brain areas traditionally considered perceptual or conceptual. Specifically, we discuss categorical perception, in which visual objects are represented according to category membership, and highlight studies showing that category knowledge can penetrate early stages of visual analysis. We next discuss the embodied account of conceptual knowledge, which holds that concepts are instantiated in the same neural regions required for specific types of perception and action, and discuss the limitations of this framework. We additionally consider studies showing that gaining abstract semantic knowledge about objects and faces leads to behavioral and electrophysiological changes that are indicative of more efficient stimulus processing. Finally, we consider the role that perceiver goals and motivation may play in shaping the interaction between conceptual and perceptual processing. We hope to demonstrate how pervasive such interactions between motivation, conceptual knowledge, and perceptual processing are to our understanding of the visual environment, and demonstrate the need for future research aimed at understanding how such interactions arise in the brain. PMID:24402731

Novel interactions of CAPS (Ca2+-dependent activator protein for secretion) with the three neuronal SNARE proteins required for vesicle fusion.

PubMed

Daily, Neil J; Boswell, Kristin L; James, Declan J; Martin, Thomas F J

2010-11-12

CAPS (aka CADPS) is required for optimal vesicle exocytosis in neurons and endocrine cells where it functions to prime the exocytic machinery for Ca(2+)-triggered fusion. Fusion is mediated by trans complexes of the SNARE proteins VAMP-2, syntaxin-1, and SNAP-25 that bridge vesicle and plasma membrane. CAPS promotes SNARE complex formation on liposomes, but the SNARE binding properties of CAPS are unknown. The current work revealed that CAPS exhibits high affinity binding to syntaxin-1 and SNAP-25 and moderate affinity binding to VAMP-2. CAPS binding is specific for a subset of exocytic SNARE protein isoforms and requires membrane integration of the SNARE proteins. SNARE protein binding by CAPS is novel and mediated by interactions with the SNARE motifs in the three proteins. The C-terminal site for CAPS binding on syntaxin-1 does not overlap the Munc18-1 binding site and both proteins can co-reside on membrane-integrated syntaxin-1. As expected for a C-terminal binding site on syntaxin-1, CAPS stimulates SNARE-dependent liposome fusion with N-terminal truncated syntaxin-1 but exhibits impaired activity with C-terminal syntaxin-1 mutants. Overall the results suggest that SNARE complex formation promoted by CAPS may be mediated by direct interactions of CAPS with each of the three SNARE proteins required for vesicle exocytosis.

Novel Interactions of CAPS (Ca2+-dependent Activator Protein for Secretion) with the Three Neuronal SNARE Proteins Required for Vesicle Fusion*

PubMed Central

Daily, Neil J.; Boswell, Kristin L.; James, Declan J.; Martin, Thomas F. J.

2010-01-01

CAPS (aka CADPS) is required for optimal vesicle exocytosis in neurons and endocrine cells where it functions to prime the exocytic machinery for Ca2+-triggered fusion. Fusion is mediated by trans complexes of the SNARE proteins VAMP-2, syntaxin-1, and SNAP-25 that bridge vesicle and plasma membrane. CAPS promotes SNARE complex formation on liposomes, but the SNARE binding properties of CAPS are unknown. The current work revealed that CAPS exhibits high affinity binding to syntaxin-1 and SNAP-25 and moderate affinity binding to VAMP-2. CAPS binding is specific for a subset of exocytic SNARE protein isoforms and requires membrane integration of the SNARE proteins. SNARE protein binding by CAPS is novel and mediated by interactions with the SNARE motifs in the three proteins. The C-terminal site for CAPS binding on syntaxin-1 does not overlap the Munc18-1 binding site and both proteins can co-reside on membrane-integrated syntaxin-1. As expected for a C-terminal binding site on syntaxin-1, CAPS stimulates SNARE-dependent liposome fusion with N-terminal truncated syntaxin-1 but exhibits impaired activity with C-terminal syntaxin-1 mutants. Overall the results suggest that SNARE complex formation promoted by CAPS may be mediated by direct interactions of CAPS with each of the three SNARE proteins required for vesicle exocytosis. PMID:20826818

Structural and motional contributions of the Bacillus subtilis ClpC N-domain in adaptor protein interactions

PubMed Central

Kojetin, Douglas J.; McLaughlin, Patrick D.; Thompson, Richele J.; Dubnau, David; Prepiak, Peter; Rance, Mark; Cavanagh, John

2009-01-01

Summary The AAA+ superfamily protein ClpC is a key regulator of cell development in Bacillus subtilis. As part of a large oligomeric complex, ClpC controls an array of cellular processes by recognizing, unfolding, and providing misfolded and aggregated proteins as substrates for the ClpP peptidase. ClpC is unique compared to other HSP100/Clp proteins, as it requires an adaptor protein for all fundamental activities. The NMR solution structure of the N-terminal repeat domain of ClpC (N-ClpCR) comprises two structural repeats of a four-helix motif. NMR experiments used to map the MecA adaptor protein interaction surface of N-ClpCR reveal that regions involved in the interaction possess conformational flexibility, as well as conformational exchange on the μs-ms time-scale. The electrostatic surface of N-ClpCR differs substantially compared to the N-domain of Escherichia coli ClpA and ClpB, suggesting that the electrostatic surface characteristics of HSP100/Clp N-domains may play a role in adaptor protein and substrate interaction specificity, and perhaps contribute to the unique adaptor protein requirement of ClpC. PMID:19361434

Interaction of proliferating cell nuclear antigen with PMS2 is required for MutLα activation and function in mismatch repair

PubMed Central

Genschel, Jochen; Kadyrova, Lyudmila Y.; Iyer, Ravi R.; Dahal, Basanta K.; Kadyrov, Farid A.; Modrich, Paul

2017-01-01

Eukaryotic MutLα (mammalian MLH1–PMS2 heterodimer; MLH1–PMS1 in yeast) functions in early steps of mismatch repair as a latent endonuclease that requires a mismatch, MutSα/β, and DNA-loaded proliferating cell nuclear antigen (PCNA) for activation. We show here that human PCNA and MutLα interact specifically but weakly in solution to form a complex of approximately 1:1 stoichiometry that depends on PCNA interaction with the C-terminal endonuclease domain of the MutLα PMS2 subunit. Amino acid substitution mutations within a PMS2 C-terminal 721QRLIAP motif attenuate or abolish human MutLα interaction with PCNA, as well as PCNA-dependent activation of MutLα endonuclease, PCNA- and DNA-dependent activation of MutLα ATPase, and MutLα function in in vitro mismatch repair. Amino acid substitution mutations within the corresponding yeast PMS1 motif (723QKLIIP) reduce or abolish mismatch repair in vivo. Coupling of a weak allele within this motif (723AKLIIP) with an exo1Δ null mutation, which individually confer only weak mutator phenotypes, inactivates mismatch repair in the yeast cell. PMID:28439008

Interaction of proliferating cell nuclear antigen with PMS2 is required for MutLα activation and function in mismatch repair.

PubMed

Genschel, Jochen; Kadyrova, Lyudmila Y; Iyer, Ravi R; Dahal, Basanta K; Kadyrov, Farid A; Modrich, Paul

2017-05-09

Eukaryotic MutLα (mammalian MLH1-PMS2 heterodimer; MLH1-PMS1 in yeast) functions in early steps of mismatch repair as a latent endonuclease that requires a mismatch, MutSα/β, and DNA-loaded proliferating cell nuclear antigen (PCNA) for activation. We show here that human PCNA and MutLα interact specifically but weakly in solution to form a complex of approximately 1:1 stoichiometry that depends on PCNA interaction with the C-terminal endonuclease domain of the MutLα PMS2 subunit. Amino acid substitution mutations within a PMS2 C-terminal 721 QRLIAP motif attenuate or abolish human MutLα interaction with PCNA, as well as PCNA-dependent activation of MutLα endonuclease, PCNA- and DNA-dependent activation of MutLα ATPase, and MutLα function in in vitro mismatch repair. Amino acid substitution mutations within the corresponding yeast PMS1 motif ( 723 QKLIIP) reduce or abolish mismatch repair in vivo. Coupling of a weak allele within this motif ( 723 AKLIIP) with an exo1 Δ null mutation, which individually confer only weak mutator phenotypes, inactivates mismatch repair in the yeast cell.

Spaceborne computer executive routine functional design specification. Volume 2: Computer executive design for space station/base

NASA Technical Reports Server (NTRS)

Kennedy, J. R.; Fitzpatrick, W. S.

1971-01-01

The computer executive functional system design concepts derived from study of the Space Station/Base are presented. Information Management System hardware configuration as directly influencing the executive design is reviewed. The hardware configuration and generic executive design requirements are considered in detail in a previous report (System Configuration and Executive Requirements Specifications for Reusable Shuttle and Space Station/Base, 9/25/70). This report defines basic system primitives and delineates processes and process control. Supervisor states are considered for describing basic multiprogramming and multiprocessing systems. A high-level computer executive including control of scheduling, allocation of resources, system interactions, and real-time supervisory functions is defined. The description is oriented to provide a baseline for a functional simulation of the computer executive system.

Tooth wear: attrition, erosion, and abrasion.

PubMed

Litonjua, Luis A; Andreana, Sebastiano; Bush, Peter J; Cohen, Robert E

2003-06-01

Attrition, erosion, and abrasion result in alterations to the tooth and manifest as tooth wear. Each classification acts through a distinct process that is associated with unique clinical characteristics. Accurate prevalence data for each classification are not available since indices do not necessarily measure one specific etiology, or the study populations may be too diverse in age and characteristics. The treatment of teeth in each classification will depend on identifying the factors associated with each etiology. Some cases may require specific restorative procedures, while others will not require treatment. A review of the literature points to the interaction of the three entities in the initiation and progression of lesions that may act synchronously or sequentially, synergistically or additively, or in conjunction with other entities to mask the true nature of tooth wear, which appears to be multifactorial.

Signaling by Antibodies: Recent Progress

PubMed Central

Bournazos, Stylianos; Wang, Taia T.; Dahan, Rony; Maamary, Jad; Ravetch, Jeffrey V.

2017-01-01

IgG antibodies mediate a diversity of immune functions by coupling of antigen specificity through the Fab domain to signal transduction via Fc-Fc receptor interactions. Indeed, balanced IgG signaling through Type I and Type II Fc receptors is required for the control of pro-inflammatory, anti-inflammatory, and immunomodulatory processes. In this review, we discuss the mechanisms that govern IgG-Fc receptor interactions, highlighting the diversity of Fc receptor-mediated effector functions that regulate immunity and inflammation, as well as determine susceptibility to infection and autoimmunity, and responsiveness to antibody-based therapeutics, and vaccine responses. PMID:28446061

A framework supporting the development of a Grid portal for analysis based on ROI.

PubMed

Ichikawa, K; Date, S; Kaishima, T; Shimojo, S

2005-01-01

In our research on brain function analysis, users require two different simultaneous types of processing: interactive processing to a specific part of data and high-performance batch processing to an entire dataset. The difference between these two types of processing is in whether or not the analysis is for data in the region of interest (ROI). In this study, we propose a Grid portal that has a mechanism to freely assign computing resources to the users on a Grid environment according to the users' two different types of processing requirements. We constructed a Grid portal which integrates interactive processing and batch processing by the following two mechanisms. First, a job steering mechanism controls job execution based on user-tagged priority among organizations with heterogeneous computing resources. Interactive jobs are processed in preference to batch jobs by this mechanism. Second, a priority-based result delivery mechanism that administrates a rank of data significance. The portal ensures a turn-around time of interactive processing by the priority-based job controlling mechanism, and provides the users with quality of services (QoS) for interactive processing. The users can access the analysis results of interactive jobs in preference to the analysis results of batch jobs. The Grid portal has also achieved high-performance computation of MEG analysis with batch processing on the Grid environment. The priority-based job controlling mechanism has been realized to freely assign computing resources to the users' requirements. Furthermore the achievement of high-performance computation contributes greatly to the overall progress of brain science. The portal has thus made it possible for the users to flexibly include the large computational power in what they want to analyze.

Robust detection of chromosomal interactions from small numbers of cells using low-input Capture-C

PubMed Central

Oudelaar, A. Marieke; Davies, James O.J.; Downes, Damien J.; Higgs, Douglas R.

2017-01-01

Abstract Chromosome conformation capture (3C) techniques are crucial to understanding tissue-specific regulation of gene expression, but current methods generally require large numbers of cells. This hampers the investigation of chromatin architecture in rare cell populations. We present a new low-input Capture-C approach that can generate high-quality 3C interaction profiles from 10 000–20 000 cells, depending on the resolution used for analysis. We also present a PCR-free, sequencing-free 3C technique based on NanoString technology called C-String. By comparing C-String and Capture-C interaction profiles we show that the latter are not skewed by PCR amplification. Furthermore, we demonstrate that chromatin interactions detected by Capture-C do not depend on the degree of cross-linking by performing experiments with varying formaldehyde concentrations. PMID:29186505

Adaptor proteins GIR1 and GIR2. II. Interaction with the co-repressor TOPLESS and promotion of histone deacetylation of target chromatin.

PubMed

Wu, Renhong; Citovsky, Vitaly

2017-07-08

Understanding how root hair development is controlled is important for understanding of many fundamental aspects of plant biology. Previously, we identified two plant-specific adaptor proteins GIR1 and GIR2 that interact with the major regulator of root hair development GL2 and suppress formation of root hair. Here, we show that GIR1 and GIR2 also interact with the co-repressor TOPLESS (TPL). This interaction required the GIR1 protein EAR motif, and was essential for the transcriptional repressor activity of GIR1. Both GIR1 and GIR2 promoted histone hypoacetylation of their target chromatin. Potentially, GIR1 and GIR2 might may link TPL to and participate in epigenetic regulation of root hair development. Copyright © 2017 Elsevier Inc. All rights reserved.

Intelligent Motion and Interaction Within Virtual Environments

NASA Technical Reports Server (NTRS)

Ellis, Stephen R. (Editor); Slater, Mel (Editor); Alexander, Thomas (Editor)

2007-01-01

What makes virtual actors and objects in virtual environments seem real? How can the illusion of their reality be supported? What sorts of training or user-interface applications benefit from realistic user-environment interactions? These are some of the central questions that designers of virtual environments face. To be sure simulation realism is not necessarily the major, or even a required goal, of a virtual environment intended to communicate specific information. But for some applications in entertainment, marketing, or aspects of vehicle simulation training, realism is essential. The following chapters will examine how a sense of truly interacting with dynamic, intelligent agents may arise in users of virtual environments. These chapters are based on presentations at the London conference on Intelligent Motion and Interaction within a Virtual Environments which was held at University College, London, U.K., 15-17 September 2003.

Targeting Allosteric Control Mechanisms in Heat Shock Protein 70 (Hsp70).

PubMed

Li, Xiaokai; Shao, Hao; Taylor, Isabelle R; Gestwicki, Jason E

2016-01-01

Heat shock protein 70 (Hsp70) is a molecular chaperone that plays critical roles in protein homeostasis. Hsp70's chaperone activity is coordinated by intra-molecular interactions between its two domains, as well as inter-molecular interactions between Hsp70 and its co-chaperones. Each of these contacts represents a potential opportunity for the development of chemical inhibitors. To illustrate this concept, we review three classes of recently identified molecules that bind distinct pockets on Hsp70. Although all three compounds share the ability to interrupt core biochemical functions of Hsp70, they stabilize different conformers. Accordingly, each compound appears to interrupt a specific subset of inter- and intra-molecular interactions. Thus, an accurate definition of an Hsp70 inhibitor may require a particularly detailed understanding of the molecule's binding site and its effects on protein-protein interactions.

Simulation modelling for new gas turbine fuel controller creation.

NASA Astrophysics Data System (ADS)

Vendland, L. E.; Pribylov, V. G.; Borisov, Yu A.; Arzamastsev, M. A.; Kosoy, A. A.

2017-11-01

State of the art gas turbine fuel flow control systems are based on throttle principle. Major disadvantage of such systems is that they require high pressure fuel intake. Different approach to fuel flow control is to use regulating compressor. And for this approach because of controller and gas turbine interaction a specific regulating compressor is required. Difficulties emerge as early as the requirement definition stage. To define requirements for new object, his properties must be known. Simulation modelling helps to overcome these difficulties. At the requirement definition stage the most simplified mathematical model is used. Mathematical models will get more complex and detailed as we advance in planned work. If future adjusting of regulating compressor physical model to work with virtual gas turbine and physical control system is planned.

Pipe Flow Simulation Software: A Team Approach to Solve an Engineering Education Problem.

ERIC Educational Resources Information Center

Engel, Renata S.; And Others

1996-01-01

A computer simulation program for use in the study of fluid mechanics is described. The package is an interactive tool to explore the fluid flow characteristics of a pipe system by manipulating the physical construction of the system. The motivation, software design requirements, and specific details on how its objectives were met are presented.…

A Transcription Factor-Binding Domain of the Coactivator CBP Is Essential for Long-Term Memory and the Expression of Specific Target Genes

ERIC Educational Resources Information Center

Oliveira, Ana M. M.; Brindle, Paul K.; Abel, Ted; Wood, Marcelo A.; Attner, Michelle A.

2006-01-01

Transcriptional activation is a key process required for long-term memory formation. Recently, the transcriptional coactivator CREB-binding protein (CBP) was shown to be critical for hippocampus-dependent long-term memory and hippocampal synaptic plasticity. As a coactivator with intrinsic histone acetyltransferase activity, CBP interacts with…

What's so Interesting Outside? A Study of Child-Initiated Interaction with Teachers in the Natural Outdoor Environment

ERIC Educational Resources Information Center

Waters, Jane; Maynard, Trisha

2010-01-01

The research question addressed in this paper is: What specific elements in the outdoor environment draw children's attention? The new approach to Welsh early years education--the "Foundation Phase: Framework for children aged 3-7 years in Wales" (DCELLS 2008)--requires the inclusion of both teacher-led and child-initiated activities and…

Improving School Climate Through School/Community Interaction: Needs Assessment Through Community Participation. Teacher Corps Developmental Training Activities.

ERIC Educational Resources Information Center

Barros, Ricardo

The initial year (1978) of the five year program, the Chama Valley Independent School District (CVISD)/University of New Mexico (UNM) Teacher Corps Project, involved an intensive collaborative effort to define and delineate specific project requirements and objectives of the remaining 4 years of the project's duration. The major tool for this…

An Analysis of Fixed Wing Tactical Airlifter Characteristics Using an Intra-Theater Airlift Computer Model

DTIC Science & Technology

1991-09-01

an Experimental Design ...... 31 Selection of Variables .................... ... 34 Defining Measures of Effectiveness ....... 37 Specification of...Required Number of Replications 44 Modification of Scenario Files ......... ... 46 Analysis of the Main Effects of a Two Level Factorial Design ...48 Analysis of the Interaction Effects of a *Two Level Factorial Design .. ............. ... 49 Yate’s Algorithm ......... ................ 50

Prioritizing parts from cutting bills when gang-ripping first

Treesearch

R. Edward Thomas

1996-01-01

Computer optimization of gang-rip-first processing is a difficult problem when working with specific cutting bills. Interactions among board grade and size, arbor setup, and part sizes and quantities greatly complicate the decision making process. Cutting the wrong parts at any moment will mean that more board footage will be required to meet the bill. Using the ROugh...

Voluntary Running Depreciates the Requirement of Ca[superscript 2+]-Stimulated cAMP Signaling in Synaptic Potentiation and Memory Formation

ERIC Educational Resources Information Center

Zheng, Fei; Zhang, Ming; Ding, Qi; Sethna, Ferzin; Yan, Lily; Moon, Changjong; Yang, Miyoung; Wang, Hongbing

2016-01-01

Mental health and cognitive functions are influenced by both genetic and environmental factors. Although having active lifestyle with physical exercise improves learning and memory, how it interacts with the specific key molecular regulators of synaptic plasticity is largely unknown. Here, we examined the effects of voluntary running on long-term…

The Tn7 transposition regulator TnsC interacts with the transposase subunit TnsB and target selector TnsD

PubMed Central

Choi, Ki Young; Spencer, Jeanelle M.; Craig, Nancy L.

2014-01-01

The excision of transposon Tn7 from a donor site and its insertion into its preferred target site, attachment site attTn7, is mediated by four Tn7-encoded transposition proteins: TnsA, TnsB, TnsC, and TnsD. Transposition requires the assembly of a nucleoprotein complex containing all four Tns proteins and the DNA substrates, the donor site containing Tn7, and the preferred target site attTn7. TnsA and TnsB together form the heteromeric Tn7 transposase, and TnsD is a target-selecting protein that binds specifically to attTn7. TnsC is the key regulator of transposition, interacting with both the TnsAB transposase and TnsD-attTn7. We show here that TnsC interacts directly with TnsB, and identify the specific region of TnsC involved in the TnsB–TnsC interaction during transposition. We also show that a TnsC mutant defective in interaction with TnsB is defective for Tn7 transposition both in vitro and in vivo. Tn7 displays cis-acting target immunity, which blocks Tn7 insertion into a target DNA that already contains Tn7. We provide evidence that the direct TnsB–TnsC interaction that we have identified also mediates cis-acting Tn7 target immunity. We also show that TnsC interacts directly with the target selector protein TnsD. PMID:24982178

A New Wnt1-CRE TomatoRosa Embryonic Stem Cell Line: A Tool for Studying Neural Crest Cell Integration Capacity.

PubMed

Acuna-Mendoza, Soledad; Martin, Sabrina; Kuchler-Bopp, Sabine; Ribes, Sandy; Thalgott, Jérémy; Chaussain, Catherine; Creuzet, Sophie; Lesot, Hervé; Lebrin, Franck; Poliard, Anne

2017-12-01

Neural crest (NC) cells are a migratory, multipotent population giving rise to numerous lineages in the embryo. Their plasticity renders attractive their use in tissue engineering-based therapies, but further knowledge on their in vivo behavior is required before clinical transfer may be envisioned. We here describe the isolation and characterization of a new mouse embryonic stem (ES) line derived from Wnt1-CRE-R26 Rosa TomatoTdv blastocyst and show that it displays the characteristics of typical ES cells. Further, these cells can be efficiently directed toward an NC stem cell-like phenotype as attested by concomitant expression of NC marker genes and Tomato fluorescence. As native NC progenitors, they are capable of differentiating toward typical derivative phenotypes and interacting with embryonic tissues to participate in the formation of neo-structures. Their specific fluorescence allows purification and tracking in vivo. This cellular tool should facilitate a better understanding of the mechanisms driving NC fate specification and help identify the key interactions developed within a tissue after in vivo implantation. Altogether, this novel model may provide important knowledge to optimize NC stem cell graft conditions, which are required for efficient tissue repair.

Morphogenic Protein RodZ Interacts with Sporulation Specific SpoIIE in Bacillus subtilis.

