Independent Transport and Sorting of Functionally Distinct Protein Families in Tetrahymena thermophila Dense Core Secretory Granules▿ †

PubMed Central

Rahaman, Abdur; Miao, Wei; Turkewitz, Aaron P.

2009-01-01

Dense core granules (DCGs) in Tetrahymena thermophila contain two protein classes. Proteins in the first class, called granule lattice (Grl), coassemble to form a crystalline lattice within the granule lumen. Lattice expansion acts as a propulsive mechanism during DCG release, and Grl proteins are essential for efficient exocytosis. The second protein class, defined by a C-terminal β/γ-crystallin domain, is poorly understood. Here, we have analyzed the function and sorting of Grt1p (granule tip), which was previously identified as an abundant protein in this family. Cells lacking all copies of GRT1, together with the closely related GRT2, accumulate wild-type levels of docked DCGs. Unlike cells disrupted in any of the major GRL genes, ΔGRT1 ΔGRT2 cells show no defect in secretion, indicating that neither exocytic fusion nor core expansion depends on GRT1. These results suggest that Grl protein sorting to DCGs is independent of Grt proteins. Consistent with this, the granule core lattice in ΔGRT1 ΔGRT2 cells appears identical to that in wild-type cells by electron microscopy, and the only biochemical component visibly absent is Grt1p itself. Moreover, gel filtration showed that Grl and Grt proteins in cell homogenates exist in nonoverlapping complexes, and affinity-isolated Grt1p complexes do not contain Grl proteins. These data demonstrate that two major classes of proteins in Tetrahymena DCGs are likely to be independently transported during DCG biosynthesis and play distinct roles in granule function. The role of Grt1p may primarily be postexocytic; consistent with this idea, DCG contents from ΔGRT1 ΔGRT2 cells appear less adhesive than those from the wild type. PMID:19684282

Independent transport and sorting of functionally distinct protein families in Tetrahymena thermophila dense core secretory granules.

PubMed

Rahaman, Abdur; Miao, Wei; Turkewitz, Aaron P

2009-10-01

Dense core granules (DCGs) in Tetrahymena thermophila contain two protein classes. Proteins in the first class, called granule lattice (Grl), coassemble to form a crystalline lattice within the granule lumen. Lattice expansion acts as a propulsive mechanism during DCG release, and Grl proteins are essential for efficient exocytosis. The second protein class, defined by a C-terminal beta/gamma-crystallin domain, is poorly understood. Here, we have analyzed the function and sorting of Grt1p (granule tip), which was previously identified as an abundant protein in this family. Cells lacking all copies of GRT1, together with the closely related GRT2, accumulate wild-type levels of docked DCGs. Unlike cells disrupted in any of the major GRL genes, DeltaGRT1 DeltaGRT2 cells show no defect in secretion, indicating that neither exocytic fusion nor core expansion depends on GRT1. These results suggest that Grl protein sorting to DCGs is independent of Grt proteins. Consistent with this, the granule core lattice in DeltaGRT1 DeltaGRT2 cells appears identical to that in wild-type cells by electron microscopy, and the only biochemical component visibly absent is Grt1p itself. Moreover, gel filtration showed that Grl and Grt proteins in cell homogenates exist in nonoverlapping complexes, and affinity-isolated Grt1p complexes do not contain Grl proteins. These data demonstrate that two major classes of proteins in Tetrahymena DCGs are likely to be independently transported during DCG biosynthesis and play distinct roles in granule function. The role of Grt1p may primarily be postexocytic; consistent with this idea, DCG contents from DeltaGRT1 DeltaGRT2 cells appear less adhesive than those from the wild type.

Overlapping and Divergent Actions of Structurally Distinct Histone Deacetylase Inhibitors in Cardiac Fibroblasts

PubMed Central

Schuetze, Katherine B.; Stratton, Matthew S.; Blakeslee, Weston W.; Wempe, Michael F.; Wagner, Florence F.; Holson, Edward B.; Kuo, Yin-Ming; Andrews, Andrew J.; Gilbert, Tonya M.; Hooker, Jacob M.

2017-01-01

Inhibitors of zinc-dependent histone deacetylases (HDACs) profoundly affect cellular function by altering gene expression via changes in nucleosomal histone tail acetylation. Historically, investigators have employed pan-HDAC inhibitors, such as the hydroxamate trichostatin A (TSA), which simultaneously targets members of each of the three zinc-dependent HDAC classes (classes I, II, and IV). More recently, class- and isoform-selective HDAC inhibitors have been developed, providing invaluable chemical biology probes for dissecting the roles of distinct HDACs in the control of various physiologic and pathophysiological processes. For example, the benzamide class I HDAC-selective inhibitor, MGCD0103 [N-(2-aminophenyl)-4-[[(4-pyridin-3-ylpyrimidin-2-yl)amino]methyl] benzamide], was shown to block cardiac fibrosis, a process involving excess extracellular matrix deposition, which often results in heart dysfunction. Here, we compare the mechanisms of action of structurally distinct HDAC inhibitors in isolated primary cardiac fibroblasts, which are the major extracellular matrix–producing cells of the heart. TSA, MGCD0103, and the cyclic peptide class I HDAC inhibitor, apicidin, exhibited a common ability to enhance histone acetylation, and all potently blocked cardiac fibroblast cell cycle progression. In contrast, MGCD0103, but not TSA or apicidin, paradoxically increased expression of a subset of fibrosis-associated genes. Using the cellular thermal shift assay, we provide evidence that the divergent effects of HDAC inhibitors on cardiac fibroblast gene expression relate to differential engagement of HDAC1- and HDAC2-containing complexes. These findings illustrate the importance of employing multiple compounds when pharmacologically assessing HDAC function in a cellular context and during HDAC inhibitor drug development. PMID:28174211

Distinct Modes of Macrophage Recognition for Apoptotic and Necrotic Cells Are Not Specified Exclusively by Phosphatidylserine Exposure

PubMed Central

Cocco, Regina E.; Ucker, David S.

2001-01-01

The distinction between physiological (apoptotic) and pathological (necrotic) cell deaths reflects mechanistic differences in cellular disintegration and is of functional significance with respect to the outcomes that are triggered by the cell corpses. Mechanistically, apoptotic cells die via an active and ordered pathway; necrotic deaths, conversely, are chaotic and passive. Macrophages and other phagocytic cells recognize and engulf these dead cells. This clearance is believed to reveal an innate immunity, associated with inflammation in cases of pathological but not physiological cell deaths. Using objective and quantitative measures to assess these processes, we find that macrophages bind and engulf native apoptotic and necrotic cells to similar extents and with similar kinetics. However, recognition of these two classes of dying cells occurs via distinct and noncompeting mechanisms. Phosphatidylserine, which is externalized on both apoptotic and necrotic cells, is not a specific ligand for the recognition of either one. The distinct modes of recognition for these different corpses are linked to opposing responses from engulfing macrophages. Necrotic cells, when recognized, enhance proinflammatory responses of activated macrophages, although they are not sufficient to trigger macrophage activation. In marked contrast, apoptotic cells profoundly inhibit phlogistic macrophage responses; this represents a cell-associated, dominant-acting anti-inflammatory signaling activity acquired posttranslationally during the process of physiological cell death. PMID:11294896

Defined types of cortical interneurone structure space and spike timing in the hippocampus

PubMed Central

Somogyi, Peter; Klausberger, Thomas

2005-01-01

The cerebral cortex encodes, stores and combines information about the internal and external environment in rhythmic activity of multiple frequency ranges. Neurones of the cortex can be defined, recognized and compared on the comprehensive application of the following measures: (i) brain area- and cell domain-specific distribution of input and output synapses, (ii) expression of molecules involved in cell signalling, (iii) membrane and synaptic properties reflecting the expression of membrane proteins, (iv) temporal structure of firing in vivo, resulting from (i)–(iii). Spatial and temporal measures of neurones in the network reflect an indivisible unity of evolutionary design, i.e. neurones do not have separate structure or function. The blueprint of this design is most easily accessible in the CA1 area of the hippocampus, where a relatively uniform population of pyramidal cells and their inputs follow an instantly recognizable laminated pattern and act within stereotyped network activity patterns. Reviewing the cell types and their spatio-temporal interactions, we suggest that CA1 pyramidal cells are supported by at least 16 distinct types of GABAergic neurone. During a given behaviour-contingent network oscillation, interneurones of a given type exhibit similar firing patterns. During different network oscillations representing two distinct brain states, interneurones of the same class show different firing patterns modulating their postsynaptic target-domain in a brain-state-dependent manner. These results suggest roles for specific interneurone types in structuring the activity of pyramidal cells via their respective target domains, and accurately timing and synchronizing pyramidal cell discharge, rather than providing generalized inhibition. Finally, interneurones belonging to different classes may fire preferentially at distinct time points during a given oscillation. As different interneurones innervate distinct domains of the pyramidal cells, the different compartments will receive GABAergic input differentiated in time. Such a dynamic, spatio-temporal, GABAergic control, which evolves distinct patterns during different brain states, is ideally suited to regulating the input integration of individual pyramidal cells contributing to the formation of cell assemblies and representations in the hippocampus and, probably, throughout the cerebral cortex. PMID:15539390

Dissecting and Targeting Latent Metastasis

DTIC Science & Technology

2015-09-01

distinct class of stem-like cancer cells , which primed to enter quiescence and evade innate immunity after infiltrating distant organs. LCC cells express...state and actively silencing WNT signaling, LCC cells can enter quiescence and evade innate immunity to remain latent for extended periods. These...mutation in Foxn1 renders the mice athymic, severely blunting the maturation of effector T cells but preserving innate immunity components including

Central Topography of Cranial Motor Nuclei Controlled by Differential Cadherin Expression

PubMed Central

Astick, Marc; Tubby, Kristina; Mubarak, Waleed M.; Guthrie, Sarah; Price, Stephen R.

2014-01-01

Summary Neuronal nuclei are prominent, evolutionarily conserved features of vertebrate central nervous system (CNS) organization [1]. Nuclei are clusters of soma of functionally related neurons and are located in highly stereotyped positions. Establishment of this CNS topography is critical to neural circuit assembly. However, little is known of either the cellular or molecular mechanisms that drive nucleus formation during development, a process termed nucleogenesis [2–5]. Brainstem motor neurons, which contribute axons to distinct cranial nerves and whose functions are essential to vertebrate survival, are organized exclusively as nuclei. Cranial motor nuclei are composed of two main classes, termed branchiomotor/visceromotor and somatomotor [6]. Each of these classes innervates evolutionarily distinct structures, for example, the branchial arches and eyes, respectively. Additionally, each class is generated by distinct progenitor cell populations and is defined by differential transcription factor expression [7, 8]; for example, Hb9 distinguishes somatomotor from branchiomotor neurons. We characterized the time course of cranial motornucleogenesis, finding that despite differences in cellular origin, segregation of branchiomotor and somatomotor nuclei occurs actively, passing through a phase of each being intermingled. We also found that differential expression of cadherin cell adhesion family members uniquely defines each motor nucleus. We show that cadherin expression is critical to nucleogenesis as its perturbation degrades nucleus topography predictably. PMID:25308074

"Subpial Fan Cell" - A Class of Calretinin Neuron in Layer 1 of Adult Monkey Prefrontal Cortex.

PubMed

Gabbott, Paul L A

2016-01-01

Layer 1 of the cortex contains populations of neurochemically distinct neurons and afferent fibers which markedly affect neural activity in the apical dendritic tufts of pyramidal cells. Understanding the causal mechanisms requires knowledge of the cellular architecture and synaptic organization of layer 1. This study has identified eight morphological classes of calretinin immunopositive (CRet+) neurons (including Cajal-Retzius cells) in layer 1 of the prefrontal cortex (PFC) in adult monkey (Macaca fasicularis), with a distinct class - termed "subpial fan (SPF) cell" - described in detail. SPF cells were rare horizontal unipolar CRet+ cells located directly beneath the pia with a single thick primary dendrite that branched into a characteristic fan-like dendritic tree tangential to the pial surface. Dendrites had spines, filamentous processes and thorny branchlets. SPF cells lay millimeters apart with intralaminar axons that ramified widely in upper layer 1. Such cells were GABA immunonegative (-) and occurred in areas beyond PFC. Interspersed amidst SPF cells displaying normal structural integrity were degenerating CRet+ neurons (including SPF cells) and clumps of lipofuscin-rich cellular debris. The number of degenerating SPF cells increased during adulthood. Ultrastructural analyses indicated SPF cell somata received asymmetric (A - presumed excitatory) and symmetric (S - presumed inhibitory) synaptic contacts. Proximal dendritic shafts received mainly S-type and distal shafts mostly A-type input. All dendritic thorns and most dendritic spines received both synapse types. The tangential areal density of SPF cell axonal varicosities varied radially from parent somata - with dense clusters in more distal zones. All boutons formed A-type contacts with CRet- structures. The main post-synaptic targets were dendritic shafts (67%; mostly spine-bearing) and dendritic spines (24%). SPF-SPF cell innervation was not observed. Morphometry of SPF cells indicated a unique class of CRet+/GABA- neuron in adult monkey PFC - possibly a subtype of persisting Cajal-Retzius cell. The distribution and connectivity of SPF cells suggest they act as integrative hubs in upper layer 1 during postnatal maturation. The main synaptic output of SPF cells likely provides a transminicolumnar excitatory influence across swathes of apical dendritic tufts - thus affecting information processing in discrete patches of layer 1 in adult monkey PFC.

Layer-specific input to distinct cell types in layer 6 of monkey primary visual cortex.

PubMed

Briggs, F; Callaway, E M

2001-05-15

Layer 6 of monkey V1 contains a physiologically and anatomically diverse population of excitatory pyramidal neurons. Distinctive arborization patterns of axons and dendrites within the functionally specialized cortical layers define eight types of layer 6 pyramidal neurons and suggest unique information processing roles for each cell type. To address how input sources contribute to cellular function, we examined the laminar sources of functional excitatory input onto individual layer 6 pyramidal neurons using scanning laser photostimulation. We find that excitatory input sources correlate with cell type. Class I neurons with axonal arbors selectively targeting magnocellular (M) recipient layer 4Calpha receive input from M-dominated layer 4B, whereas class I neurons whose axonal arbors target parvocellular (P) recipient layer 4Cbeta receive input from P-dominated layer 2/3. Surprisingly, these neuronal types do not differ significantly in the inputs they receive directly from layers 4Calpha or 4Cbeta. Class II cells, which lack dense axonal arbors within layer 4C, receive excitatory input from layers targeted by their local axons. Specifically, type IIA cells project axons to and receive input from the deep but not superficial layers. Type IIB neurons project to and receive input from the deepest and most superficial, but not middle layers. Type IIC neurons arborize throughout the cortical layers and tend to receive inputs from all cortical layers. These observations have implications for the functional roles of different layer 6 cell types in visual information processing.

Crystal structure of the Streptomyces coelicolor sortase E1 transpeptidase provides insight into the binding mode of the novel class E sorting signal

DOE PAGES

Kattke, Michele D.; Chan, Albert H.; Duong, Andrew; ...

2016-12-09

Here, many species of Gram-positive bacteria use sortase transpeptidases to covalently affix proteins to their cell wall or to assemble pili. Sortase-displayed proteins perform critical and diverse functions for cell survival, including cell adhesion, nutrient acquisition, and morphological development, among others. Based on their amino acid sequences, there are at least six types of sortases (class A to F enzymes); however, class E enzymes have not been extensively studied. Class E sortases are used by soil and freshwater-dwelling Actinobacteria to display proteins that contain a non-canonical LAXTG sorting signal, which differs from 90% of known sorting signals by substitution ofmore » alanine for proline. Here we report the first crystal structure of a class E sortase, the 1.93 Å resolution structure of the SrtE1 enzyme from Streptomyces coelicolor. The active site is bound to a tripeptide, providing insight into the mechanism of substrate binding. SrtE1 possesses β3/β4 and β6/β7 active site loops that contact the LAXTG substrate and are structurally distinct from other classes. We propose that SrtE1 and other class E sortases employ a conserved tyrosine residue within their β3/β4 loop to recognize the amide nitrogen of alanine at position P3 of the sorting signal through a hydrogen bond, as seen here. Incapability of hydrogen-bonding with canonical proline-containing sorting signals likely contributes to class E substrate specificity. Furthermore, we demonstrate that surface anchoring of proteins involved in aerial hyphae formation requires an N-terminal segment in SrtE1 that is presumably positioned within the cytoplasm. Combined, our results reveal unique features within class E enzymes that enable them to recognize distinct sorting signals, and could facilitate the development of substrate-based inhibitors of this important enzyme family.« less

Overlapping and Divergent Actions of Structurally Distinct Histone Deacetylase Inhibitors in Cardiac Fibroblasts.

PubMed

Schuetze, Katherine B; Stratton, Matthew S; Blakeslee, Weston W; Wempe, Michael F; Wagner, Florence F; Holson, Edward B; Kuo, Yin-Ming; Andrews, Andrew J; Gilbert, Tonya M; Hooker, Jacob M; McKinsey, Timothy A

2017-04-01

Inhibitors of zinc-dependent histone deacetylases (HDACs) profoundly affect cellular function by altering gene expression via changes in nucleosomal histone tail acetylation. Historically, investigators have employed pan-HDAC inhibitors, such as the hydroxamate trichostatin A (TSA), which simultaneously targets members of each of the three zinc-dependent HDAC classes (classes I, II, and IV). More recently, class- and isoform-selective HDAC inhibitors have been developed, providing invaluable chemical biology probes for dissecting the roles of distinct HDACs in the control of various physiologic and pathophysiological processes. For example, the benzamide class I HDAC-selective inhibitor, MGCD0103 [ N -(2-aminophenyl)-4-[[(4-pyridin-3-ylpyrimidin-2-yl)amino]methyl] benzamide], was shown to block cardiac fibrosis, a process involving excess extracellular matrix deposition, which often results in heart dysfunction. Here, we compare the mechanisms of action of structurally distinct HDAC inhibitors in isolated primary cardiac fibroblasts, which are the major extracellular matrix-producing cells of the heart. TSA, MGCD0103, and the cyclic peptide class I HDAC inhibitor, apicidin, exhibited a common ability to enhance histone acetylation, and all potently blocked cardiac fibroblast cell cycle progression. In contrast, MGCD0103, but not TSA or apicidin, paradoxically increased expression of a subset of fibrosis-associated genes. Using the cellular thermal shift assay, we provide evidence that the divergent effects of HDAC inhibitors on cardiac fibroblast gene expression relate to differential engagement of HDAC1- and HDAC2-containing complexes. These findings illustrate the importance of employing multiple compounds when pharmacologically assessing HDAC function in a cellular context and during HDAC inhibitor drug development. Copyright © 2017 by The American Society for Pharmacology and Experimental Therapeutics.

Non-equivalent antigen presenting capabilities of dendritic cells and macrophages in generating brain-infiltrating CD8 + T cell responses.

PubMed

Malo, Courtney S; Huggins, Matthew A; Goddery, Emma N; Tolcher, Heather M A; Renner, Danielle N; Jin, Fang; Hansen, Michael J; Pease, Larry R; Pavelko, Kevin D; Johnson, Aaron J

2018-02-12

The contribution of antigen-presenting cell (APC) types in generating CD8 + T cell responses in the central nervous system (CNS) is not fully defined, limiting the development of vaccines and understanding of immune-mediated neuropathology. Here, we generate a transgenic mouse that enables cell-specific deletion of the H-2Kb MHC class I molecule. By deleting H-2K b on dendritic cells and macrophages, we compare the effect of each APC in three distinct models of neuroinflammation: picornavirus infection, experimental cerebral malaria, and a syngeneic glioma. Dendritic cells and macrophages both activate CD8 + T cell responses in response to these CNS immunological challenges. However, the extent to which each of these APCs contributes to CD8 + T cell priming varies. These findings reveal distinct functions for dendritic cells and macrophages in generating CD8 + T cell responses to neurological disease.

Modeling alternative binding registers of a minimal immunogenic peptide on two class II major histocompatibility complex (MHC II) molecules predicts polarized T-cell receptor (TCR) contact positions.

PubMed

Murray, J S; Fois, S D S; Schountz, T; Ford, S R; Tawde, M D; Brown, J C; Siahaan, T J

2002-03-01

Several major histocompatibility complex class II (MHC II) complexes with known minimal immunogenic peptides have now been solved by X-ray crystallography. Specificity pockets within the MHC II binding groove provide distinct peptide contacts that influence peptide conformation and define the binding register within different allelic MHC II molecules. Altering peptide ligands with respect to the residues that contact the T-cell receptor (TCR) can drastically change the nature of the ensuing immune response. Here, we provide an example of how MHC II (I-A) molecules may indirectly effect TCR contacts with a peptide and drive functionally distinct immune responses. We modeled the same immunogenic 12-amino acid peptide into the binding grooves of two allelic MHC II molecules linked to distinct cytokine responses against the peptide. Surprisingly, the favored conformation of the peptide in each molecule was distinct with respect to the exposure of the N- or C-terminus of the peptide above the MHC II binding groove. T-cell clones derived from each allelic MHC II genotype were found to be allele-restricted with respect to the recognition of these N- vs. C-terminal residues on the bound peptide. Taken together, these data suggest that MHC II alleles may influence T-cell functions by restricting TCR access to specific residues of the I-A-bound peptide. Thus, these data are of significance to diseases that display genetic linkage to specific MHC II alleles, e.g. type 1 diabetes and rheumatoid arthritis.

Serotonergic Modulation Differentially Targets Distinct Network Elements within the Antennal Lobe of Drosophila melanogaster

PubMed Central

Sizemore, Tyler R.; Dacks, Andrew M.

2016-01-01

Neuromodulation confers flexibility to anatomically-restricted neural networks so that animals are able to properly respond to complex internal and external demands. However, determining the mechanisms underlying neuromodulation is challenging without knowledge of the functional class and spatial organization of neurons that express individual neuromodulatory receptors. Here, we describe the number and functional identities of neurons in the antennal lobe of Drosophila melanogaster that express each of the receptors for one such neuromodulator, serotonin (5-HT). Although 5-HT enhances odor-evoked responses of antennal lobe projection neurons (PNs) and local interneurons (LNs), the receptor basis for this enhancement is unknown. We used endogenous reporters of transcription and translation for each of the five 5-HT receptors (5-HTRs) to identify neurons, based on cell class and transmitter content, that express each receptor. We find that specific receptor types are expressed by distinct combinations of functional neuronal classes. For instance, the excitatory PNs express the excitatory 5-HTRs, while distinct classes of LNs each express different 5-HTRs. This study therefore provides a detailed atlas of 5-HT receptor expression within a well-characterized neural network, and enables future dissection of the role of serotonergic modulation of olfactory processing. PMID:27845422

Identification and differentiation of hepatic stem cells during liver development.

PubMed

Kamiya, Akihide; Gonzalez, Frank J; Nakauchi, Hiromitsu

2006-05-01

Stem cells responsible for maintenance and repair of tissues are found in a number of organs. The liver's remarkable capacity to regenerate after hepatectomy or chemical-induced injury does not involve proliferation of stem cells. However, recent studies suggest that liver stem cells exist in both embryonic and adult livers. Using fluorescence-activated cell sorting and a culture system in which primitive hepatic progenitor cells form colonies, a novel class of cells with the marker profile c-Met(+)CD49f(+/low)c-Kit(-)CD45(-)TER119(-) was found in the developing liver. This class apparently represents the population of cells that form colonies containing distinct hepatocytes and cholangiocytes. When cells in this class are transplanted into the spleen or liver of mice subjected to liver injury, the cells migrate and differentiate into liver parenchymal cells and cholangiocytes that are morphologically and functionally indistinguishable from their native counterparts. During mid-gestation, hematopoietic cells migrate into the liver from a region bounded by aorta, gonad, and mesonephros and produce oncostatin M (OSM). In combination with glucocorticoid hormones, OSM induces maturation of liver stem and progenitor cells, including those of the c-Met(+)CD49f(+/low)c-Kit(-)CD45(-)TER119(-) class. The ability to manipulate the proliferation and differentiation of liver stem cells in vitro will greatly aid in analyzing mechanisms of liver development and offers promise in stem cell therapy of liver diseases.

Switch Transcripts in Immunoglobulin Class Switching

NASA Astrophysics Data System (ADS)

Lorenz, Matthias; Jung, Steffen; Radbruch, Andreas

1995-03-01

B cells can exchange gene segments for the constant region of the immunoglobulin heavy chain, altering the class and effector function of the antibodies that they produce. Class switching is directed to distinct classes by cytokines, which induce transcription of the targeted DNA sequences. These transcripts are processed, resulting in spliced "switch" transcripts. Switch recombination can be directed to immunoglobulin G1 (IgG1) by the heterologous human metallothionein II_A promoter in mutant mice. Induction of the structurally conserved, spliced switch transcripts is sufficient to target switch recombination to IgG1, whereas transcription alone is not.

Somatostatin-Expressing Inhibitory Interneurons in Cortical Circuits

PubMed Central

Yavorska, Iryna; Wehr, Michael

2016-01-01

Cortical inhibitory neurons exhibit remarkable diversity in their morphology, connectivity, and synaptic properties. Here, we review the function of somatostatin-expressing (SOM) inhibitory interneurons, focusing largely on sensory cortex. SOM neurons also comprise a number of subpopulations that can be distinguished by their morphology, input and output connectivity, laminar location, firing properties, and expression of molecular markers. Several of these classes of SOM neurons show unique dynamics and characteristics, such as facilitating synapses, specific axonal projections, intralaminar input, and top-down modulation, which suggest possible computational roles. SOM cells can be differentially modulated by behavioral state depending on their class, sensory system, and behavioral paradigm. The functional effects of such modulation have been studied with optogenetic manipulation of SOM cells, which produces effects on learning and memory, task performance, and the integration of cortical activity. Different classes of SOM cells participate in distinct disinhibitory circuits with different inhibitory partners and in different cortical layers. Through these disinhibitory circuits, SOM cells help encode the behavioral relevance of sensory stimuli by regulating the activity of cortical neurons based on subcortical and intracortical modulatory input. Associative learning leads to long-term changes in the strength of connectivity of SOM cells with other neurons, often influencing the strength of inhibitory input they receive. Thus despite their heterogeneity and variability across cortical areas, current evidence shows that SOM neurons perform unique neural computations, forming not only distinct molecular but also functional subclasses of cortical inhibitory interneurons. PMID:27746722

Elevation of c-MYC Disrupts HLA Class II-mediated Immune Recognition of Human B-cell Tumors1

PubMed Central

God, Jason M.; Cameron, Christine; Figueroa, Janette; Amria, Shereen; Hossain, Azim; Kempkes, Bettina; Bornkamm, Georg W.; Stuart, Robert K.; Blum, Janice S.; Haque, Azizul

2014-01-01

Elevated levels of the transcription factor c-myc are strongly associated with various cancers, and in particular B-cell lymphomas. While many of c-MYC’s functions have been elucidated, its effect on the presentation of antigen (Ag) through the HLA class II pathway has not previously been reported. This is an issue of considerable importance, given the low immunogenicity of many c-MYC-positive tumors. We report here that increased c-MYC expression has a negative effect on the ability of B-cell lymphomas to functionally present Ags/peptides to CD4+ T cells. This defect was associated with alterations in the expression of distinct co-factors as well as interactions of antigenic peptides with class II molecules required for the presentation of class II-peptide complexes and T cell engagement. Using early passage Burkitt’s lymphoma (BL) tumors and transformed cells, we show that compared to B-lymphoblasts, BL cells express decreased levels of the class II editor HLA-DM, lysosomal thiol-reductase GILT, and a 47kDa enolase-like protein. Functional Ag presentation was partially restored in BL cells treated with a c-MYC inhibitor, demonstrating the impact of this oncogene on Ag recognition. This restoration of HLA class II-mediated Ag presentation in early passage BL tumors/cells was linked to enhanced HLA-DM expression and a concurrent decrease in HLA-DO in BL cells. Taken together, these results reveal c-MYC exerts suppressive effects at several critical checkpoints in Ag presentation which contribute to the immunoevasive properties of BL tumors. PMID:25595783

Th-1 polarization is regulated by dendritic-cell comparison of MHC class I and class II antigens

PubMed Central

Xing, Dongxia; Li, Sufang; Robinson, Simon N.; Yang, Hong; Steiner, David; Komanduri, Krishna V.; Shpall, Elizabeth J.

2009-01-01

In the control of T-helper type I (Th-1) polarization, dendritic cells (DCs) must interpret a complex array of stimuli, many of which are poorly understood. Here we demonstrate that Th-1 polarization is heavily influenced by DC-autonomous phenomena triggered by the loading of DCs with antigenically matched major histocompatibility complex (MHC) class I and class II determinants, that is, class I and II peptide epitopes exhibiting significant amino acid sequence overlap (such as would be physiologically present during infectious processes requiring Th-1 immunity for clearance). Data were derived from 13 independent antigenic models including whole-cell systems, single-protein systems, and 3 different pairs of overlapping class I and II binding epitopes. Once loaded with matched class I and II antigens, these “Th-1 DCs” exhibited differential cytokine secretion and surface marker expression, a distinct transcriptional signature, and acquired the ability to enhance generation of CD8+ T lymphocytes. Mechanistically, tRNA-synthetases were implicated as components of a putative sensor complex involved in the comparison of class I and II epitopes. These data provide rigorous conceptual explanations for the process of Th-1 polarization and the antigenic specificity of cognate T-cell help, enhance the understanding of Th-1 responses, and should contribute to the formulation of more effective vaccination strategies. PMID:19171878

A computational neural approach to support the discovery of gene function and classes of cancer.

PubMed

Azuaje, F

2001-03-01

Advances in molecular classification of tumours may play a central role in cancer treatment. Here, a novel approach to genome expression pattern interpretation is described and applied to the recognition of B-cell malignancies as a test set. Using cDNA microarrays data generated by a previous study, a neural network model known as simplified fuzzy ARTMAP is able to identify normal and diffuse large B-cell lymphoma (DLBCL) patients. Furthermore, it discovers the distinction between patients with molecularly distinct forms of DLBCL without previous knowledge of those subtypes.

Parvalbumin+ Neurons and Npas1+ Neurons Are Distinct Neuron Classes in the Mouse External Globus Pallidus

PubMed Central

Hernández, Vivian M.; Hegeman, Daniel J.; Cui, Qiaoling; Kelver, Daniel A.; Fiske, Michael P.; Glajch, Kelly E.; Pitt, Jason E.; Huang, Tina Y.; Justice, Nicholas J.

2015-01-01

Compelling evidence suggests that pathological activity of the external globus pallidus (GPe), a nucleus in the basal ganglia, contributes to the motor symptoms of a variety of movement disorders such as Parkinson's disease. Recent studies have challenged the idea that the GPe comprises a single, homogenous population of neurons that serves as a simple relay in the indirect pathway. However, we still lack a full understanding of the diversity of the neurons that make up the GPe. Specifically, a more precise classification scheme is needed to better describe the fundamental biology and function of different GPe neuron classes. To this end, we generated a novel multicistronic BAC (bacterial artificial chromosome) transgenic mouse line under the regulatory elements of the Npas1 gene. Using a combinatorial transgenic and immunohistochemical approach, we discovered that parvalbumin-expressing neurons and Npas1-expressing neurons in the GPe represent two nonoverlapping cell classes, amounting to 55% and 27% of the total GPe neuron population, respectively. These two genetically identified cell classes projected primarily to the subthalamic nucleus and to the striatum, respectively. Additionally, parvalbumin-expressing neurons and Npas1-expressing neurons were distinct in their autonomous and driven firing characteristics, their expression of intrinsic ion conductances, and their responsiveness to chronic 6-hydroxydopamine lesion. In summary, our data argue that parvalbumin-expressing neurons and Npas1-expressing neurons are two distinct functional classes of GPe neurons. This work revises our understanding of the GPe, and provides the foundation for future studies of its function and dysfunction. SIGNIFICANCE STATEMENT Until recently, the heterogeneity of the constituent neurons within the external globus pallidus (GPe) was not fully appreciated. We addressed this knowledge gap by discovering two principal GPe neuron classes, which were identified by their nonoverlapping expression of the markers parvalbumin and Npas1. Our study provides evidence that parvalbumin and Npas1 neurons have different topologies within the basal ganglia. PMID:26311767

Parvalbumin+ Neurons and Npas1+ Neurons Are Distinct Neuron Classes in the Mouse External Globus Pallidus.

PubMed

Hernández, Vivian M; Hegeman, Daniel J; Cui, Qiaoling; Kelver, Daniel A; Fiske, Michael P; Glajch, Kelly E; Pitt, Jason E; Huang, Tina Y; Justice, Nicholas J; Chan, C Savio

2015-08-26

Compelling evidence suggests that pathological activity of the external globus pallidus (GPe), a nucleus in the basal ganglia, contributes to the motor symptoms of a variety of movement disorders such as Parkinson's disease. Recent studies have challenged the idea that the GPe comprises a single, homogenous population of neurons that serves as a simple relay in the indirect pathway. However, we still lack a full understanding of the diversity of the neurons that make up the GPe. Specifically, a more precise classification scheme is needed to better describe the fundamental biology and function of different GPe neuron classes. To this end, we generated a novel multicistronic BAC (bacterial artificial chromosome) transgenic mouse line under the regulatory elements of the Npas1 gene. Using a combinatorial transgenic and immunohistochemical approach, we discovered that parvalbumin-expressing neurons and Npas1-expressing neurons in the GPe represent two nonoverlapping cell classes, amounting to 55% and 27% of the total GPe neuron population, respectively. These two genetically identified cell classes projected primarily to the subthalamic nucleus and to the striatum, respectively. Additionally, parvalbumin-expressing neurons and Npas1-expressing neurons were distinct in their autonomous and driven firing characteristics, their expression of intrinsic ion conductances, and their responsiveness to chronic 6-hydroxydopamine lesion. In summary, our data argue that parvalbumin-expressing neurons and Npas1-expressing neurons are two distinct functional classes of GPe neurons. This work revises our understanding of the GPe, and provides the foundation for future studies of its function and dysfunction. Until recently, the heterogeneity of the constituent neurons within the external globus pallidus (GPe) was not fully appreciated. We addressed this knowledge gap by discovering two principal GPe neuron classes, which were identified by their nonoverlapping expression of the markers parvalbumin and Npas1. Our study provides evidence that parvalbumin and Npas1 neurons have different topologies within the basal ganglia. Copyright © 2015 the authors 0270-6474/15/3511830-18$15.00/0.

High-content image informatics of the structural nuclear protein NuMA parses trajectories for stem/progenitor cell lineages and oncogenic transformation

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

Vega, Sebastián L.; Liu, Er; Arvind, Varun

Stem and progenitor cells that exhibit significant regenerative potential and critical roles in cancer initiation and progression remain difficult to characterize. Cell fates are determined by reciprocal signaling between the cell microenvironment and the nucleus; hence parameters derived from nuclear remodeling are ideal candidates for stem/progenitor cell characterization. Here we applied high-content, single cell analysis of nuclear shape and organization to examine stem and progenitor cells destined to distinct differentiation endpoints, yet undistinguishable by conventional methods. Nuclear descriptors defined through image informatics classified mesenchymal stem cells poised to either adipogenic or osteogenic differentiation, and oligodendrocyte precursors isolated from different regionsmore » of the brain and destined to distinct astrocyte subtypes. Nuclear descriptors also revealed early changes in stem cells after chemical oncogenesis, allowing the identification of a class of cancer-mitigating biomaterials. To capture the metrology of nuclear changes, we developed a simple and quantitative “imaging-derived” parsing index, which reflects the dynamic evolution of the high-dimensional space of nuclear organizational features. A comparative analysis of parsing outcomes via either nuclear shape or textural metrics of the nuclear structural protein NuMA indicates the nuclear shape alone is a weak phenotypic predictor. In contrast, variations in the NuMA organization parsed emergent cell phenotypes and discerned emergent stages of stem cell transformation, supporting a prognosticating role for this protein in the outcomes of nuclear functions. - Highlights: • High-content analysis of nuclear shape and organization classify stem and progenitor cells poised for distinct lineages. • Early oncogenic changes in mesenchymal stem cells (MSCs) are also detected with nuclear descriptors. • A new class of cancer-mitigating biomaterials was identified based on image informatics. • Textural metrics of the nuclear structural protein NuMA are sufficient to parse emergent cell phenotypes.« less

The leader peptide of mutacin 1140 has distinct structural components compared to related class I lantibiotics

PubMed Central

Escano, Jerome; Stauffer, Byron; Brennan, Jacob; Bullock, Monica; Smith, Leif

2014-01-01

Lantibiotics are ribosomally synthesized peptide antibiotics composed of an N-terminal leader peptide that promotes the core peptide's interaction with the post translational modification (PTM) enzymes. Following PTMs, mutacin 1140 is transported out of the cell and the leader peptide is cleaved to yield the antibacterial peptide. Mutacin 1140 leader peptide is structurally unique compared to other class I lantibiotic leader peptides. Herein, we further our understanding of the structural differences of mutacin 1140 leader peptide with regard to other class I leader peptides. We have determined that the length of the leader peptide is important for the biosynthesis of mutacin 1140. We have also determined that mutacin 1140 leader peptide contains a novel four amino acid motif compared to related lantibiotics. PTM enzyme recognition of the leader peptide appears to be evolutionarily distinct from related class I lantibiotics. Our study on mutacin 1140 leader peptide provides a basis for future studies aimed at understanding its interaction with the PTM enzymes. PMID:25400246

In-Depth Analysis of Citrulline-Specific CD4 T-Cells in Rheumatoid Arthritis

DTIC Science & Technology

2016-01-01

1 AWARD NUMBER: W81XWH-15-1-0004 TITLE: In-Depth Analysis of Citrulline-Specific CD4 T-Cells in Rheumatoid Arthritis PRINCIPAL INVESTIGATOR...present in rheumatoid arthritis (RA) patients exhibit a distinct cell surface phenotype and transcriptional signature that could be used to predict...and are on track to achieve our Year 2 goals 15. SUBJECT TERMS Rheumatoid arthritis ; CD4 T cells; citrulline; HLA class II tetramers; RNAseq

Cellulose synthase 'class specific regions' are intrinsically disordered and functionally undifferentiated.

PubMed

Scavuzzo-Duggan, Tess R; Chaves, Arielle M; Singh, Abhishek; Sethaphong, Latsavongsakda; Slabaugh, Erin; Yingling, Yaroslava G; Haigler, Candace H; Roberts, Alison W

2018-06-01

Cellulose synthases (CESAs) are glycosyltransferases that catalyze formation of cellulose microfibrils in plant cell walls. Seed plant CESA isoforms cluster in six phylogenetic clades, whose non-interchangeable members play distinct roles within cellulose synthesis complexes (CSCs). A 'class specific region' (CSR), with higher sequence similarity within versus between functional CESA classes, has been suggested to contribute to specific activities or interactions of different isoforms. We investigated CESA isoform specificity in the moss, Physcomitrella patens (Hedw.) B. S. G. to gain evolutionary insights into CESA structure/function relationships. Like seed plants, P. patens has oligomeric rosette-type CSCs, but the PpCESAs diverged independently and form a separate CESA clade. We showed that P. patens has two functionally distinct CESAs classes, based on the ability to complement the gametophore-negative phenotype of a ppcesa5 knockout line. Thus, non-interchangeable CESA classes evolved separately in mosses and seed plants. However, testing of chimeric moss CESA genes for complementation demonstrated that functional class-specificity is not determined by the CSR. Sequence analysis and computational modeling showed that the CSR is intrinsically disordered and contains predicted molecular recognition features, consistent with a possible role in CESA oligomerization and explaining the evolution of class-specific sequences without selection for class-specific function. © 2018 Institute of Botany, Chinese Academy of Sciences.

Activation of raphe nuclei triggers rapid and distinct effects on parallel olfactory bulb output channels

PubMed Central

Kapoor, Vikrant; Provost, Allison; Agarwal, Prateek; Murthy, Venkatesh N.