PubMed

Muchová, Katarína; Chromiková, Zuzana; Bradshaw, Niels; Wilkinson, Anthony J; Barák, Imrich

2016-01-01

The first landmark in sporulation of Bacillus subtilis is the formation of an asymmetric septum followed by selective activation of the transcription factor σF in the resulting smaller cell. How the morphological transformations that occur during sporulation are coupled to cell-specific activation of transcription is largely unknown. The membrane protein SpoIIE is a constituent of the asymmetric sporulation septum and is a crucial determinant of σF activation. Here we report that the morphogenic protein, RodZ, which is essential for cell shape determination, is additionally required for asymmetric septum formation and sporulation. In cells depleted of RodZ, formation of asymmetric septa is disturbed and σF activation is perturbed. During sporulation, we found that SpoIIE recruits RodZ to the asymmetric septum. Moreover, we detected a direct interaction between SpoIIE and RodZ in vitro and in vivo, indicating that SpoIIE-RodZ may form a complex to coordinate asymmetric septum formation and σF activation. We propose that RodZ could provide a link between the cell shape machinery and the coordinated morphological and developmental transitions required to form a resistant spore.

Application of surface plasmon resonance for the detection of carbohydrates, glycoconjugates, and measurement of the carbohydrate-specific interactions: a comparison with conventional analytical techniques. A critical review.

PubMed

Safina, Gulnara

2012-01-27

Carbohydrates (glycans) and their conjugates with proteins and lipids contribute significantly to many biological processes. That makes these compounds important targets to be detected, monitored and identified. The identification of the carbohydrate content in their conjugates with proteins and lipids (glycoforms) is often a challenging task. Most of the conventional instrumental analytical techniques are time-consuming and require tedious sample pretreatment and utilising various labeling agents. Surface plasmon resonance (SPR) has been intensively developed during last two decades and has received the increasing attention for different applications, from the real-time monitoring of affinity bindings to biosensors. SPR does not require any labels and is capable of direct measurement of biospecific interaction occurring on the sensing surface. This review provides a critical comparison of modern analytical instrumental techniques with SPR in terms of their analytical capabilities to detect carbohydrates, their conjugates with proteins and lipids and to study the carbohydrate-specific bindings. A few selected examples of the SPR approaches developed during 2004-2011 for the biosensing of glycoforms and for glycan-protein affinity studies are comprehensively discussed. Copyright © 2011 Elsevier B.V. All rights reserved.

Display rules versus display autonomy: emotion regulation, emotional exhaustion, and task performance in a call center simulation.

PubMed

Goldberg, Lori Sideman; Grandey, Alicia A

2007-07-01

"Service with a smile" is satisfying for the customer, but such display rules may be costly to the employee and the organization. Most previous research on such costs has used self-reported and cross-sectional designs. The authors use an experimental approach to test tenets of resource depletion theories; specifically, whether the self-regulation of emotions required by display rules depletes energy and attentional resources during a service encounter. Using a call center simulation with three "customer" interactions, the authors found that participants given positive display rules (e.g., be enthusiastic and hide frustration) reported more postsimulation exhaustion and made more errors on the order form compared to those with display autonomy. Customer hostility during one of the calls also increased exhaustion overall and the number of errors during that specific call, though proposed interactions with display rules were not supported. Surface-level emotion regulation, but not deep-level, was the mechanism for the energy depletion effect of display rules, while display rules had a direct effect on performance decrements. Theoretical and practical implications for display rules as part of job requirements are discussed. Copyright 2007 APA

Drosophila Vps16A is required for trafficking to lysosomes and biogenesis of pigment granules.

PubMed

Pulipparacharuvil, Suprabha; Akbar, Mohammed Ali; Ray, Sanchali; Sevrioukov, Evgueny A; Haberman, Adam S; Rohrer, Jack; Krämer, Helmut

2005-08-15

Mutations that disrupt trafficking to lysosomes and lysosome-related organelles cause multiple diseases, including Hermansky-Pudlak syndrome. The Drosophila eye is a model system for analyzing such mutations. The eye-color genes carnation and deep orange encode two subunits of the Vps-C protein complex required for endosomal trafficking and pigment-granule biogenesis. Here we demonstrate that dVps16A (CG8454) encodes another Vps-C subunit. Biochemical experiments revealed a specific interaction between the dVps16A C-terminus and the Sec1/Munc18 homolog Carnation but not its closest homolog, dVps33B. Instead, dVps33B interacted with a related protein, dVps16B (CG18112). Deep orange bound both Vps16 homologs. Like a deep orange null mutation, eye-specific RNAi-induced knockdown of dVps16A inhibited lysosomal delivery of internalized ligands and interfered with biogenesis of pigment granules. Ubiquitous knockdown of dVps16A was lethal. Together, these findings demonstrate that Drosophila Vps16A is essential for lysosomal trafficking. Furthermore, metazoans have two types of Vps-C complexes with non-redundant functions.

The Drosophila Juvenile Hormone Receptor Candidates Methoprene-tolerant (MET) and Germ Cell-expressed (GCE) Utilize a Conserved LIXXL Motif to Bind the FTZ-F1 Nuclear Receptor*

PubMed Central

Bernardo, Travis J.; Dubrovsky, Edward B.

2012-01-01

Juvenile hormone (JH) has been implicated in many developmental processes in holometabolous insects, but its mechanism of signaling remains controversial. We previously found that in Drosophila Schneider 2 cells, the nuclear receptor FTZ-F1 is required for activation of the E75A gene by JH. Here, we utilized insect two-hybrid assays to show that FTZ-F1 interacts with two JH receptor candidates, the bHLH-PAS paralogs MET and GCE, in a JH-dependent manner. These interactions are severely reduced when helix 12 of the FTZ-F1 activation function 2 (AF2) is removed, implicating AF2 as an interacting site. Through homology modeling, we found that MET and GCE possess a C-terminal α-helix featuring a conserved motif LIXXL that represents a novel nuclear receptor (NR) box. Docking simulations supported by two-hybrid experiments revealed that FTZ-F1·MET and FTZ-F1·GCE heterodimer formation involves a typical NR box-AF2 interaction but does not require the canonical charge clamp residues of FTZ-F1 and relies primarily on hydrophobic contacts, including a unique interaction with helix 4. Moreover, we identified paralog-specific features, including a secondary interaction site found only in MET. Our findings suggest that a novel NR box enables MET and GCE to interact JH-dependently with the AF2 of FTZ-F1. PMID:22249180

Evaluation of galectin binding by frontal affinity chromatography (FAC).

PubMed

Iwaki, Jun; Hirabayashi, Jun

2015-01-01

Frontal affinity chromatography (FAC) is a simple and versatile procedure enabling quantitative determination of diverse biological interactions in terms of dissociation constants (K d), even though these interactions are relatively weak. The method is best applied to glycans and their binding proteins, with the analytical system operating on the basis of highly reproducible isocratic elution by liquid chromatography. Its application to galectins has been successfully developed to characterize their binding specificities in detail. As a result, their minimal requirements for recognition of disaccharides, i.e., β-galactosides, as well as characteristic features of individual galectins, have been elucidated. In this chapter, we describe standard procedures to determine the K d's for interactions between a series of standard glycans and various galectins.

The Ca{sup 2+} channel TRPML3 specifically interacts with the mammalian ATG8 homologue GATE16 to regulate autophagy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Choi, Suzy; Kim, Hyun Jin, E-mail: kimhyunjin@skku.edu

2014-01-03

Highlights: •Split-ubiquitin MY2H screen identified GATE16 as an interacting protein of TRPML3. •TRPML3 specifically binds to a mammalian ATG8 homologue GATE16, not to LC3B. •The interaction of TRPML3 with GATE16 facilitates autophagosome formation. •GATE16 is expressed in both autophagosome and extra-autophagosomal compartments. -- Abstract: TRPML3 is a Ca{sup 2+} permeable cation channel expressed in multiple intracellular compartments. Although TRPML3 is implicated in autophagy, how TRPML3 can regulate autophagy is not understood. To search interacting proteins with TRPML3 in autophagy, we performed split-ubiquitin membrane yeast two-hybrid (MY2H) screening with TRPML3-loop as a bait and identified GATE16, a mammalian ATG8 homologue. GSTmore » pull-down assay revealed that TRPML3 and TRPML3-loop specifically bind to GATE16, not to LC3B. Co-immunoprecipitation (co-IP) experiments showed that TRPML3 and TRPML3-loop pull down only the lipidated form of GATE16, indicating that the interaction occurs exclusively at the organellar membrane. The interaction of TRPML3 with GATE16 and GATE16-positive vesicle formation were increased in starvation induced autophagy, suggesting that the interaction facilitates the function of GATE16 in autophagosome formation. However, GATE16 was not required for TRPML3 trafficking to autophagosomes. Experiments using dominant-negative (DN) TRPML3(D458K) showed that GATE16 is localized not only in autophagosomes but also in extra-autophagosomal compartments, by contrast with LC3B. Since GATE16 acts at a later stage of the autophagosome biogenesis, our results suggest that TRPML3 plays a role in autophagosome maturation through the interaction with GATE16, by providing Ca{sup 2+} in the fusion process.« less

Simulated tri-trophic networks reveal complex relationships between species diversity and interaction diversity

PubMed Central

Lumpkin, Will; Hurtado, Paul J.; Dyer, Lee A.

2018-01-01

Most of earth’s biodiversity is comprised of interactions among species, yet it is unclear what causes variation in interaction diversity across space and time. We define interaction diversity as the richness and relative abundance of interactions linking species together at scales from localized, measurable webs to entire ecosystems. Large-scale patterns suggest that two basic components of interaction diversity differ substantially and predictably between different ecosystems: overall taxonomic diversity and host specificity of consumers. Understanding how these factors influence interaction diversity, and quantifying the causes and effects of variation in interaction diversity are important goals for community ecology. While previous studies have examined the effects of sampling bias and consumer specialization on determining patterns of ecological networks, these studies were restricted to two trophic levels and did not incorporate realistic variation in species diversity and consumer diet breadth. Here, we developed a food web model to generate tri-trophic ecological networks, and evaluated specific hypotheses about how the diversity of trophic interactions and species diversity are related under different scenarios of species richness, taxonomic abundance, and consumer diet breadth. We investigated the accumulation of species and interactions and found that interactions accumulate more quickly; thus, the accumulation of novel interactions may require less sampling effort than sampling species in order to get reliable estimates of either type of diversity. Mean consumer diet breadth influenced the correlation between species and interaction diversity significantly more than variation in both species richness and taxonomic abundance. However, this effect of diet breadth on interaction diversity is conditional on the number of observed interactions included in the models. The results presented here will help develop realistic predictions of the relationships between consumer diet breadth, interaction diversity, and species diversity within multi-trophic communities, which is critical for the conservation of biodiversity in this period of accelerated global change. PMID:29579077

Simulated tri-trophic networks reveal complex relationships between species diversity and interaction diversity.

PubMed

Pardikes, Nicholas A; Lumpkin, Will; Hurtado, Paul J; Dyer, Lee A

2018-01-01

Most of earth's biodiversity is comprised of interactions among species, yet it is unclear what causes variation in interaction diversity across space and time. We define interaction diversity as the richness and relative abundance of interactions linking species together at scales from localized, measurable webs to entire ecosystems. Large-scale patterns suggest that two basic components of interaction diversity differ substantially and predictably between different ecosystems: overall taxonomic diversity and host specificity of consumers. Understanding how these factors influence interaction diversity, and quantifying the causes and effects of variation in interaction diversity are important goals for community ecology. While previous studies have examined the effects of sampling bias and consumer specialization on determining patterns of ecological networks, these studies were restricted to two trophic levels and did not incorporate realistic variation in species diversity and consumer diet breadth. Here, we developed a food web model to generate tri-trophic ecological networks, and evaluated specific hypotheses about how the diversity of trophic interactions and species diversity are related under different scenarios of species richness, taxonomic abundance, and consumer diet breadth. We investigated the accumulation of species and interactions and found that interactions accumulate more quickly; thus, the accumulation of novel interactions may require less sampling effort than sampling species in order to get reliable estimates of either type of diversity. Mean consumer diet breadth influenced the correlation between species and interaction diversity significantly more than variation in both species richness and taxonomic abundance. However, this effect of diet breadth on interaction diversity is conditional on the number of observed interactions included in the models. The results presented here will help develop realistic predictions of the relationships between consumer diet breadth, interaction diversity, and species diversity within multi-trophic communities, which is critical for the conservation of biodiversity in this period of accelerated global change.

Handling Qualities Specifications for U.S. Military Helicopters

NASA Technical Reports Server (NTRS)

Key, David L.

1982-01-01

Inadequacies in the military specification for helicopter handling qualities, MIL-H-8501A, have long been recognized, and the latest procurements by the U.S. Army used special Prime Item Development Specifications (PIDS). This paper assesses the efficacy of these PIDS and suggests that changes should be made. In particular, the structure developed in MIL-F-8785B (ASG) (the specification for flying qualities of piloted airplanes) should be incorporated. Improved requirements must be based on a systematic data base and concentrated on topics most important in preliminary design: static and dynamic stability, control power and sensitivity, and interaction with controllers and displays. Emphasis should be on current military helicopter missions and helicopter idiosyncrasies such as cross-coupling, nonlinearities, and higher-order dynamics.

Solid-phase synthesis and screening of N-acylated polyamine (NAPA) combinatorial libraries for protein binding.

PubMed

Iera, Jaclyn A; Jenkins, Lisa M Miller; Kajiyama, Hiroshi; Kopp, Jeffrey B; Appella, Daniel H

2010-11-15

Inhibitors for protein-protein interactions are challenging to design, in part due to the unique and complex architectures of each protein's interaction domain. Most approaches to develop inhibitors for these interactions rely on rational design, which requires prior structural knowledge of the target and its ligands. In the absence of structural information, a combinatorial approach may be the best alternative to finding inhibitors of a protein-protein interaction. Current chemical libraries, however, consist mostly of molecules designed to inhibit enzymes. In this manuscript, we report the synthesis and screening of a library based on an N-acylated polyamine (NAPA) scaffold that we designed to have specific molecular features necessary to inhibit protein-protein interactions. Screens of the library identified a member with favorable binding properties to the HIV viral protein R (Vpr), a regulatory protein from HIV, that is involved in numerous interactions with other proteins critical for viral replication. Published by Elsevier Ltd.

Heparin/heparan sulfate 6-O-sulfatase from Flavobacterium heparinum: integrated structural and biochemical investigation of enzyme active site and substrate specificity.

PubMed

Myette, James R; Soundararajan, Venkataramanan; Shriver, Zachary; Raman, Rahul; Sasisekharan, Ram

2009-12-11

Heparin and heparan sulfate glycosaminoglycans (HSGAGs) comprise a chemically heterogeneous class of sulfated polysaccharides. The development of structure-activity relationships for this class of polysaccharides requires the identification and characterization of degrading enzymes with defined substrate specificity and enzymatic activity. Toward this end, we report here the molecular cloning and extensive structure-function analysis of a 6-O-sulfatase from the Gram-negative bacterium Flavobacterium heparinum. In addition, we report the recombinant expression of this enzyme in Escherichia coli in a soluble, active form and identify it as a specific HSGAG sulfatase. We further define the mechanism of action of the enzyme through biochemical and structural studies. Through the use of defined substrates, we investigate the kinetic properties of the enzyme. This analysis was complemented by homology-based molecular modeling studies that sought to rationalize the substrate specificity of the enzyme and mode of action through an analysis of the active-site topology of the enzyme including identifying key enzyme-substrate interactions and assigning key amino acids within the active site of the enzyme. Taken together, our structural and biochemical studies indicate that 6-O-sulfatase is a predominantly exolytic enzyme that specifically acts on N-sulfated or N-acetylated 6-O-sulfated glucosamines present at the non-reducing end of HSGAG oligosaccharide substrates. This requirement for the N-acetyl or N-sulfo groups on the glucosamine substrate can be explained through eliciting favorable interactions with key residues within the active site of the enzyme. These findings provide a framework that enables the use of 6-O-sulfatase as a tool for HSGAG structure-activity studies as well as expand our biochemical and structural understanding of this important class of enzymes.

Transcriptional control of the tissue-specific, developmentally regulated osteocalcin gene requires a binding motif for the Msx family of homeodomain proteins.

PubMed

Hoffmann, H M; Catron, K M; van Wijnen, A J; McCabe, L R; Lian, J B; Stein, G S; Stein, J L

1994-12-20

The OC box of the rat osteocalcin promoter (nt -99 to -76) is the principal proximal regulatory element contributing to both tissue-specific and developmental control of osteocalcin gene expression. The central motif of the OC box includes a perfect consensus DNA binding site for certain homeodomain proteins. Homeodomain proteins are transcription factors that direct proper development by regulating specific temporal and spatial patterns of gene expression. We therefore addressed the role of the homeodomain binding motif in the activity of the OC promoter. In this study, by the combined application of mutagenesis and site-specific protein recognition analysis, we examined interactions of ROS 17/2.8 osteosarcoma cell nuclear proteins and purified Msx-1 homeodomain protein with the OC box. We detected a series of related specific protein-DNA interactions, a subset of which were inhibited by antibodies directed against the Msx-1 homeodomain but which also recognize the Msx-2 homeodomain. Our results show that the sequence requirements for binding the Msx-1 or Msx-2 homeodomain closely parallel those necessary for osteocalcin gene promoter activity in vivo. This functional relationship was demonstrated by transient expression in ROS 17/2.8 osteosarcoma cells of a series of osteocalcin promoter (nt -1097 to +24)-reporter gene constructs containing mutations within and flanking the homeodomain binding site of the OC box. Northern blot analysis of several bone-related cell types showed that all of the cells expressed msx-1, whereas msx-2 expression was restricted to cells transcribing osteocalcin. Taken together, our results suggest a role for Msx-1 and -2 or related homeodomain proteins in transcription of the osteocalcin gene.

Use of Phage Display to Identify Novel Mineralocorticoid Receptor-Interacting Proteins

PubMed Central

Yang, Jun; Fuller, Peter J.; Morgan, James; Shibata, Hirotaka; McDonnell, Donald P.; Clyne, Colin D.

2014-01-01

The mineralocorticoid receptor (MR) plays a central role in salt and water homeostasis via the kidney; however, inappropriate activation of the MR in the heart can lead to heart failure. A selective MR modulator that antagonizes MR signaling in the heart but not the kidney would provide the cardiovascular protection of current MR antagonists but allow for normal electrolyte balance. The development of such a pharmaceutical requires an understanding of coregulators and their tissue-selective interactions with the MR, which is currently limited by the small repertoire of MR coregulators described in the literature. To identify potential novel MR coregulators, we used T7 phage display to screen tissue-selective cDNA libraries for MR-interacting proteins. Thirty MR binding peptides were identified, from which three were chosen for further characterization based on their nuclear localization and their interaction with other MR-interacting proteins or, in the case of x-ray repair cross-complementing protein 6, its known status as an androgen receptor coregulator. Eukaryotic elongation factor 1A1, structure-specific recognition protein 1, and x-ray repair cross-complementing protein 6 modulated MR-mediated transcription in a ligand-, cell- and/or promoter-specific manner and colocalized with the MR upon agonist treatment when imaged using immunofluorescence microscopy. These results highlight the utility of phage display for rapid and sensitive screening of MR binding proteins and suggest that eukaryotic elongation factor 1A1, structure-specific recognition protein 1, and x-ray repair cross-complementing protein 6 may be potential MR coactivators whose activity is dependent on the ligand, cellular context, and target gene promoter. PMID:25000480

Visualization of protein interactions in living Drosophila embryos by the bimolecular fluorescence complementation assay.

PubMed

Hudry, Bruno; Viala, Séverine; Graba, Yacine; Merabet, Samir

2011-01-28

Protein interactions control the regulatory networks underlying developmental processes. The understanding of developmental complexity will, therefore, require the characterization of protein interactions within their proper environment. The bimolecular fluorescence complementation (BiFC) technology offers this possibility as it enables the direct visualization of protein interactions in living cells. However, its potential has rarely been applied in embryos of animal model organisms and was only performed under transient protein expression levels. Using a Hox protein partnership as a test case, we investigated the suitability of BiFC for the study of protein interactions in the living Drosophila embryo. Importantly, all BiFC parameters were established with constructs that were stably expressed under the control of endogenous promoters. Under these physiological conditions, we showed that BiFC is specific and sensitive enough to analyse dynamic protein interactions. We next used BiFC in a candidate interaction screen, which led to the identification of several Hox protein partners. Our results establish the general suitability of BiFC for revealing and studying protein interactions in their physiological context during the rapid course of Drosophila embryonic development.

Species-Specific Elements in the Large T-Antigen J Domain Are Required for Cellular Transformation and DNA Replication by Simian Virus 40

PubMed Central

Sullivan, Christopher S.; Tremblay, James D.; Fewell, Sheara W.; Lewis, John A.; Brodsky, Jeffrey L.; Pipas, James M.

2000-01-01

The J domain of simian virus 40 (SV40) large T antigen is required for efficient DNA replication and transformation. Despite previous reports demonstrating the promiscuity of J domains in heterologous systems, results presented here show the requirement for specific J-domain sequences in SV40 large-T-antigen-mediated activities. In particular, chimeric-T-antigen constructs in which the SV40 T-antigen J domain was replaced with that from the yeast Ydj1p or Escherichia coli DnaJ proteins failed to replicate in BSC40 cells and did not transform REF52 cells. However, T antigen containing the JC virus J domain was functional in these assays, although it was less efficient than the wild type. The inability of some large-T-antigen chimeras to promote DNA replication and elicit cellular transformation was not due to a failure to interact with hsc70, since a nonfunctional chimera, containing the DnaJ J domain, bound hsc70. However, this nonfunctional chimeric T antigen was reduced in its ability to stimulate hsc70 ATPase activity and unable to liberate E2F from p130, indicating that transcriptional activation of factors required for cell growth and DNA replication may be compromised. Our data suggest that the T-antigen J domain harbors species-specific elements required for viral activities in vivo. PMID:10891510

Identification of amino acids in the transmembrane and juxtamembrane domains of the platelet-derived growth factor receptor required for productive interaction with the bovine papillomavirus E5 protein.

PubMed

Petti, L M; Reddy, V; Smith, S O; DiMaio, D

1997-10-01

The bovine papillomavirus E5 protein forms a stable complex with the cellular platelet-derived growth factor (PDGF) beta receptor, resulting in receptor activation and cell transformation. Amino acids in both the putative transmembrane domain and extracytoplasmic carboxyl-terminal domain of the E5 protein appear important for PDGF receptor binding and activation. Previous analysis indicated that the transmembrane domain of the receptor was also required for complex formation and receptor activation. Here we analyzed receptor chimeras and point mutants to identify specific amino acids in the PDGF beta receptor required for productive interaction with the E5 protein. These receptor mutants were analyzed in murine Ba/F3 cells, which do not express endogenous receptor. Our results confirmed the importance of the transmembrane domain of the receptor for complex formation, receptor tyrosine phosphorylation, and mitogenic signaling in response to the E5 protein and established that the threonine residue in this domain is required for these activities. In addition, a positive charge in the extracellular juxtamembrane domain of the receptor was required for E5 interaction and signaling, whereas replacement of the wild-type lysine with either a neutral or acidic amino acid inhibited E5-induced receptor activation and transformation. All of the receptor mutants defective for activation by the E5 protein responded to acute treatment with PDGF and to stable expression of v-Sis, a form of PDGF. The required juxtamembrane lysine and transmembrane threonine are predicted to align precisely on the same face of an alpha helix packed in a left-handed coiled-coil geometry. These results establish that the E5 protein and v-Sis recognize distinct binding sites on the PDGF beta receptor and further clarify the nature of the interaction between the viral transforming protein and its cellular target.