2015-01-01

The serotonergic raphe nuclei are involved in regulating brain states over time-scales of minutes and hours. We examined more rapid effects of serotonergic activation on two classes of principal neurons in the mouse olfactory bulb, mitral and tufted cells, which send olfactory information to distinct targets. Brief stimulation of the raphe nuclei led to excitation of tufted cells at rest and potentiation of their odor responses. While mitral cells at rest were also excited by raphe activation, their odor responses were bidirectionally modulated, leading to improved pattern separation of odors. In vitro whole-cell recordings revealed that specific optogenetic activation of raphe axons affected bulbar neurons through dual release of serotonin and glutamate. Therefore, the raphe nuclei, in addition to their role in neuromodulation of brain states, are also involved in fast, sub-second top-down modulation, similar to cortical feedback. This modulation can selectively and differentially sensitize or decorrelate distinct output channels. PMID:26752161

Two Polyhydroxyalkanoate Synthases from Distinct Classes from the Aromatic Degrader Cupriavidus pinatubonensis JMP134 Exhibit the Same Substrate Preference.

PubMed

Jiang, Xuan; Luo, Xi; Zhou, Ning-Yi

2015-01-01

Cupriavidus pinatubonensis JMP134 utilizes a variety of aromatic substrates as sole carbon sources, including meta-nitrophenol (MNP). Two polyhydroxyalkanoate (PHA) synthase genes, phaC1 and phaC2, were annotated and categorized as class I and class II PHA synthase genes, respectively. In this study, both His-tagged purified PhaC1 and PhaC2 were shown to exhibit typical class I PHA synthase substrate specificity to make short-chain-length (SCL) PHA from 3-hydroxybutyryl-CoA and failed to make medium-chain-length (MCL) PHA from 3-hydroxyoctanoyl-CoA. The phaC1 or phaC2 deletion strain could also produce SCL PHA when grown in fructose or octanoate, but the double mutant of phaC1 and phaC2 lost this ability. The PhaC2 also exhibited substrate preference towards SCL substrates when expressed in Pseudomonas aeruginosa PAO1 phaC mutant strain. On the other hand, the transcriptional level of phaC1 was 70-fold higher than that of phaC2 in MNP-grown cells, but 240-fold lower in octanoate-grown cells. Further study demonstrated that only phaC1 was involved in PHA synthesis in MNP-grown cells. These findings suggested that phaC1 and phaC2 genes were differentially regulated under different growth conditions in this strain. Within the phaC2-containing gene cluster, a single copy of PHA synthase gene was present clustering with genes encoding enzymes in the biosynthesis of PHA precursors. This is markedly different from the genetic organization of all other previously reported class II PHA synthase gene clusters and this cluster likely comes from a distinct evolutionary path.

Two Polyhydroxyalkanoate Synthases from Distinct Classes from the Aromatic Degrader Cupriavidus pinatubonensis JMP134 Exhibit the Same Substrate Preference

PubMed Central

Jiang, Xuan; Luo, Xi; Zhou, Ning-Yi

2015-01-01

Cupriavidus pinatubonensis JMP134 utilizes a variety of aromatic substrates as sole carbon sources, including meta-nitrophenol (MNP). Two polyhydroxyalkanoate (PHA) synthase genes, phaC1 and phaC2, were annotated and categorized as class I and class II PHA synthase genes, respectively. In this study, both His-tagged purified PhaC1 and PhaC2 were shown to exhibit typical class I PHA synthase substrate specificity to make short-chain-length (SCL) PHA from 3-hydroxybutyryl-CoA and failed to make medium-chain-length (MCL) PHA from 3-hydroxyoctanoyl-CoA. The phaC1 or phaC2 deletion strain could also produce SCL PHA when grown in fructose or octanoate, but the double mutant of phaC1 and phaC2 lost this ability. The PhaC2 also exhibited substrate preference towards SCL substrates when expressed in Pseudomonas aeruginosa PAO1 phaC mutant strain. On the other hand, the transcriptional level of phaC1 was 70-fold higher than that of phaC2 in MNP-grown cells, but 240-fold lower in octanoate-grown cells. Further study demonstrated that only phaC1 was involved in PHA synthesis in MNP-grown cells. These findings suggested that phaC1 and phaC2 genes were differentially regulated under different growth conditions in this strain. Within the phaC2-containing gene cluster, a single copy of PHA synthase gene was present clustering with genes encoding enzymes in the biosynthesis of PHA precursors. This is markedly different from the genetic organization of all other previously reported class II PHA synthase gene clusters and this cluster likely comes from a distinct evolutionary path. PMID:26544851

“Subpial Fan Cell” — A Class of Calretinin Neuron in Layer 1 of Adult Monkey Prefrontal Cortex

PubMed Central

Gabbott, Paul L. A.

2016-01-01

Layer 1 of the cortex contains populations of neurochemically distinct neurons and afferent fibers which markedly affect neural activity in the apical dendritic tufts of pyramidal cells. Understanding the causal mechanisms requires knowledge of the cellular architecture and synaptic organization of layer 1. This study has identified eight morphological classes of calretinin immunopositive (CRet+) neurons (including Cajal-Retzius cells) in layer 1 of the prefrontal cortex (PFC) in adult monkey (Macaca fasicularis), with a distinct class — termed “subpial fan (SPF) cell” — described in detail. SPF cells were rare horizontal unipolar CRet+ cells located directly beneath the pia with a single thick primary dendrite that branched into a characteristic fan-like dendritic tree tangential to the pial surface. Dendrites had spines, filamentous processes and thorny branchlets. SPF cells lay millimeters apart with intralaminar axons that ramified widely in upper layer 1. Such cells were GABA immunonegative (-) and occurred in areas beyond PFC. Interspersed amidst SPF cells displaying normal structural integrity were degenerating CRet+ neurons (including SPF cells) and clumps of lipofuscin-rich cellular debris. The number of degenerating SPF cells increased during adulthood. Ultrastructural analyses indicated SPF cell somata received asymmetric (A — presumed excitatory) and symmetric (S — presumed inhibitory) synaptic contacts. Proximal dendritic shafts received mainly S-type and distal shafts mostly A-type input. All dendritic thorns and most dendritic spines received both synapse types. The tangential areal density of SPF cell axonal varicosities varied radially from parent somata — with dense clusters in more distal zones. All boutons formed A-type contacts with CRet- structures. The main post-synaptic targets were dendritic shafts (67%; mostly spine-bearing) and dendritic spines (24%). SPF-SPF cell innervation was not observed. Morphometry of SPF cells indicated a unique class of CRet+/GABA- neuron in adult monkey PFC — possibly a subtype of persisting Cajal-Retzius cell. The distribution and connectivity of SPF cells suggest they act as integrative hubs in upper layer 1 during postnatal maturation. The main synaptic output of SPF cells likely provides a transminicolumnar excitatory influence across swathes of apical dendritic tufts — thus affecting information processing in discrete patches of layer 1 in adult monkey PFC. PMID:27147978

The morphologies of breast cancer cell lines in three-dimensionalassays correlate with their profiles of gene expression

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

Kenny, Paraic A.; Lee, Genee Y.; Myers, Connie A.

2007-01-31

3D cell cultures are rapidly becoming the method of choice for the physiologically relevant modeling of many aspects of non-malignant and malignant cell behavior ex vivo. Nevertheless, only a limited number of distinct cell types have been evaluated in this assay to date. Here we report the first large scale comparison of the transcriptional profiles and 3D cell culture phenotypes of a substantial panel of human breast cancer cell lines. Each cell line adopts a colony morphology of one of four main classes in 3D culture. These morphologies reflect, at least in part, the underlying gene expression profile and proteinmore » expression patterns of the cell lines, and distinct morphologies were also associated with tumor cell invasiveness and with cell lines originating from metastases. We further demonstrate that consistent differences in genes encoding signal transduction proteins emerge when even tumor cells are cultured in 3D microenvironments.« less

Automated cell-type classification in intact tissues by single-cell molecular profiling

PubMed Central

2018-01-01

A major challenge in biology is identifying distinct cell classes and mapping their interactions in vivo. Tissue-dissociative technologies enable deep single cell molecular profiling but do not provide spatial information. We developed a proximity ligation in situ hybridization technology (PLISH) with exceptional signal strength, specificity, and sensitivity in tissue. Multiplexed data sets can be acquired using barcoded probes and rapid label-image-erase cycles, with automated calculation of single cell profiles, enabling clustering and anatomical re-mapping of cells. We apply PLISH to expression profile ~2900 cells in intact mouse lung, which identifies and localizes known cell types, including rare ones. Unsupervised classification of the cells indicates differential expression of ‘housekeeping’ genes between cell types, and re-mapping of two sub-classes of Club cells highlights their segregated spatial domains in terminal airways. By enabling single cell profiling of various RNA species in situ, PLISH can impact many areas of basic and medical research. PMID:29319504

Three separate classes of bacterial ice nucleation structures

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

Turner, M.A.; Arellano, F.; Kozloff, L.M.

1990-05-01

Studies of the properties of the ice nucleation structure exposed on the surfaces of various bacteria such as Pseudomonas syringae, Erwinia herbicola, or various strains of Ice+ recombinant Escherichia coli have shown that there are clearly three major related but chemically distinct types of structures on these cells. First, the ability of Ice+ cells to nucleate super-cooled D2O has been examined, and it has been found that this ability (relative to the ability of the same cells to nucleate super-cooled H2O) exhibited three characteristic nucleating patterns. The rarest structure, called class A, is found on only a small fraction ofmore » cells in a culture, nucleates H2O at temperatures above -4.4 degrees C, and is an effective nucleator of super-cooled D2O. A second class of structure, called class B, is found on a larger portion of the cells, nucleates H2O between -4.8 and -5.7 degrees C, and is a relatively poor nucleator of super-cooled D2O. The class C structure is found on almost all cells and nucleates at -7.6 degrees C or colder. These three classes of structures were also differentiated by their sensitivities to low concentrations of water-miscible organic solvents such as dioxane or dimethyl sulfoxide. Depending on the specific bacterial strain, the addition of these solvents to bacterial suspensions lowered the nucleation activity of the class A structure by 1,000-fold or more. The nucleation activities of class B structures in the same culture were highly resistant to these compounds and were lowered only by 20 to 40%.« less

HLA Engineering of Human Pluripotent Stem Cells

PubMed Central

Riolobos, Laura; Hirata, Roli K; Turtle, Cameron J; Wang, Pei-Rong; Gornalusse, German G; Zavajlevski, Maja; Riddell, Stanley R; Russell, David W

2013-01-01

The clinical use of human pluripotent stem cells and their derivatives is limited by the rejection of transplanted cells due to differences in their human leukocyte antigen (HLA) genes. This has led to the proposed use of histocompatible, patient-specific stem cells; however, the preparation of many different stem cell lines for clinical use is a daunting task. Here, we develop two distinct genetic engineering approaches that address this problem. First, we use a combination of gene targeting and mitotic recombination to derive HLA-homozygous embryonic stem cell (ESC) subclones from an HLA-heterozygous parental line. A small bank of HLA-homozygous stem cells with common haplotypes would match a significant proportion of the population. Second, we derive HLA class I–negative cells by targeted disruption of both alleles of the Beta-2 Microglobulin (B2M) gene in ESCs. Mixed leukocyte reactions and peptide-specific HLA-restricted CD8+ T cell responses were reduced in class I–negative cells that had undergone differentiation in embryoid bodies. These B2M−/− ESCs could act as universal donor cells in applications where the transplanted cells do not express HLA class II genes. Both approaches used adeno-associated virus (AAV) vectors for efficient gene targeting in the absence of potentially genotoxic nucleases, and produced pluripotent, transgene-free cell lines. PMID:23629003

HLA engineering of human pluripotent stem cells.

PubMed

Riolobos, Laura; Hirata, Roli K; Turtle, Cameron J; Wang, Pei-Rong; Gornalusse, German G; Zavajlevski, Maja; Riddell, Stanley R; Russell, David W

2013-06-01

The clinical use of human pluripotent stem cells and their derivatives is limited by the rejection of transplanted cells due to differences in their human leukocyte antigen (HLA) genes. This has led to the proposed use of histocompatible, patient-specific stem cells; however, the preparation of many different stem cell lines for clinical use is a daunting task. Here, we develop two distinct genetic engineering approaches that address this problem. First, we use a combination of gene targeting and mitotic recombination to derive HLA-homozygous embryonic stem cell (ESC) subclones from an HLA-heterozygous parental line. A small bank of HLA-homozygous stem cells with common haplotypes would match a significant proportion of the population. Second, we derive HLA class I-negative cells by targeted disruption of both alleles of the Beta-2 Microglobulin (B2M) gene in ESCs. Mixed leukocyte reactions and peptide-specific HLA-restricted CD8(+) T cell responses were reduced in class I-negative cells that had undergone differentiation in embryoid bodies. These B2M(-/-) ESCs could act as universal donor cells in applications where the transplanted cells do not express HLA class II genes. Both approaches used adeno-associated virus (AAV) vectors for efficient gene targeting in the absence of potentially genotoxic nucleases, and produced pluripotent, transgene-free cell lines.

SOX2 expression levels distinguish between neural progenitor populations of the developing dorsal telencephalon.

PubMed

Hutton, Scott R; Pevny, Larysa H

2011-04-01

The HMG-Box transcription factor SOX2 is expressed in neural progenitor populations throughout the developing and adult central nervous system and is necessary to maintain their progenitor identity. However, it is unclear whether SOX2 levels are uniformly expressed across all neural progenitor populations. In the developing dorsal telencephalon, two distinct populations of neural progenitors, radial glia and intermediate progenitor cells, are responsible for generating a majority of excitatory neurons found in the adult neocortex. Here we demonstrate, using both cellular and molecular analyses, that SOX2 is differentially expressed between radial glial and intermediate progenitor populations. Moreover, utilizing a SOX2(EGFP) mouse line, we show that this differential expression can be used to prospectively isolate distinct, viable populations of radial glia and intermediate cells for in vitro analysis. Given the limited repertoire of cell-surface markers currently available for neural progenitor cells, this provides an invaluable tool for prospectively identifying and isolating distinct classes of neural progenitor cells from the central nervous system. Copyright © 2011 Elsevier Inc. All rights reserved.

Distinct cellular properties of oncogenic KIT receptor tyrosine kinase mutants enable alternative courses of cancer cell inhibition

PubMed Central

Shi, Xiarong; Sousa, Leiliane P.; Mandel-Bausch, Elizabeth M.; Tome, Francisco; Reshetnyak, Andrey V.; Hadari, Yaron; Schlessinger, Joseph; Lax, Irit

2016-01-01

Large genomic sequencing analysis as part of precision medicine efforts revealed numerous activating mutations in receptor tyrosine kinases, including KIT. Unfortunately, a single approach is not effective for inhibiting cancer cells or treating cancers driven by all known oncogenic KIT mutants. Here, we show that each of the six major KIT oncogenic mutants exhibits different enzymatic, cellular, and dynamic properties and responds distinctly to different KIT inhibitors. One class of KIT mutants responded well to anti-KIT antibody treatment alone or in combination with a low dose of tyrosine kinase inhibitors (TKIs). A second class of KIT mutants, including a mutant resistant to imatinib treatment, responded well to a combination of TKI with anti-KIT antibodies or to anti-KIT toxin conjugates, respectively. We conclude that the preferred choice of precision medicine treatments for cancers driven by activated KIT and other RTKs may rely on clear understanding of the dynamic properties of oncogenic mutants. PMID:27482095

Diverse modes of synaptic signaling, regulation, and plasticity distinguish two classes of C. elegans glutamatergic neurons.

PubMed

Ventimiglia, Donovan; Bargmann, Cornelia I

2017-11-21

Synaptic vesicle release properties vary between neuronal cell types, but in most cases the molecular basis of this heterogeneity is unknown. Here, we compare in vivo synaptic properties of two neuronal classes in the C. elegans central nervous system, using VGLUT-pHluorin to monitor synaptic vesicle exocytosis and retrieval in intact animals. We show that the glutamatergic sensory neurons AWC ON and ASH have distinct synaptic dynamics associated with tonic and phasic synaptic properties, respectively. Exocytosis in ASH and AWC ON is differentially affected by SNARE-complex regulators that are present in both neurons: phasic ASH release is strongly dependent on UNC-13, whereas tonic AWC ON release relies upon UNC-18 and on the protein kinase C homolog PKC-1. Strong stimuli that elicit high calcium levels increase exocytosis and retrieval rates in AWC ON , generating distinct tonic and evoked synaptic modes. These results highlight the differential deployment of shared presynaptic proteins in neuronal cell type-specific functions.

Diverse modes of synaptic signaling, regulation, and plasticity distinguish two classes of C. elegans glutamatergic neurons

PubMed Central

Ventimiglia, Donovan

2017-01-01

Synaptic vesicle release properties vary between neuronal cell types, but in most cases the molecular basis of this heterogeneity is unknown. Here, we compare in vivo synaptic properties of two neuronal classes in the C. elegans central nervous system, using VGLUT-pHluorin to monitor synaptic vesicle exocytosis and retrieval in intact animals. We show that the glutamatergic sensory neurons AWCON and ASH have distinct synaptic dynamics associated with tonic and phasic synaptic properties, respectively. Exocytosis in ASH and AWCON is differentially affected by SNARE-complex regulators that are present in both neurons: phasic ASH release is strongly dependent on UNC-13, whereas tonic AWCON release relies upon UNC-18 and on the protein kinase C homolog PKC-1. Strong stimuli that elicit high calcium levels increase exocytosis and retrieval rates in AWCON, generating distinct tonic and evoked synaptic modes. These results highlight the differential deployment of shared presynaptic proteins in neuronal cell type-specific functions. PMID:29160768

Distinct cellular pathways select germline-encoded and somatically mutated antibodies into immunological memory

PubMed Central

Kaji, Tomohiro; Ishige, Akiko; Hikida, Masaki; Taka, Junko; Hijikata, Atsushi; Kubo, Masato; Nagashima, Takeshi; Takahashi, Yoshimasa; Kurosaki, Tomohiro; Okada, Mariko; Ohara, Osamu

2012-01-01

One component of memory in the antibody system is long-lived memory B cells selected for the expression of somatically mutated, high-affinity antibodies in the T cell–dependent germinal center (GC) reaction. A puzzling observation has been that the memory B cell compartment also contains cells expressing unmutated, low-affinity antibodies. Using conditional Bcl6 ablation, we demonstrate that these cells are generated through proliferative expansion early after immunization in a T cell–dependent but GC-independent manner. They soon become resting and long-lived and display a novel distinct gene expression signature which distinguishes memory B cells from other classes of B cells. GC-independent memory B cells are later joined by somatically mutated GC descendants at roughly equal proportions and these two types of memory cells efficiently generate adoptive secondary antibody responses. Deletion of T follicular helper (Tfh) cells significantly reduces the generation of mutated, but not unmutated, memory cells early on in the response. Thus, B cell memory is generated along two fundamentally distinct cellular differentiation pathways. One pathway is dedicated to the generation of high-affinity somatic antibody mutants, whereas the other preserves germ line antibody specificities and may prepare the organism for rapid responses to antigenic variants of the invading pathogen. PMID:23027924

Utility of temporally distinct baculovirus promoters for constitutive and baculovirus-inducible transgene expression in transformed insect cells.

PubMed

Lin, Chi-Hung; Jarvis, Donald L

2013-05-10

Genetically transformed lepidopteran insect cell lines have biotechnological applications as constitutive recombinant protein production platforms and improved hosts for baculovirus-mediated recombinant protein production. Insect cell transformation is often accomplished with a DNA construct(s) encoding a foreign protein(s) under the transcriptional control of a baculovirus immediate early promoter, such as the ie1 promoter. However, the potential utility of increasingly stronger promoters from later baculovirus gene classes, such as delayed early (39K), late (p6.9), and very late (polh), has not been systematically assessed. Hence, we produced DNA constructs encoding secreted alkaline phosphatase (SEAP) under the transcriptional control of each of the four temporally distinct classes of baculovirus promoters, used them to transform insect cells, and compared the levels of SEAP RNA and protein production obtained before and after baculovirus infection. The ie1 construct was the only one that supported SEAP protein production by transformed insect cells prior to baculovirus infection, confirming that only immediate early promoters can be used to isolate transformed insect cells for constitutive recombinant protein production. However, baculovirus infection activated transgene expression by all four classes of baculovirus promoters. After infection, cells transformed with the very late (polh) and late (p6.9) promoter constructs produced the highest levels of SEAP RNA, but only low levels of SEAP protein. Conversely, cells transformed with the immediate early (ie1) and delayed early (39K) promoter constructs produced lower levels of RNA, but equal or higher levels of SEAP protein. Unexpectedly, the 39K promoter construct provided tightly regulated, baculovirus-inducible protein production at higher levels than the later promoter constructs. Thus, this study demonstrated the utility of the 39K promoter for insect cell engineering, particularly when one requires higher levels of effector protein production than obtained with ie1 and/or when constitutive transgene expression adversely impacts host cell fitness and/or genetic stability. Copyright © 2013 Elsevier B.V. All rights reserved.

Synaptic communication between neurons and NG2+ cells.

PubMed

Paukert, Martin; Bergles, Dwight E

2006-10-01

Chemical synaptic transmission provides the basis for much of the rapid signaling that occurs within neuronal networks. However, recent studies have provided compelling evidence that synapses are not used exclusively for communication between neurons. Physiological and anatomical studies indicate that a distinct class of glia known as NG2(+) cells also forms direct synaptic junctions with both glutamatergic and GABAergic neurons. Glutamatergic signaling can influence intracellular Ca(2+) levels in NG2(+) cells by activating Ca(2+) permeable AMPA receptors, and these inputs can be potentiated through high frequency stimulation. Although the significance of this highly differentiated form of communication remains to be established, these neuro-glia synapses might enable neurons to influence rapidly the behavior of this ubiquitous class of glial progenitors.

Adhesion, invasion and evasion: the many functions of the surface proteins of Staphylococcus aureus

PubMed Central

Foster, Timothy J.; Geoghegan, Joan A.; Ganesh, Vannakambadi K.; Höök, Magnus

2014-01-01

Staphylococcus aureus is an important opportunistic pathogen and persistently colonizes about 20% of the human population. Its surface is ‘decorated’ with proteins that are covalently anchored to the cell wall peptidoglycan. Structural and functional analysis has identified four distinct classes of surface proteins, of which microbial surface component recognizing adhesive matrix molecules (MSCRAMMs) are the largest class. These surface proteins have numerous functions, including adhesion to and invasion of host cells and tissues, evasion of immune responses and biofilm formation. Thus, cell wall-anchored proteins are essential virulence factors for the survival of S. aureus in the commensal state and during invasive infections, and targeting them with vaccines could combat S. aureus infections. PMID:24336184

Notch ligands Delta1 and Jagged1 transmit distinct signals to T-cell precursors

PubMed Central

Lehar, Sophie M.; Dooley, James; Farr, Andrew G.; Bevan, Michael J.

2009-01-01

Signaling through the Notch pathway plays an essential role in inducing T-lineage commitment and promoting the maturation of immature thymocytes. Using an in vitro culture system, we show that 2 different classes of Notch ligands, Jagged1 or Delta1, transmit distinct signals to T-cell progenitors. OP9 stromal cells expressing either Jagged1 or Delta1 inhibit the differentiation of DN1 thymocytes into the B-cell lineage, but only the Delta1-expressing stromal cells promote the proliferation and maturation of T-cell progenitors through the early double-negative (DN) stages of thymocyte development. Whereas the majority of bone marrow-derived stem cells do not respond to Jagged1 signals, T-cell progenitors respond to Jagged1 signals during a brief window of their development between the DN1 and DN3 stages of thymic development. During these stages, Jagged1 signals can influence the differentiation of immature thymocytes along the natural killer (NK) and γδ T-cell lineages. PMID:15486060

Bap31 enhances the ER export and quality control of human class I MHC molecules

PubMed Central

Ladasky, John J.; Boyle, Sarah; Seth, Malini; Li, Hewang; Pentcheva, Tsvetelina; Abe, Fumiyoshi; Steinberg, Steven J.; Edidin, Michael

2006-01-01

The assembly of class I MHC molecules and their export from the endoplasmic reticulum is governed by chaperones and accessory proteins. We present evidence that the putative cargo receptor protein Bap31 participates in the transport and the quality control of human class I molecules. Transfection of the human adenocarcinoma cell line HeLa with YFP-Bap31 chimeras increased surface levels of class I in a dose-dependent manner, by as much as 3.7-fold. The increase in surface class I resulted from an increase in the rate of export of newly-synthesized class I molecules to the cell surface and from an increase in the stability of the exported molecules. We propose that Bap31 performs quality control on class I molecules in two distinct phases: first, by exporting peptide-loaded class I molecules to the ERGIC and second, by retrieving class I molecules which have lost peptides in the acidic post-ER environment. This function of Bap31 is conditional or redundant, since we find that Bap31 deficiency does not reduce surface class I levels. Overexpression of the Bap31 homolog, Bap29, decreases surface class levels in HeLa, indicating that it does not substitute for Bap31. PMID:17056546

The leader peptide of mutacin 1140 has distinct structural components compared to related class I lantibiotics.

PubMed

Escano, Jerome; Stauffer, Byron; Brennan, Jacob; Bullock, Monica; Smith, Leif

2014-12-01

Lantibiotics are ribosomally synthesized peptide antibiotics composed of an N-terminal leader peptide that promotes the core peptide's interaction with the post translational modification (PTM) enzymes. Following PTMs, mutacin 1140 is transported out of the cell and the leader peptide is cleaved to yield the antibacterial peptide. Mutacin 1140 leader peptide is structurally unique compared to other class I lantibiotic leader peptides. Herein, we further our understanding of the structural differences of mutacin 1140 leader peptide with regard to other class I leader peptides. We have determined that the length of the leader peptide is important for the biosynthesis of mutacin 1140. We have also determined that mutacin 1140 leader peptide contains a novel four amino acid motif compared to related lantibiotics. PTM enzyme recognition of the leader peptide appears to be evolutionarily distinct from related class I lantibiotics. Our study on mutacin 1140 leader peptide provides a basis for future studies aimed at understanding its interaction with the PTM enzymes. © 2014 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.

Quantitating protein synthesis, degradation, and endogenous antigen processing.

PubMed

Princiotta, Michael F; Finzi, Diana; Qian, Shu-Bing; Gibbs, James; Schuchmann, Sebastian; Buttgereit, Frank; Bennink, Jack R; Yewdell, Jonathan W

2003-03-01

Using L929 cells, we quantitated the macroeconomics of protein synthesis and degradation and the microeconomics of producing MHC class I associated peptides from viral translation products. To maintain a content of 2.6 x 10(9) proteins, each cell's 6 x 10(6) ribosomes produce 4 x 10(6) proteins min(-1). Each of the cell's 8 x 10(5) proteasomes degrades 2.5 substrates min(-1), creating one MHC class I-peptide complex for each 500-3000 viral translation products degraded. The efficiency of complex formation is similar in dendritic cells and macrophages, which play a critical role in activating T cells in vivo. Proteasomes create antigenic peptides at different efficiencies from two distinct substrate pools: rapidly degraded newly synthesized proteins that clearly represent defective ribosomal products (DRiPs) and a less rapidly degraded pool in which DRiPs may also predominate.

Distinctive CD8+ T cell and MHC class I signatures in polycythemia vera patients.

PubMed

Cardoso, Elsa M; Esgalhado, André J; Patrão, Luís; Santos, Mónica; Neves, Vasco Pinto; Martinez, Jorge; Patto, Maria Assunção Vaz; Silva, Helena; Arosa, Fernando A

2018-05-22

Polycythemia vera (PV) is a myeloproliferative neoplasm characterized by overproduction of red blood cells. We have performed a comprehensive characterization of blood immune cells for expression of naïve and memory receptors as well as β 2 m-associated and β 2 m-free MHC class I heavy chains, also known as closed and open conformers, respectively, in PV patients and age-matched controls (CTR). We show that the peripheral CD3 + CD8 + T cell pool in PV patients is clearly divided into two discrete populations, a more granular CD3 + CD8 high T cell population enriched in effector-memory CD45RA + T cells (CD8 + TEMRA) when compared to CTR (P < 0.001), and a less granular CD3 + CD8 int T cell population that is completely absent in the CTR group (78 vs. 0%, P < 0.001) and is a mixture of naïve (CD8 + T N ) and CD8 + TEMRA cells expressing intermediate levels of CD28, i.e., CD3 + CD8 int CD28 int . While the percentage of CD3 + CD8 int TN cells correlated positively with the number of erythrocytes, the percentage of CD3 + CD8 int TEMRA correlated negatively with the number of platelets. Finally, we report that PV patients' lymphocytes and monocytes display lower levels of closed (W6/32 + ) MHC-I conformers at the cell surface while exhibiting increased amounts of open (HC-10 + ) MHC-I conformers. The implications of this distinctive immune signature are discussed.

Differential Alterations in Excitatory and Inhibitory Networks Involving Dentate Granule Cells Following Chronic Treatment with Distinct Classes of NMDAR Antagonists in Hippocampal Slice Cultures

DTIC Science & Technology

2010-03-08

1992; Jung and McNaughton, 1993); (2) low incidence of recurrent excitatory synapses between granule cells (Molnar and Nadler, 1999; Okazaki et al...neurons, dentate granule cells have a relatively more negative resting membrane potential and exhibit low-frequency firing (Staley et al., 1992; Jung ...inhibition plays a dual role in brain function and possibly seizure occurrence through balancing excitation and synchronizing neuronal firing. An

An incoherent regulatory network architecture that orchestrates B cell diversification in response to antigen signaling

PubMed Central

Sciammas, Roger; Li, Ying; Warmflash, Aryeh; Song, Yiqiang; Dinner, Aaron R; Singh, Harinder

2011-01-01

The B-lymphocyte lineage is a leading system for analyzing gene regulatory networks (GRNs) that orchestrate distinct cell fate transitions. Upon antigen recognition, B cells can diversify their immunoglobulin (Ig) repertoire via somatic hypermutation (SHM) and/or class switch DNA recombination (CSR) before differentiating into antibody-secreting plasma cells. We construct a mathematical model for a GRN underlying this developmental dynamic. The intensity of signaling through the Ig receptor is shown to control the bimodal expression of a pivotal transcription factor, IRF-4, which dictates B cell fate outcomes. Computational modeling coupled with experimental analysis supports a model of ‘kinetic control', in which B cell developmental trajectories pass through an obligate transient state of variable duration that promotes diversification of the antibody repertoire by SHM/CSR in direct response to antigens. More generally, this network motif could be used to translate a morphogen gradient into developmental inductive events of varying time, thereby enabling the specification of distinct cell fates. PMID:21613984

Classification of rocky headlands in California with relevance to littoral cell boundary delineation

USGS Publications Warehouse

George, Douglas A.; Largier, John L.; Storlazzi, Curt D.; Barnard, Patrick L.

2015-01-01

Despite extensive studies of hydrodynamics and sediment flux along beaches, there is little information on the processes, pathways and timing of water and sediment transport around rocky headlands. In this study, headlands along the California coast are classified to advance understanding of headland dynamics and littoral cell boundaries in support of improved coastal management decisions. Geomorphological parameters for 78 headlands were quantified from geological maps, remote-sensing imagery, navigational charts, and shoreline geospatial databases. K-means cluster analysis grouped the headlands into eight distinct classes based on headland perimeter, bathymetric slope ratio, and the headland apex angle. Wave data were used to investigate the potential for sediment transport around the headland types and determine the efficacy of the headland as a littoral cell boundary. Four classes of headland appear to function well as littoral cell boundaries, with headland size (e.g., perimeter or area) and a marked change in nearshore bathymetry across the headland being relevant attributes. About half of the traditional California littoral cell boundaries align with headland classes that are expected to perform poorly in blocking alongshore sediment transport, calling into question these boundaries. Better definition of these littoral cell boundaries is important for regional sediment management decisions.

A novel class of small RNAs bind to MILI protein in mouse testes.

PubMed

Aravin, Alexei; Gaidatzis, Dimos; Pfeffer, Sébastien; Lagos-Quintana, Mariana; Landgraf, Pablo; Iovino, Nicola; Morris, Patricia; Brownstein, Michael J; Kuramochi-Miyagawa, Satomi; Nakano, Toru; Chien, Minchen; Russo, James J; Ju, Jingyue; Sheridan, Robert; Sander, Chris; Zavolan, Mihaela; Tuschl, Thomas

2006-07-13

Small RNAs bound to Argonaute proteins recognize partially or fully complementary nucleic acid targets in diverse gene-silencing processes. A subgroup of the Argonaute proteins--known as the 'Piwi family'--is required for germ- and stem-cell development in invertebrates, and two Piwi members--MILI and MIWI--are essential for spermatogenesis in mouse. Here we describe a new class of small RNAs that bind to MILI in mouse male germ cells, where they accumulate at the onset of meiosis. The sequences of the over 1,000 identified unique molecules share a strong preference for a 5' uridine, but otherwise cannot be readily classified into sequence families. Genomic mapping of these small RNAs reveals a limited number of clusters, suggesting that these RNAs are processed from long primary transcripts. The small RNAs are 26-31 nucleotides (nt) in length--clearly distinct from the 21-23 nt of microRNAs (miRNAs) or short interfering RNAs (siRNAs)--and we refer to them as 'Piwi-interacting RNAs' or piRNAs. Orthologous human chromosomal regions also give rise to small RNAs with the characteristics of piRNAs, but the cloned sequences are distinct. The identification of this new class of small RNAs provides an important starting point to determine the molecular function of Piwi proteins in mammalian spermatogenesis.

Imbalanced PTEN and Phosphoinositide 3-kinase signaling impairs class switch recombination1

PubMed Central

Chen, Xiaomi; Dollin, Yonatan; Cambier, John C.; Wang, Jing H.

2015-01-01

Class switch recombination (CSR) generates isotype-switched antibodies with distinct effector functions. B cells express phosphatase and tensin homolog (PTEN) and multiple isoforms of class IA phosphoinositide 3-kinase (PI3K) catalytic subunits, including p110α and p110δ, whose roles in CSR remain unknown or controversial. Here, we demonstrate a direct effect of PTEN on CSR signaling by acute deletion of Pten specifically in mature B cells, thereby excluding the developmental impact of Pten deletion. We show that mature B cell-specific PTEN overexpression enhances CSR. More importantly, we establish a critical role of p110α in CSR. Furthermore, we identify a cooperative role of p110α and p110δ in suppressing CSR. Mechanistically, dysregulation of p110α or PTEN reversely affects activation-induced deaminase expression via modulating AKT activity. Thus, our study reveals that a signaling balance between PTEN and PI3K isoforms is essential to maintain normal CSR. PMID:26500350

Beyond the frontiers of neuronal types

PubMed Central

Battaglia, Demian; Karagiannis, Anastassios; Gallopin, Thierry; Gutch, Harold W.; Cauli, Bruno

2012-01-01

Cortical neurons and, particularly, inhibitory interneurons display a large diversity of morphological, synaptic, electrophysiological, and molecular properties, as well as diverse embryonic origins. Various authors have proposed alternative classification schemes that rely on the concomitant observation of several multimodal features. However, a broad variability is generally observed even among cells that are grouped into a same class. Furthermore, the attribution of specific neurons to a single defined class is often difficult, because individual properties vary in a highly graded fashion, suggestive of continua of features between types. Going beyond the description of representative traits of distinct classes, we focus here on the analysis of atypical cells. We introduce a novel paradigm for neuronal type classification, assuming explicitly the existence of a structured continuum of diversity. Our approach, grounded on the theory of fuzzy sets, identifies a small optimal number of model archetypes. At the same time, it quantifies the degree of similarity between these archetypes and each considered neuron. This allows highlighting archetypal cells, which bear a clear similarity to a single model archetype, and edge cells, which manifest a convergence of traits from multiple archetypes. PMID:23403725

Postaccess processes in the open vs. closed class distinction.

PubMed

Matthei, E H; Kean, M L

1989-02-01

We present the results of two auditory lexical decision experiments in which we attempted to replicate findings originally presented in Bradley (1978, Computational distinctions of vocabulary type, Ph.D. dissertation, MIT). The results obtained by Bradley were used as evidence for a processing distinction between the open and the closed class vocabularies; this distinction then used as part of an explanation for agrammatism in the comprehension and production of Broca's aphasics. In our first experiment we failed to replicate Bradley's result of frequency insensitivity in the closed class. Our second experiment, however, replicates Bradley's finding that closed class based nonwords (e.g., thanage) fail to induce interference effects in nonword decisions. We argue that our results, together with the various other reported failures to replicate Bradley's frequency insensitivity result, indicate that the open and closed classes may play distinct roles in postaccess phenomena involving the processing of morphological information but that such studies cannot address the question of whether the open vs. closed class distinction plays a role in syntactic processing.

Major histocompatibility complex class I-deficient NOD-B2mnull mice are diabetes and insulitis resistant.

PubMed

Serreze, D V; Leiter, E H; Christianson, G J; Greiner, D; Roopenian, D C

1994-03-01

Specific allelic combinations within the class II region of the major histocompatibility complex (MHC) represent a major genetic component for susceptibility to autoimmune insulin-dependent diabetes mellitus (IDDM) in humans. We produced and used a stock of NOD/Lt mice congenic for a functionally inactivated beta 2-microglobulin (B2mnull) locus to assess whether there was an absolute requirement for MHC class I expression and/or CD8+ T-cells in diabetogenesis. These NOD-B2mnull mice do not express cell surface MHC class I molecules or produce detectable levels of CD8+ T-cells and are diabetes and insulitis resistant. Previous results from transgenic mouse models indicated that intracellular accumulation of MHC class I molecules negatively affects pancreatic beta-cell function and can result in the development of nonautoimmune insulin-dependent diabetes mellitus (IDDM). MHC class I molecules have been shown to accumulate intracellularly in the presence of a disrupted B2m locus, but this mutation does not negatively affect plasma insulin levels in either NOD/Lt mice or in those of a mixed 129 and C57BL/6 genetic background. Interestingly, 14% of the male mice in this mixed background did develop hyperinsulinemia (> 1,500 pM) independent of the disrupted B2m locus, suggesting that these mice could conceivably develop insulin-resistant diabetes. However, none of these mice became diabetic at up to 22 months of age. Thus, elimination of cell surface MHC class I expression with a disrupted B2m gene blocks autoimmune diabetes in NOD/Lt mice, without engendering a separate, distinct form of glucose intolerance.

Purinergic receptor immunoreactivity in the rostral ventromedial medulla.

PubMed

Close, L N; Cetas, J S; Heinricher, M M; Selden, N R

2009-01-23

The rostral ventromedial medulla (RVM) has long been recognized to play a pivotal role in nociceptive modulation. Pro-nociception within the RVM is associated with a distinct functional class of neurons, ON-cells that begin to discharge immediately before nocifensive reflexes. Anti-nociceptive function within the RVM, including the analgesic response to opiates, is associated with another distinct class, OFF-cells, which pause immediately prior to nocifensive reflexes. A third class of RVM neurons, NEUTRAL-cells, does not alter firing in association with nocifensive reflexes. ON-, OFF- and NEUTRAL-cells show differential responsiveness to various behaviorally relevant neuromodulators, including purinergic ligands. Iontophoresis of semi-selective P2X ligands, which are associated with nociceptive transmission in the spinal cord and dorsal root ganglia, preferentially activate ON-cells. By contrast, P2Y ligands activate OFF-cells and P1 ligands suppress the firing of NEUTRAL cells. The current study investigates the distribution of P2X, P2Y and P1 receptor immunoreactivity in RVM neurons of Sprague-Dawley rats. Co-localization with tryptophan hydroxylase (TPH), a well-established marker for serotonergic neurons was also studied. Immunoreactivity for the four purinergic receptor subtypes examined was abundant in all anatomical subdivisions of the RVM. By contrast, TPH-immunoreactivity was restricted to a relatively small subset of RVM neurons concentrated in the nucleus raphe magnus and pallidus, as expected. There was a significant degree of co-localization of each purinergic receptor subtype with TPH-immunoreactivity. This co-localization was most pronounced for P2Y1 receptor immunoreactivity, although this was the least abundant among the different purinergic receptor subtypes examined. Immunoreactivity for multiple purinergic receptor subtypes was often co-localized in single neurons. These results confirm the physiological finding that purinergic receptors are widely expressed in the RVM. Purinergic neurotransmission in this region may play an important role in nociception and/or nociceptive modulation, as at other levels of the neuraxis.