The Formal Semantics of PVS

NASA Technical Reports Server (NTRS)

Owre, Sam; Shankar, Natarajan

1999-01-01

A specification language is a medium for expressing what is computed rather than how it is computed. Specification languages share some features with programming languages but are also different in several important ways. For our purpose, a specification language is a logic within which the behavior of computational systems can be formalized. Although a specification can be used to simulate the behavior of such systems, we mainly use specifications to state and prove system properties with mechanical assistance. We present the formal semantics of the specification language of SRI's Prototype Verification System (PVS). This specification language is based on the simply typed lambda calculus. The novelty in PVS is that it contains very expressive language features whose static analysis (e.g., typechecking) requires the assistance of a theorem prover. The formal semantics illuminates several of the design considerations underlying PVS, the interaction between theorem proving and typechecking.

Specificity and non-specificity in RNA–protein interactions

PubMed Central

Jankowsky, Eckhard; Harris, Michael E.

2016-01-01

Gene expression is regulated by complex networks of interactions between RNAs and proteins. Proteins that interact with RNA have been traditionally viewed as either specific or non-specific; specific proteins interact preferentially with defined RNA sequence or structure motifs, whereas non-specific proteins interact with RNA sites devoid of such characteristics. Recent studies indicate that the binary “specific vs. non-specific” classification is insufficient to describe the full spectrum of RNA–protein interactions. Here, we review new methods that enable quantitative measurements of protein binding to large numbers of RNA variants, and the concepts aimed as describing resulting binding spectra: affinity distributions, comprehensive binding models and free energy landscapes. We discuss how these new methodologies and associated concepts enable work towards inclusive, quantitative models for specific and non-specific RNA–protein interactions. PMID:26285679

Co-stimulatory function in primary germinal center responses: CD40 and B7 are required on distinct antigen-presenting cells.

PubMed

Watanabe, Masashi; Fujihara, Chiharu; Radtke, Andrea J; Chiang, Y Jeffrey; Bhatia, Sumeena; Germain, Ronald N; Hodes, Richard J

2017-09-04

T cell-dependent germinal center (GC) responses require coordinated interactions of T cells with two antigen-presenting cell (APC) populations, B cells and dendritic cells (DCs), in the presence of B7- and CD40-dependent co-stimulatory pathways. Contrary to the prevailing paradigm, we found unique cellular requirements for B7 and CD40 expression in primary GC responses to vaccine immunization with protein antigen and adjuvant: B7 was required on DCs but was not required on B cells, whereas CD40 was required on B cells but not on DCs in the generation of antigen-specific follicular helper T cells, antigen-specific GC B cells, and high-affinity class-switched antibody production. There was, in fact, no requirement for coexpression of B7 and CD40 on the same cell in these responses. Our findings support a substantially revised model for co-stimulatory function in the primary GC response, with crucial and distinct contributions of B7- and CD40-dependent pathways expressed by different APC populations and with important implications for understanding how to optimize vaccine responses or limit autoimmunity. This is a work of the U.S. Government and is not subject to copyright protection in the United States. Foreign copyrights may apply.

Sequence-dependent catalytic regulation of the SpoIIIE motor activity ensures directionality of DNA translocation.

PubMed

Chara, Osvaldo; Borges, Augusto; Milhiet, Pierre-Emmanuel; Nöllmann, Marcelo; Cattoni, Diego I

2018-03-27

Transport of cellular cargo by molecular motors requires directionality to ensure proper biological functioning. During sporulation in Bacillus subtilis, directionality of chromosome transport is mediated by the interaction between the membrane-bound DNA translocase SpoIIIE and specific octameric sequences (SRS). Whether SRS regulate directionality by recruiting and orienting SpoIIIE or by simply catalyzing its translocation activity is still unclear. By using atomic force microscopy and single-round fast kinetics translocation assays we determined the localization and dynamics of diffusing and translocating SpoIIIE complexes on DNA with or without SRS. Our findings combined with mathematical modelling revealed that SpoIIIE directionality is not regulated by protein recruitment to SRS but rather by a fine-tuned balance among the rates governing SpoIIIE-DNA interactions and the probability of starting translocation modulated by SRS. Additionally, we found that SpoIIIE can start translocation from non-specific DNA, providing an alternative active search mechanism for SRS located beyond the exploratory length defined by 1D diffusion. These findings are relevant in vivo in the context of chromosome transport through an open channel, where SpoIIIE can rapidly explore DNA while directionality is modulated by the probability of translocation initiation upon interaction with SRS versus non-specific DNA.

Cellular projections from sensory hair cells form polarity-specific scaffolds during synaptogenesis

PubMed Central

Dow, Eliot; Siletti, Kimberly

2015-01-01

The assembly of a nervous system requires the extension of axons and dendrites to specific regions where they are matched with appropriate synaptic targets. Although the cues that guide long-range outgrowth have been characterized extensively, additional mechanisms are required to explain short-range guidance in neural development. Using a complementary combination of time-lapse imaging by fluorescence confocal microscopy and serial block-face electron microscopy, we identified a novel type of presynaptic projection that participates in the assembly of the vertebrate nervous system. Synapse formation by each hair cell of the zebrafish's lateral line occurs during a particular interval after the cell's birth. During the same period, projections emerge from the cellular soma, extending toward a specific subpopulation of mature hair cells and interacting with polarity-specific afferent nerve terminals. The terminals then extend along the projections to reach appropriately matched presynaptic sites, after which the projections recede. Our results suggest that presynaptic projections act as transient scaffolds for short-range partner matching, a mechanism that may occur elsewhere in the nervous system. PMID:25995190

Tyr721 regulates specific binding of the CSF-1 receptor kinase insert to PI 3'-kinase SH2 domains: a model for SH2-mediated receptor-target interactions.

PubMed Central

Reedijk, M; Liu, X; van der Geer, P; Letwin, K; Waterfield, M D; Hunter, T; Pawson, T

1992-01-01

Efficient binding of active phosphatidylinositol (PI) 3'-kinase to the autophosphorylated macrophage colony stimulating factor receptor (CSF-1R) requires the noncatalytic kinase insert (KI) region of the receptor. To test whether this region could function independently to bind PI 3'-kinase, the isolated CSF-1R KI was expressed in Escherichia coli, and was inducibly phosphorylated on tyrosine. The tyrosine phosphorylated form of the CSF-1R KI bound PI 3'-kinase in vitro, whereas the unphosphorylated form had no binding activity. The p85 alpha subunit of PI 3'-kinase contains two Src homology (SH)2 domains, which are implicated in the interactions of signalling proteins with activated receptors. Bacterially expressed p85 alpha SH2 domains complexed in vitro with the tyrosine phosphorylated CSF-1R KI. Binding of the CSF-1R KI to PI 3'-kinase activity, and to the p85 alpha SH2 domains, required phosphorylation of Tyr721 within the KI domain, but was independent of phosphorylation at Tyr697 and Tyr706. Tyr721 was also critical for the association of activated CSF-1R with PI 3'-kinase in mammalian cells. Complex formation between the CSF-1R and PI 3'-kinase can therefore be reconstructed in vitro in a specific interaction involving the phosphorylated receptor KI and the SH2 domains of p85 alpha. Images PMID:1314163

Transcriptional regulation of Drosophila gonad formation.

PubMed

Tripathy, Ratna; Kunwar, Prabhat S; Sano, Hiroko; Renault, Andrew D

2014-08-15

The formation of the Drosophila embryonic gonad, involving the fusion of clusters of somatic gonadal precursor cells (SGPs) and their ensheathment of germ cells, provides a simple and genetically tractable model for the interplay between cells during organ formation. In a screen for mutants affecting gonad formation we identified a SGP cell autonomous role for Midline (Mid) and Longitudinals lacking (Lola). These transcriptional factors are required for multiple aspects of SGP behaviour including SGP cluster fusion, germ cell ensheathment and gonad compaction. The lola locus encodes more than 25 differentially spliced isoforms and we have identified an isoform specific requirement for lola in the gonad which is distinct from that in nervous system development. Mid and Lola work in parallel in gonad formation and surprisingly Mid overexpression in a lola background leads to additional SGPs at the expense of fat body cells. Our findings support the idea that although the transcription factors required by SGPs can ostensibly be assigned to those being required for either SGP specification or behaviour, they can also interact to impinge on both processes. Copyright © 2014 Elsevier Inc. All rights reserved.

Segmental expression of Hoxb2 in r4 requires two separate sites that integrate cooperative interactions between Prep1, Pbx and Hox proteins.

PubMed

Ferretti, E; Marshall, H; Pöpperl, H; Maconochie, M; Krumlauf, R; Blasi, F

2000-01-01

Direct auto- and cross-regulatory interactions between Hox genes serve to establish and maintain segmentally restricted patterns in the developing hindbrain. Rhombomere r4-specific expression of both Hoxb1 and Hoxb2 depends upon bipartite cis Hox response elements for the group 1 paralogous proteins, Hoxal and Hoxbl. The DNA-binding ability and selectivity of these proteins depend upon the formation of specific heterodimeric complexes with members of the PBC homeodomain protein family (Pbx genes). The r4 enhancers from Hoxb1 and Hoxb2 have the same activity, but differ with respect to the number and organisation of bipartite Pbx/Hox (PH) sites required, suggesting the intervention of other components/sequences. We report here that another family of homeodomain proteins (TALE, Three-Amino acids-Loop-Extension: Prep1, Meis, HTH), capable of dimerizing with Pbx/EXD, is involved in the mechanisms of r4-restricted expression. We show that: (1) the r4-specific Hoxb1 and Hoxb2 enhancers are complex elements containing separate PH and Prep/Meis (PM) sites; (2) the PM site of the Hoxb2, but not Hoxb1, enhancer is essential in vivo for r4 expression and also influences other sites of expression; (3) both PM and PH sites are required for in vitro binding of Prepl-Pbx and formation and binding of a ternary Hoxbl-Pbxla (or 1b)-Prepl complex. (4) A similar ternary association forms in nuclear extracts from embryonal P19 cells, but only upon retinoic acid induction. This requires synthesis of Hoxbl and also contains Pbx with either Prepl or Meisl. Together these findings highlight the fact that PM sites are found in close proximity to bipartite PH motifs in several Hox responsive elements shown to be important in vivo and that such sites play an essential role in potentiating regulatory activity in combination with the PH motifs.

Three input concepts for flight crew interaction with information presented on a large-screen electronic cockpit display

NASA Technical Reports Server (NTRS)

Jones, Denise R.

1990-01-01

A piloted simulation study was conducted comparing three different input methods for interfacing to a large-screen, multiwindow, whole-flight-deck display for management of transport aircraft systems. The thumball concept utilized a miniature trackball embedded in a conventional side-arm controller. The touch screen concept provided data entry through a capacitive touch screen. The voice concept utilized a speech recognition system with input through a head-worn microphone. No single input concept emerged as the most desirable method of interacting with the display. Subjective results, however, indicate that the voice concept was the most preferred method of data entry and had the most potential for future applications. The objective results indicate that, overall, the touch screen concept was the most effective input method. There was also significant differences between the time required to perform specific tasks and the input concept employed, with each concept providing better performance relative to a specific task. These results suggest that a system combining all three input concepts might provide the most effective method of interaction.

Ligand Recognition by A-Class Eph Receptors: Crystal Structures of the EphA2 Ligand-Binding Domain and the EphA2/ephrin-A1 Complex

DOE Office of Scientific and Technical Information (OSTI.GOV)

Himanen, J.; Goldgur, Y; Miao, H

2009-01-01

Ephrin (Eph) receptor tyrosine kinases fall into two subclasses (A and B) according to preferences for their ephrin ligands. All published structural studies of Eph receptor/ephrin complexes involve B-class receptors. Here, we present the crystal structures of an A-class complex between EphA2 and ephrin-A1 and of unbound EphA2. Although these structures are similar overall to their B-class counterparts, they reveal important differences that define subclass specificity. The structures suggest that the A-class Eph receptor/ephrin interactions involve smaller rearrangements in the interacting partners, better described by a 'lock-and-key'-type binding mechanism, in contrast to the 'induced fit' mechanism defining the B-class molecules.more » This model is supported by structure-based mutagenesis and by differential requirements for ligand oligomerization by the two subclasses in cell-based Eph receptor activation assays. Finally, the structure of the unligated receptor reveals a homodimer assembly that might represent EphA2-specific homotypic cell adhesion interactions.« less

Plant-Aphid Interactions Under Elevated CO2: Some Cues from Aphid Feeding Behavior.

PubMed

Sun, Yucheng; Guo, Huijuan; Ge, Feng

2016-01-01

Although the increasing concentration of atmospheric carbon dioxide (CO2) accelerates the accumulation of carbohydrates and increases the biomass and yield of C3 crop plants, it also reduces their nitrogen concentration. The consequent changes in primary and secondary metabolites affect the palatability of host plants and the feeding of herbivorous insects. Aphids are phloem feeders and are considered the only feeding guild that positively responds to elevated CO2. In this review, we consider how elevated CO2 modifies host defenses, nutrients, and water-use efficiency by altering concentrations of the phytohormones jasmonic acid, salicylic acid, ethylene, and abscisic acid. We will describe how these elevated CO2-induced changes in defenses, nutrients, and water statusfacilitate specific stages of aphid feeding, including penetration, phloem-feeding, and xylem absorption. We conclude that a better understanding of the effects of elevated CO2 on aphids and on aphid damage to crop plants will require research on the molecular aspects of the interaction between plant and aphid but also research on aphid interactions with their intra- and inter-specific competitors and with their natural enemies.

Probing the dynamics of restriction endonuclease NgoMIV-DNA interaction by single-molecule FRET.

PubMed

Tutkus, Marijonas; Sasnauskas, Giedrius; Rutkauskas, Danielis

2017-12-01

Many type II restriction endonucleases require two copies of their recognition sequence for optimal activity. Concomitant binding of two DNA sites by such an enzyme produces a DNA loop. Here we exploit single-molecule Förster resonance energy transfer (smFRET) of surface-immobilized DNA fragments to study the dynamics of DNA looping induced by tetrameric endonuclease NgoMIV. We have employed a DNA fragment with two NgoMIV recognition sites and a FRET dye pair such that upon protein-induced DNA looping the dyes are brought to close proximity resulting in a FRET signal. The dynamics of DNA-NgoMIV interactions proved to be heterogeneous, with individual smFRET trajectories exhibiting broadly different average looped state durations. Distinct types of the dynamics were attributed to different types of DNA-protein complexes, mediated either by one NgoMIV tetramer simultaneously bound to two specific sites ("slow" trajectories) or by semi-specific interactions of two DNA-bound NgoMIV tetramers ("fast" trajectories), as well as to conformational heterogeneity of individual NgoMIV molecules. © 2017 Wiley Periodicals, Inc.

Numerical Simulation of Two Phase Flows

NASA Technical Reports Server (NTRS)

Liou, Meng-Sing

2001-01-01

Two phase flows can be found in broad situations in nature, biology, and industry devices and can involve diverse and complex mechanisms. While the physical models may be specific for certain situations, the mathematical formulation and numerical treatment for solving the governing equations can be general. Hence, we will require information concerning each individual phase as needed in a single phase. but also the interactions between them. These interaction terms, however, pose additional numerical challenges because they are beyond the basis that we use to construct modern numerical schemes, namely the hyperbolicity of equations. Moreover, due to disparate differences in time scales, fluid compressibility and nonlinearity become acute, further complicating the numerical procedures. In this paper, we will show the ideas and procedure how the AUSM-family schemes are extended for solving two phase flows problems. Specifically, both phases are assumed in thermodynamic equilibrium, namely, the time scales involved in phase interactions are extremely short in comparison with those in fluid speeds and pressure fluctuations. Details of the numerical formulation and issues involved are discussed and the effectiveness of the method are demonstrated for several industrial examples.

DNA binding by FOXP3 domain-swapped dimer suggests mechanisms of long-range chromosomal interactions

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chen, Y.; Chen, C.; Zhang, Z.

2015-01-07

FOXP3 is a lineage-specific transcription factor that is required for regulatory T cell development and function. In this study, we determined the crystal structure of the FOXP3 forkhead domain bound to DNA. The structure reveals that FOXP3 can form a stable domain-swapped dimer to bridge DNA in the absence of cofactors, suggesting that FOXP3 may play a role in long-range gene interactions. To test this hypothesis, we used circular chromosome conformation capture coupled with high throughput sequencing (4C-seq) to analyze FOXP3-dependent genomic contacts around a known FOXP3-bound locus, Ptpn22. Our studies reveal that FOXP3 induces significant changes in the chromatinmore » contacts between the Ptpn22 locus and other Foxp3-regulated genes, reflecting a mechanism by which FOXP3 reorganizes the genome architecture to coordinate the expression of its target genes. Our results suggest that FOXP3 mediates long-range chromatin interactions as part of its mechanisms to regulate specific gene expression in regulatory T cells.« less

Hyperbranched polyglycerols at the biointerface

NASA Astrophysics Data System (ADS)

Moore, Eli; Thissen, Helmut; Voelcker, Nicolas H.

2013-08-01

The control over biointerfacial interactions is the key to a broad range of biomedical applications, ranging from implantable devices to drug delivery and nanomedicine. In many of these applications, coatings are required that reduce or prevent non-specific interactions with the biological environment, while at the same time presenting specific bioactive signals. Whilst surface coatings based on polymers such as poly(ethylene glycol) (PEG) have been used successfully, many limitations persist in regard to the biocompatibility, stability and functionality of state-of-the-art polymer coatings. Most of these limitations are related to the fact that, typically, linear polymers are used with associated limited chemical functionality. Here, we examine the development of hyperbranched polyglycerols (HPGs) as promising candidates for the replacement of traditional linear polymers, such as the chemically analogous PEG, for the control of biointerfacial interactions. HPGs are highly branched globular molecules that exhibit a high valency, allow easy access to a variety of functionalities and can present biologically active signals. In this review, a comprehensive overview is provided with respect to the history, synthetic strategies, modifications and applications of HPGs.

Chimeric Saccharomyces cerevisiae Msh6 protein with an Msh3 mispair-binding domain combines properties of both proteins.

PubMed

Shell, Scarlet S; Putnam, Christopher D; Kolodner, Richard D

2007-06-26

Msh2-Msh3 and Msh2-Msh6 are two partially redundant mispair-recognition complexes that initiate mismatch repair in eukaryotes. Crystal structures of the prokaryotic homolog MutS suggest the mechanism by which Msh6 interacts with mispairs because key mispair-contacting residues are conserved in these two proteins. Because Msh3 lacks these conserved residues, we constructed a series of mutants to investigate the requirements for mispair interaction by Msh3. We found that a chimeric protein in which the mispair-binding domain (MBD) of Msh6 was replaced by the equivalent domain of Msh3 was functional for mismatch repair. This chimera possessed the mispair-binding specificity of Msh3 and revealed that communication between the MBD and the ATPase domain is conserved between Msh2-Msh3 and Msh2-Msh6. Further, the chimeric protein retained Msh6-like properties with respect to genetic interactions with the MutL homologs and an Msh2 MBD deletion mutant, indicating that Msh3-like behaviors beyond mispair specificity are not features controlled by the MBD.

The Nuclear Pore-Associated TREX-2 Complex Employs Mediator to Regulate Gene Expression

PubMed Central

Schneider, Maren; Hellerschmied, Doris; Schubert, Tobias; Amlacher, Stefan; Vinayachandran, Vinesh; Reja, Rohit; Pugh, B. Franklin; Clausen, Tim; Köhler, Alwin

2015-01-01

Summary Nuclear pore complexes (NPCs) influence gene expression besides their established function in nuclear transport. The TREX-2 complex localizes to the NPC basket and affects gene-NPC interactions, transcription, and mRNA export. How TREX-2 regulates the gene expression machinery is unknown. Here, we show that TREX-2 interacts with the Mediator complex, an essential regulator of RNA Polymerase (Pol) II. Structural and biochemical studies identify a conserved region on TREX-2, which directly binds the Mediator Med31/Med7N submodule. TREX-2 regulates assembly of Mediator with the Cdk8 kinase and is required for recruitment and site-specific phosphorylation of Pol II. Transcriptome and phenotypic profiling confirm that TREX-2 and Med31 are functionally interdependent at specific genes. TREX-2 additionally uses its Mediator-interacting surface to regulate mRNA export suggesting a mechanism for coupling transcription initiation and early steps of mRNA processing. Our data provide mechanistic insight into how an NPC-associated adaptor complex accesses the core transcription machinery. PMID:26317468

Decoy Strategies: The Structure of TL1A:DcR3 Complex

DOE Office of Scientific and Technical Information (OSTI.GOV)

C Zhan; Y Patskovsky; Q Yan

2011-12-31

Decoy Receptor 3 (DcR3), a secreted member of the Tumor Necrosis Factor (TNF) receptor superfamily, neutralizes three different TNF ligands: FasL, LIGHT, and TL1A. Each of these ligands engages unique signaling receptors which direct distinct and critical immune responses. We report the crystal structures of the unliganded DcR3 ectodomain and its complex with TL1A, as well as complementary mutagenesis and biochemical studies. These analyses demonstrate that DcR3 interacts with invariant backbone and side-chain atoms in the membrane-proximal half of TL1A which supports recognition of its three distinct TNF ligands. Additional features serve as antideterminants that preclude interaction with other membersmore » of the TNF superfamily. This mode of interaction is unique among characterized TNF:TNFR family members and provides a mechanistic basis for the broadened specificity required to support the decoy function of DcR3, as well as for the rational manipulation of specificity and affinity of DcR3 and its ligands.« less

Plant–Aphid Interactions Under Elevated CO2: Some Cues from Aphid Feeding Behavior

PubMed Central

Sun, Yucheng; Guo, Huijuan; Ge, Feng

2016-01-01

Although the increasing concentration of atmospheric carbon dioxide (CO2) accelerates the accumulation of carbohydrates and increases the biomass and yield of C3 crop plants, it also reduces their nitrogen concentration. The consequent changes in primary and secondary metabolites affect the palatability of host plants and the feeding of herbivorous insects. Aphids are phloem feeders and are considered the only feeding guild that positively responds to elevated CO2. In this review, we consider how elevated CO2 modifies host defenses, nutrients, and water-use efficiency by altering concentrations of the phytohormones jasmonic acid, salicylic acid, ethylene, and abscisic acid. We will describe how these elevated CO2-induced changes in defenses, nutrients, and water statusfacilitate specific stages of aphid feeding, including penetration, phloem-feeding, and xylem absorption. We conclude that a better understanding of the effects of elevated CO2 on aphids and on aphid damage to crop plants will require research on the molecular aspects of the interaction between plant and aphid but also research on aphid interactions with their intra- and inter-specific competitors and with their natural enemies. PMID:27148325

HaloTag Technology: A Versatile Platform for Biomedical Applications

PubMed Central

2015-01-01

Exploration of protein function and interaction is critical for discovering links among genomics, proteomics, and disease state; yet, the immense complexity of proteomics found in biological systems currently limits our investigational capacity. Although affinity and autofluorescent tags are widely employed for protein analysis, these methods have been met with limited success because they lack specificity and require multiple fusion tags and genetic constructs. As an alternative approach, the innovative HaloTag protein fusion platform allows protein function and interaction to be comprehensively analyzed using a single genetic construct with multiple capabilities. This is accomplished using a simplified process, in which a variable HaloTag ligand binds rapidly to the HaloTag protein (usually linked to the protein of interest) with high affinity and specificity. In this review, we examine all current applications of the HaloTag technology platform for biomedical applications, such as the study of protein isolation and purification, protein function, protein–protein and protein–DNA interactions, biological assays, in vitro cellular imaging, and in vivo molecular imaging. In addition, novel uses of the HaloTag platform are briefly discussed along with potential future applications. PMID:25974629

Ultrasonic power measurement system based on acousto-optic interaction.