Co-evolution of MHC class I and variable NK cell receptors in placental mammals.

PubMed

Guethlein, Lisbeth A; Norman, Paul J; Hilton, Hugo G; Parham, Peter

2015-09-01

Shaping natural killer (NK) cell functions in human immunity and reproduction are diverse killer cell immunoglobulin-like receptors (KIRs) that recognize polymorphic MHC class I determinants. A survey of placental mammals suggests that KIRs serve as variable NK cell receptors only in certain primates and artiodactyls. Divergence of the functional and variable KIRs in primates and artiodactyls predates placental reproduction. Among artiodactyls, cattle but not pigs have diverse KIRs. Catarrhine (humans, apes, and Old World monkeys) and platyrrhine (New World monkeys) primates, but not prosimians, have diverse KIRs. Platyrrhine and catarrhine systems of KIR and MHC class I are highly diverged, but within the catarrhines, a stepwise co-evolution of MHC class I and KIR is discerned. In Old World monkeys, diversification focuses on MHC-A and MHC-B and their cognate lineage II KIR. With evolution of C1-bearing MHC-C from MHC-B, as informed by orangutan, the focus changes to MHC-C and its cognate lineage III KIR. Evolution of C2 from C1 and fixation of MHC-C drove further elaboration of MHC-C-specific KIR, as exemplified by chimpanzee. In humans, the evolutionary trajectory changes again. Emerging from reorganization of the KIR locus and selective attenuation of KIR avidity for MHC class I are the functionally distinctive KIR A and KIR B haplotypes. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Distinct learning-induced changes in stimulus selectivity and interactions of GABAergic interneuron classes in visual cortex.

PubMed

Khan, Adil G; Poort, Jasper; Chadwick, Angus; Blot, Antonin; Sahani, Maneesh; Mrsic-Flogel, Thomas D; Hofer, Sonja B

2018-06-01

How learning enhances neural representations for behaviorally relevant stimuli via activity changes of cortical cell types remains unclear. We simultaneously imaged responses of pyramidal cells (PYR) along with parvalbumin (PV), somatostatin (SOM), and vasoactive intestinal peptide (VIP) inhibitory interneurons in primary visual cortex while mice learned to discriminate visual patterns. Learning increased selectivity for task-relevant stimuli of PYR, PV and SOM subsets but not VIP cells. Strikingly, PV neurons became as selective as PYR cells, and their functional interactions reorganized, leading to the emergence of stimulus-selective PYR-PV ensembles. Conversely, SOM activity became strongly decorrelated from the network, and PYR-SOM coupling before learning predicted selectivity increases in individual PYR cells. Thus, learning differentially shapes the activity and interactions of multiple cell classes: while SOM inhibition may gate selectivity changes, PV interneurons become recruited into stimulus-specific ensembles and provide more selective inhibition as the network becomes better at discriminating behaviorally relevant stimuli.

Phenotype and function of T cells infiltrating visceral metastases from gastrointestinal cancers and melanoma: implications for adoptive cell transfer therapy.

PubMed

Turcotte, Simon; Gros, Alena; Hogan, Katherine; Tran, Eric; Hinrichs, Christian S; Wunderlich, John R; Dudley, Mark E; Rosenberg, Steven A

2013-09-01

Adoptive cell transfer of tumor-infiltrating lymphocytes (TILs) can mediate cancer regression in patients with metastatic melanoma, but whether this approach can be applied to common epithelial malignancies remains unclear. In this study, we compared the phenotype and function of TILs derived from liver and lung metastases from patients with gastrointestinal (GI) cancers (n = 14) or melanoma (n = 42). Fewer CD3(+) T cells were found to infiltrate GI compared with melanoma metastases, but the proportions of CD8(+) cells, T cell differentiation stage, and expression of costimulatory molecules were similar for both tumor types. Clinical-scale expansion up to ~50 × 10(9) T cells on average was obtained for all patients with GI cancer and melanoma. From GI tumors, however, TIL outgrowth in high-dose IL-2 yielded 22 ± 1.4% CD3(+)CD8(+) cells compared with 63 ± 2.4% from melanoma (p < 0.001). IFN-γ ELISA demonstrated MHC class I-mediated reactivity of TIL against autologous tumor in 5 of 7 GI cancer patients tested (9% of 188 distinct TIL cultures) and in 9 of 10 melanoma patients (43% of 246 distinct TIL cultures). In these assays, MHC class I-mediated up-regulation of CD137 (4-1BB) expression on CD8(+) cells suggested that 0-3% of TILs expanded from GI cancer metastases were tumor-reactive. This study implies that the main challenge to the development of TIL adoptive cell transfer for metastatic GI cancers may not be the in vitro expansion of bulk TILs, but the ability to select and enrich for tumor-reactive T cells.

Phenotype and Function of T Cells Infiltrating Visceral Metastases from Gastrointestinal Cancers and Melanoma: Implications for Adoptive Cell Transfer Therapy

PubMed Central

Turcotte, Simon; Gros, Alena; Hogan, Katherine; Tran, Eric; Hinrichs, Christian S.; Wunderlich, John R.; Dudley, Mark E.

2013-01-01

Adoptive cell transfer of tumor-infiltrating lymphocytes (TILs) can mediate cancer regression in patients with metastatic melanoma, but whether this approach can be applied to common epithelial malignancies remains unclear. In this study, we compared the phenotype and function of TILs derived from liver and lung metastases from patients with gastrointestinal (GI) cancers (n = 14) or melanoma (n = 42). Fewer CD3+ T cells were found to infiltrate GI compared with melanoma metastases, but the proportions of CD8+ cells, T cell differentiation stage, and expression of costimulatory molecules were similar for both tumor types. Clinical-scale expansion up to ∼50 × 109 T cells on average was obtained for all patients with GI cancer and melanoma. From GI tumors, however, TIL outgrowth in high-dose IL-2 yielded 22 ± 1.4% CD3+CD8+ cells compared with 63 ± 2.4% from melanoma (p < 0.001). IFN-γ ELISA demonstrated MHC class I–mediated reactivity of TIL against autologous tumor in 5 of 7 GI cancer patients tested (9% of 188 distinct TIL cultures) and in 9 of 10 melanoma patients (43% of 246 distinct TIL cultures). In these assays, MHC class I–mediated up-regulation of CD137 (4-1BB) expression on CD8+ cells suggested that 0–3% of TILs expanded from GI cancer metastases were tumor-reactive. This study implies that the main challenge to the development of TIL adoptive cell transfer for metastatic GI cancers may not be the in vitro expansion of bulk TILs, but the ability to select and enrich for tumor-reactive T cells. PMID:23904171

Voltage-Gated Ion Channels in Cancer Cell Proliferation

PubMed Central

Rao, Vidhya R.; Perez-Neut, Mathew; Kaja, Simon; Gentile, Saverio

2015-01-01

Changes of the electrical charges across the surface cell membrane are absolutely necessary to maintain cellular homeostasis in physiological as well as in pathological conditions. The opening of ion channels alter the charge distribution across the surface membrane as they allow the diffusion of ions such as K+, Ca++, Cl−, Na+. Traditionally, voltage-gated ion channels (VGIC) are known to play fundamental roles in controlling rapid bioelectrical signaling including action potential and/or contraction. However, several investigations have revealed that these classes of proteins can also contribute significantly to cell mitotic biochemical signaling, cell cycle progression, as well as cell volume regulation. All these functions are critically important for cancer cell proliferation. Interestingly, a variety of distinct VGICs are expressed in different cancer cell types, including metastasis but not in the tissues from which these tumors were generated. Given the increasing evidence suggesting that VGIC play a major role in cancer cell biology, in this review we discuss the role of distinct VGIC in cancer cell proliferation and possible therapeutic potential of VIGC pharmacological manipulation. PMID:26010603

Ionotropic and metabotropic mechanisms in chemoreception: 'chance or design'?

PubMed

Silbering, Ana Florencia; Benton, Richard

2010-03-01

Chemosensory receptors convert an enormous diversity of chemical signals from the external world into a common language of electrical activity in the brain. Mammals and insects use several families of transmembrane receptor proteins to recognize distinct classes of volatile and non-volatile chemicals that are produced by conspecifics or other environmental sources. A comparison of the signalling mechanisms of mammalian and insect receptors has revealed an unexpected functional distinction: mammals rely almost exclusively on metabotropic ligand-binding receptors, which use second messenger signalling cascades to indirectly activate ion channels, whereas insects use ionotropic receptors, which are gated directly by chemical stimuli, thereby leading to neuronal depolarization. In this review, we consider possible reasons for this dichotomy, taking into account biophysical, cell biological, ecological and evolutionary influences on how information is extracted from chemosensory cues by these animal classes.

Ionotropic and metabotropic mechanisms in chemoreception: ‘chance or design'?

PubMed Central

Silbering, Ana Florencia; Benton, Richard

2010-01-01

Chemosensory receptors convert an enormous diversity of chemical signals from the external world into a common language of electrical activity in the brain. Mammals and insects use several families of transmembrane receptor proteins to recognize distinct classes of volatile and non-volatile chemicals that are produced by conspecifics or other environmental sources. A comparison of the signalling mechanisms of mammalian and insect receptors has revealed an unexpected functional distinction: mammals rely almost exclusively on metabotropic ligand-binding receptors, which use second messenger signalling cascades to indirectly activate ion channels, whereas insects use ionotropic receptors, which are gated directly by chemical stimuli, thereby leading to neuronal depolarization. In this review, we consider possible reasons for this dichotomy, taking into account biophysical, cell biological, ecological and evolutionary influences on how information is extracted from chemosensory cues by these animal classes. PMID:20111052

Cuticular Waxes of Arabidopsis thaliana Shoots: Cell-Type-Specific Composition and Biosynthesis

PubMed Central

Hegebarth, Daniela; Jetter, Reinhard

2017-01-01

It is generally assumed that all plant epidermis cells are covered with cuticles, and the distinct surface geometries of pavement cells, guard cells, and trichomes imply functional differences and possibly different wax compositions. However, experiments probing cell-type-specific wax compositions and biosynthesis have been lacking until recently. This review summarizes new evidence showing that Arabidopsis trichomes have fewer wax compound classes than pavement cells, and higher amounts of especially long-chain hydrocarbons. The biosynthesis machinery generating this characteristic surface coating is discussed. Interestingly, wax compounds with similar, long hydrocarbon chains had been identified previously in some unrelated species, not all of them bearing trichomes. PMID:28686187

Peripheral blood mononuclear cells analysis in microfluidic flow by coherent imaging tools

NASA Astrophysics Data System (ADS)

Dannhauser, David; Rossi, Domenico; Memmolo, Pasquale; Causa, Filippo; Finizio, Andrea; Ferraro, Pietro; Netti, Paolo A.

2017-06-01

Cell of human blood stream are divided into two groups: Red Blood Cells (RBC) and White Blood Cells (WBC). RBC have a peculiar biconcave disk shape and they are responsible for the delivering of O2 and CO2 through the body. WBC are a more widespread class of cell ensuring immunity against pathogens. They can be divided in two main classes: granulocyte cells and A-granulocyte cells. Neutrophils, basophils and eosinophils belong to the granulocyte cell class, while lymphocytes and monocytes belong to A-granulocyte. Both in RBC and WBC, the intrinsic physical properties of a cell are indicators of cell condition and, furthermore, of the overall human body state. Thus, the accurate comprehension of the physiological structure of WBCs is fundamental to recognize diseases. Here we show the possibility to simple and straightforwardly characterize the physical properties of individual RBC and mononuclear WBC in a microfluidic context, using a wide angle light scattering apparatus and a corresponding theoretical simulation of Optical Signature (OS). A non-Newtonian polymer alignment solution for cell is used to ensure an individual cell alignment in the microfluidic flow, thus permitting a precise investigation. Additionally, Quantitative Phase Imaging (QPI) holographic measurements are performed to estimate cell morphometric features, such as their refractive index. We analyzed more than 200 WBCs and 100 RBCs of three different probands. Results showed distinct cell populations according to their measured dimensions and shape, which can be associated to the presence of RBC, lymphocytes and monocytes.

Antagonism of human CC-chemokine receptor 4 can be achieved through three distinct binding sites on the receptor

PubMed Central

Slack, Robert J; Russell, Linda J; Barton, Nick P; Weston, Cathryn; Nalesso, Giovanna; Thompson, Sally-Anne; Allen, Morven; Chen, Yu Hua; Barnes, Ashley; Hodgson, Simon T; Hall, David A

2013-01-01

Chemokine receptor antagonists appear to access two distinct binding sites on different members of this receptor family. One class of CCR4 antagonists has been suggested to bind to a site accessible from the cytoplasm while a second class did not bind to this site. In this report, we demonstrate that antagonists representing a variety of structural classes bind to two distinct allosteric sites on CCR4. The effects of pairs of low-molecular weight and/or chemokine CCR4 antagonists were evaluated on CCL17- and CCL22-induced responses of human CCR4+ T cells. This provided an initial grouping of the antagonists into sets which appeared to bind to distinct binding sites. Binding studies were then performed with radioligands from each set to confirm these groupings. Some novel receptor theory was developed to allow the interpretation of the effects of the antagonist combinations. The theory indicates that, generally, the concentration-ratio of a pair of competing allosteric modulators is maximally the sum of their individual effects while that of two modulators acting at different sites is likely to be greater than their sum. The low-molecular weight antagonists could be grouped into two sets on the basis of the functional and binding experiments. The antagonistic chemokines formed a third set whose behaviour was consistent with that of simple competitive antagonists. These studies indicate that there are two allosteric regulatory sites on CCR4. PMID:25505571

Genomic profiling of rice sperm cell transcripts reveals conserved and distinct elements in the flowering plant male germ lineage.

PubMed

Russell, Scott D; Gou, Xiaoping; Wong, Chui E; Wang, Xinkun; Yuan, Tong; Wei, Xiaoping; Bhalla, Prem L; Singh, Mohan B

2012-08-01

Genomic assay of sperm cell RNA provides insight into functional control, modes of regulation, and contributions of male gametes to double fertilization. Sperm cells of rice (Oryza sativa) were isolated from field-grown, disease-free plants and RNA was processed for use with the full-genome Affymetrix microarray. Comparison with Gene Expression Omnibus (GEO) reference arrays confirmed expressionally distinct gene profiles. A total of 10,732 distinct gene sequences were detected in sperm cells, of which 1668 were not expressed in pollen or seedlings. Pathways enriched in male germ cells included ubiquitin-mediated pathways, pathways involved in chromatin modeling including histones, histone modification and nonhistone epigenetic modification, and pathways related to RNAi and gene silencing. Genome-wide expression patterns in angiosperm sperm cells indicate common and divergent themes in the male germline that appear to be largely self-regulating through highly up-regulated chromatin modification pathways. A core of highly conserved genes appear common to all sperm cells, but evidence is still emerging that another class of genes have diverged in expression between monocots and dicots since their divergence. Sperm cell transcripts present at fusion may be transmitted through plasmogamy during double fertilization to effect immediate post-fertilization expression of early embryo and (or) endosperm development. © 2012 The Authors. New Phytologist © 2012 New Phytologist Trust.

A distinct class of dominant negative Ras mutants: cytosolic GTP-bound Ras effector domain mutants that inhibit Ras signaling and transformation and enhance cell adhesion.

PubMed

Fiordalisi, James J; Holly, Stephen P; Johnson, Ronald L; Parise, Leslie V; Cox, Adrienne D

2002-03-29

Cytosolic GTP-bound Ras has been shown to act as a dominant negative (DN) inhibitor of Ras by sequestering Raf in non-productive cytosolic complexes. Nevertheless, this distinct class of DN mutants has been neither well characterized nor extensively used to analyze Ras signaling. In contrast, DN Ras17N, which functions by blocking Ras guanine nucleotide exchange factors, has been well characterized and is widely used. Cytosolic GTP-bound Ras mutants could be used to inhibit particular Ras effectors by introducing additional mutations (T35S, E37G or Y40C) that permit them to associate selectively with and inhibit Raf, RalGDS, or phosphoinositide 3-kinase, respectively. When the wild-type Ras effector binding region is used, cytosolic Ras should associate with all Ras effectors, even those that are not yet identified, making these DN Ras mutants effective inhibitors of multiple Ras functions. We generated cytosolic GTP-bound H-, N-, and K-Ras, and we assessed their ability to inhibit Ras-induced phenotypes. In fibroblasts, cytosolic H-, N-, and K-Ras inhibited Ras-induced Elk-1 activation and focus formation, induced a flattened cell morphology, and increased adhesion to fibronectin through modulation of a beta(1)-subunit-containing integrin, thereby demonstrating that DN activity is not limited to a subset of Ras isoforms. We also generated cytosolic GTP-bound Ras effector domain mutants (EDMs), each of which reduced the ability of cytosolic GTP-bound Ras proteins to inhibit Elk-1 activation and to induce cell flattening, implicating multiple pathways in these phenotypes. In contrast, Ras-induced focus formation, platelet-derived growth factor (PDGF)-, or Ras-induced phospho-Akt levels and cell adhesion to fibronectin were affected by T35S and Y40C EDMs, whereas PDGF- or Ras-induced phospho-Erk levels were affected only by the T35S EDM, implying that a more limited set of Ras-mediated pathways participate in these phenotypes. These data constitute the first extensive characterization of this functionally distinct class of DN Ras inhibitor proteins.

Incidence and management of toxicity associated with ibrutinib and idelalisib: a practical approach

PubMed Central

de Weerdt, Iris; Koopmans, Suzanne M.; Kater, Arnon P.; van Gelder, Michel

2017-01-01

The use of novel B-cell receptor signaling inhibitors results in high response rates and long progression-free survival in patients with indolent B-cell malignancies, such as chronic lymphocytic leukemia, follicular lymphoma, mantle cell lymphoma and Waldenström macroglobulinemia. Ibrutinib, the first-in-class inhibitor of Bruton tyrosine kinase, and idelalisib, the first-in-class inhibitor of phosphatidylinositol 3-kinase δ, have recently been approved for the treatment of several indolent B-cell malignancies. These drugs are especially being used for previously unmet needs, i.e., for patients with relapsed or refractory disease, high-risk cytogenetic or molecular abnormalities, or with comorbidities. Treatment with ibrutinib and idelalisib is generally well tolerated, even by elderly patients. However, the use of these drugs may come with toxicities that are distinct from the side effects of immunochemotherapy. In this review we discuss the most commonly reported and/or most clinically relevant adverse events associated with these B-cell receptor inhibitors, with special emphasis on recommendations for their management. PMID:28775119

Plant vegetative and animal cytoplasmic actins share functional competence for spatial development with protists.

PubMed

Kandasamy, Muthugapatti K; McKinney, Elizabeth C; Roy, Eileen; Meagher, Richard B

2012-05-01

Actin is an essential multifunctional protein encoded by two distinct ancient classes of genes in animals (cytoplasmic and muscle) and plants (vegetative and reproductive). The prevailing view is that each class of actin variants is functionally distinct. However, we propose that the vegetative plant and cytoplasmic animal variants have conserved functional competence for spatial development inherited from an ancestral protist actin sequence. To test this idea, we ectopically expressed animal and protist actins in Arabidopsis thaliana double vegetative actin mutants that are dramatically altered in cell and organ morphologies. We found that expression of cytoplasmic actins from humans and even a highly divergent invertebrate Ciona intestinalis qualitatively and quantitatively suppressed the root cell polarity and organ defects of act8 act7 mutants and moderately suppressed the root-hairless phenotype of act2 act8 mutants. By contrast, human muscle actins were unable to support prominently any aspect of plant development. Furthermore, actins from three protists representing Choanozoa, Archamoeba, and green algae efficiently suppressed all the phenotypes of both the plant mutants. Remarkably, these data imply that actin's competence to carry out a complex suite of processes essential for multicellular development was already fully developed in single-celled protists and evolved nonprogressively from protists to plants and animals.

Plant Vegetative and Animal Cytoplasmic Actins Share Functional Competence for Spatial Development with Protists[W][OA

PubMed Central

Kandasamy, Muthugapatti K.; McKinney, Elizabeth C.; Roy, Eileen; Meagher, Richard B.

2012-01-01

Actin is an essential multifunctional protein encoded by two distinct ancient classes of genes in animals (cytoplasmic and muscle) and plants (vegetative and reproductive). The prevailing view is that each class of actin variants is functionally distinct. However, we propose that the vegetative plant and cytoplasmic animal variants have conserved functional competence for spatial development inherited from an ancestral protist actin sequence. To test this idea, we ectopically expressed animal and protist actins in Arabidopsis thaliana double vegetative actin mutants that are dramatically altered in cell and organ morphologies. We found that expression of cytoplasmic actins from humans and even a highly divergent invertebrate Ciona intestinalis qualitatively and quantitatively suppressed the root cell polarity and organ defects of act8 act7 mutants and moderately suppressed the root-hairless phenotype of act2 act8 mutants. By contrast, human muscle actins were unable to support prominently any aspect of plant development. Furthermore, actins from three protists representing Choanozoa, Archamoeba, and green algae efficiently suppressed all the phenotypes of both the plant mutants. Remarkably, these data imply that actin’s competence to carry out a complex suite of processes essential for multicellular development was already fully developed in single-celled protists and evolved nonprogressively from protists to plants and animals. PMID:22589468

A caspase-2-RFXANK interaction and its implication for MHC class II expression.

PubMed

Forsberg, Jeremy; Li, Xinge; Akpinar, Birce; Salvatori, Roger; Ott, Martin; Zhivotovsky, Boris; Olsson, Magnus

2018-01-23

Despite recent achievements implicating caspase-2 in tumor suppression, the enzyme stands out from the apoptotic caspase family as a factor whose function requires further clarification. To specify enzyme characteristics through the definition of interacting proteins in apoptotic or non-apoptotic settings, a yeast 2-hybrid (Y2H) screen was performed using the full-length protein as bait. The current report describes the analysis of a captured prey and putative novel caspase-2 interacting factor, the regulatory factor X-associated ankyrin-containing protein (RFXANK), previously associated with CIITA, the transactivator regulating cell-type specificity and inducibility of MHC class II gene expression. The interaction between caspase-2 and RFXANK was verified by co-immunoprecipitations using both exogenous and endogenous proteins, where the latter approach suggested that binding of the components occurs in the cytoplasm. Cellular co-localization was confirmed by transfection of fluorescently conjugated proteins. Enhanced caspase-2 processing in RFXANK-overexpressing HEK293T cells treated with chemotherapeutic agents further supported Y2H data. Yet, no distinct differences with respect to MHC class II expression were observed in plasma membranes of antigen-presenting cells derived from wild type and caspase-2 -/- mice. In contrast, increased levels of the total MHC class II protein was evident in protein lysates from caspase-2 RNAi-silenced leukemia cell lines and B-cells isolated from gene-targeted mice. Together, these data identify a novel caspase-2-interacting factor, RFXANK, and indicate a potential non-apoptotic role for the enzyme in the control of MHC class II gene regulation.

Brain oxygen tension controls the expansion of outer subventricular zone-like basal progenitors in the developing mouse brain.

PubMed

Wagenführ, Lisa; Meyer, Anne K; Braunschweig, Lena; Marrone, Lara; Storch, Alexander

2015-09-01

The mammalian neocortex shows a conserved six-layered structure that differs between species in the total number of cortical neurons produced owing to differences in the relative abundance of distinct progenitor populations. Recent studies have identified a new class of proliferative neurogenic cells in the outer subventricular zone (OSVZ) in gyrencephalic species such as primates and ferrets. Lissencephalic brains of mice possess fewer OSVZ-like progenitor cells and these do not constitute a distinct layer. Most in vitro and in vivo studies have shown that oxygen regulates the maintenance, proliferation and differentiation of neural progenitor cells. Here we dissect the effects of fetal brain oxygen tension on neural progenitor cell activity using a novel mouse model that allows oxygen tension to be controlled within the hypoxic microenvironment in the neurogenic niche of the fetal brain in vivo. Indeed, maternal oxygen treatment of 10%, 21% and 75% atmospheric oxygen tension for 48 h translates into robust changes in fetal brain oxygenation. Increased oxygen tension in fetal mouse forebrain in vivo leads to a marked expansion of a distinct proliferative cell population, basal to the SVZ. These cells constitute a novel neurogenic cell layer, similar to the OSVZ, and contribute to corticogenesis by heading for deeper cortical layers as a part of the cortical plate. © 2015. Published by The Company of Biologists Ltd.

Class Matters

ERIC Educational Resources Information Center

Valdata, Patricia

2005-01-01

Ever since George Washington opted for the title of president rather than king, Americans have been uncomfortable with the idea of class distinctions. This article presents an interview with Dr. Janet Galligani Casey regarding the idea of class distinctions. Galligani Casey, who grew up in a working-class neighborhood in Somerville, Massachusetts,…

Exosomal microRNA Biomarkers: Emerging Frontiers in Colorectal and Other Human Cancers

PubMed Central

Goel, Ajay; Tovar-Camargo, Oscar A; Toden, Shusuke

2016-01-01

Diagnostic strategies, particularly non-invasive blood-based screening approaches, are gaining increased attention for the early detection and attenuation of mortality associated with colorectal cancer (CRC). However, the majority of current screening approaches are inadequate at replacing the conventional CRC diagnostic procedures. Yet, due to technological advances and a better understanding of molecular events underlying human cancer, a new category of biomarkers are on the horizon. Recent evidence indicates that cells release a distinct class of small vesicles called ‘exosomes’, which contain nucleic acids and proteins that reflect and typify host-cell molecular architecture. Intriguingly, exosomes released from cancer cells have a distinct genetic and epigenetic makeup, which allows them to undertake their tumorigenic function. From a clinical standpoint, these unique cancer-specific fingerprints present in exosomes appear to be detectable in a small amount of blood, making them very attractive substrates for developing cancer biomarkers, particularly noninvasive diagnostic approaches. PMID:26892862

Differential response of cancer cells to HDAC inhibitors trichostatin A and depsipeptide.

PubMed

Chang, J; Varghese, D S; Gillam, M C; Peyton, M; Modi, B; Schiltz, R L; Girard, L; Martinez, E D

2012-01-03

Over the last decade, several drugs that inhibit class I and/or class II histone deacetylases (HDACs) have been identified, including trichostatin A, the cyclic depsipeptide FR901228 and the antibiotic apicidin. These compounds have had immediate application in cancer research because of their ability to reactivate aberrantly silenced tumour suppressor genes and/or block tumour cell growth. Although a number of HDAC inhibitors are being evaluated in preclinical cancer models and in clinical trials, little is known about the differences in their specific mechanism of action and about the unique determinants of cancer cell sensitivity to each of these inhibitors. Using a combination of cell viability assays, HDAC enzyme activity measurements, western blots for histone modifications, microarray gene expression analysis and qRT-PCR, we have characterised differences in trichostatin A vs depsipeptide-induced phenotypes in lung cancer, breast cancer and skin cancer cells and in normal cells and have then expanded these studies to other HDAC inhibitors. Cell viability profiles across panels of lung cancer, breast cancer and melanoma cell lines showed distinct sensitivities to the pan-inhibitor TSA compared with the class 1 selective inhibitor depsipeptide. In several instances, the cell lines most sensitive to one inhibitor were most resistant to the other inhibitor, demonstrating these drugs act on at least some non-overlapping cellular targets. These differences were not explained by the HDAC selectivity of these inhibitors alone since apicidin, which is a class 1 selective compound similar to depsipeptide, also showed a unique drug sensitivity profile of its own. TSA had greater specificity for cancer vs normal cells compared with other HDAC inhibitors. In addition, at concentrations that blocked cancer cell viability, TSA effectively inhibited purified recombinant HDACs 1, 2 and 5 and moderately inhibited HDAC8, while depsipeptide did not inhibit the activity of purified HDACs in vitro but did in cellular extracts, suggesting a potentially indirect action of this drug. Although both depsipeptide and TSA increased levels of histone acetylation in cancer cells, only depsipeptide decreased global levels of transcriptionally repressive histone methylation marks. Analysis of gene expression profiles of an isogenic cell line pair that showed discrepant sensitivity to depsipeptide, suggested that resistance to this inhibitor may be mediated by increased expression of multidrug resistance genes triggered by exposure to chemotherapy as was confirmed by verapamil studies. Although generally thought to have similar activities, the HDAC modulators trichostatin A and depsipeptide demonstrated distinct phenotypes in the inhibition of cancer cell viability and of HDAC activity, in their selectivity for cancer vs normal cells, and in their effects on histone modifications. These differences in mode of action may bear on the future therapeutic and research application of these inhibitors.

The use of ebselen for radioprotection in cultured cells and mice.

PubMed

Tak, Jean Kyoung; Park, Jeen-Woo

2009-04-15

Ionizing radiation induces the production of reactive oxygen species (ROS), which play an important causative role in cell death. Therefore, compounds that control the level of ROS may confer radioprotective effects. Ebselen, a seleno-organic compound, has been shown to protect against cell injury caused by ROS. The objective of this study was to examine the effects of ebselen on radiation-dependent toxicity. We investigated the protective role of ebselen against ionizing radiation in U937 cells and mice. Upon exposure to 20 Gy of gamma-irradiation, there was a distinct difference between untreated cells and the cells pretreated with 5 microM ebselen for 2 h with respect to viability, cellular redox status, and oxidative damage to cells. When cells were exposed to 2 Gy of gamma-irradiation, there was a distinct difference between the untreated cells and the cells pretreated with ebselen with respect to apoptotic features and mitochondrial function. Ebselen administration for 14 days at a daily dosage of 10 mg/kg provided substantial protection against killing and oxidative damage to mice exposed to whole-body irradiation. These data indicate that ebselen may have great potential as a new class of in vivo, non-sulfur-containing radiation protector.

Generation and preservation of the slow underlying membrane potential oscillation in model bursting neurons.

PubMed

Franklin, Clarence C; Ball, John M; Schulz, David J; Nair, Satish S

2010-09-01

The underlying membrane potential oscillation of both forced and endogenous slow-wave bursting cells affects the number of spikes per burst, which in turn affects outputs downstream. We use a biophysical model of a class of slow-wave bursting cells with six active currents to investigate and generalize correlations among maximal current conductances that might generate and preserve its underlying oscillation. We propose three phases for the underlying oscillation for this class of cells: generation, maintenance, and termination and suggest that different current modules coregulate to preserve the characteristics of each phase. Coregulation of I(Burst) and I(A) currents within distinct boundaries maintains the dynamics during the generation phase. Similarly, coregulation of I(CaT) and I(Kd) maintains the peak and duration of the underlying oscillation, whereas the calcium-activated I(KCa) ensures appropriate termination of the oscillation and adjusts the duration independent of peak.

Serum angiotensin-1 converting enzyme activity processes a human immunodeficiency virus 1 gp160 peptide for presentation by major histocompatibility complex class I molecules

PubMed Central

1992-01-01

T cell stimulation by the human immunodeficiency virus 1 gp160-derived peptide p18 presented by H-2Dd class I major histocompatibility complex molecules in a cell-free system was found to require proteolytic cleavage. This extracellular processing was mediated by peptidases present in fetal calf serum. In vitro processing of p18 resulted in a distinct reverse phase high performance liquid chromatography profile, from which a biologically active product was isolated and sequenced. This peptide processing can be specifically blocked by the angiotensin- 1 converting enzyme (ACE) inhibitor captopril, and can occur by exposing p18 to purified ACE. The ability of naturally occurring extracellular proteases to convert inactive peptides to T cell antigens has important implications for understanding cytotoxic T lymphocyte responses in vivo, and for rational peptide vaccine design. PMID:1316930

CIITA promoter I CARD-deficient mice express functional MHC class II genes in myeloid and lymphoid compartments.

PubMed

Zinzow-Kramer, W M; Long, A B; Youngblood, B A; Rosenthal, K M; Butler, R; Mohammed, A-U-R; Skountzou, I; Ahmed, R; Evavold, B D; Boss, J M

2012-06-01

Three distinct promoters control the master regulator of major histocompatibility complex (MHC) class II expression, class II transactivator (CIITA), in a cell type-specific manner. Promoter I (pI) CIITA, expressed primarily by dendritic cells (DCs) and macrophages, expresses a unique isoform that contains a caspase-recruitment domain (CARD). The activity and function of this isoform are not understood, but are believed to enhance the function of CIITA in antigen-presenting cells. To determine whether isoform I of CIITA has specific functions, CIITA mutant mice were created in which isoform I was replaced with isoform III sequences. Mice in which pI and the CARD-encoding exon were deleted were also created. No defect in the formation of CD4 T cells, the ability to respond to a model antigen or bacterial or viral challenge was observed in mice lacking CIITA isoform I. Although CIITA and MHC-II expression was decreased in splenic DCs, pI knockout animals expressed CIITA from downstream promoters, suggesting that control of pI activity is mediated by unknown distal elements that could act at pIII, the B-cell promoter. Thus, no critical function is linked to the CARD domain of CIITA isoform I with respect to basic immune system development, function and challenge.

The Drosophila blood-brain barrier: development and function of a glial endothelium.

PubMed

Limmer, Stefanie; Weiler, Astrid; Volkenhoff, Anne; Babatz, Felix; Klämbt, Christian

2014-01-01

The efficacy of neuronal function requires a well-balanced extracellular ion homeostasis and a steady supply with nutrients and metabolites. Therefore, all organisms equipped with a complex nervous system developed a so-called blood-brain barrier, protecting it from an uncontrolled entry of solutes, metabolites or pathogens. In higher vertebrates, this diffusion barrier is established by polarized endothelial cells that form extensive tight junctions, whereas in lower vertebrates and invertebrates the blood-brain barrier is exclusively formed by glial cells. Here, we review the development and function of the glial blood-brain barrier of Drosophila melanogaster. In the Drosophila nervous system, at least seven morphologically distinct glial cell classes can be distinguished. Two of these glial classes form the blood-brain barrier. Perineurial glial cells participate in nutrient uptake and establish a first diffusion barrier. The subperineurial glial (SPG) cells form septate junctions, which block paracellular diffusion and thus seal the nervous system from the hemolymph. We summarize the molecular basis of septate junction formation and address the different transport systems expressed by the blood-brain barrier forming glial cells.

Burkitt Lymphoma: Pathogenesis and Immune Evasion

PubMed Central

God, Jason M.; Haque, Azizul

2010-01-01

B-cell lymphomas arise at distinct stages of cellular development and maturation, potentially influencing antigen (Ag) presentation and T-cell recognition. Burkitt lymphoma (BL) is a highly malignant B-cell tumor associated with Epstein-Barr Virus (EBV) infection. Although BL can be effectively treated in adults and children, leading to high survival rates, its ability to mask itself from the immune system makes BL an intriguing disease to study. In this paper, we will provide an overview of BL and its association with EBV and the c-myc oncogene. The contributions of EBV and c-myc to B-cell transformation, proliferation, or attenuation of cellular network and immune recognition or evasion will be summarized. We will also discuss the various pathways by which BL escapes immune detection by inhibiting both HLA class I- and II-mediated Ag presentation to T cells. Finally, we will provide an overview of recent developments suggesting the existence of BL-associated inhibitory molecules that may block HLA class II-mediated Ag presentation to CD4+ T cells, facilitating immune escape of BL. PMID:20953370

A Remarkably Simple Class of Imidazolium-Based Lipids and Their Biological Properties.

PubMed

Wang, Da; Richter, Christian; Rühling, Andreas; Drücker, Patrick; Siegmund, Daniel; Metzler-Nolte, Nils; Glorius, Frank; Galla, Hans-Joachim

2015-10-19

A series of imidazolium salts bearing two alkyl chains in the backbone of the imidazolium core were synthesized, resembling the structure of lipids. Their antibacterial activity and cytotoxicity were evaluated using Gram-positive and Gram-negative bacteria and eukaryotic cell lines including tumor cells. It is shown that the length of alkyl chains in the backbone is vital for the antibiofilm activities of these lipid-mimicking components. In addition to their biological activity, their surface activity and their membrane interactions are shown by film balance and quartz crystal microbalance (QCM) measurements. The structure-activity relationship indicates that the distinctive chemical structure contributes considerably to the biological activities of this novel class of lipids. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Evidence to Support a Contribution of Polyreactive Antibodies to HLA Serum Reactivity.

PubMed

Gao, Baoshan; Rong, Chunshu; Porcheray, Fabrice; Moore, Carolina; Girouard, Timothy C; Saidman, Susan L; Wong, Waichi; Fu, Yaowen; Zorn, Emmanuel

2016-01-01

Assessing the serum reactivity to HLA is essential for the evaluation of transplant candidates and the follow-up of allograft recipients. In this study, we look for evidence at the clonal level that polyreactive antibodies cross-reactive to apoptotic cells and multiple autoantigens can also react to HLA and contribute to the overall serum reactivity. We immortalized B cell clones from the blood of 2 kidney transplant recipients and characterized their reactivity to self-antigens, apoptotic cells as well as native, denatured, and cryptic HLA determinants using enzyme-linked immunosorbent assay (ELISA), immunofluorescence, flow cytometry and Luminex assays. We also assessed the reactivity of 300 pretransplant serum specimens to HLA and apoptotic cells. We report here 4 distinct B cell clones cross-reactive to self and HLA class I. All 4 clones reacted to numerous HLA class I alleles but did not appear to target canonical "shared" epitopes. In parallel experiments, we observed a strong correlation between IgG reactivity to HLA and apoptotic cells in pretransplant serum samples collected from 300 kidney transplant recipients. Further analysis revealed that samples with higher reactivity to apoptotic cells displayed significantly higher class I percent panel-reactive antibodies compared to samples with low reactivity to apoptotic cells. We provide here (1) proof of principle at the clonal level that human polyreactive antibodies can cross-react to HLA, multiple self-antigens and apoptotic cells and (2) supportive evidence that polyreactive antibodies contribute to overall HLA reactivity in the serum of patients awaiting kidney transplant.

Proliferative and transcriptional identity of distinct classes of neural precursors in the mammalian olfactory epithelium.

PubMed

Tucker, Eric S; Lehtinen, Maria K; Maynard, Tom; Zirlinger, Mariela; Dulac, Catherine; Rawson, Nancy; Pevny, Larysa; Lamantia, Anthony-Samuel

2010-08-01

Neural precursors in the developing olfactory epithelium (OE) give rise to three major neuronal classes - olfactory receptor (ORNs), vomeronasal (VRNs) and gonadotropin releasing hormone (GnRH) neurons. Nevertheless, the molecular and proliferative identities of these precursors are largely unknown. We characterized two precursor classes in the olfactory epithelium (OE) shortly after it becomes a distinct tissue at midgestation in the mouse: slowly dividing self-renewing precursors that express Meis1/2 at high levels, and rapidly dividing neurogenic precursors that express high levels of Sox2 and Ascl1. Precursors expressing high levels of Meis genes primarily reside in the lateral OE, whereas precursors expressing high levels of Sox2 and Ascl1 primarily reside in the medial OE. Fgf8 maintains these expression signatures and proliferative identities. Using electroporation in the wild-type embryonic OE in vitro as well as Fgf8, Sox2 and Ascl1 mutant mice in vivo, we found that Sox2 dose and Meis1 - independent of Pbx co-factors - regulate Ascl1 expression and the transition from lateral to medial precursor state. Thus, we have identified proliferative characteristics and a dose-dependent transcriptional network that define distinct OE precursors: medial precursors that are most probably transit amplifying neurogenic progenitors for ORNs, VRNs and GnRH neurons, and lateral precursors that include multi-potent self-renewing OE neural stem cells.