PubMed

He, Liping; Zhu, Fulong; Chen, Yanming; Duan, Ke; Lin, Xinxin; Pan, Yongjun; Tao, Jiaquan

2016-05-01

Ultrasonic waves are widely used, with applications including the medical, military, and chemical fields. However, there are currently no effective methods for ultrasonic power measurement. Previously, ultrasonic power measurement has been reliant on mechanical methods such as hydrophones and radiation force balances. This paper deals with ultrasonic power measurement based on an unconventional method: acousto-optic interaction. Compared with mechanical methods, the optical method has a greater ability to resist interference and also has reduced environmental requirements. Therefore, this paper begins with an experimental determination of the acoustic power in water contained in a glass tank using a set of optical devices. Because the light intensity of the diffraction image generated by acousto-optic interaction contains the required ultrasonic power information, specific software was written to extract the light intensity information from the image through a combination of filtering, binarization, contour extraction, and other image processing operations. The power value can then be obtained rapidly by processing the diffraction image using a computer. The results of this work show that the optical method offers advantages that include accuracy, speed, and a noncontact measurement method.

Ultrasonic power measurement system based on acousto-optic interaction

NASA Astrophysics Data System (ADS)

He, Liping; Zhu, Fulong; Chen, Yanming; Duan, Ke; Lin, Xinxin; Pan, Yongjun; Tao, Jiaquan

2016-05-01

Ultrasonic waves are widely used, with applications including the medical, military, and chemical fields. However, there are currently no effective methods for ultrasonic power measurement. Previously, ultrasonic power measurement has been reliant on mechanical methods such as hydrophones and radiation force balances. This paper deals with ultrasonic power measurement based on an unconventional method: acousto-optic interaction. Compared with mechanical methods, the optical method has a greater ability to resist interference and also has reduced environmental requirements. Therefore, this paper begins with an experimental determination of the acoustic power in water contained in a glass tank using a set of optical devices. Because the light intensity of the diffraction image generated by acousto-optic interaction contains the required ultrasonic power information, specific software was written to extract the light intensity information from the image through a combination of filtering, binarization, contour extraction, and other image processing operations. The power value can then be obtained rapidly by processing the diffraction image using a computer. The results of this work show that the optical method offers advantages that include accuracy, speed, and a noncontact measurement method.

Issues in Conducting Epidemiologic Research Among Elders: Lessons From The MOBILIZE Boston Study

PubMed Central

Kelsey, Jennifer L.; Kiel, Douglas P.; Roman, Anthony M.; Cupples, L. Adrienne; Freeman, Marcie B.; Jones, Richard N.; Hannan, Marian T.; Leveille, Suzanne G.; Gagnon, Margaret M.; Lipsitz, Lewis A.

2008-01-01

Conducting research in elderly populations is important, but challenging. In this paper, the authors describe specific challenges that have arisen and solutions that have been used in carrying out The MOBILIZE Boston Study, a community-based, prospective cohort study in Massachusetts focusing on falls among 765 participants aged 70 years or older enrolled during 2005–2007. To recruit older individuals, face-to-face interactions are more effective than less personal approaches. Use of a board of community leaders facilitated community acceptance of the research. Establishing eligibility for potential participants required several interactions, so resources must be anticipated in advance. Assuring a safe and warm environment for elderly participants and offering a positive experience are a vital priority. Adequate funding, planning, and monitoring are required to provide transportation and a fully accessible environment in which to conduct study procedures as well as to select personnel highly skilled in interacting with elders. It is hoped that this paper will encourage and inform future epidemiologic research in this important segment of the population. PMID:18953059

Predicting the clinical efficacy and potential adverse effects of a humanized anticocaine monoclonal antibody

PubMed Central

Norman, Andrew B; Ball, William J

2012-01-01

The effects of a humanized monoclonal antibody (mAb) having high affinity and specificity for cocaine in animal models are reviewed. The mAb reduced the concentration of cocaine in the brain of mice after intravenous injection of cocaine. In addition, the mAb increased the concentration of cocaine required to reinstate cocaine self-administration. These effects may predict clinical efficacy of a passive immunotherapy for reducing the probability of cocaine-induced relapse. However, in the presence of the mAb, once cocaine self-administration was reinstated, the consumption rate of cocaine was increased. This effect is hypothesized to result from a pharmacokinetic/pharmacodynamic interaction. A humanized mAb should minimize adverse events related to the immunogenicity of the mAb protein, and the specificity for cocaine should avoid adverse events related to interactions with physiologically relevant endogenous proteins. PMID:22401638

Bmp2 and Notch cooperate to pattern the embryonic endocardium.

PubMed

Papoutsi, T; Luna-Zurita, L; Prados, B; Zaffran, S; de la Pompa, J L

2018-05-31

Signaling interactions between myocardium and endocardium pattern embryonic cardiac regions, instructing their development to fulfill specific functions in the mature heart. We show that ectopic Bmp2 expression in the mouse chamber myocardium changes the transcriptional signature of adjacent chamber endocardial cells into valve tissue, and enables them to undergo epithelial-mesenchyme transition. This induction is independent of valve myocardium specification and requires high levels of Notch1 activity. Biochemical experiments suggest that Bmp2-mediated Notch1 induction is achieved through transcriptional activation of the Notch ligand Jag1, and physical interaction of Smad1/5 with the intracellular domain of the Notch1 receptor. Thus, widespread myocardial Bmp2 and endocardial Notch signaling drive presumptive ventricular endocardium to differentiate into valve endocardium. Understanding the molecular basis of valve development is instrumental to designing therapeutic strategies for congenital heart valve defects. © 2018. Published by The Company of Biologists Ltd.

Effect of mutation at the interface of Trp-repressor dimeric protein: a steered molecular dynamics simulation.

PubMed

Miño, German; Baez, Mauricio; Gutierrez, Gonzalo

2013-09-01

The strength of key interfacial contacts that stabilize protein-protein interactions have been studied by computer simulation. Experimentally, changes in the interface are evaluated by generating specific mutations at one or more points of the protein structure. Here, such an evaluation is performed by means of steered molecular dynamics and use of a dimeric model of tryptophan repressor and in-silico mutants as a test case. Analysis of four particular cases shows that, in principle, it is possible to distinguish between wild-type and mutant forms by examination of the total energy and force-extension profiles. In particular, detailed atomic level structural analysis indicates that specific mutations at the interface of the dimeric model (positions 19 and 39) alter interactions that appear in the wild-type form of tryptophan repressor, reducing the energy and force required to separate both subunits.

Interactive information processing for NASA's mesoscale analysis and space sensor program

NASA Technical Reports Server (NTRS)

Parker, K. G.; Maclean, L.; Reavis, N.; Wilson, G.; Hickey, J. S.; Dickerson, M.; Karitani, S.; Keller, D.

1985-01-01

The Atmospheric Sciences Division (ASD) of the Systems Dynamics Laboratory at NASA's Marshall Space Flight Center (MSFC) is currently involved in interactive information processing for the Mesoscale Analysis and Space Sensor (MASS) program. Specifically, the ASD is engaged in the development and implementation of new space-borne remote sensing technology to observe and measure mesoscale atmospheric processes. These space measurements and conventional observational data are being processed together to gain an improved understanding of the mesoscale structure and the dynamical evolution of the atmosphere relative to cloud development and precipitation processes. To satisfy its vast data processing requirements, the ASD has developed a Researcher Computer System consiting of three primary computer systems which provides over 20 scientists with a wide range of capabilities for processing and displaying a large volumes of remote sensing data. Each of the computers performs a specific function according to its unique capabilities.

Electrostatically Accelerated Coupled Binding and Folding of Intrinsically Disordered Proteins

PubMed Central

Ganguly, Debabani; Otieno, Steve; Waddell, Brett; Iconaru, Luigi; Kriwacki, Richard W.; Chen, Jianhan

2012-01-01

Intrinsically disordered proteins (IDPs) are now recognized to be prevalent in biology, and many potential functional benefits have been discussed. However, the frequent requirement of peptide folding in specific interactions of IDPs could impose a kinetic bottleneck, which could be overcome only by efficient folding upon encounter. Intriguingly, existing kinetic data suggest that specific binding of IDPs is generally no slower than that of globular proteins. Here, we exploited the cell cycle regulator p27Kip1 (p27) as a model system to understand how IDPs might achieve efficient folding upon encounter for facile recognition. Combining experiments and coarse-grained modeling, we demonstrate that long-range electrostatic interactions between enriched charges on p27 and near its binding site on cyclin A not only enhance the encounter rate (i.e., electrostatic steering), but also promote folding-competent topologies in the encounter complexes, allowing rapid subsequent formation of short-range native interactions en route to the specific complex. In contrast, nonspecific hydrophobic interactions, while hardly affecting the encounter rate, can significantly reduce the efficiency of folding upon encounter and lead to slower binding kinetics. Further analysis of charge distributions in a set of known IDP complexes reveals that, although IDP binding sites tend to be more hydrophobic compared to the rest of the target surface, their vicinities are frequently enriched with charges to complement those on IDPs. This observation suggests that electrostatically accelerated encounter and induced folding might represent a prevalent mechanism for promoting facile IDP recognition. PMID:22721951

Genetic Dissection of Photoreceptor Subtype Specification by the Drosophila melanogaster Zinc Finger Proteins Elbow and No ocelli

PubMed Central

Wernet, Mathias F.; Meier, Kerstin M.; Baumann-Klausener, Franziska; Dorfman, Ruslan; Weihe, Ulrich; Labhart, Thomas; Desplan, Claude

2014-01-01

The elbow/no ocelli (elb/noc) complex of Drosophila melanogaster encodes two paralogs of the evolutionarily conserved NET family of zinc finger proteins. These transcriptional repressors share a conserved domain structure, including a single atypical C2H2 zinc finger. In flies, Elb and Noc are important for the development of legs, eyes and tracheae. Vertebrate NET proteins play an important role in the developing nervous system, and mutations in the homolog ZNF703 human promote luminal breast cancer. However, their interaction with transcriptional regulators is incompletely understood. Here we show that loss of both Elb and Noc causes mis-specification of polarization-sensitive photoreceptors in the ‘dorsal rim area’ (DRA) of the fly retina. This phenotype is identical to the loss of the homeodomain transcription factor Homothorax (Hth)/dMeis. Development of DRA ommatidia and expression of Hth are induced by the Wingless/Wnt pathway. Our data suggest that Elb/Noc genetically interact with Hth, and we identify two conserved domains crucial for this function. Furthermore, we show that Elb/Noc specifically interact with the transcription factor Orthodenticle (Otd)/Otx, a crucial regulator of rhodopsin gene transcription. Interestingly, different Elb/Noc domains are required to antagonize Otd functions in transcriptional activation, versus transcriptional repression. We propose that similar interactions between vertebrate NET proteins and Meis and Otx factors might play a role in development and disease. PMID:24625735

Arabidopsis VARIEGATED 3 encodes a chloroplast-targeted, zinc-finger protein required for chloroplast and palisade cell development.

PubMed

Naested, Henrik; Holm, Agnethe; Jenkins, Tom; Nielsen, H Bjørn; Harris, Cassandra A; Beale, Michael H; Andersen, Mathias; Mant, Alexandra; Scheller, Henrik; Camara, Bilal; Mattsson, Ole; Mundy, John

2004-09-15

The stable, recessive Arabidopsis variegated 3 (var3) mutant exhibits a variegated phenotype due to somatic areas lacking or containing developmentally retarded chloroplasts and greatly reduced numbers of palisade cells. The VAR3 gene, isolated by transposon tagging, encodes the 85.9 kDa VAR3 protein containing novel repeats and zinc fingers described as protein interaction domains. VAR3 interacts specifically in yeast and in vitro with NCED4, a putative polyene chain or carotenoid dioxygenase, and both VAR3 and NCED4 accumulate in the chloroplast stroma. Metabolic profiling demonstrates that pigment profiles are qualitatively similar in wild type and var3, although var3 accumulates lower levels of chlorophylls and carotenoids. These results indicate that VAR3 is a part of a protein complex required for normal chloroplast and palisade cell development.

Active learning in forensic science using Brownfield Action in a traditional or hybrid course in earth, environmental, or engineering sciences

NASA Astrophysics Data System (ADS)

Bower, P.; Liddicoat (2), J.

2009-04-01

Brownfield Action (BA - http://www.brownfieldaction.org) is a web-based, interactive, three-dimensional digital space and learning simulation in which students form geotechnical consulting companies and work collaboratively to explore and solve problems in environmental forensics. BA is being used in the United States at 10 colleges and universities in earth, environmental, or engineering sciences undergraduate and graduate courses. As a semester-long activity or done in modular form for specific topics, BA encourages active learning that requires attention to detail, intuition, and positive interaction between peers that results in Phase 1 and Phase 2 Environmental Site Assessments. Besides use in higher education courses, BA also can be adapted for instruction to local, state, and federal governmental employees, and employees in industry where brownfields need to be investigated or require remediation.

Fumed silica nanoparticle mediated biomimicry for optimal cell-material interactions for artificial organ development.

PubMed

de Mel, Achala; Ramesh, Bala; Scurr, David J; Alexander, Morgan R; Hamilton, George; Birchall, Martin; Seifalian, Alexander M

2014-03-01

Replacement of irreversibly damaged organs due to chronic disease, with suitable tissue engineered implants is now a familiar area of interest to clinicians and multidisciplinary scientists. Ideal tissue engineering approaches require scaffolds to be tailor made to mimic physiological environments of interest with specific surface topographical and biological properties for optimal cell-material interactions. This study demonstrates a single-step procedure for inducing biomimicry in a novel nanocomposite base material scaffold, to re-create the extracellular matrix, which is required for stem cell integration and differentiation to mature cells. Fumed silica nanoparticle mediated procedure of scaffold functionalization, can be potentially adapted with multiple bioactive molecules to induce cellular biomimicry, in the development human organs. The proposed nanocomposite materials already in patients for number of implants, including world first synthetic trachea, tear ducts and vascular bypass graft. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Mathematical Models of Continuous Flow Electrophoresis

NASA Technical Reports Server (NTRS)

Saville, D. A.; Snyder, R. S.

1985-01-01

Development of high resolution continuous flow electrophoresis devices ultimately requires comprehensive understanding of the ways various phenomena and processes facilitate or hinder separation. A comprehensive model of the actual three dimensional flow, temperature and electric fields was developed to provide guidance in the design of electrophoresis chambers for specific tasks and means of interpreting test data on a given chamber. Part of the process of model development includes experimental and theoretical studies of hydrodynamic stability. This is necessary to understand the origin of mixing flows observed with wide gap gravitational effects. To insure that the model accurately reflects the flow field and particle motion requires extensive experimental work. Another part of the investigation is concerned with the behavior of concentrated sample suspensions with regard to sample stream stability particle-particle interactions which might affect separation in an electric field, especially at high field strengths. Mathematical models will be developed and tested to establish the roles of the various interactions.

Subunit-specific rules governing AMPA receptor trafficking to synapses in hippocampal pyramidal neurons.

PubMed

Shi, S; Hayashi, Y; Esteban, J A; Malinow, R

2001-05-04

AMPA-type glutamate receptors (AMPA-Rs) mediate a majority of excitatory synaptic transmission in the brain. In hippocampus, most AMPA-Rs are hetero-oligomers composed of GluR1/GluR2 or GluR2/GluR3 subunits. Here we show that these AMPA-R forms display different synaptic delivery mechanisms. GluR1/GluR2 receptors are added to synapses during plasticity; this requires interactions between GluR1 and group I PDZ domain proteins. In contrast, GluR2/GluR3 receptors replace existing synaptic receptors continuously; this occurs only at synapses that already have AMPA-Rs and requires interactions by GluR2 with NSF and group II PDZ domain proteins. The combination of regulated addition and continuous replacement of synaptic receptors can stabilize long-term changes in synaptic efficacy and may serve as a general model for how surface receptor number is established and maintained.

MyDas, an Extensible Java DAS Server

PubMed Central

Jimenez, Rafael C.; Quinn, Antony F.; Jenkinson, Andrew M.; Mulder, Nicola; Martin, Maria; Hunter, Sarah; Hermjakob, Henning

2012-01-01

A large number of diverse, complex, and distributed data resources are currently available in the Bioinformatics domain. The pace of discovery and the diversity of information means that centralised reference databases like UniProt and Ensembl cannot integrate all potentially relevant information sources. From a user perspective however, centralised access to all relevant information concerning a specific query is essential. The Distributed Annotation System (DAS) defines a communication protocol to exchange annotations on genomic and protein sequences; this standardisation enables clients to retrieve data from a myriad of sources, thus offering centralised access to end-users. We introduce MyDas, a web server that facilitates the publishing of biological annotations according to the DAS specification. It deals with the common functionality requirements of making data available, while also providing an extension mechanism in order to implement the specifics of data store interaction. MyDas allows the user to define where the required information is located along with its structure, and is then responsible for the communication protocol details. PMID:23028496

CDE-1 affects chromosome segregation through uridylation of CSR-1-bound siRNAs.

PubMed

van Wolfswinkel, Josien C; Claycomb, Julie M; Batista, Pedro J; Mello, Craig C; Berezikov, Eugene; Ketting, René F

2009-10-02

We have studied the function of a conserved germline-specific nucleotidyltransferase protein, CDE-1, in RNAi and chromosome segregation in C. elegans. CDE-1 localizes specifically to mitotic chromosomes in embryos. This localization requires the RdRP EGO-1, which physically interacts with CDE-1, and the Argonaute protein CSR-1. We found that CDE-1 is required for the uridylation of CSR-1 bound siRNAs, and that in the absence of CDE-1 these siRNAs accumulate to inappropriate levels, accompanied by defects in both meiotic and mitotic chromosome segregation. Elevated siRNA levels are associated with erroneous gene silencing, most likely through the inappropriate loading of CSR-1 siRNAs into other Argonaute proteins. We propose a model in which CDE-1 restricts specific EGO-1-generated siRNAs to the CSR-1 mediated, chromosome associated RNAi pathway, thus separating it from other endogenous RNAi pathways. The conserved nature of CDE-1 suggests that similar sorting mechanisms may operate in other animals, including mammals.

MyDas, an extensible Java DAS server.

PubMed

Salazar, Gustavo A; García, Leyla J; Jones, Philip; Jimenez, Rafael C; Quinn, Antony F; Jenkinson, Andrew M; Mulder, Nicola; Martin, Maria; Hunter, Sarah; Hermjakob, Henning

2012-01-01

A large number of diverse, complex, and distributed data resources are currently available in the Bioinformatics domain. The pace of discovery and the diversity of information means that centralised reference databases like UniProt and Ensembl cannot integrate all potentially relevant information sources. From a user perspective however, centralised access to all relevant information concerning a specific query is essential. The Distributed Annotation System (DAS) defines a communication protocol to exchange annotations on genomic and protein sequences; this standardisation enables clients to retrieve data from a myriad of sources, thus offering centralised access to end-users.We introduce MyDas, a web server that facilitates the publishing of biological annotations according to the DAS specification. It deals with the common functionality requirements of making data available, while also providing an extension mechanism in order to implement the specifics of data store interaction. MyDas allows the user to define where the required information is located along with its structure, and is then responsible for the communication protocol details.

Kif3a Controls Murine Nephron Number Via GLI3 Repressor, Cell Survival, and Gene Expression in a Lineage-Specific Manner

PubMed Central

Chi, Lijun; Galtseva, Alevtina; Chen, Lin; Mo, Rong; Hui, Chi-chung; Rosenblum, Norman D.

2013-01-01

The primary cilium is required during early embryo patterning, epithelial tubulogenesis, and growth factor-dependent signal transduction. The requirement for primary cilia during renal epithelial-mesenchymal tissue interactions that give rise to nephrons is undefined. Here, we used Cre-mediated recombination to generate mice with Kif3a deficiency targeted to the ureteric and/or metanephric mesenchyme cell lineages in the embryonic kidney. Gradual loss of primary cilia in either lineage leads to a phenotype of reduced nephron number. Remarkably, in addition to cyst formation, loss of primary cilia in the ureteric epithelial cell leads to decreased expression of Wnt11 and Ret and reduced ureteric branching. Constitutive expression of GLI3 repressor (Gli3Δ699/+) rescues these abnormalities. In embryonic metanephric mesenchyme cells, Kif3a deficiency limits survival of nephrogenic progenitor cells and expression of genes required for nephron formation. Together, our data demonstrate that Kif3a controls nephron number via distinct cell lineage-specific mechanisms. PMID:23762375

PHD domain-mediated E3 ligase activity directs intramolecular sumoylation of an adjacent bromodomain required for gene silencing.

PubMed

Ivanov, Alexey V; Peng, Hongzhuang; Yurchenko, Vyacheslav; Yap, Kyoko L; Negorev, Dmitri G; Schultz, David C; Psulkowski, Elyse; Fredericks, William J; White, David E; Maul, Gerd G; Sadofsky, Moshe J; Zhou, Ming-Ming; Rauscher, Frank J

2007-12-14

Tandem PHD and bromodomains are often found in chromatin-associated proteins and have been shown to cooperate in gene silencing. Each domain can bind specifically modified histones: the mechanisms of cooperation between these domains are unknown. We show that the PHD domain of the KAP1 corepressor functions as an intramolecular E3 ligase for sumoylation of the adjacent bromodomain. The RING finger-like structure of the PHD domain is required for both Ubc9 binding and sumoylation and directs modification to specific lysine residues in the bromodomain. Sumoylation is required for KAP1-mediated gene silencing and functions by directly recruiting the SETDB1 histone methyltransferase and the CHD3/Mi2 component of the NuRD complex via SUMO-interacting motifs. Sumoylated KAP1 stimulates the histone methyltransferase activity of SETDB1. These data provide a mechanistic explanation for the cooperation of PHD and bromodomains in gene regulation and describe a function of the PHD domain as an intramolecular E3 SUMO ligase.

Cryogenic fluid management in space

NASA Technical Reports Server (NTRS)

Antar, Basil N.

1988-01-01

Many future space based vehicles and satellites will require on orbit refuelling procedures. Cryogenic fluid management technology is being developed to assess the requirements of such procedures as well as to aid in the design and development of these vehicles. Cryogenic fluid management technology for this application could be divided into two areas of study, one is concerned with fluid transfer process and the other with cryogenic liquid storage. This division is based upon the needed technology for the development of each area. In the first, the interaction of fluid dynamics with thermodynamics is essential, while in the second only thermodynamic analyses are sufficient to define the problem. The following specific process related to the liquid transfer area are discussed: tank chilldown and fill; tank pressurization; liquid positioning; and slosh dynamics and control. These specific issues are discussed in relation with the required technology for their development in the low gravity application area. In each process the relevant physics controlling the technology is identified and methods for resolving some of the basic questions are discussed.