Proliferative and transcriptional identity of distinct classes of neural precursors in the mammalian olfactory epithelium

PubMed Central

Tucker, Eric S.; Lehtinen, Maria K.; Maynard, Tom; Zirlinger, Mariela; Dulac, Catherine; Rawson, Nancy; Pevny, Larysa; LaMantia, Anthony-Samuel

2010-01-01

Neural precursors in the developing olfactory epithelium (OE) give rise to three major neuronal classes – olfactory receptor (ORNs), vomeronasal (VRNs) and gonadotropin releasing hormone (GnRH) neurons. Nevertheless, the molecular and proliferative identities of these precursors are largely unknown. We characterized two precursor classes in the olfactory epithelium (OE) shortly after it becomes a distinct tissue at midgestation in the mouse: slowly dividing self-renewing precursors that express Meis1/2 at high levels, and rapidly dividing neurogenic precursors that express high levels of Sox2 and Ascl1. Precursors expressing high levels of Meis genes primarily reside in the lateral OE, whereas precursors expressing high levels of Sox2 and Ascl1 primarily reside in the medial OE. Fgf8 maintains these expression signatures and proliferative identities. Using electroporation in the wild-type embryonic OE in vitro as well as Fgf8, Sox2 and Ascl1 mutant mice in vivo, we found that Sox2 dose and Meis1 – independent of Pbx co-factors – regulate Ascl1 expression and the transition from lateral to medial precursor state. Thus, we have identified proliferative characteristics and a dose-dependent transcriptional network that define distinct OE precursors: medial precursors that are most probably transit amplifying neurogenic progenitors for ORNs, VRNs and GnRH neurons, and lateral precursors that include multi-potent self-renewing OE neural stem cells. PMID:20573694

A Distinctive Cytoplasmic Tail Contributes to Low Surface Expression and Intracellular Retention of the Patr-AL MHC class I molecule1

PubMed Central

Goyos, Ana; Guethlein, Lisbeth A.; Horowitz, Amir; Hilton, Hugo G.; Gleimer, Michael; Brodsky, Frances M.; Parham, Peter

2015-01-01

Chimpanzees have orthologs of the six, fixed, functional human MHC class I genes. But in addition, the chimpanzee has a seventh functional gene, Patr-AL, which is not polymorphic but contributes substantially to population diversity by its presence on only 50% of MHC haplotypes. The ancestral AL gene emerged long before the separation of human and chimpanzee ancestors and then subsequently and specifically lost function during human evolution, but was maintained in chimpanzees. Patr-AL is an alloantigen that participates in negative and positive selection of the T-cell repertoire. The three-dimensional structure and the peptide-binding repertoire of Patr-AL and HLA-A*02 are surprisingly similar. In contrast, the expression of these two molecules is very different as shown using specific monoclonal and polyclonal antibodies made against Patr-AL. Peripheral blood cells and B cell lines express low levels of Patr-AL at the cell surface. Higher levels are seen for 221-cell transfectants expressing Patr-AL, but in these cells a large majority of Patr-AL molecules are retained in the early compartments of the secretory pathway: mainly the endoplasmic reticulum but also cis-Golgi. Replacing the cytoplasmic tail of Patr-AL with that of HLA-A*02 increased the cell-surface expression of Patr-AL substantially. Four substitutions distinguish the Patr-AL and HLA-A*02 cytoplasmic tails. Systematic mutagenesis showed that each substitution contributes changes in cell-surface expression. The combination of residues present in Patr-AL appears unique, but each individual residue is present in other primate MHC class I molecules, notably MHC-E, the most ancient of the functional human MHC class I molecules. PMID:26371256

HIV-1 Envelope Resistance to Proteasomal Cleavage: Implications for Vaccine Induced Immune Responses

PubMed Central

Steers, Nicholas J.; Ratto-Kim, Silvia; de Souza, Mark S.; Currier, Jeffrey R.; Kim, Jerome H.; Michael, Nelson L.; Alving, Carl R.; Rao, Mangala

2012-01-01

Background Antigen processing involves many proteolytic enzymes such as proteasomes and cathepsins. The processed antigen is then presented on the cell surface bound to either MHC class I or class II molecules and induces/interacts with antigen-specific CD8+ and CD4+ T-cells, respectively. Preliminary immunological data from the RV144 phase III trial indicated that the immune responses were biased towards the Env antigen with a dominant CD4+ T-cell response. Methods In this study, we examined the susceptibility of HIV-1 Env-A244 gp120 protein, one of the protein boost subunits of the RV144 Phase III vaccine trial, to proteasomes and cathepsins and identified the generated peptide epitope repertoire by mass spectrometry. The peptide fragments were tested for cytokine production in CD4+ T-cell lines derived from RV144 volunteers. Results Env-A244 was resistant to proteasomes, thus diminishing the possibility of the generation of class I epitopes by the classical MHC class I pathway. However, Env-A244 was efficiently cleaved by cathepsins generating peptide arrays identified by mass spectrometry that contained both MHC class I and class II epitopes as reported in the Los Alamos database. Each of the cathepsins generated distinct degradation patterns containing regions of light and dense epitope clusters. The sequence DKKQKVHALF that is part of the V2 loop of gp120 produced by cathepsins induced a polyfunctional cytokine response including the generation of IFN-γ from CD4+ T-cell lines-derived from RV144 vaccinees. This sequence is significant since antibodies to the V1/V2-loop region correlated inversely with HIV-1 infection in the RV144 trial. Conclusions Based on our results, the susceptibility of Env-A244 to cathepsins and not to proteasomes suggests a possible mechanism for the generation of Env-specific CD4+T cell and antibody responses in the RV144 vaccinees. PMID:22880042

Leptospira interrogans causes quantitative and morphological disturbances in adherens junctions and other biological groups of proteins in human endothelial cells

PubMed Central

Sato, Hiromi

2017-01-01

Pathogenic Leptospira transmits from animals to humans, causing the zoonotic life-threatening infection called leptospirosis. This infection is reported worldwide with higher risk in tropical regions. Symptoms of leptospirosis range from mild illness to severe illness such as liver damage, kidney failure, respiratory distress, meningitis, and fatal hemorrhagic disease. Invasive species of Leptospira rapidly disseminate to multiple tissues where this bacterium damages host endothelial cells, increasing vascular permeability. Despite the burden in humans and animals, the pathogenic mechanisms of Leptospira infection remain to be elucidated. The pathogenic leptospires adhere to endothelial cells and permeabilize endothelial barriers in vivo and in vitro. In this study, human endothelial cells were infected with the pathogenic L. interrogans serovar Copenhageni or the saprophyte L. biflexa serovar Patoc to investigate morphological changes and other distinctive phenotypes of host cell proteins by fluorescence microscopy. Among those analyzed, 17 proteins from five biological classes demonstrated distinctive phenotypes in morphology and/or signal intensity upon infection with Leptospira. The affected biological groups include: 1) extracellular matrix, 2) intercellular adhesion molecules and cell surface receptors, 3) intracellular proteins, 4) cell-cell junction proteins, and 5) a cytoskeletal protein. Infection with the pathogenic strain most profoundly disturbed the biological structures of adherens junctions (VE-cadherin and catenins) and actin filaments. Our data illuminate morphological disruptions and reduced signals of cell-cell junction proteins and filamentous actin in L. interrogans-infected endothelial cells. In addition, Leptospira infection, regardless of pathogenic status, influenced other host proteins belonging to multiple biological classes. Our data suggest that this zoonotic agent may damage endothelial cells via multiple cascades or pathways including endothelial barrier damage and inflammation, potentially leading to vascular hyperpermeability and severe illness in vivo. This work provides new insights into the pathophysiological mechanisms of Leptospira infection. PMID:28750011

Leptospira interrogans causes quantitative and morphological disturbances in adherens junctions and other biological groups of proteins in human endothelial cells.

PubMed

Sato, Hiromi; Coburn, Jenifer

2017-07-01

Pathogenic Leptospira transmits from animals to humans, causing the zoonotic life-threatening infection called leptospirosis. This infection is reported worldwide with higher risk in tropical regions. Symptoms of leptospirosis range from mild illness to severe illness such as liver damage, kidney failure, respiratory distress, meningitis, and fatal hemorrhagic disease. Invasive species of Leptospira rapidly disseminate to multiple tissues where this bacterium damages host endothelial cells, increasing vascular permeability. Despite the burden in humans and animals, the pathogenic mechanisms of Leptospira infection remain to be elucidated. The pathogenic leptospires adhere to endothelial cells and permeabilize endothelial barriers in vivo and in vitro. In this study, human endothelial cells were infected with the pathogenic L. interrogans serovar Copenhageni or the saprophyte L. biflexa serovar Patoc to investigate morphological changes and other distinctive phenotypes of host cell proteins by fluorescence microscopy. Among those analyzed, 17 proteins from five biological classes demonstrated distinctive phenotypes in morphology and/or signal intensity upon infection with Leptospira. The affected biological groups include: 1) extracellular matrix, 2) intercellular adhesion molecules and cell surface receptors, 3) intracellular proteins, 4) cell-cell junction proteins, and 5) a cytoskeletal protein. Infection with the pathogenic strain most profoundly disturbed the biological structures of adherens junctions (VE-cadherin and catenins) and actin filaments. Our data illuminate morphological disruptions and reduced signals of cell-cell junction proteins and filamentous actin in L. interrogans-infected endothelial cells. In addition, Leptospira infection, regardless of pathogenic status, influenced other host proteins belonging to multiple biological classes. Our data suggest that this zoonotic agent may damage endothelial cells via multiple cascades or pathways including endothelial barrier damage and inflammation, potentially leading to vascular hyperpermeability and severe illness in vivo. This work provides new insights into the pathophysiological mechanisms of Leptospira infection.

Immunoproteomic analysis of house dust mite antigens reveals distinct classes of dominant T cell antigens according to function and serological reactivity

PubMed Central

Westernberg, Luise; Pham, John; Lane, Jerome; Paul, Sinu; Greenbaum, Jason; Stranzl, Thomas; Lund, Gitte; Hoof, Ilka; Holm, Jens; Würtzen, Peter A; Meno, Kåre H.; Frazier, April; Schulten, Veronique; Andersen, Peter S.; Peters, Bjoern; Sette, Alessandro

2016-01-01

BACKGROUND House dust mite (HDM) allergens are a common cause of allergy and allergic asthma. A comprehensive analysis of proteins targeted by T cells, which are implicated in the development and regulation of allergic disease independent of their antibody reactivity, is still lacking. OBJECTIVE To comprehensively analyze the HDM-derived protein targets of T cell responses in HDM-allergic individuals, and investigate their correlation with IgE/IgG responses and protein function. METHODS Proteomic analysis (liquid chromatography tandem mass spectrometry) of HDM extracts identified 90 distinct protein clusters, corresponding to 29 known allergens and 61 novel proteins. Peripheral blood mononuclear cells (PBMC) from 20 HDM-allergic individuals were stimulated with HDM extracts and assayed with a set of ~2500 peptides derived from these 90 protein clusters and predicted to bind the most common HLA class II types. 2D immunoblots were made in parallel to elucidate IgE and IgG reactivity and putative function analyses were performed in silico according to gene ontology (GO) annotations. RESULTS Analysis of T cell reactivity revealed a large number of T cell epitopes. Overall response magnitude and frequency was comparable for known and novel proteins, with 15 antigens (nine of which were novel) dominating the total T cell response. Most of the known allergens that were dominant at the T cell level were also IgE-reactive, as expected, while few novel dominant T cell antigens were IgE reactive. Among known allergens, hydrolase activity and detectable IgE/IgG reactivity are strongly correlated, while no protein function correlates with immunogenicity of novel proteins. A total of 106 epitopes accounted for half of the total T-cell response, underlining the heterogeneity of T cell responses to HDM allergens. CONCLUSIONS AND CLINICAL RELEVANCE Herein, we define the T cell targets for both known allergens and novel proteins, which may inform future diagnostics and immunotherapeutics for allergy to HDM. PMID:27684489

Inhibition of Voltage-Gated Calcium Channels as Common Mode of Action for (Mixtures of) Distinct Classes of Insecticides

PubMed Central

Meijer, Marieke; Dingemans, Milou M.L.; van den Berg, Martin; Westerink, Remco H.S.

2014-01-01

Humans are exposed to distinct structural classes of insecticides with different neurotoxic modes of action. Because calcium homeostasis is essential for proper neuronal function and development, we investigated the effects of insecticides from different classes (pyrethroid: (α-)cypermethrin; organophosphate: chlorpyrifos; organochlorine: endosulfan; neonicotinoid: imidacloprid) and mixtures thereof on the intracellular calcium concentration ([Ca2+]i). Effects of acute (20 min) exposure to (mixtures of) insecticides on basal and depolarization-evoked [Ca2+]i were studied in vitro with Fura-2-loaded PC12 cells and high resolution single-cell fluorescence microscopy. The data demonstrate that cypermethrin, α-cypermethrin, endosulfan, and chlorpyrifos concentration-dependently decreased depolarization-evoked [Ca2+]i, with 50% (IC50) at 78nM, 239nM, 250nM, and 899nM, respectively. Additionally, acute exposure to chlorpyrifos or endosulfan (10μM) induced a modest increase in basal [Ca2+]i, amounting to 68 ± 8nM and 53 ± 8nM, respectively. Imidacloprid did not disturb basal or depolarization-evoked [Ca2+]i at 10μM. Following exposure to binary mixtures, effects on depolarization-evoked [Ca2+]i were within the expected effect additivity range, whereas the effect of the tertiary mixture was less than this expected additivity effect range. These results demonstrate that different types of insecticides inhibit depolarization-evoked [Ca2+]i in PC12 cells by inhibiting voltage-gated calcium channels (VGCCs) in vitro at concentrations comparable with human occupational exposure levels. Moreover, the effective concentrations in this study are below those for earlier described modes of action. Because inhibition of VGCCs appears to be a common and potentially additive mode of action of several classes of insecticides, this target should be considered in neurotoxicity risk assessment studies. PMID:24913802

Phosphatidylinositol 3-phosphates-at the interface between cell signalling and membrane traffic.

PubMed

Marat, Andrea L; Haucke, Volker

2016-03-15

Phosphoinositides (PIs) form a minor class of phospholipids with crucial functions in cell physiology, ranging from cell signalling and motility to a role as signposts of compartmental membrane identity. Phosphatidylinositol 3-phosphates are present at the plasma membrane and within the endolysosomal system, where they serve as key regulators of both cell signalling and of intracellular membrane traffic. Here, we provide an overview of the metabolic pathways that regulate cellular synthesis of PI 3-phosphates at distinct intracellular sites and discuss the mechanisms by which these lipids regulate cell signalling and membrane traffic. Finally, we provide a framework for how PI 3-phosphate metabolism is integrated into the cellular network. © 2016 The Authors.

27 CFR 4.62 - Mandatory statements.

Code of Federal Regulations, 2010 CFR

2010-04-01

..., type, and distinctive designation. The advertisement shall contain a conspicuous statement of the class, type, or distinctive designation to which the product belongs, corresponding with the statement of class, type, or distinctive designation which is required to appear on the label of the product. (c...

O-GlcNAc transferase regulates transcriptional activity of human Oct4.

PubMed

Constable, Sandii; Lim, Jae-Min; Vaidyanathan, Krithika; Wells, Lance

2017-10-01

O-linked β-N-acetylglucosamine (O-GlcNAc) is a single sugar modification found on many different classes of nuclear and cytoplasmic proteins. Addition of this modification, by the enzyme O-linked N-acetylglucosamine transferase (OGT), is dynamic and inducible. One major class of proteins modified by O-GlcNAc is transcription factors. O-GlcNAc regulates transcription factor properties through a variety of different mechanisms including localization, stability and transcriptional activation. Maintenance of embryonic stem (ES) cell pluripotency requires tight regulation of several key transcription factors, many of which are modified by O-GlcNAc. Octamer-binding protein 4 (Oct4) is one of the key transcription factors required for pluripotency of ES cells and more recently, the generation of induced pluripotent stem (iPS) cells. The action of Oct4 is modulated by the addition of several post-translational modifications, including O-GlcNAc. Previous studies in mice found a single site of O-GlcNAc addition responsible for transcriptional regulation. This study was designed to determine if this mechanism is conserved in humans. We mapped 10 novel sites of O-GlcNAc attachment on human Oct4, and confirmed a role for OGT in transcriptional activation of Oct4 at a site distinct from that found in mouse that allows distinction between different Oct4 target promoters. Additionally, we uncovered a potential new role for OGT that does not include its catalytic function. These results confirm that human Oct4 activity is being regulated by OGT by a mechanism that is distinct from mouse Oct4. © The Author 2017. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Characterization of distinct classes of differential gene expression in osteoblast cultures from non-syndromic craniosynostosis bone.

PubMed

Rojas-Peña, Monica L; Olivares-Navarrete, Rene; Hyzy, Sharon; Arafat, Dalia; Schwartz, Zvi; Boyan, Barbara D; Williams, Joseph; Gibson, Greg

2014-01-01

Craniosynostosis, the premature fusion of one or more skull sutures, occurs in approximately 1 in 2500 infants, with the majority of cases non-syndromic and of unknown etiology. Two common reasons proposed for premature suture fusion are abnormal compression forces on the skull and rare genetic abnormalities. Our goal was to evaluate whether different sub-classes of disease can be identified based on total gene expression profiles. RNA-Seq data were obtained from 31 human osteoblast cultures derived from bone biopsy samples collected between 2009 and 2011, representing 23 craniosynostosis fusions and 8 normal cranial bones or long bones. No differentiation between regions of the skull was detected, but variance component analysis of gene expression patterns nevertheless supports transcriptome-based classification of craniosynostosis. Cluster analysis showed 4 distinct groups of samples; 1 predominantly normal and 3 craniosynostosis subtypes. Similar constellations of sub-types were also observed upon re-analysis of a similar dataset of 199 calvarial osteoblast cultures. Annotation of gene function of differentially expressed transcripts strongly implicates physiological differences with respect to cell cycle and cell death, stromal cell differentiation, extracellular matrix (ECM) components, and ribosomal activity. Based on these results, we propose non-syndromic craniosynostosis cases can be classified by differences in their gene expression patterns and that these may provide targets for future clinical intervention.

Characterization of Distinct Classes of Differential Gene Expression in Osteoblast Cultures from Non-Syndromic Craniosynostosis Bone

PubMed Central

Rojas-Peña, Monica L.; Olivares-Navarrete, Rene; Hyzy, Sharon; Arafat, Dalia; Schwartz, Zvi; Boyan, Barbara D.; Williams, Joseph; Gibson, Greg

2014-01-01

Craniosynostosis, the premature fusion of one or more skull sutures, occurs in approximately 1 in 2500 infants, with the majority of cases non-syndromic and of unknown etiology. Two common reasons proposed for premature suture fusion are abnormal compression forces on the skull and rare genetic abnormalities. Our goal was to evaluate whether different sub-classes of disease can be identified based on total gene expression profiles. RNA-Seq data were obtained from 31 human osteoblast cultures derived from bone biopsy samples collected between 2009 and 2011, representing 23 craniosynostosis fusions and 8 normal cranial bones or long bones. No differentiation between regions of the skull was detected, but variance component analysis of gene expression patterns nevertheless supports transcriptome-based classification of craniosynostosis. Cluster analysis showed 4 distinct groups of samples; 1 predominantly normal and 3 craniosynostosis subtypes. Similar constellations of sub-types were also observed upon re-analysis of a similar dataset of 199 calvarial osteoblast cultures. Annotation of gene function of differentially expressed transcripts strongly implicates physiological differences with respect to cell cycle and cell death, stromal cell differentiation, extracellular matrix (ECM) components, and ribosomal activity. Based on these results, we propose non-syndromic craniosynostosis cases can be classified by differences in their gene expression patterns and that these may provide targets for future clinical intervention. PMID:25184005

Distinct T helper cell dependence of memory B-cell proliferation versus plasma cell differentiation.

PubMed

Zabel, Franziska; Fettelschoss, Antonia; Vogel, Monique; Johansen, Pål; Kündig, Thomas M; Bachmann, Martin F

2017-03-01

Several memory B-cell subclasses with distinct functions have been described, of which the most effective is the class-switched (CS) memory B-cell population. We have previously shown, using virus-like particles (VLPs), that the proliferative potential of these CS memory B cells is limited and they fail to re-enter germinal centres (GCs). However, VLP-specific memory B cells quickly differentiated into secondary plasma cells (PCs) with the virtue of elevated antibody production compared with primary PCs. Whereas the induction of VLP + memory B cells was strongly dependent on T helper cells, we were wondering whether re-stimulation of VLP + memory B cells and their differentiation into secondary PCs would also require T helper cells. Global absence of T helper cells led to strongly impaired memory B cell proliferation and PC differentiation. In contrast, lack of interleukin-21 receptor-dependent follicular T helper cells or CD40 ligand signalling strongly affected proliferation of memory B cells, but differentiation into mature secondary PCs exhibiting increased antibody production was essentially normal. This contrasts with primary B-cell responses, where a strong dependence on CD40 ligand but limited importance of interleukin-21 receptor was seen. Hence, T helper cell dependence differs between primary and secondary B-cell responses as well as between memory B-cell proliferation and PC differentiation. © 2016 John Wiley & Sons Ltd.