The C-terminal region of lymphocytic choriomeningitis virus nucleoprotein contains distinct and segregable functional domains involved in NP-Z interaction and counteraction of the type I interferon response.

PubMed

Ortiz-Riaño, Emilio; Cheng, Benson Yee Hin; de la Torre, Juan Carlos; Martínez-Sobrido, Luis

2011-12-01

Several arenaviruses cause hemorrhagic fever (HF) disease in humans that is associated with high morbidity and significant mortality. Arenavirus nucleoprotein (NP), the most abundant viral protein in infected cells and virions, encapsidates the viral genome RNA, and this NP-RNA complex, together with the viral L polymerase, forms the viral ribonucleoprotein (vRNP) that directs viral RNA replication and gene transcription. Formation of infectious arenavirus progeny requires packaging of vRNPs into budding particles, a process in which arenavirus matrix-like protein (Z) plays a central role. In the present study, we have characterized the NP-Z interaction for the prototypic arenavirus lymphocytic choriomeningitis virus (LCMV). The LCMV NP domain that interacted with Z overlapped with a previously documented C-terminal domain that counteracts the host type I interferon (IFN) response. However, we found that single amino acid mutations that affect the anti-IFN function of LCMV NP did not disrupt the NP-Z interaction, suggesting that within the C-terminal region of NP different amino acid residues critically contribute to these two distinct and segregable NP functions. A similar NP-Z interaction was confirmed for the HF arenavirus Lassa virus (LASV). Notably, LCMV NP interacted similarly with both LCMV Z and LASV Z, while LASV NP interacted only with LASV Z. Our results also suggest the presence of a conserved protein domain within NP but with specific amino acid residues playing key roles in determining the specificity of NP-Z interaction that may influence the viability of reassortant arenaviruses. In addition, this NP-Z interaction represents a potential target for the development of antiviral drugs to combat human-pathogenic arenaviruses.

A male-specific QTL for social interaction behavior in mice mapped with automated pattern detection by a hidden Markov model incorporated into newly developed freeware.

PubMed

Arakawa, Toshiya; Tanave, Akira; Ikeuchi, Shiho; Takahashi, Aki; Kakihara, Satoshi; Kimura, Shingo; Sugimoto, Hiroki; Asada, Nobuhiko; Shiroishi, Toshihiko; Tomihara, Kazuya; Tsuchiya, Takashi; Koide, Tsuyoshi

2014-08-30

Owing to their complex nature, social interaction tests normally require the observation of video data by a human researcher, and thus are difficult to use in large-scale studies. We previously established a statistical method, a hidden Markov model (HMM), which enables the differentiation of two social states ("interaction" and "indifference"), and three social states ("sniffing", "following", and "indifference"), automatically in silico. Here, we developed freeware called DuoMouse for the rapid evaluation of social interaction behavior. This software incorporates five steps: (1) settings, (2) video recording, (3) tracking from the video data, (4) HMM analysis, and (5) visualization of the results. Using DuoMouse, we mapped a genetic locus related to social interaction. We previously reported that a consomic strain, B6-Chr6C(MSM), with its chromosome 6 substituted for one from MSM/Ms, showed more social interaction than C57BL/6 (B6). We made four subconsomic strains, C3, C5, C6, and C7, each of which has a shorter segment of chromosome 6 derived from B6-Chr6C, and conducted social interaction tests on these strains. DuoMouse indicated that C6, but not C3, C5, and C7, showed higher interaction, sniffing, and following than B6, specifically in males. The data obtained by human observation showed high concordance to those from DuoMouse. The results indicated that the MSM-derived chromosomal region present in C6-but not in C3, C5, and C7-associated with increased social behavior. This method to analyze social interaction will aid primary screening for difference in social behavior in mice. Copyright © 2014 Elsevier B.V. All rights reserved.

A novel TPR–BEN domain interaction mediates PICH–BEND3 association

PubMed Central

Pitchai, Ganesha P.; Kaulich, Manuel; Mesa, Pablo; Yao, Qi; Sarlos, Kata; Streicher, Werner W.; Nigg, Erich A.

2017-01-01

Abstract PICH is a DNA translocase required for the maintenance of chromosome stability in human cells. Recent data indicate that PICH co-operates with topoisomerase IIα to suppress pathological chromosome missegregation through promoting the resolution of ultra-fine anaphase bridges (UFBs). Here, we identify the BEN domain-containing protein 3 (BEND3) as an interaction partner of PICH in human cells in mitosis. We have purified full length PICH and BEND3 and shown that they exhibit a functional biochemical interaction in vitro. We demonstrate that the PICH–BEND3 interaction occurs via a novel interface between a TPR domain in PICH and a BEN domain in BEND3, and have determined the crystal structure of this TPR–BEN complex at 2.2 Å resolution. Based on the structure, we identified amino acids important for the TPR–BEN domain interaction, and for the functional interaction of the full-length proteins. Our data reveal a proposed new function for BEND3 in association with PICH, and the first example of a specific protein–protein interaction mediated by a BEN domain. PMID:28977671

Classroom Interaction Studies as a Source for Teacher Competencies: The Use of Case Studies with Multiple Instruments for Studying Teacher Competencies in Multicultural Classes

ERIC Educational Resources Information Center

den Brok, Perry; van Eerde, Dolly; Hajer, Maaike

2010-01-01

Dealing with ethnic and linguistic diversity is one of the major challenges in today's education. Therefore, the formulation of competencies for teachers and teacher training should take into account the specific requirements of teaching in multicultural schools. In 2002, a series of small-scale studies were conducted to identify and formulate…

An Analysis of TPACK Integration into English Language Teacher Education Programs: A Comparative Study of Turkey and Denmark

ERIC Educational Resources Information Center

Turgut, Yildiz

2017-01-01

Technology has always been a part of teaching process and being able to teach with technology requires an understanding of how technology, pedagogy, and content interact to support student learning. Recently, studies on developing TPACK skills of teachers have been emerged; however, studies on TPACK in pre-service education on a specific domain,…

Enabling Learning through the Assessment Process

DTIC Science & Technology

2010-04-08

Software, 47. 32 a specific pattern over time.”98 Johnson provides an example of this when discussing the computer simulation of slime mold growth. He...asserts that since the designers understood the underlying interactions between the individual slime molds , they could increase or decrease the...density of individual mold cells and the aggregating chemical that is required for the molds to group together. Furthermore, Johnson suggests that this

Factors Affecting the Participation of the Deaf and Hard of Hearing in e-Learning and Their Satisfaction: A Quantitative Study

ERIC Educational Resources Information Center

Ferreiro-Lago, Emilio; Osuna-Acedo, Sara

2017-01-01

Today the digital divide is measured not so much by access to the internet but by people's ability to interact in cyberspace, which calls for specific digital communication skills. These skills are also required for participation in e-learning, bearing in mind that the deaf and hard of hearing have certain characteristics related to their…

Technical aspects of contrast-enhanced ultrasound (CEUS) examinations: tips and tricks.

PubMed

Greis, C

2014-01-01

Ultrasound contrast agents have substantially extended the clinical value of ultrasound, allowing the assessment of blood flow and distribution in real-time down to microcapillary level. Selective imaging of contrast agent signals requires a contrast-specific imaging mode on the ultrasound scanner, allowing real-time separation of tissue and contrast agent signals. The creation of a contrast image requires a specific interaction between the insonated ultrasound wave and the contrast agent microbubbles, leading to persistent oscillation of the bubbles. Several technical and procedural parameters have a significant influence on the quality of CEUS images and should be controlled carefully to obtain good image quality and a reliable diagnosis. Achieving the proper balance between the respective parameters is a matter of technical knowledge and experience. Appropriate training and education should be mandatory for every investigator performing CEUS examinations.

The MINERVA Software Development Process

NASA Technical Reports Server (NTRS)

Narkawicz, Anthony; Munoz, Cesar A.; Dutle, Aaron M.

2017-01-01

This paper presents a software development process for safety-critical software components of cyber-physical systems. The process is called MINERVA, which stands for Mirrored Implementation Numerically Evaluated against Rigorously Verified Algorithms. The process relies on formal methods for rigorously validating code against its requirements. The software development process uses: (1) a formal specification language for describing the algorithms and their functional requirements, (2) an interactive theorem prover for formally verifying the correctness of the algorithms, (3) test cases that stress the code, and (4) numerical evaluation on these test cases of both the algorithm specifications and their implementations in code. The MINERVA process is illustrated in this paper with an application to geo-containment algorithms for unmanned aircraft systems. These algorithms ensure that the position of an aircraft never leaves a predetermined polygon region and provide recovery maneuvers when the region is inadvertently exited.

Transformations of software design and code may lead to reduced errors

NASA Technical Reports Server (NTRS)

Connelly, E. M.

1983-01-01

The capability of programmers and non-programmers to specify problem solutions by developing example-solutions and also for the programmers by writing computer programs was investigated; each method of specification was accomplished at various levels of problem complexity. The level of difficulty of each problem was reflected by the number of steps needed by the user to develop a solution. Machine processing of the user inputs permitted inferences to be developed about the algorithms required to solve a particular problem. The interactive feedback of processing results led users to a more precise definition of the desired solution. Two participant groups (programmers and bookkeepers/accountants) working with three levels of problem complexity and three levels of processor complexity were used. The experimental task employed required specification of a logic for solution of a Navy task force problem.

Tuning the specificity of a Two-in-One Fab against three angiogenic antigens by fully utilizing the information of deep mutational scanning.

PubMed

Koenig, Patrick; Sanowar, Sarah; Lee, Chingwei V; Fuh, Germaine

Monoclonal antibodies developed for therapeutic or diagnostic purposes need to demonstrate highly defined binding specificity profiles. Engineering of an antibody to enhance or reduce binding to related antigens is often needed to achieve the desired biologic activity without safety concern. Here, we describe a deep sequencing-aided engineering strategy to fine-tune the specificity of an angiopoietin-2 (Ang2)/vascular endothelial growth factor (VEGF) dual action Fab, 5A12.1 for the treatment of age-related macular degeneration. This antibody utilizes overlapping complementarity-determining region (CDR) sites for dual Ang2/VEGF interaction with K D in the sub-nanomolar range. However, it also exhibits significant (K D of 4 nM) binding to angiopoietin-1, which has high sequence identity with Ang2. We generated a large phage-displayed library of 5A12.1 Fab variants with all possible single mutations in the 6 CDRs. By tracking the change of prevalence of each mutation during various selection conditions, we identified 35 mutations predicted to decrease the affinity for Ang1 while maintaining the affinity for Ang2 and VEGF. We confirmed the specificity profiles for 25 of these single mutations as Fab protein. Structural analysis showed that some of the Fab mutations cluster near a potential Ang1/2 epitope residue that differs in the 2 proteins, while others are up to 15 Å away from the antigen-binding site and likely influence the binding interaction remotely. The approach presented here provides a robust and efficient method for specificity engineering that does not require prior knowledge of the antigen antibody interaction and can be broadly applied to antibody specificity engineering projects.

Trust and Its Role in the Medical Encounter.

PubMed

Holland, Stephen; Stocks, David

2017-09-01

This paper addresses two research questions. The first is theoretical: What is trust? In the first half of this paper we present a distinctive tripartite analysis. We describe three attitudes, here called reliance, specific trust and general trust, each of which is characterised and illustrated. We argue that these attitudes are related, but not reducible, to one another. We suggest that the current impasse in the analysis of trust is in part due to the fact that some writers allude to these distinctions, but unclearly so, whilst others elide them altogether. The second research question focuses on doctor-patient interaction. Trust is often said to be central in medical encounters but this strikes us as too vague. The success of doctor-patient relations in part depends on adopting the most appropriate of the three attitudes we delineate. We argue that reliance is the appropriate attitude for most medical encounters. When circumstances do require trust, the distinction between specific trust and general trust is crucial. We describe medical encounters requiring specific trust. General trust is less often required in medicine; but it is appropriate in some cases and, when called for, it is called for strongly.

A Unified Approach to Modeling Multidisciplinary Interactions

NASA Technical Reports Server (NTRS)

Samareh, Jamshid A.; Bhatia, Kumar G.

2000-01-01

There are a number of existing methods to transfer information among various disciplines. For a multidisciplinary application with n disciplines, the traditional methods may be required to model (n(exp 2) - n) interactions. This paper presents a unified three-dimensional approach that reduces the number of interactions from (n(exp 2) - n) to 2n by using a computer-aided design model. The proposed modeling approach unifies the interactions among various disciplines. The approach is independent of specific discipline implementation, and a number of existing methods can be reformulated in the context of the proposed unified approach. This paper provides an overview of the proposed unified approach and reformulations for two existing methods. The unified approach is specially tailored for application environments where the geometry is created and managed through a computer-aided design system. Results are presented for a blended-wing body and a high-speed civil transport.

A systems model for immune cell interactions unravels the mechanism of inflammation in human skin.

PubMed

Valeyev, Najl V; Hundhausen, Christian; Umezawa, Yoshinori; Kotov, Nikolay V; Williams, Gareth; Clop, Alex; Ainali, Crysanthi; Ouzounis, Christos; Tsoka, Sophia; Nestle, Frank O

2010-12-02

Inflammation is characterized by altered cytokine levels produced by cell populations in a highly interdependent manner. To elucidate the mechanism of an inflammatory reaction, we have developed a mathematical model for immune cell interactions via the specific, dose-dependent cytokine production rates of cell populations. The model describes the criteria required for normal and pathological immune system responses and suggests that alterations in the cytokine production rates can lead to various stable levels which manifest themselves in different disease phenotypes. The model predicts that pairs of interacting immune cell populations can maintain homeostatic and elevated extracellular cytokine concentration levels, enabling them to operate as an immune system switch. The concept described here is developed in the context of psoriasis, an immune-mediated disease, but it can also offer mechanistic insights into other inflammatory pathologies as it explains how interactions between immune cell populations can lead to disease phenotypes.

Targeting Allosteric Control Mechanisms in Heat Shock Protein 70 (Hsp70)

PubMed Central

Li, Xiaokai; Shao, Hao; Taylor, Isabelle R.; Gestwicki, Jason E.

2017-01-01

Heat shock protein 70 (Hsp70) is a molecular chaperone that plays critical roles in protein homeostasis. Hsp70’s chaperone activity is coordinated by intra-molecular interactions between its two domains, as well as inter-molecular interactions between Hsp70 and its co-chaperones. Each of these contacts represents a potential opportunity for the development of chemical inhibitors. To illustrate this concept, we review three classes of recently identified molecules that bind distinct pockets on Hsp70. Although all three compounds share the ability to interrupt core biochemical functions of Hsp70, they stabilize different conformers. Accordingly, each compound appears to interrupt a specific subset of inter- and intra-molecular interactions. Thus, an accurate definition of an Hsp70 inhibitor may require a particularly detailed understanding of the molecule’s binding site and its effects on protein-protein interactions. PMID:27072701

Autoantibody-induced internalization of CNS AQP4 water channel and EAAT2 glutamate transporter requires astrocytic Fc receptor.

PubMed

Hinson, Shannon R; Clift, Ian C; Luo, Ningling; Kryzer, Thomas J; Lennon, Vanda A

2017-05-23

Aquaporin-4 (AQP4) water channel-specific IgG distinguishes neuromyelitis optica (NMO) from multiple sclerosis and causes characteristic immunopathology in which central nervous system (CNS) demyelination is secondary. Early events initiating the pathophysiological outcomes of IgG binding to astrocytic AQP4 are poorly understood. CNS lesions reflect events documented in vitro following IgG interaction with AQP4: AQP4 internalization, attenuated glutamate uptake, intramyelinic edema, interleukin-6 release, complement activation, inflammatory cell recruitment, and demyelination. Here, we demonstrate that AQP4 internalization requires AQP4-bound IgG to engage an astrocytic Fcγ receptor (FcγR). IgG-lacking Fc redistributes AQP4 within the plasma membrane and induces interleukin-6 release. However, AQP4 endocytosis requires an activating FcγR's gamma subunit and involves astrocytic membrane loss of an inhibitory FcγR, CD32B. Interaction of the IgG-AQP4 complex with FcγRs triggers coendocytosis of the excitatory amino acid transporter 2 (EAAT2). Requirement of FcγR engagement for internalization of two astrocytic membrane proteins critical to CNS homeostasis identifies a complement-independent, upstream target for potential early therapeutic intervention in NMO.

A learning controller for nonrepetitive robotic operation

NASA Technical Reports Server (NTRS)

Miller, W. T., III

1987-01-01

A practical learning control system is described which is applicable to complex robotic and telerobotic systems involving multiple feedback sensors and multiple command variables. In the controller, the learning algorithm is used to learn to reproduce the nonlinear relationship between the sensor outputs and the system command variables over particular regions of the system state space, rather than learning the actuator commands required to perform a specific task. The learned information is used to predict the command signals required to produce desired changes in the sensor outputs. The desired sensor output changes may result from automatic trajectory planning or may be derived from interactive input from a human operator. The learning controller requires no a priori knowledge of the relationships between the sensor outputs and the command variables. The algorithm is well suited for real time implementation, requiring only fixed point addition and logical operations. The results of learning experiments using a General Electric P-5 manipulator interfaced to a VAX-11/730 computer are presented. These experiments involved interactive operator control, via joysticks, of the position and orientation of an object in the field of view of a video camera mounted on the end of the robot arm.

Analysis of the DNA-Binding Activities of the Arabidopsis R2R3-MYB Transcription Factor Family by One-Hybrid Experiments in Yeast

PubMed Central

Kelemen, Zsolt; Sebastian, Alvaro; Xu, Wenjia; Grain, Damaris; Salsac, Fabien; Avon, Alexandra; Berger, Nathalie; Tran, Joseph; Dubreucq, Bertrand; Lurin, Claire; Lepiniec, Loïc; Contreras-Moreira, Bruno; Dubos, Christian

2015-01-01

The control of growth and development of all living organisms is a complex and dynamic process that requires the harmonious expression of numerous genes. Gene expression is mainly controlled by the activity of sequence-specific DNA binding proteins called transcription factors (TFs). Amongst the various classes of eukaryotic TFs, the MYB superfamily is one of the largest and most diverse, and it has considerably expanded in the plant kingdom. R2R3-MYBs have been extensively studied over the last 15 years. However, DNA-binding specificity has been characterized for only a small subset of these proteins. Therefore, one of the remaining challenges is the exhaustive characterization of the DNA-binding specificity of all R2R3-MYB proteins. In this study, we have developed a library of Arabidopsis thaliana R2R3-MYB open reading frames, whose DNA-binding activities were assayed in vivo (yeast one-hybrid experiments) with a pool of selected cis-regulatory elements. Altogether 1904 interactions were assayed leading to the discovery of specific patterns of interactions between the various R2R3-MYB subgroups and their DNA target sequences and to the identification of key features that govern these interactions. The present work provides a comprehensive in vivo analysis of R2R3-MYB binding activities that should help in predicting new DNA motifs and identifying new putative target genes for each member of this very large family of TFs. In a broader perspective, the generated data will help to better understand how TF interact with their target DNA sequences. PMID:26484765

Suppressor mutations identify amino acids in PAA-1/PR65 that facilitate regulatory RSA-1/B″ subunit targeting of PP2A to centrosomes in C. elegans

PubMed Central

Lange, Karen I.; Heinrichs, Jeffrey; Cheung, Karen; Srayko, Martin

2013-01-01

Summary Protein phosphorylation and dephosphorylation is a key mechanism for the spatial and temporal regulation of many essential developmental processes and is especially prominent during mitosis. The multi-subunit protein phosphatase 2A (PP2A) enzyme plays an important, yet poorly characterized role in dephosphorylating proteins during mitosis. PP2As are heterotrimeric complexes comprising a catalytic, structural, and regulatory subunit. Regulatory subunits are mutually exclusive and determine subcellular localization and substrate specificity of PP2A. At least 3 different classes of regulatory subunits exist (termed B, B′, B″) but there is no obvious similarity in primary sequence between these classes. Therefore, it is not known how these diverse regulatory subunits interact with the same holoenzyme to facilitate specific PP2A functions in vivo. The B″ family of regulatory subunits is the least understood because these proteins lack conserved structural domains. RSA-1 (regulator of spindle assembly) is a regulatory B″ subunit required for mitotic spindle assembly in Caenorhabditis elegans. In order to address how B″ subunits interact with the PP2A core enzyme, we focused on a conditional allele, rsa-1(or598ts), and determined that this mutation specifically disrupts the protein interaction between RSA-1 and the PP2A structural subunit, PAA-1. Through genetic screening, we identified a putative interface on the PAA-1 structural subunit that interacts with a defined region of RSA-1/B″. In the context of previously published results, these data propose a mechanism of how different PP2A B-regulatory subunit families can bind the same holoenzyme in a mutually exclusive manner, to perform specific tasks in vivo. PMID:23336080

Suppressor mutations identify amino acids in PAA-1/PR65 that facilitate regulatory RSA-1/B″ subunit targeting of PP2A to centrosomes in C. elegans.

PubMed

Lange, Karen I; Heinrichs, Jeffrey; Cheung, Karen; Srayko, Martin

2013-01-15

Protein phosphorylation and dephosphorylation is a key mechanism for the spatial and temporal regulation of many essential developmental processes and is especially prominent during mitosis. The multi-subunit protein phosphatase 2A (PP2A) enzyme plays an important, yet poorly characterized role in dephosphorylating proteins during mitosis. PP2As are heterotrimeric complexes comprising a catalytic, structural, and regulatory subunit. Regulatory subunits are mutually exclusive and determine subcellular localization and substrate specificity of PP2A. At least 3 different classes of regulatory subunits exist (termed B, B', B″) but there is no obvious similarity in primary sequence between these classes. Therefore, it is not known how these diverse regulatory subunits interact with the same holoenzyme to facilitate specific PP2A functions in vivo. The B″ family of regulatory subunits is the least understood because these proteins lack conserved structural domains. RSA-1 (regulator of spindle assembly) is a regulatory B″ subunit required for mitotic spindle assembly in Caenorhabditis elegans. In order to address how B″ subunits interact with the PP2A core enzyme, we focused on a conditional allele, rsa-1(or598ts), and determined that this mutation specifically disrupts the protein interaction between RSA-1 and the PP2A structural subunit, PAA-1. Through genetic screening, we identified a putative interface on the PAA-1 structural subunit that interacts with a defined region of RSA-1/B″. In the context of previously published results, these data propose a mechanism of how different PP2A B-regulatory subunit families can bind the same holoenzyme in a mutually exclusive manner, to perform specific tasks in vivo.

The Caenorhabditis elegans NF2/Merlin Molecule NFM-1 Nonautonomously Regulates Neuroblast Migration and Interacts Genetically with the Guidance Cue SLT-1/Slit.

PubMed

Josephson, Matthew P; Aliani, Rana; Norris, Megan L; Ochs, Matthew E; Gujar, Mahekta; Lundquist, Erik A

2017-02-01

During nervous system development, neurons and their progenitors migrate to their final destinations. In Caenorhabditis elegans, the bilateral Q neuroblasts and their descendants migrate long distances in opposite directions, despite being born in the same posterior region. QR on the right migrates anteriorly and generates the AQR neuron positioned near the head, and QL on the left migrates posteriorly, giving rise to the PQR neuron positioned near the tail. In a screen for genes required for AQR and PQR migration, we identified an allele of nfm-1, which encodes a molecule similar to vertebrate NF2/Merlin, an important tumor suppressor in humans. Mutations in NF2 lead to neurofibromatosis type II, characterized by benign tumors of glial tissues. Here we demonstrate that in C. elegans, nfm-1 is required for the ability of Q cells and their descendants to extend protrusions and to migrate, but is not required for direction of migration. Using a combination of mosaic analysis and cell-specific expression, we show that NFM-1 is required nonautonomously, possibly in muscles, to promote Q lineage migrations. We also show a genetic interaction between nfm-1 and the C. elegans Slit homolog slt-1, which encodes a conserved secreted guidance cue. Our results suggest that NFM-1 might be involved in the generation of an extracellular cue that promotes Q neuroblast protrusion and migration that acts with or in parallel to SLT-1 In vertebrates, NF2 and Slit2 interact in axon pathfinding, suggesting a conserved interaction of NF2 and Slit2 in regulating migratory events. Copyright © 2017 by the Genetics Society of America.

The Caenorhabditis elegans NF2/Merlin Molecule NFM-1 Nonautonomously Regulates Neuroblast Migration and Interacts Genetically with the Guidance Cue SLT-1/Slit

PubMed Central

Josephson, Matthew P.; Aliani, Rana; Norris, Megan L.; Ochs, Matthew E.; Gujar, Mahekta; Lundquist, Erik A.

2017-01-01

During nervous system development, neurons and their progenitors migrate to their final destinations. In Caenorhabditis elegans, the bilateral Q neuroblasts and their descendants migrate long distances in opposite directions, despite being born in the same posterior region. QR on the right migrates anteriorly and generates the AQR neuron positioned near the head, and QL on the left migrates posteriorly, giving rise to the PQR neuron positioned near the tail. In a screen for genes required for AQR and PQR migration, we identified an allele of nfm-1, which encodes a molecule similar to vertebrate NF2/Merlin, an important tumor suppressor in humans. Mutations in NF2 lead to neurofibromatosis type II, characterized by benign tumors of glial tissues. Here we demonstrate that in C. elegans, nfm-1 is required for the ability of Q cells and their descendants to extend protrusions and to migrate, but is not required for direction of migration. Using a combination of mosaic analysis and cell-specific expression, we show that NFM-1 is required nonautonomously, possibly in muscles, to promote Q lineage migrations. We also show a genetic interaction between nfm-1 and the C. elegans Slit homolog slt-1, which encodes a conserved secreted guidance cue. Our results suggest that NFM-1 might be involved in the generation of an extracellular cue that promotes Q neuroblast protrusion and migration that acts with or in parallel to SLT-1. In vertebrates, NF2 and Slit2 interact in axon pathfinding, suggesting a conserved interaction of NF2 and Slit2 in regulating migratory events. PMID:27913619

A novel requirement for C. elegans Alix/ALX-1 in RME-1 mediated membrane transport

PubMed Central

Shi, Anbing; Pant, Saumya; Balklava, Zita; Chen, Carlos Chih-Hsiung; Figueroa, Vanesa; Grant, Barth D.

2007-01-01

Summary Background Alix/Bro1p family proteins have recently been identified as important components of multivesicular endosomes (MVEs) involved in the sorting of endocytosed integral membrane proteins, interacting with components of the ESCRT complex, the unconventional phospholipid LBPA, and other known endocytosis regulators. During infection Alix can be co-opted by enveloped retroviruses, including HIV, providing an important function during virus budding from the plasma membrane. In addition Alix is associated with the actin cytoskeleton and may regulate cytoskeletal dynamics. Results Here we demonstrate a novel physical interaction between the only apparent Alix/Bro1p family protein in C. elegans, ALX-1, and a key regulator of receptor recycling from endosomes to the plasma membrane called RME-1. Analysis of alx-1 mutants indicates that ALX-1 is required for endocytic recycling of specific basolateral cargo in the C. elegans intestine, a pathway previously defined by analysis of rme-1 mutants. Expression of truncated human Alix in HeLa cells disrupts recycling of MHCI, a known Ehd1/RME-1 dependent transport step, suggesting phylogenetic conservation of this function. We show that the interaction of ALX-1 with RME-1 in C. elegans, mediated by RME-1/YPSL and ALX-1/NPF motifs, is required for this recycling process. In the C. elegans intestine ALX-1 localizes to both recycling endosomes and MVEs, but the ALX-1/RME-1 interaction appears dispensable for ALX-1 function in MVEs/late endosomes. Conclusions This work provides the first demonstration of a requirement for an Alix/Bro1p family member in the endocytic recycling pathway in association with the recycling regulator RME-1. PMID:17997305

Redox potential tuning through differential quinone binding in the photosynthetic reaction center of Rhodobacter sphaeroides

DOE PAGES

Vermaas, Josh V.; Taguchi, Alexander T.; Dikanov, Sergei A.; ...

2015-03-03

Ubiquinone forms an integral part of the electron transport chain in cellular respiration and photosynthesis across a vast number of organisms. Prior experimental results have shown that the photosynthetic reaction center (RC) from Rhodobacter sphaeroides is only fully functional with a limited set of methoxy-bearing quinones, suggesting that specific interactions with this substituent are required to drive electron transport and the formation of quinol. The nature of these interactions has yet to be determined. Through parameterization of a CHARMM-compatible quinone force field and subsequent molecular dynamics simulations of the quinone-bound RC, in this paper we have investigated and characterized themore » interactions of the protein with the quinones in the Q A and Q B sites using both equilibrium simulation and thermodynamic integration. In particular, we identify a specific interaction between the 2-methoxy group of ubiquinone in the Q B site and the amide nitrogen of GlyL225 that we implicate in locking the orientation of the 2-methoxy group, thereby tuning the redox potential difference between the quinones occupying the Q A and Q B sites. Finally, disruption of this interaction leads to weaker binding in a ubiquinone analogue that lacks a 2-methoxy group, a finding supported by reverse electron transfer electron paramagnetic resonance experiments of the Q A–Q B– biradical and competitive binding assays.« less

Rapid surface-biostructure interaction analysis using strong metal-based nanomagnets.

PubMed

Rotzetter, Aline C C; Schumacher, Christoph M; Zako, Tamotsu; Stark, Wendelin J; Maeda, Mizuo

2013-11-19

Nanomaterials are increasingly suggested for the selective adsorption and extraction of complex compounds in biomedicine. Binding of the latter requires specific surface modifications of the nanostructures. However, even complicated macromolecules such as proteins can afford affinities toward basic surface characteristics such as hydrophobicity, topology, and electrostatic charge. In this study, we address these more basic physical interactions. In a model system, the interaction of bovine serum albumin and amyloid β 42 fibrillar aggregates with carbon-coated cobalt nanoparticles, functionalized with various polymers differing in character, was studied. The possibility of rapid magnetic separation upon binding to the surface represents a valuable tool for studying surface interactions and selectivities. We find that the surface interaction of Aβ 42 fibrillar aggregates is mostly hydrophobic in nature. Because bovine serum albumin (BSA) is conformationally adaptive, it is known to bind surfaces with widely differing properties (charge, topology, and hydrophobicity). However, the rate of tight binding (no desorption upon washing) can vary largely depending on the extent of necessary conformational changes for a specific surface. We found that BSA can only bind slowly to polyethylenimine-coated nanomagnets. Under competitive conditions (high excess BSA compared to that for β 42 fibrillar aggregates), this effect is beneficial for targeting the fibrillar species. These findings highlight the possibility of selective extractions from complex media when advantageous basic physical surface properties are chosen.

Redox potential tuning through differential quinone binding in the photosynthetic reaction center of Rhodobacter sphaeroides.

PubMed

Vermaas, Josh V; Taguchi, Alexander T; Dikanov, Sergei A; Wraight, Colin A; Tajkhorshid, Emad

2015-03-31

Ubiquinone forms an integral part of the electron transport chain in cellular respiration and photosynthesis across a vast number of organisms. Prior experimental results have shown that the photosynthetic reaction center (RC) from Rhodobacter sphaeroides is only fully functional with a limited set of methoxy-bearing quinones, suggesting that specific interactions with this substituent are required to drive electron transport and the formation of quinol. The nature of these interactions has yet to be determined. Through parameterization of a CHARMM-compatible quinone force field and subsequent molecular dynamics simulations of the quinone-bound RC, we have investigated and characterized the interactions of the protein with the quinones in the Q(A) and Q(B) sites using both equilibrium simulation and thermodynamic integration. In particular, we identify a specific interaction between the 2-methoxy group of ubiquinone in the Q(B) site and the amide nitrogen of GlyL225 that we implicate in locking the orientation of the 2-methoxy group, thereby tuning the redox potential difference between the quinones occupying the Q(A) and Q(B) sites. Disruption of this interaction leads to weaker binding in a ubiquinone analogue that lacks a 2-methoxy group, a finding supported by reverse electron transfer electron paramagnetic resonance experiments of the Q(A)⁻Q(B)⁻ biradical and competitive binding assays.

Heterodimerization of Msx and Dlx homeoproteins results in functional antagonism.

PubMed

Zhang, H; Hu, G; Wang, H; Sciavolino, P; Iler, N; Shen, M M; Abate-Shen, C

1997-05-01

Protein-protein interactions are known to be essential for specifying the transcriptional activities of homeoproteins. Here we show that representative members of the Msx and Dlx homeoprotein families form homo- and heterodimeric complexes. We demonstrate that dimerization by Msx and Dlx proteins is mediated through their homeodomains and that the residues required for this interaction correspond to those necessary for DNA binding. Unlike most other known examples of homeoprotein interactions, association of Msx and Dlx proteins does not promote cooperative DNA binding; instead, dimerization and DNA binding are mutually exclusive activities. In particular, we show that Msx and Dlx proteins interact independently and noncooperatively with homeodomain DNA binding sites and that dimerization is specifically blocked by the presence of such DNA sites. We further demonstrate that the transcriptional properties of Msx and Dlx proteins display reciprocal inhibition. Specifically, Msx proteins act as transcriptional repressors and Dlx proteins act as activators, while in combination, Msx and Dlx proteins counteract each other's transcriptional activities. Finally, we show that the expression patterns of representative Msx and Dlx genes (Msx1, Msx2, Dlx2, and Dlx5) overlap in mouse embryogenesis during limb bud and craniofacial development, consistent with the potential for their protein products to interact in vivo. Based on these observations, we propose that functional antagonism through heterodimer formation provides a mechanism for regulating the transcriptional actions of Msx and Dlx homeoproteins in vivo.

HRI usability evaluation of interaction modes for a teleoperated agricultural robotic sprayer.

PubMed

Adamides, George; Katsanos, Christos; Parmet, Yisrael; Christou, Georgios; Xenos, Michalis; Hadzilacos, Thanasis; Edan, Yael

2017-07-01

Teleoperation of an agricultural robotic system requires effective and efficient human-robot interaction. This paper investigates the usability of different interaction modes for agricultural robot teleoperation. Specifically, we examined the overall influence of two types of output devices (PC screen, head mounted display), two types of peripheral vision support mechanisms (single view, multiple views), and two types of control input devices (PC keyboard, PS3 gamepad) on observed and perceived usability of a teleoperated agricultural sprayer. A modular user interface for teleoperating an agricultural robot sprayer was constructed and field-tested. Evaluation included eight interaction modes: the different combinations of the 3 factors. Thirty representative participants used each interaction mode to navigate the robot along a vineyard and spray grape clusters based on a 2 × 2 × 2 repeated measures experimental design. Objective metrics of the effectiveness and efficiency of the human-robot collaboration were collected. Participants also completed questionnaires related to their user experience with the system in each interaction mode. Results show that the most important factor for human-robot interface usability is the number and placement of views. The type of robot control input device was also a significant factor in certain dependents, whereas the effect of the screen output type was only significant on the participants' perceived workload index. Specific recommendations for mobile field robot teleoperation to improve HRI awareness for the agricultural spraying task are presented. Copyright © 2017 Elsevier Ltd. All rights reserved.

Direct interaction of the bacteriophage SPP1 packaging ATPase with the portal protein.

PubMed

Oliveira, Leonor; Cuervo, Ana; Tavares, Paulo

2010-03-05

DNA packaging in tailed bacteriophages and other viruses requires assembly of a complex molecular machine at a specific vertex of the procapsid. This machine is composed of the portal protein that provides a tunnel for DNA entry, an ATPase that fuels DNA translocation (large terminase subunit), and most frequently, a small terminase subunit. Here we characterized the interaction between the terminase ATPase subunit of bacteriophage SPP1 (gp2) and the procapsid portal vertex. We found, by affinity pulldown assays with purified proteins, that gp2 interacts with the portal protein, gp6, independently of the terminase small subunit gp1, DNA, or ATP. The gp2-procapsid interaction via the portal protein depends on gp2 concentration and requires the presence of divalent cations. Competition experiments showed that isolated gp6 can only inhibit gp2-procapsid interactions and DNA packaging at gp6:procapsid molar ratios above 10-fold. Assays with gp6 carrying mutations in distinct regions of its structure that affect the portal-induced stimulation of ATPase and DNA packaging revealed that none of these mutations impedes gp2-gp6 binding. Our results demonstrate that the SPP1 packaging ATPase binds directly to the portal and that the interaction is stronger with the portal embedded in procapsids. Identification of mutations in gp6 that allow for assembly of the ATPase-portal complex but impair DNA packaging support an intricate cross-talk between the two proteins for activity of the DNA translocation motor.

The 727 approach energy management system avionics specification (preliminary)

NASA Technical Reports Server (NTRS)

Jackson, D. O.; Lambregts, A. A.

1976-01-01

Hardware and software requirements for an Approach Energy Management System (AEMS) consisting of an airborne digital computer and cockpit displays are presented. The displays provide the pilot with a visual indication of when to manually operate the gear, flaps, and throttles during a delayed flap approach so as to reduce approach time, fuel consumption, and community noise. The AEMS is an independent system that does not interact with other navigation or control systems, and is compatible with manually flown or autopilot coupled approaches. Operational use of the AEMS requires a DME ground station colocated with the flight path reference.

The Ames Virtual Environment Workstation: Implementation issues and requirements

NASA Technical Reports Server (NTRS)

Fisher, Scott S.; Jacoby, R.; Bryson, S.; Stone, P.; Mcdowall, I.; Bolas, M.; Dasaro, D.; Wenzel, Elizabeth M.; Coler, C.; Kerr, D.

1991-01-01

This presentation describes recent developments in the implementation of a virtual environment workstation in the Aerospace Human Factors Research Division of NASA's Ames Research Center. Introductory discussions are presented on the primary research objectives and applications of the system and on the system's current hardware and software configuration. Principle attention is then focused on unique issues and problems encountered in the workstation's development with emphasis on its ability to meet original design specifications for computational graphics performance and for associated human factors requirements necessary to provide compelling sense of presence and efficient interaction in the virtual environment.

Allosteric properties of hemoglobin and the plasma membrane of the erythrocyte: new insights in gas transport and metabolic modulation.

PubMed

De Rosa, Maria Cristina; Carelli Alinovi, Cristiana; Galtieri, Antonio; Russo, Annamaria; Giardina, Bruno

2008-02-01

Within the red blood cell the hemoglobin molecule is subjected to modulation mechanisms, namely homo- and heterotropic interactions, which optimize its functional behavior to the specific physiological requirements. At the cellular level, these modulation mechanisms are utilized to perform a number of other functions that are not minor with respect to the basic function of oxygen transport. Here we report some key examples concerning: (i) the interaction of hemoglobin with band 3 and its influence on glucose metabolism; (ii) the role of the ligand-linked quaternary transition of hemoglobin in the control of "NO bioactivity" and of gas diffusion; (iii) the interaction of plasma membrane with the various oxidative derivatives of the hemoglobin molecule. (c) 2008 IUBMB.

Structural insight into the interaction of proteins containing NPF, DPF, and GPF motifs with the C-terminal EH-domain of EHD1

PubMed Central

Kieken, Fabien; Jović, Marko; Tonelli, Marco; Naslavsky, Naava; Caplan, Steve; Sorgen, Paul L

2009-01-01

Eps15 homology (EH)-domain containing proteins are regulators of endocytic membrane trafficking. EH-domain binding to proteins containing the tripeptide NPF has been well characterized, but recent studies have shown that EH-domains are also able to interact with ligands containing DPF or GPF motifs. We demonstrate that the three motifs interact in a similar way with the EH-domain of EHD1, with the NPF motif having the highest affinity due to the presence of an intermolecular hydrogen bond. The weaker affinity for the DPF and GPF motifs suggests that if complex formation occurs in vivo, they may require high ligand concentrations, the presence of successive motifs and/or specific flanking residues. PMID:19798736

Design of interactive health drama built on social realism.

PubMed

Graspemo, Gabriella; Hassling, Linda; Nordfeldt, Sam; Eriksson, Hendrik; Timpka, Toomas

2004-01-01

There are many psychosocial aspects of chronic diseases, such as diabetes. Educational multimedia can support patients with chronic diseases and their families by communicating narratives based on social realism. The production of such socio-realistic interactive health dramas requires systematic methods, especially for the identification of significant stories. The aim of this study is to explore the use of self-documentary video in the design of an Interactive Health Drama environment to support diabetic adolescents and their families. In particular, the potential of the self-documentary video for story development in combination with qualitative interviews were explored. The proposed approach, when further developed, is intended to enable all types of chronic disease patients to work with their specific psychosocial problems in a supportive and stimulating environment adapted to their personality and preferences.

Complete analysis of steady and transient missile aerodynamic/propulsive/plume flowfield interactions

NASA Astrophysics Data System (ADS)

York, B. J.; Sinha, N.; Dash, S. M.; Hosangadi, A.; Kenzakowski, D. C.; Lee, R. A.

1992-07-01

The analysis of steady and transient aerodynamic/propulsive/plume flowfield interactions utilizing several state-of-the-art computer codes (PARCH, CRAFT, and SCHAFT) is discussed. These codes have been extended to include advanced turbulence models, generalized thermochemistry, and multiphase nonequilibrium capabilities. Several specialized versions of these codes have been developed for specific applications. This paper presents a brief overview of these codes followed by selected cases demonstrating steady and transient analyses of conventional as well as advanced missile systems. Areas requiring upgrades include turbulence modeling in a highly compressible environment and the treatment of particulates in general. Recent progress in these areas are highlighted.

Meeting the Deadline: Why, When and How

NASA Technical Reports Server (NTRS)

Dignum, Frank; Broersen, Jan; Dignum, Virginia; Meyer, John-Jules

2004-01-01

A normative system is defined as any set of interacting agents whose behavior can usefully be regarded as norm-directed. Most organizations, and more specifically institutions, fall under this definition. Interactions in these normative systems are regulated by normative templates that describe desired behavior in terms of deontic concepts (obligations, prohibitions and permissions), deadlines, violations and sanctions. Agreements between agents, and between an agent and the society, can then be specified by means of contracts. Contracts provide flexible but verifiable means to integrate society requirements and agent autonomy. and are an adequate means for the explicit specification of interactions. From the society perspective, it is important that these contracts adhere to the specifications described in the model of the organization. If we want to automate such verifications, we have to formalize the languages used for contracts and for the specification of organizations. The logic LCR is based on deontic temporal logic. LCR is an expressive language for describing interaction in multi-agent systems, including obligations with deadlines. Deadlines are important norms in most interactions between agents. Intuitively, a deadline states that an agent should perform an action before a certain point in time. The obligation to perform the action starts at the moment the deadline becomes active. E.g. when a contract is signed or approved. If the action is not performed in time a violation of the deadline occurs. It can be specified independently what measure has to be taken in this case. In this paper we investigate the deadline concept in more detail. The paper is organized as follows. Section 2 defines the variant of CTL we use. In section 3, we discuss the basic intuitions of deadlines. Section 4 presents a first intuitive formalization for deadlines. In section 5, we look at a more complex model for deadlines trying to catch some more practical aspects. Finally, in section 6 we present issues for future work and our conciusions.

Plasmacytoid Dendritic Cells Require Direct Infection To Sustain the Pulmonary Influenza A Virus-Specific CD8 T Cell Response.

PubMed

Hemann, Emily A; Sjaastad, Louisa E; Langlois, Ryan A; Legge, Kevin L

2015-12-30

Following influenza A virus (IAV) infection, development of a robust IAV-specific CD8 T cell response is required for clearance of primary infection and enhances memory protection. Following IAV infection, plasmacytoid dendritic cells (pDC) or CD8α(+) DC regulate pulmonary effector CD8 T cell responses within the lung. Without this DC-T cell interaction, insufficient effector CD8 T cells are maintained in the lungs, leading to enhanced morbidity and mortality. Previous studies have demonstrated that pDC are capable of classical presentation or cross-presentation of IAV antigens and could potentially regulate IAV-specific CD8 T cell responses through either mechanism. Our results demonstrate that pDC from the lungs of donor mice infected with an IAV that is not able to replicate in hematopoietic cells (142t-IAV), unlike donor pDC isolated from the lungs of control infected mice, are not able to rescue the host IAV-specific CD8 T cell response from apoptosis. This indicates that pDC must utilize the direct presentation pathway for this rescue. This inability of pDC from 142t-IAV donors to rescue the IAV-specific CD8 T cell response is not due to differences in the overall ability of 142t-IAV to replicate within the lungs or generate defective viral genomes or to differences in levels of costimulatory molecules required for this interaction. We further demonstrate that bypassing the antigen presentation pathway by coating the 142t-IAV pDC with IAV peptide epitopes restores their ability to rescue the IAV-specific CD8 T cell response. IAV continues to be a global health burden, infecting 5 to 20% of the global population annually. Continued investigation into the mechanisms that mediate protective immune responses against IAV is important to improving current vaccination and antiviral strategies antagonistic toward IAV. Our findings presented herein demonstrate a key requirement for pDC promotion of effector CD8 T cell survival: that rather than utilizing cross-presentation, pDC must be infected and utilize the endogenous pathway for presentation of antigens to CD8 T cells during in vivo IAV infections. This suggests that targeting presentation via the endogenous pathway in pDC could be important for the development of unique antiviral cellular therapies. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Drug-device combination products: regulatory landscape and market growth.

PubMed

Bayarri, L

2015-08-01

Combination products are therapeutic and diagnostic products that combine drugs, devices and/or biological products, leading to safer and more effective treatments thanks to careful and precise drug targeting, local administration and individualized therapy. These technologies can especially benefit patients suffering from serious diseases and conditions such as cancer, heart disease, multiple sclerosis and diabetes, among others. On the other hand, drug-device combination products have also introduced a new dynamic in medical product development, regulatory approval and corporate interaction. Due to the increasing integration of drugs and devices observed in the latest generation of combination products, regulatory agencies have developed specific competences and regulations over the last decade. Manufacturers are required to fully understand the specific requirements in each country in order to ensure timely and accurate market access of new combination products, and the development of combination products involves a very specific pattern of interactions between manufacturers and regulatory agencies. The increased sophistication of the products brought to market over the last couple of decades has accentuated the need to develop drugs and devices collaboratively using resources from both industries, fostering the need of business partnering and technology licensing. This review will provide a global overview of the market trends, as well as (in the last section) an analysis of the drug-device combination products approved by the FDA during the latest 5 years. Copyright 2015 Prous Science, S.A.U. or its licensors. All rights reserved.

Structural Insights into the Assembly of the Adeno-associated Virus Type 2 Rep68 Protein on the Integration Site AAVS1*

PubMed Central

Musayev, Faik N.; Zarate-Perez, Francisco; Bishop, Clayton; Burgner, John W.; Escalante, Carlos R.

2015-01-01

Adeno-associated virus (AAV) is the only eukaryotic virus with the property of establishing latency by integrating site-specifically into the human genome. The integration site known as AAVS1 is located in chromosome 19 and contains multiple GCTC repeats that are recognized by the AAV non-structural Rep proteins. These proteins are multifunctional, with an N-terminal origin-binding domain (OBD) and a helicase domain joined together by a short linker. As a first step to understand the process of site-specific integration, we proceeded to characterize the recognition and assembly of Rep68 onto the AAVS1 site. We first determined the x-ray structure of AAV-2 Rep68 OBD in complex with the AAVS1 DNA site. Specificity is achieved through the interaction of a glycine-rich loop that binds the major groove and an α-helix that interacts with a downstream minor groove on the same face of the DNA. Although the structure shows a complex with three OBD molecules bound to the AAVS1 site, we show by using analytical centrifugation and electron microscopy that the full-length Rep68 forms a heptameric complex. Moreover, we determined that a minimum of two direct repeats is required to form a stable complex and to melt DNA. Finally, we show that although the individual domains bind DNA poorly, complex assembly requires oligomerization and cooperation between its OBD, helicase, and the linker domains. PMID:26370092

Developmental and transcriptional consequences of mutations in Drosophila TAF(II)60.

PubMed

Aoyagi, N; Wassarman, D A

2001-10-01

In vitro, the TAF(II)60 component of the TFIID complex contributes to RNA polymerase II transcription initiation by serving as a coactivator that interacts with specific activator proteins and possibly as a promoter selectivity factor that interacts with the downstream promoter element. In vivo roles for TAF(II)60 in metazoan transcription are not as clear. Here we have investigated the developmental and transcriptional requirements for TAF(II)60 by analyzing four independent Drosophila melanogaster TAF(II)60 mutants. Loss-of-function mutations in Drosophila TAF(II)60 result in lethality, indicating that TAF(II)60 provides a nonredundant function in vivo. Molecular analysis of TAF(II)60 alleles revealed that essential TAF(II)60 functions are provided by two evolutionarily conserved regions located in the N-terminal half of the protein. TAF(II)60 is required at all stages of Drosophila development, in both germ cells and somatic cells. Expression of TAF(II)60 from a transgene rescued the lethality of TAF(II)60 mutants and exposed requirements for TAF(II)60 during imaginal development, spermatogenesis, and oogenesis. Phenotypes of rescued TAF(II)60 mutant flies implicate TAF(II)60 in transcriptional mechanisms that regulate cell growth and cell fate specification and suggest that TAF(II)60 is a limiting component of the machinery that regulates the transcription of dosage-sensitive genes. Finally, TAF(II)60 plays roles in developmental regulation of gene expression that are distinct from those of other TAF(II) proteins.

Conserved sequence-specific lincRNA-steroid receptor interactions drive transcriptional repression and direct cell fate

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hudson, William H.; Pickard, Mark R.; de Vera, Ian Mitchelle S.

2014-12-23

The majority of the eukaryotic genome is transcribed, generating a significant number of long intergenic noncoding RNAs (lincRNAs). Although lincRNAs represent the most poorly understood product of transcription, recent work has shown lincRNAs fulfill important cellular functions. In addition to low sequence conservation, poor understanding of structural mechanisms driving lincRNA biology hinders systematic prediction of their function. Here we report the molecular requirements for the recognition of steroid receptors (SRs) by the lincRNA growth arrest-specific 5 (Gas5), which regulates steroid-mediated transcriptional regulation, growth arrest and apoptosis. We identify the functional Gas5-SR interface and generate point mutations that ablate the SR-Gas5more » lincRNA interaction, altering Gas5-driven apoptosis in cancer cell lines. Further, we find that the Gas5 SR-recognition sequence is conserved among haplorhines, with its evolutionary origin as a splice acceptor site. This study demonstrates that lincRNAs can recognize protein targets in a conserved, sequence-specific manner in order to affect critical cell functions.« less

A FYVE zinc finger domain protein specifically links mRNA transport to endosome trafficking

PubMed Central

Pohlmann, Thomas; Baumann, Sebastian; Haag, Carl; Albrecht, Mario; Feldbrügge, Michael

2015-01-01

An emerging theme in cellular logistics is the close connection between mRNA and membrane trafficking. A prominent example is the microtubule-dependent transport of mRNAs and associated ribosomes on endosomes. This coordinated process is crucial for correct septin filamentation and efficient growth of polarised cells, such as fungal hyphae. Despite detailed knowledge on the key RNA-binding protein and the molecular motors involved, it is unclear how mRNAs are connected to membranes during transport. Here, we identify a novel factor containing a FYVE zinc finger domain for interaction with endosomal lipids and a new PAM2-like domain required for interaction with the MLLE domain of the key RNA-binding protein. Consistently, loss of this FYVE domain protein leads to specific defects in mRNA, ribosome, and septin transport without affecting general functions of endosomes or their movement. Hence, this is the first endosomal component specific for mRNP trafficking uncovering a new mechanism to couple mRNPs to endosomes. DOI: http://dx.doi.org/10.7554/eLife.06041.001 PMID:25985087

Single Molecule Junctions: A Laboratory for Chemistry, Mechanics and Bond Rupture

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hybertsen M. S.

Simultaneous measurement [1] of junction conductance and sustained force in single molecule junctions bridging metal electrodes provides a powerful tool in the quantitative study of the character of molecule-metal bonds. In this talk I will discuss three topics. First, I will describe chemical trends in link bond strength based on experiments and Density Functional Theory based calculations. Second, I will focus on the specific case of pyridine-linked junctions. Bond rupture from the high conductance junction structure shows a requires a force that exceeds the rupture force of gold point contacts and clearly indicates the role of additional forces, beyond themore » specific N-Au donor acceptor bond. DFT-D2 calculations with empirical addition of dispersion interactions illustrates the interplay between the donor-acceptor bonding and the non-specific van der Waals interactions between the pyridine rings and Au asperities. Third, I will describe recent efforts to characterize the diversity of junction structures realized in break-junction experiments with suitable models for the potential surfaces that are observed. [1] Venkataraman Group, Columbia University.« less

Long-range population dynamics of anatomically defined neocortical networks

PubMed Central

Chen, Jerry L; Voigt, Fabian F; Javadzadeh, Mitra; Krueppel, Roland; Helmchen, Fritjof

2016-01-01

The coordination of activity across neocortical areas is essential for mammalian brain function. Understanding this process requires simultaneous functional measurements across the cortex. In order to dissociate direct cortico-cortical interactions from other sources of neuronal correlations, it is furthermore desirable to target cross-areal recordings to neuronal subpopulations that anatomically project between areas. Here, we combined anatomical tracers with a novel multi-area two-photon microscope to perform simultaneous calcium imaging across mouse primary (S1) and secondary (S2) somatosensory whisker cortex during texture discrimination behavior, specifically identifying feedforward and feedback neurons. We find that coordination of S1-S2 activity increases during motor behaviors such as goal-directed whisking and licking. This effect was not specific to identified feedforward and feedback neurons. However, these mutually projecting neurons especially participated in inter-areal coordination when motor behavior was paired with whisker-texture touches, suggesting that direct S1-S2 interactions are sensory-dependent. Our results demonstrate specific functional coordination of anatomically-identified projection neurons across sensory cortices. DOI: http://dx.doi.org/10.7554/eLife.14679.001 PMID:27218452

Obscurin is required for ankyrinB-dependent dystrophin localization and sarcolemma integrity

PubMed Central

Randazzo, Davide; Giacomello, Emiliana; Lorenzini, Stefania; Rossi, Daniela; Pierantozzi, Enrico; Blaauw, Bert; Reggiani, Carlo; Lange, Stephan; Peter, Angela K.; Chen, Ju

2013-01-01

Obscurin is a large myofibrillar protein that contains several interacting modules, one of which mediates binding to muscle-specific ankyrins. Interaction between obscurin and the muscle-specific ankyrin sAnk1.5 regulates the organization of the sarcoplasmic reticulum in striated muscles. Additional muscle-specific ankyrin isoforms, ankB and ankG, are localized at the subsarcolemma level, at which they contribute to the organization of dystrophin and β-dystroglycan at costameres. In this paper, we report that in mice deficient for obscurin, ankB was displaced from its localization at the M band, whereas localization of ankG at the Z disk was not affected. In obscurin KO mice, localization at costameres of dystrophin, but not of β-dystroglycan, was altered, and the subsarcolemma microtubule cytoskeleton was disrupted. In addition, these mutant mice displayed marked sarcolemmal fragility and reduced muscle exercise tolerance. Altogether, the results support a model in which obscurin, by targeting ankB at the M band, contributes to the organization of subsarcolemma microtubules, localization of dystrophin at costameres, and maintenance of sarcolemmal integrity. PMID:23420875

Yersinia pestis targets neutrophils via complement receptor 3

PubMed Central

Merritt, Peter M.; Nero, Thomas; Bohman, Lesley; Felek, Suleyman; Krukonis, Eric S.; Marketon, Melanie M.

2015-01-01

Yersinia species display a tropism for lymphoid tissues during infection, and the bacteria select innate immune cells for delivery of cytotoxic effectors by the type III secretion system. Yet the mechanism for target cell selection remains a mystery. Here we investigate the interaction of Yersinia pestis with murine splenocytes to identify factors that participate in the targeting process. We find that interactions with primary immune cells rely on multiple factors. First, the bacterial adhesin Ail is required for efficient targeting of neutrophils in vivo. However, Ail does not appear to directly mediate binding to a specific cell type. Instead, we find that host serum factors direct Y. pestis to specific innate immune cells, particularly neutrophils. Importantly, specificity towards neutrophils was increased in the absence of bacterial adhesins due to reduced targeting of other cell types, but this phenotype was only visible in the presence of mouse serum. Addition of antibodies against complement receptor 3 and CD14 blocked target cell selection, suggesting that a combination of host factors participate in steering bacteria toward neutrophils during plague infection. PMID:25359083

PROTEIN TARGETING TO STARCH Is Required for Localising GRANULE-BOUND STARCH SYNTHASE to Starch Granules and for Normal Amylose Synthesis in Arabidopsis

PubMed Central

Seung, David; Soyk, Sebastian; Coiro, Mario; Maier, Benjamin A.; Eicke, Simona; Zeeman, Samuel C.

2015-01-01

The domestication of starch crops underpinned the development of human civilisation, yet we still do not fully understand how plants make starch. Starch is composed of glucose polymers that are branched (amylopectin) or linear (amylose). The amount of amylose strongly influences the physico-chemical behaviour of starchy foods during cooking and of starch mixtures in non-food manufacturing processes. The GRANULE-BOUND STARCH SYNTHASE (GBSS) is the glucosyltransferase specifically responsible for elongating amylose polymers and was the only protein known to be required for its biosynthesis. Here, we demonstrate that PROTEIN TARGETING TO STARCH (PTST) is also specifically required for amylose synthesis in Arabidopsis. PTST is a plastidial protein possessing an N-terminal coiled coil domain and a C-terminal carbohydrate binding module (CBM). We discovered that Arabidopsis ptst mutants synthesise amylose-free starch and are phenotypically similar to mutants lacking GBSS. Analysis of granule-bound proteins showed a dramatic reduction of GBSS protein in ptst mutant starch granules. Pull-down assays with recombinant proteins in vitro, as well as immunoprecipitation assays in planta, revealed that GBSS physically interacts with PTST via a coiled coil. Furthermore, we show that the CBM domain of PTST, which mediates its interaction with starch granules, is also required for correct GBSS localisation. Fluorescently tagged Arabidopsis GBSS, expressed either in tobacco or Arabidopsis leaves, required the presence of Arabidopsis PTST to localise to starch granules. Mutation of the CBM of PTST caused GBSS to remain in the plastid stroma. PTST fulfils a previously unknown function in targeting GBSS to starch. This sheds new light on the importance of targeting biosynthetic enzymes to sub-cellular sites where their action is required. Importantly, PTST represents a promising new gene target for the biotechnological modification of starch composition, as it is exclusively involved in amylose synthesis. PMID:25710501

Meis3 synergizes with Pbx4 and Hoxb1b in promoting hindbrain fates in the zebrafish.

PubMed

Vlachakis, N; Choe, S K; Sagerström, C G

2001-04-01

Many Hox proteins are thought to require Pbx and Meis co-factors to specify cell identity during embryogenesis. Here we demonstrate that Meis3 synergizes with Pbx4 and Hoxb1b in promoting hindbrain fates in the zebrafish. We find that Hoxb1b and Pbx4 act together to induce ectopic hoxb1a expression in rhombomere 2 of the hindbrain. In contrast, Hoxb1b and Pbx4 acting together with Meis3 induce hoxb1a, hoxb2, krox20 and valentino expression rostrally and cause extensive transformation of forebrain and midbrain fates to hindbrain fates, including differentiation of excess rhombomere 4-specific Mauthner neurons. This synergistic effect requires that Hoxb1b and Meis3 have intact Pbx-interaction domains, suggesting that their in vivo activity is dependent on binding to Pbx4. In the case of Meis3, binding to Pbx4 is also required for nuclear access. Our results are consistent with Hoxb1b and Meis3 interacting with Pbx4 to form complexes that regulate hindbrain development during zebrafish embryogenesis.

Med-psych units. Financial viability and quality assurance.

PubMed

Fogel, B S

1989-01-01

Although medical-psychiatric units may have unique advantages for treating patients with combined medical and psychiatric illness, they may be costly to run, and their success requires a sound financial basis. This begins with filling beds and instituting a waiting list, and then setting admission priorities to regulate case mix deliberately to address financial as well as ethical and clinical considerations. Development of short-stay geropsychiatric evaluation services may offset financial problems associated with long stays of elderly patients requiring definitive treatment for complex conditions. Data are presented to show the effectiveness of deliberate regulation of case mix. Regarding quality assurance, key issues include maintaining documentation to meet HCFA standards for DRG exemption, and effectively integrating physical and psychiatric care, with a special focus on drug interactions and psychiatric toxicities of medical drugs. Effective multidisciplinary treatment planning meeting help in this effort, as do periodic walking rounds focusing specifically on pharmacologic issues. Denials of payment by third parties are most likely to be a problem when both the medical and the psychiatric illness are subacute but their interaction requires conjoint inpatient treatment. Prospective work with PROs can minimize retrospective denials.

Poverty, AIDS and child health: identifying highest-risk children in South Africa.

PubMed

Cluver, Lucie; Boyes, Mark; Orkin, Mark; Sherr, Lorraine

2013-10-11

Identifying children at the highest risk of negative health effects is a prerequisite to effective public health policies in Southern Africa. A central ongoing debate is whether poverty, orphanhood or parental AIDS most reliably indicates child health risks. Attempts to address this key question have been constrained by a lack of data allowing distinction of AIDS-specific parental death or morbidity from other causes of orphanhood and chronic illness. To examine whether household poverty, orphanhood and parental illness (by AIDS or other causes) independently or interactively predict child health, developmental and HIV-infection risks. We interviewed 6 002 children aged 10 - 17 years in 2009 - 2011, using stratified random sampling in six urban and rural sites across three South African provinces. Outcomes were child mental health risks, educational risks and HIV-infection risks. Regression models that controlled for socio-demographic co-factors tested potential impacts and interactions of poverty, AIDS-specific and other orphanhood and parental illness status. Household poverty independently predicted child mental health and educational risks, AIDS orphanhood independently predicted mental health risks and parental AIDS illness independently predicted mental health, educational and HIV-infection risks. Interaction effects of poverty with AIDS orphanhood and parental AIDS illness were found across all outcomes. No effects, or interactions with poverty, were shown by AIDS-unrelated orphanhood or parental illness. The identification of children at highest risk requires recognition and measurement of both poverty and parental AIDS. This study shows negative impacts of poverty and AIDS-specific vulnerabilities distinct from orphanhood and adult illness more generally. Additionally, effects of interaction between family AIDS and poverty suggest that, where these co-exist, children are at highest risk of all.

Functional implications of an intermeshing cogwheel-like interaction between TolC and MacA in the action of macrolide-specific efflux pump MacAB-TolC.

PubMed

Xu, Yongbin; Song, Saemee; Moeller, Arne; Kim, Nahee; Piao, Shunfu; Sim, Se-Hoon; Kang, Mooseok; Yu, Wookyung; Cho, Hyun-Soo; Chang, Iksoo; Lee, Kangseok; Ha, Nam-Chul

2011-04-15

Macrolide-specific efflux pump MacAB-TolC has been identified in diverse gram-negative bacteria including Escherichia coli. The inner membrane transporter MacB requires the outer membrane factor TolC and the periplasmic adaptor protein MacA to form a functional tripartite complex. In this study, we used a chimeric protein containing the tip region of the TolC α-barrel to investigate the role of the TolC α-barrel tip region with regard to its interaction with MacA. The chimeric protein formed a stable complex with MacA, and the complex formation was abolished by substitution at the functionally essential residues located at the MacA α-helical tip region. Electron microscopic study delineated that this complex was made by tip-to-tip interaction between the tip regions of the α-barrels of TolC and MacA, which correlated well with the TolC and MacA complex calculated by molecular dynamics. Taken together, our results demonstrate that the MacA hexamer interacts with TolC in a tip-to-tip manner, and implies the manner by which MacA induces opening of the TolC channel.

Functional Implications of an Intermeshing Cogwheel-like Interaction between TolC and MacA in the Action of Macrolide-specific Efflux Pump MacAB-TolC*

PubMed Central

Xu, Yongbin; Song, Saemee; Moeller, Arne; Kim, Nahee; Piao, Shunfu; Sim, Se-Hoon; Kang, Mooseok; Yu, Wookyung; Cho, Hyun-Soo; Chang, Iksoo; Lee, Kangseok; Ha, Nam-Chul

2011-01-01

Macrolide-specific efflux pump MacAB-TolC has been identified in diverse Gram-negative bacteria including Escherichia coli. The inner membrane transporter MacB requires the outer membrane factor TolC and the periplasmic adaptor protein MacA to form a functional tripartite complex. In this study, we used a chimeric protein containing the tip region of the TolC α-barrel to investigate the role of the TolC α-barrel tip region with regard to its interaction with MacA. The chimeric protein formed a stable complex with MacA, and the complex formation was abolished by substitution at the functionally essential residues located at the MacA α-helical tip region. Electron microscopic study delineated that this complex was made by tip-to-tip interaction between the tip regions of the α-barrels of TolC and MacA, which correlated well with the TolC and MacA complex calculated by molecular dynamics. Taken together, our results demonstrate that the MacA hexamer interacts with TolC in a tip-to-tip manner, and implies the manner by which MacA induces opening of the TolC channel. PMID:21325274

The Binding Mode Prediction and Similar Ligand Potency in the Active Site of Vitamin D Receptor with QM/MM Interaction, MESP, and MD Simulation.

PubMed

Selvaraman, Nagamani; Selvam, Saravana Kumar; Muthusamy, Karthikeyan

2016-08-01

Non-secosteroidal ligands are well-known vitamin D receptor (VDR) agonists. In this study, we described a combined QM/MM to define the protein-ligand interaction energy a strong positive correlation in both QM-MM interaction energy and binding free energy against the biological activity. The molecular dynamics simulation study was performed, and specific interactions were extensively studied. The molecular docking results and surface analysis shed light on steric and electrostatic complementarities of these non-secosteroidal ligands to VDR. Finally, the drug likeness properties were also calculated and found within the acceptable range. The results show that bulky group substitutions in side chain decrease the VDR activity, whereas a small substitution increased it. Functional analyses of H393A and H301A mutations substantiate their roles in the VDR agonistic and antagonistic activities. Apart from the His393 and His301, two other amino acids in the hinge region viz. Ser233 and Arg270 acted as an electron donor/acceptor specific to the agonist in the distinct ligand potency. The results from this study disclose the binding mechanism of VDR agonists and structural modifications required to improve the selectivity. © 2016 John Wiley & Sons A/S.

Functional relevance of G-protein-coupled-receptor-associated proteins, exemplified by receptor-activity-modifying proteins (RAMPs).

PubMed

Fischer, J A; Muff, R; Born, W

2002-08-01

The calcitonin (CT) receptor (CTR) and the CTR-like receptor (CRLR) are close relatives within the type II family of G-protein-coupled receptors, demonstrating sequence identity of 50%. Unlike the interaction between CT and CTR, receptors for the related hormones and neuropeptides amylin, CT-gene-related peptide (CGRP) and adrenomedullin (AM) require one of three accessory receptor-activity-modifying proteins (RAMPs) for ligand recognition. An amylin/CGRP receptor is revealed when CTR is co-expressed with RAMP1. When complexed with RAMP3, CTR interacts with amylin alone. CRLR, initially classed as an orphan receptor, is a CGRP receptor when co-expressed with RAMP1. The same receptor is specific for AM in the presence of RAMP2. Together with human RAMP3, CRLR defines an AM receptor, and with mouse RAMP3 it is a low-affinity CGRP/AM receptor. CTR-RAMP1, antagonized preferentially by salmon CT-(8-32) and not by CGRP-(8-37), and CRLR-RAMP1, antagonized by CGRP-(8-37), are two CGRP receptor isotypes. Thus amylin and CGRP interact specifically with heterodimeric complexes between CTR and RAMP1 or RAMP3, and CGRP and AM interact with complexes between CRLR and RAMP1, RAMP2 or RAMP3.

Characterization of the Grp94/OS-9 chaperone-lectin complex

PubMed Central

Seidler, Paul M.; Shinsky, Stephen A.; Hong, Feng; Li, Zihai; Cosgrove, Michael S.; Gewirth, Daniel T.

2014-01-01

Grp94 is a macromolecular chaperone belonging to the hsp90 family and is the most abundant glycoprotein in the endoplasmic reticulum of mammals. In addition to its essential role in protein folding, Grp94 was proposed to participate in the ER associated degradation (ERAD) quality control pathway by interacting with the lectin OS-9, a sensor for terminally misfolded proteins (TMPs). To understand how OS-9 interacts with ER chaperone proteins, we mapped its interaction with Grp94. Glycosylation of the full length Grp94 protein was essential for OS-9 binding, although deletion of the Grp94 N-terminal domain relieved this requirement suggesting that the effect was allosteric rather than direct. Although yeast OS-9 is composed of a well-established N-terminal MRH lectin domain and a C-terminal dimerization domain, we find that the C-terminal domain of OS-9 in higher eukaryotes contains ‘mammalian-specific insets’ that are specifically recognized by the middle and C-terminal domains of Grp94. Additionally, the Grp94 binding domain in OS-9 was found to be intrinsically disordered. The biochemical analysis of the interacting regions provides insight into the manner by which the two associate, and additionally hints at a plausible biological role for the Grp94/OS-9 complex. PMID:25193139

Challenges and dreams: physics of weak interactions essential to life.

PubMed

Chien, Peter; Gierasch, Lila M

2014-11-05

Biological systems display stunning capacities to self-organize. Moreover, their subcellular architectures are dynamic and responsive to changing needs and conditions. Key to these properties are manifold weak "quinary" interactions that have evolved to create specific spatial networks of macromolecules. These specific arrangements of molecules enable signals to be propagated over distances much greater than molecular dimensions, create phase separations that define functional regions in cells, and amplify cellular responses to changes in their environments. A major challenge is to develop biochemical tools and physical models to describe the panoply of weak interactions operating in cells. We also need better approaches to measure the biases in the spatial distributions of cellular macromolecules that result from the integrated action of multiple weak interactions. Partnerships between cell biologists, biochemists, and physicists are required to deploy these methods. Together these approaches will help us realize the dream of understanding the biological "glue" that sustains life at a molecular and cellular level. © 2014 Chien and Gierasch. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

DOE Office of Scientific and Technical Information (OSTI.GOV)

Son, Ora; Kim, Sunghan; Hur, Yoon-Sun

TOR (target of rapamycin) kinase signaling plays central role as a regulator of growth and proliferation in all eukaryotic cells and its key signaling components and effectors are also conserved in plants. Unlike the mammalian and yeast counterparts, however, we found through yeast two-hybrid analysis that multiple regions of the Arabidopsis Raptor (regulatory associated protein of TOR) are required for binding to its substrate. We also identified that a 44-amino acid region at the N-terminal end of Arabidopsis ribosomal S6 kinase 1 (AtS6K1) specifically interacted with AtRaptor1, indicating that this region may contain a functional equivalent of the TOS (TOR-Signaling)more » motif present in the mammalian TOR substrates. Transient over-expression of this 44-amino acid fragment in Arabidopsis protoplasts resulted in significant decrease in rDNA transcription, demonstrating a feasibility of developing a new plant-specific TOR signaling inhibitor based upon perturbation of the Raptor-substrate interaction. - Highlights: • Multiple regions on the Arabidopsis Raptor protein were found to be involved in substrate binding. • N-terminal end of the Arabidopsis ribosomal S6 kinase 1 (AtS6K1) was responsible for interacting with AtRaptor1. • The Raptor-interacting fragment of AtS6K1 could be utilized as an effective inhibitor of plant TOR signaling.« less

Emotional valence and physical space: limits of interaction.

PubMed

de la Vega, Irmgard; de Filippis, Mónica; Lachmair, Martin; Dudschig, Carolin; Kaup, Barbara

2012-04-01

According to the body-specificity hypothesis, people associate positive things with the side of space that corresponds to their dominant hand and negative things with the side corresponding to their nondominant hand. Our aim was to find out whether this association holds also true for a response time study using linguistic stimuli, and whether such an association is activated automatically. Four experiments explored this association using positive and negative words. In Exp. 1, right-handers made a lexical judgment by pressing a left or right key. Attention was not explicitly drawn to the valence of the stimuli. No valence-by-side interaction emerged. In Exp. 2 and 3, right-handers and left-handers made a valence judgment by pressing a left or a right key. A valence-by-side interaction emerged: For positive words, responses were faster when participants responded with their dominant hand, whereas for negative words, responses were faster for the nondominant hand. Exp. 4 required a valence judgment without stating an explicit mapping of valence and side. No valence-by-side interaction emerged. The experiments provide evidence for an association between response side and valence, which, however, does not seem to be activated automatically but rather requires a task with an explicit response mapping to occur.

The role of gut microbiota in health and disease: In vitro modeling of host-microbe interactions at the aerobe-anaerobe interphase of the human gut.

PubMed

von Martels, Julius Z H; Sadaghian Sadabad, Mehdi; Bourgonje, Arno R; Blokzijl, Tjasso; Dijkstra, Gerard; Faber, Klaas Nico; Harmsen, Hermie J M

2017-04-01

The microbiota of the gut has many crucial functions in human health. Dysbiosis of the microbiota has been correlated to a large and still increasing number of diseases. Recent studies have mostly focused on analyzing the associations between disease and an aberrant microbiota composition. Functional studies using (in vitro) gut models are required to investigate the precise interactions that occur between specific bacteria (or bacterial mixtures) and gut epithelial cells. As most gut bacteria are obligate or facultative anaerobes, studying their effect on oxygen-requiring human gut epithelial cells is technically challenging. Still, several (anaerobic) bacterial-epithelial co-culture systems have recently been developed that mimic host-microbe interactions occurring in the human gut, including 1) the Transwell "apical anaerobic model of the intestinal epithelial barrier", 2) the Host-Microbiota Interaction (HMI) module, 3) the "Human oxygen-Bacteria anaerobic" (HoxBan) system, 4) the human gut-on-a-chip and 5) the HuMiX model. This review discusses the role of gut microbiota in health and disease and gives an overview of the characteristics and applications of these novel host-microbe co-culture systems. Copyright © 2017 Elsevier Ltd. All rights reserved.

Discriminating toxicant classes by mode of action. 1. (Eco)toxicity profiles.

PubMed

Nendza, Monika; Wenzel, Andrea

2006-05-01

Predictive toxicology, particularly quantitative structure-activity relationships (QSARs), require classification of chemicals by mode of action (MOA). MOA is, however, not a constant property of a compound but it varies between species and may change with concentration and duration of exposure. A battery of MOA-specific in-vitro and low-complexity assays, featuring biomolecular targets for major classes of environmental pollutants, provides characteristic responses for (1.) classification of chemicals by MOA, (2.) identification of (eco)toxicity profiles of chemicals, (3.) identification of chemicals with specific MOAs, (4.) indication of most sensitive species, (5.) identification of chemicals that are outliers in QSARs and (6.) selection of appropriate QSARs for predictions. Chemicals covering nine distinct modes of toxic action (non-polar non-specific toxicants (n=14), polar non-specific toxicants (n=18), uncouplers of oxidative phosphorylation (n=25), inhibitors of photosynthesis (n=15), inhibitors of acetylcholinesterase (n=14), inhibitors of respiration (n=3), thiol-alkylating agents (n=9), reactives (irritants) (n=8), estrogen receptor agonists (n=9)) were tested for cytotoxicity in the neutralred assay, oxygen consumption in isolated mitochondria, oxygen production in algae, inhibition of AChE, reaction with GSH and activity in the yeast estrogen receptor assay. Data on in-vivo aquatic toxicity (LC50, EC50) towards fish, daphnids, algae and bacteria were collected from the literature for reasons of comparison and reference scaling. In the MOA-specific in-vitro test battery, most test chemicals are specifically active at low concentrations, though multiple effects do occur. Graphical and statistical evaluation of the individual classes versus MOA 1 (non-polar non-specific toxicants) identifies interactions related to predominant MOA. Discriminant analyses (DA) on subsets of the data revealed correct classifications between 70% (in-vivo data) and >90% (in-vitro data). Functional similarity of chemical substances is defined in terms of their (eco)toxicity profiles. Within each MOA class, the compounds share some properties related to the rate-limiting interactions, e.g., steric fit to the target site and/or reactivity with target biomolecules, revealing a specific pattern (fingerprint) of characteristic effects. The successful discrimination of toxicant classes by MOA is based on comprehensive characterization of test chemicals' properties related to interactions with target sites. The suite of aquatic in-vivo tests using fish, daphnids, algae and bacteria covers most acute effects, whilst long-term (latent) impacts are generally neglected. With the MOA-specific in-vitro test battery such distinctions are futile, because it focuses on isolated targets, i.e. it indicates the possible targets of a chemical regardless of the timescale of effects. The data analysis indicates that the in-vitro battery covers most effects in vivo and moreover provides additional aspects of the compounds' MOA. Translating in-vitro effects to in-vivo toxicity requires combining physiological and chemical knowledge about underlying processes. Comparison of the specific in-vitro effects of a compound with the respective sensitivities of aquatic organisms indicates particularly sensitive species. Classifications of toxicants by MOA based on physicochemical descriptors provides insight to interactions and directs to mechanistic QSARs.

A novel ICOS-independent, but CD28- and SAP-dependent, pathway of T cell-dependent, polysaccharide-specific humoral immunity in response to intact Streptococcus pneumoniae versus pneumococcal conjugate vaccine

PubMed Central

Chen, Quanyi; Cannons, Jennifer L.; Paton, James C.; Akiba, Hisaya; Schwartzberg, Pamela L.; Snapper, Clifford M.

2010-01-01

Polysaccharide (PS)- and protein-specific murine IgG responses to intact Streptococcus pneumoniae (Pn) are both dependent upon CD4+ T cell help, B7-dependent costimulation, and CD40/CD40-ligand interactions. However, the primary PS-, relative to protein-specific, IgG response terminates more rapidly, requires a shorter period of T cell help and B7-dependent costimulation, and fails to generate memory. In light of the critical role for ICOS/ICOS-ligand interactions in sustaining T cell-dependent Ig responses and promoting germinal center reactions, we hypothesized that this interaction was non-essential for PS-specific IgG responses to Pn. We now demonstrate that ICOS-/-, relative to WT, mice elicit a normal PS-specific IgG isotype response to Pn, despite marked inhibition of both the primary and secondary IgG anti-protein (i.e. PspA, PspC, and PsaA) response. A blocking anti-ICOS-ligand mAb injected during primary Pn immunization inhibits both the primary anti-protein response and the generation of protein-specific memory, but has no effect when injected during secondary immunization. In contrast to Pn, both PS- and protein-specific IgG responses to a pneumococcal conjugate vaccine are inhibited in ICOS-/- mice. ICOS-/- mice immunized with intact Pn or conjugate exhibit nearly complete abrogation in germinal center formation. Finally, although mice that lack the adaptor molecule SAP resemble ICOS-/- mice (and can exhibit decreased ICOS expression), we observe that the PS-, as well as protein-specific IgG responses to both Pn and conjugate are markedly defective in SAP-/- mice. These data define a novel T cell-, SAP-, and B7-dependent, but ICOS-independent, extrafollicular pathway of Ig induction. PMID:19050242

Modular biological function is most effectively captured by combining molecular interaction data types.

PubMed

Ames, Ryan M; Macpherson, Jamie I; Pinney, John W; Lovell, Simon C; Robertson, David L

2013-01-01

Large-scale molecular interaction data sets have the potential to provide a comprehensive, system-wide understanding of biological function. Although individual molecules can be promiscuous in terms of their contribution to function, molecular functions emerge from the specific interactions of molecules giving rise to modular organisation. As functions often derive from a range of mechanisms, we demonstrate that they are best studied using networks derived from different sources. Implementing a graph partitioning algorithm we identify subnetworks in yeast protein-protein interaction (PPI), genetic interaction and gene co-regulation networks. Among these subnetworks we identify cohesive subgraphs that we expect to represent functional modules in the different data types. We demonstrate significant overlap between the subgraphs generated from the different data types and show these overlaps can represent related functions as represented by the Gene Ontology (GO). Next, we investigate the correspondence between our subgraphs and the Gene Ontology. This revealed varying degrees of coverage of the biological process, molecular function and cellular component ontologies, dependent on the data type. For example, subgraphs from the PPI show enrichment for 84%, 58% and 93% of annotated GO terms, respectively. Integrating the interaction data into a combined network increases the coverage of GO. Furthermore, the different annotation types of GO are not predominantly associated with one of the interaction data types. Collectively our results demonstrate that successful capture of functional relationships by network data depends on both the specific biological function being characterised and the type of network data being used. We identify functions that require integrated information to be accurately represented, demonstrating the limitations of individual data types. Combining interaction subnetworks across data types is therefore essential for fully understanding the complex and emergent nature of biological function.

Fundamentals of Focused Ion Beam Nanostructural Processing: Below, At, and Above the Surface

DOE PAGES

MoberlyChan, Warren J.; Adams, David P.; Aziz, Michael J.; ...

2007-05-01

This paper considers the fundamentals of what happens in a solid when it is impacted by a medium-energy gallium ion. The study of the ion/sample interaction at the nanometer scale is applicable to most focused ion beam (FIB)–based work even if the FIB/sample interaction is only a step in the process, for example, micromachining or microelectronics device processing. Whereas the objective in other articles in this issue is to use the FIB tool to characterize a material or to machine a device or transmission electron microscopy sample, the goal of the FIB in this article is to have the FIB/samplemore » interaction itself become the product. To that end, the FIB/sample interaction is considered in three categories according to geometry: below, at, and above the surface. First, the FIB ions can penetrate the top atom layer(s) and interact below the surface. Ion implantation and ion damage on flat surfaces have been comprehensively examined; however, FIB applications require the further investigation of high doses in three-dimensional profiles. Second, the ions can interact at the surface, where a morphological instability can lead to ripples and surface self-organization, which can depend on boundary conditions for site-specific and compound FIB processing. Third, the FIB may interact above the surface (and/or produce secondary particles that interact above the surface). Such ion beam–assisted deposition, FIB–CVD (chemical vapor deposition), offers an elaborate complexity in three dimensions with an FIB using a gas injection system. Finally, at the nanometer scale, these three regimes—below, at, and above the surface—can require an interdependent understanding to be judiciously controlled by the FIB.« less

DnaA protein DNA-binding domain binds to Hda protein to promote inter-AAA+ domain interaction involved in regulatory inactivation of DnaA.

PubMed

Keyamura, Kenji; Katayama, Tsutomu

2011-08-19

Chromosomal replication is initiated from the replication origin oriC in Escherichia coli by the active ATP-bound form of DnaA protein. The regulatory inactivation of DnaA (RIDA) system, a complex of the ADP-bound Hda and the DNA-loaded replicase clamp, represses extra initiations by facilitating DnaA-bound ATP hydrolysis, yielding the inactive ADP-bound form of DnaA. However, the mechanisms involved in promoting the DnaA-Hda interaction have not been determined except for the involvement of an interaction between the AAA+ domains of the two. This study revealed that DnaA Leu-422 and Pro-423 residues within DnaA domain IV, including a typical DNA-binding HTH motif, are specifically required for RIDA-dependent ATP hydrolysis in vitro and that these residues support efficient interaction with the DNA-loaded clamp·Hda complex and with Hda in vitro. Consistently, substitutions of these residues caused accumulation of ATP-bound DnaA in vivo and oriC-dependent inhibition of cell growth. Leu-422 plays a more important role in these activities than Pro-423. By contrast, neither of these residues is crucial for DNA replication from oriC, although they are highly conserved in DnaA orthologues. Structural analysis of a DnaA·Hda complex model suggested that these residues make contact with residues in the vicinity of the Hda AAA+ sensor I that participates in formation of a nucleotide-interacting surface. Together, the results show that functional DnaA-Hda interactions require a second interaction site within DnaA domain IV in addition to the AAA+ domain and suggest that these interactions are crucial for the formation of RIDA complexes that are active for DnaA-ATP hydrolysis.

DnaA Protein DNA-binding Domain Binds to Hda Protein to Promote Inter-AAA+ Domain Interaction Involved in Regulatory Inactivation of DnaA*

PubMed Central

Keyamura, Kenji; Katayama, Tsutomu

2011-01-01

Chromosomal replication is initiated from the replication origin oriC in Escherichia coli by the active ATP-bound form of DnaA protein. The regulatory inactivation of DnaA (RIDA) system, a complex of the ADP-bound Hda and the DNA-loaded replicase clamp, represses extra initiations by facilitating DnaA-bound ATP hydrolysis, yielding the inactive ADP-bound form of DnaA. However, the mechanisms involved in promoting the DnaA-Hda interaction have not been determined except for the involvement of an interaction between the AAA+ domains of the two. This study revealed that DnaA Leu-422 and Pro-423 residues within DnaA domain IV, including a typical DNA-binding HTH motif, are specifically required for RIDA-dependent ATP hydrolysis in vitro and that these residues support efficient interaction with the DNA-loaded clamp·Hda complex and with Hda in vitro. Consistently, substitutions of these residues caused accumulation of ATP-bound DnaA in vivo and oriC-dependent inhibition of cell growth. Leu-422 plays a more important role in these activities than Pro-423. By contrast, neither of these residues is crucial for DNA replication from oriC, although they are highly conserved in DnaA orthologues. Structural analysis of a DnaA·Hda complex model suggested that these residues make contact with residues in the vicinity of the Hda AAA+ sensor I that participates in formation of a nucleotide-interacting surface. Together, the results show that functional DnaA-Hda interactions require a second interaction site within DnaA domain IV in addition to the AAA+ domain and suggest that these interactions are crucial for the formation of RIDA complexes that are active for DnaA-ATP hydrolysis. PMID:21708944

Cellular Protein WDR11 Interacts with Specific Herpes Simplex Virus Proteins at the trans-Golgi Network To Promote Virus Replication

PubMed Central

Taylor, Kathryne E.

2015-01-01

ABSTRACT It has recently been proposed that the herpes simplex virus (HSV) protein ICP0 has cytoplasmic roles in blocking antiviral signaling and in promoting viral replication in addition to its well-known proteasome-dependent functions in the nucleus. However, the mechanisms through which it produces these effects remain unclear. While investigating this further, we identified a novel cytoplasmic interaction between ICP0 and the poorly characterized cellular protein WDR11. During an HSV infection, WDR11 undergoes a dramatic change in localization at late times in the viral replication cycle, moving from defined perinuclear structures to a dispersed cytoplasmic distribution. While this relocation was not observed during infection with viruses other than HSV-1 and correlated with efficient HSV-1 replication, the redistribution was found to occur independently of ICP0 expression, instead requiring viral late gene expression. We demonstrate for the first time that WDR11 is localized to the trans-Golgi network (TGN), where it interacts specifically with some, but not all, HSV virion components, in addition to ICP0. Knockdown of WDR11 in cultured human cells resulted in a modest but consistent decrease in yields of both wild-type and ICP0-null viruses, in the supernatant and cell-associated fractions, without affecting viral gene expression. Although further study is required, we propose that WDR11 participates in viral assembly and/or secondary envelopment. IMPORTANCE While the TGN has been proposed to be the major site of HSV-1 secondary envelopment, this process is incompletely understood, and in particular, the role of cellular TGN components in this pathway is unknown. Additionally, little is known about the cellular functions of WDR11, although the disruption of this protein has been implicated in multiple human diseases. Therefore, our finding that WDR11 is a TGN-resident protein that interacts with specific viral proteins to enhance viral yields improves both our understanding of basic cellular biology as well as how this protein is co-opted by HSV. PMID:26178983

Study and selection of in vivo protein interactions by coupling bimolecular fluorescence complementation and flow cytometry.

PubMed

Morell, Montse; Espargaro, Alba; Aviles, Francesc Xavier; Ventura, Salvador

2008-01-01

We present a high-throughput approach to study weak protein-protein interactions by coupling bimolecular fluorescent complementation (BiFC) to flow cytometry (FC). In BiFC, the interaction partners (bait and prey) are fused to two rationally designed fragments of a fluorescent protein, which recovers its function upon the binding of the interacting proteins. For weak protein-protein interactions, the detected fluorescence is proportional to the interaction strength, thereby allowing in vivo discrimination between closely related binders with different affinity for the bait protein. FC provides a method for high-speed multiparametric data acquisition and analysis; the assay is simple, thousands of cells can be analyzed in seconds and, if required, selected using fluorescence-activated cell sorting (FACS). The combination of both methods (BiFC-FC) provides a technically straightforward, fast and highly sensitive method to validate weak protein interactions and to screen and identify optimal ligands in biologically synthesized libraries. Once plasmids encoding the protein fusions have been obtained, the evaluation of a specific interaction, the generation of a library and selection of active partners using BiFC-FC can be accomplished in 5 weeks.

Visualization of protein interactions in living Drosophila embryos by the bimolecular fluorescence complementation assay

PubMed Central

2011-01-01

Background Protein interactions control the regulatory networks underlying developmental processes. The understanding of developmental complexity will, therefore, require the characterization of protein interactions within their proper environment. The bimolecular fluorescence complementation (BiFC) technology offers this possibility as it enables the direct visualization of protein interactions in living cells. However, its potential has rarely been applied in embryos of animal model organisms and was only performed under transient protein expression levels. Results Using a Hox protein partnership as a test case, we investigated the suitability of BiFC for the study of protein interactions in the living Drosophila embryo. Importantly, all BiFC parameters were established with constructs that were stably expressed under the control of endogenous promoters. Under these physiological conditions, we showed that BiFC is specific and sensitive enough to analyse dynamic protein interactions. We next used BiFC in a candidate interaction screen, which led to the identification of several Hox protein partners. Conclusion Our results establish the general suitability of BiFC for revealing and studying protein interactions in their physiological context during the rapid course of Drosophila embryonic development. PMID:21276241

A preventive groupwork intervention with new immigrants to Israel.

PubMed

Roskin, M

1986-03-01

The act of immigration brings with it numerous major life changes and requires considerable social readjustment. Immigrants are, however, seldom offered specific training in how to effectively interact in a new culture. This paper describes an initial preventive groupwork effort to help English-speaking immigrants living in an Israeli absorption center to adjust successfully to life in a new culture. Suggestions for future preventive interventions with immigrants are considered.

Caring communications: how technology enhances interpersonal relations, Part II.

PubMed

Simpson, Roy L

2008-01-01

Part I of this 2-part series about technology's role in interpersonal communications examined how humans interact; proposed a caring theory of communication, collaboration, and conflict resolution; and delineated ways that technology--in general--supports this carative model of interpersonal relations. Part II will examine the barriers to adoption of carative technologies, describe the core capabilities required to overcome them, and discuss specific technologies that can support carative interpersonal relationships.

A Hypermedia Training Module for the Navy’s P-3C Armament System

DTIC Science & Technology

1993-07-01

procedures. Both aural , and visual cues are used throughout the program as necessary to alert the learner to specific items requiring his attention... learner the opportunity for a great deal of interactivity and feedback. The project is divided into five chapters including an introduction, review of the...literature, methodology, program description, and summary and conclusions. The literature review concentrates on the foliowing topics: adult learners

Detectors for Linear Colliders: Tracking and Vertexing (2/4)

ScienceCinema

Battaglia, Marco

2018-04-16

Efficient and precise determination of the flavour of partons in multi-hadron final states is essential to the anticipated LC physics program. This makes tracking in the vicinity of the interaction region of great importance. Tracking extrapolation and momentum resolution are specified by precise physics requirements. The R&D; towards detectors able to meet these specifications will be discussed, together with some of their application beyond particle physics.

Identification of Potential Therapeutic Mechanisms for HIP1 Inhibition in Breast Cancer

DTIC Science & Technology

2006-05-01

repressing transcription when knocked down using a silencing RNA approach and activating tran- scription when overexpressed. We have also identified a...HIP1, huntingtin interacting protein 1; PSA, prostate-specific antigen; siRNA, silencing RNA . The online version of this article includes supplemental...We examined whether endogenous HIP1 was also required to sustain AR transcriptional activity by taking a silencing RNA (siRNA) approach to knockdown