In Vitro Evidence of the Presence of Mesenchymal Stromal Cells in Cervical Cancer and Their Role in Protecting Cancer Cells from Cytotoxic T Cell Activity

PubMed Central

Montesinos, Juan J.; Mora-García, María de L.; Mayani, Héctor; Flores-Figueroa, Eugenia; García-Rocha, Rosario; Fajardo-Orduña, Guadalupe R.; Castro-Manrreza, Marta E.; Weiss-Steider, Benny

2013-01-01

Mesenchymal stromal cells (MSCs) have been isolated from different tumors and it has been suggested that they support tumor growth through immunosuppression processes that favor tumor cell evasion from the immune system. To date, however, the presence of MSCs in cervical cancer (CeCa) and their possible role in tumor growth remains unknown. Herein we report on the presence of MSCs in cervical tissue, both in normal conditions (NCx-MSCs) and in CeCa (CeCa-MSCs), and described several biological properties of such cells. Our study showed similar patterns of cell surface antigen expression, but distinct differentiation potentials, when we compared both cervical MSC populations to MSCs from normal bone marrow (BM-MSCs, the gold standard). Interestingly, CeCa-MSCs were negative for the presence of human papiloma virus, indicating that these cells are not infected by such a viral agent. Also, interestingly, and in contrast to NCx-MSCs, CeCa-MSCs induced significant downregulation of surface HLA class I molecules (HLA-A*0201) on CaSki cells and other CeCa cell lines. We further observed that CeCa-MSCs inhibited antigen-specific T cell recognition of CaSki cells by cytotoxic T lymphocytes (CTLs). HLA class I downregulation on CeCa cells correlated with the production of IL-10 in cell cocultures. Importantly, this cytokine strongly suppressed recognition of CeCa cells by CTLs. In summary, this study demonstrates the presence of MSCs in CeCa and suggests that tumor-derived MSCs may provide immune protection to tumor cells by inducing downregulation of HLA class I molecules. This mechanism may have important implications in tumor growth. PMID:23656504

SHIP1 intrinsically regulates NK cell signaling and education, resulting in tolerance of an MHC class I-mismatched bone marrow graft in mice.

PubMed

Gumbleton, Matthew; Vivier, Eric; Kerr, William G

2015-03-15

NK cells are an important component of host immune defense against malignancy and infection. NK cells are educated by MHC class I ligands to ensure self-tolerance while also promoting lytic competency against altered self and damaged self targets. However, the intracellular molecular events that culminate in tolerance and functional competency of educated NK cells remain undefined. Mice with germline deficiency in SHIP1 were shown to have a defective NK cell compartment. However, SHIP1 is expressed in all hematopoietic lineages, and consequently several hematolymphoid phenotypes have already been identified in certain cell types that are the result of SHIP1 deficiency in cells in separate and distinct lineages, that is, cell-extrinsic phenotypes. Thus, it was previously impossible to determine the NK cell-intrinsic role of SHIP1. In the present study, through the creation of an NK cell-specific deletion mouse model of SHIP1, we show that SHIP1 plays a profound NK lineage-intrinsic role in NK cell homeostasis, development, education, and cytokine production. Moreover, we show SHIP1 expression by NK cells is required for in vivo-mismatched bone marrow allograft rejection as well as for NK memory responses to hapten. Copyright © 2015 by The American Association of Immunologists, Inc.

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.

NKT Cell Subsets Can Exert Opposing Effects in Autoimmunity, Tumor Surveillance and Inflammation

PubMed Central

Viale, Rachael; Ware, Randle; Maricic, Igor; Chaturvedi, Varun; Kumar, Vipin

2014-01-01

The innate-like natural killer T (NKT) cells are essential regulators of immunity. These cells comprise at least two distinct subsets and recognize different lipid antigens presented by the MHC class I like molecules CD1d. The CD1d-dependent recognition pathway of NKT cells is highly conserved from mouse to humans. While most type I NKT cells can recognize αGalCer and express a semi-invariant T cell receptor (TCR), a major population of type II NKT cells reactive to sulfatide utilizes an oligoclonal TCR. Furthermore TCR recognition features of NKT subsets are also distinctive with almost parallel as opposed to perpendicular footprints on the CD1d molecules for the type I and type II NKT cells respectively. Here we present a view based upon the recent studies in different clinical and experimental settings that while type I NKT cells are more often pathogenic, they may also be regulatory. On the other hand, sulfatide-reactive type II NKT cells mostly play an inhibitory role in the control of autoimmune and inflammatory diseases. Since the activity and cytokine secretion profiles of NKT cell subsets can be modulated differently by lipid ligands or their analogs, novel immunotherapeutic strategies are being developed for their differential activation for potential intervention in inflammatory diseases. PMID:25288922

Two distinct HLA-A0201-presented epitopes of the Wilms tumor antigen 1 can function as targets for leukemia-reactive CTL.

PubMed

Bellantuono, Ilaria; Gao, Liquan; Parry, Suzanne; Marley, Steve; Dazzi, Francesco; Apperley, Jane; Goldman, John M; Stauss, Hans J

2002-11-15

Using the allo-restricted T-cell approach to circumvent tolerance, we have previously identified a cytotoxic T-lymphocyte (CTL) epitope in the transcription factor Wilms tumor antigen 1 (WT1) presented by HLA-A0201 (A2) class I molecules. Here we describe an additional A2-presented epitope and show that CTLs against both epitopes kill WT1-expressing leukemia cell lines. Colony-forming assays demonstrated that both types of CTL killed CD34(+) progenitor cells from A2(+) leukemia patients, but not from A2(+) healthy individuals. The long-term culture-initiating cell (LTC-IC) assay was used to analyze the killing activity of WT1-specific CTLs against the more immature fraction of CD34(+) cells. The CTLs killed LTC-ICs of patients with chronic myelogenous leukemia (CML), whereas the function of normal CD34(+) progenitor/stem cells was not inhibited. Together, the data show that CTLs specific for 2 distinct peptide epitopes of WT1 can discriminate between normal and leukemia LTC-ICs, suggesting that such CTLs have the potential to selectively kill CML progenitor/stem cells.

Meiotic recombination generates rich diversity in NK cell receptor genes, alleles, and haplotypes

PubMed Central

Norman, Paul J.; Abi-Rached, Laurent; Gendzekhadze, Ketevan; Hammond, John A.; Moesta, Achim K.; Sharma, Deepti; Graef, Thorsten; McQueen, Karina L.; Guethlein, Lisbeth A.; Carrington, Christine V.F.; Chandanayingyong, Dasdayanee; Chang, Yih-Hsin; Crespí, Catalina; Saruhan-Direskeneli, Güher; Hameed, Kamran; Kamkamidze, Giorgi; Koram, Kwadwo A.; Layrisse, Zulay; Matamoros, Nuria; Milà, Joan; Park, Myoung Hee; Pitchappan, Ramasamy M.; Ramdath, D. Dan; Shiau, Ming-Yuh; Stephens, Henry A.F.; Struik, Siske; Tyan, Dolly; Verity, David H.; Vaughan, Robert W.; Davis, Ronald W.; Fraser, Patricia A.; Riley, Eleanor M.; Ronaghi, Mostafa; Parham, Peter

2009-01-01

Natural killer (NK) cells contribute to the essential functions of innate immunity and reproduction. Various genes encode NK cell receptors that recognize the major histocompatibility complex (MHC) Class I molecules expressed by other cells. For primate NK cells, the killer-cell immunoglobulin-like receptors (KIR) are a variable and rapidly evolving family of MHC Class I receptors. Studied here is KIR3DL1/S1, which encodes receptors for highly polymorphic human HLA-A and -B and comprises three ancient allelic lineages that have been preserved by balancing selection throughout human evolution. While the 3DS1 lineage of activating receptors has been conserved, the two 3DL1 lineages of inhibitory receptors were diversified through inter-lineage recombination with each other and with 3DS1. Prominent targets for recombination were D0-domain polymorphisms, which modulate enhancer function, and dimorphism at position 283 in the D2 domain, which influences inhibitory function. In African populations, unequal crossing over between the 3DL1 and 3DL2 genes produced a deleted KIR haplotype in which the telomeric “half” was reduced to a single fusion gene with functional properties distinct from its 3DL1 and 3DL2 parents. Conversely, in Eurasian populations, duplication of the KIR3DL1/S1 locus by unequal crossing over has enabled individuals to carry and express alleles of all three KIR3DL1/S1 lineages. These results demonstrate how meiotic recombination combines with an ancient, preserved diversity to create new KIR phenotypes upon which natural selection acts. A consequence of such recombination is to blur the distinction between alleles and loci in the rapidly evolving human KIR gene family. PMID:19411600

Generation of large numbers of dendritic cells from mouse bone marrow cultures supplemented with granulocyte/macrophage colony-stimulating factor

PubMed Central

1992-01-01

Antigen-presenting, major histocompatibility complex (MHC) class II- rich dendritic cells are known to arise from bone marrow. However, marrow lacks mature dendritic cells, and substantial numbers of proliferating less-mature cells have yet to be identified. The methodology for inducing dendritic cell growth that was recently described for mouse blood now has been modified to MHC class II- negative precursors in marrow. A key step is to remove the majority of nonadherent, newly formed granulocytes by gentle washes during the first 2-4 d of culture. This leaves behind proliferating clusters that are loosely attached to a more firmly adherent "stroma." At days 4-6 the clusters can be dislodged, isolated by 1-g sedimentation, and upon reculture, large numbers of dendritic cells are released. The latter are readily identified on the basis of their distinct cell shape, ultrastructure, and repertoire of antigens, as detected with a panel of monoclonal antibodies. The dendritic cells express high levels of MHC class II products and act as powerful accessory cells for initiating the mixed leukocyte reaction. Neither the clusters nor mature dendritic cells are generated if macrophage colony-stimulating factor rather than granulocyte/macrophage colony-stimulating factor (GM-CSF) is applied. Therefore, GM-CSF generates all three lineages of myeloid cells (granulocytes, macrophages, and dendritic cells). Since > 5 x 10(6) dendritic cells develop in 1 wk from precursors within the large hind limb bones of a single animal, marrow progenitors can act as a major source of dendritic cells. This feature should prove useful for future molecular and clinical studies of this otherwise trace cell type. PMID:1460426

Long-Peptide Cross-Presentation by Human Dendritic Cells Occurs in Vacuoles by Peptide Exchange on Nascent MHC Class I Molecules.

PubMed

Ma, Wenbin; Zhang, Yi; Vigneron, Nathalie; Stroobant, Vincent; Thielemans, Kris; van der Bruggen, Pierre; Van den Eynde, Benoît J

2016-02-15

Cross-presentation enables dendritic cells to present on their MHC class I molecules antigenic peptides derived from exogenous material, through a mechanism that remains partly unclear. It is particularly efficient with long peptides, which are used in cancer vaccines. We studied the mechanism of long-peptide cross-presentation using human dendritic cells and specific CTL clones against melanoma Ags gp100 and Melan-A/MART1. We found that cross-presentation of those long peptides does not depend on the proteasome or the transporter associated with Ag processing, and therefore follows a vacuolar pathway. We also observed that it makes use of newly synthesized MHC class I molecules, through peptide exchange in vesicles distinct from the endoplasmic reticulum and classical secretory pathway, in an SEC22b- and CD74-independent manner. Our results indicate a nonclassical secretion pathway followed by nascent HLA-I molecules that are used for cross-presentation of those long melanoma peptides in the vacuolar pathway. Our results may have implications for the development of vaccines based on long peptides. Copyright © 2016 by The American Association of Immunologists, Inc.

Structural insights into ligand recognition and selectivity for class A, B, and C GPCRs

PubMed Central

Lee, Sang-Min; Booe, Jason M.; Pioszak, Augen A.

2015-01-01

The G protein-coupled receptor (GPCR) superfamily constitutes the largest collection of cell surface signaling proteins with approximately 800 members in the human genome. GPCRs regulate virtually all aspects of physiology and they are an important class of drug targets with ~30% of drugs on the market targeting a GPCR. Breakthroughs in GPCR structural biology in recent years have significantly expanded our understanding of GPCR structure and function and ushered in a new era of structure-based drug design for GPCRs. Crystal structures for nearly thirty distinct GPCRs are now available including receptors from each of the major classes, A, B, C, and F. These structures provide a foundation for understanding the molecular basis of GPCR pharmacology. Here, we review structural mechanisms of ligand recognition and selectivity of GPCRs with a focus on selected examples from classes A, B, and C, and we highlight major unresolved questions for future structural studies. PMID:25981303

Evidence to support a contribution of polyreactive antibodies to HLA serum reactivity

PubMed Central

Gao, Baoshan; Rong, Chunshu; Porcheray, Fabrice; Moore, Carolina; Girouard, Timothy C.; Saidman, Susan L.; Wong, Waichi; Fu, Yaowen; Zorn, Emmanuel

2015-01-01

Background Assessing the serum reactivity to HLA is essential for the evaluation of transplant candidates and the follow-up of allograft recipients. In this study, we look for evidence at the clonal level that polyreactive antibodies cross-reactive to apoptotic cells and multiple autoantigens can also react to HLA and contribute to the overall serum reactivity. Methods We immortalized B cell clones from the blood of two kidney transplant recipients and characterized their reactivity to self-antigens, apoptotic cells as well as native, denatured and cryptic HLA determinants using ELISA, immunofluorescence, flow cytometry and Luminex assays. We also assessed the reactivity of 300 pre-transplant serum specimens to HLA and apoptotic cells. Results We report here 4 distinct B cell clones cross-reactive to self and HLA class I. All 4 clones reacted to numerous HLA class I alleles but did not appear to target canonical “shared” epitopes. In parallel experiments, we observed a strong correlation between IgG reactivity to HLA and apoptotic cells in pre-transplant serum samples collected from 300 kidney transplant recipients. Further analysis revealed that samples with higher reactivity to apoptotic cells displayed significantly higher class I percent PRA compared to samples with low reactivity to apoptotic cells. Conclusions We provide here 1) proof of principle at the clonal level that human polyreactive antibodies can cross-react to HLA, multiple self-antigens and apoptotic cells and 2) supportive evidence that polyreactive antibodies contribute to overall HLA reactivity in the serum of patients awaiting kidney transplant. PMID:26285015

DREAMs make plant cells to cycle or to become quiescent.

PubMed

Magyar, Zoltán; Bögre, László; Ito, Masaki

2016-12-01

Cell cycle phase specific oscillation of gene transcription has long been recognized as an underlying principle for ordered processes during cell proliferation. The G1/S-specific and G2/M-specific cohorts of genes in plants are regulated by the E2F and the MYB3R transcription factors. Mutant analysis suggests that activator E2F functions might not be fully required for cell cycle entry. In contrast, the two activator-type MYB3Rs are part of positive feedback loops to drive the burst of mitotic gene expression, which is necessary at least to accomplish cytokinesis. Repressor MYB3Rs act outside the mitotic time window during cell cycle progression, and are important for the shutdown of mitotic genes to impose quiescence in mature organs. The two distinct classes of E2Fs and MYB3Rs together with the RETINOBLATOMA RELATED are part of multiprotein complexes that may be evolutionary related to what is known as DREAM complex in animals. In plants, there are multiple such complexes with distinct compositions and functions that may be involved in the coordinated cell cycle and developmental regulation of E2F targets and mitotic genes. Copyright © 2016 Elsevier Ltd. All rights reserved.

Devescovinid trichomonad with axostyle-based rotary motor ("Rubberneckia"): taxonomic assignment as Caduceia versatilis sp. nov

NASA Technical Reports Server (NTRS)

d'Ambrosio, U.; Dolan, M.; Wier, A. M.; Margulis, L.

1999-01-01

An amitochondriate trichomonad cell of the family Devescovinidae (Class Parabasalia), helped demonstrate the fluid model of lipoprotein cell membranes. This wood-ingesting symbiont in the hindgut of the dry wood-eating termite Cryptotermes cavifrons is informally known to cell biologists as "Rubberneckia". As the microtubular axo-style complex generates force causing clockwise movement of the entire anterior portion of the cell at the shear zone the protist displays "head" rotation. Studies by phase contrast and videomicroscopy of live cells, of whole mounts by scanning, and thin sections by transmission electron microscopy extend the observations of Tamm and Tamm [24-26] and Tamm [19-23]. Habitat, cell shape, size, nuclear features, parabasal apparatus and other morphological details permit the assignment of "Rubberneckia" to Kirby's cosmopolitan genus Caduceia. This large-sized devescovinid has distinctive parabasal gyres, an axostylar rotary, motor, and regularly-associated nonflagellated, fusiform and flagellated rod epibiotic surface bacteria. In addition to regularly aligned epibionts intranuclear and endocytoplasmic bacteria are abundant and hydrogenosomes are Present. "Rubberneckia" is compared here to the other seven species of Caduceia. Since it is clearly sufficiently distinctive to warrant new species status, we named it C. versatilis.

Major Histocompatibility Complex (MHC) Class I and MHC Class II Proteins: Conformational Plasticity in Antigen Presentation

PubMed Central

Wieczorek, Marek; Abualrous, Esam T.; Sticht, Jana; Álvaro-Benito, Miguel; Stolzenberg, Sebastian; Noé, Frank; Freund, Christian

2017-01-01

Antigen presentation by major histocompatibility complex (MHC) proteins is essential for adaptive immunity. Prior to presentation, peptides need to be generated from proteins that are either produced by the cell’s own translational machinery or that are funneled into the endo-lysosomal vesicular system. The prolonged interaction between a T cell receptor and specific pMHC complexes, after an extensive search process in secondary lymphatic organs, eventually triggers T cells to proliferate and to mount a specific cellular immune response. Once processed, the peptide repertoire presented by MHC proteins largely depends on structural features of the binding groove of each particular MHC allelic variant. Additionally, two peptide editors—tapasin for class I and HLA-DM for class II—contribute to the shaping of the presented peptidome by favoring the binding of high-affinity antigens. Although there is a vast amount of biochemical and structural information, the mechanism of the catalyzed peptide exchange for MHC class I and class II proteins still remains controversial, and it is not well understood why certain MHC allelic variants are more susceptible to peptide editing than others. Recent studies predict a high impact of protein intermediate states on MHC allele-specific peptide presentation, which implies a profound influence of MHC dynamics on the phenomenon of immunodominance and the development of autoimmune diseases. Here, we review the recent literature that describe MHC class I and II dynamics from a theoretical and experimental point of view and we highlight the similarities between MHC class I and class II dynamics despite the distinct functions they fulfill in adaptive immunity. PMID:28367149

Comprehensive Morpho-Electrotonic Analysis Shows 2 Distinct Classes of L2 and L3 Pyramidal Neurons in Human Temporal Cortex

PubMed Central

Deitcher, Yair; Eyal, Guy; Kanari, Lida; Verhoog, Matthijs B; Atenekeng Kahou, Guy Antoine; Mansvelder, Huibert D; de Kock, Christiaan P J; Segev, Idan

2017-01-01

Abstract There have been few quantitative characterizations of the morphological, biophysical, and cable properties of neurons in the human neocortex. We employed feature-based statistical methods on a rare data set of 60 3D reconstructed pyramidal neurons from L2 and L3 in the human temporal cortex (HL2/L3 PCs) removed after brain surgery. Of these cells, 25 neurons were also characterized physiologically. Thirty-two morphological features were analyzed (e.g., dendritic surface area, 36 333 ± 18 157 μm2; number of basal trees, 5.55 ± 1.47; dendritic diameter, 0.76 ± 0.28 μm). Eighteen features showed a significant gradual increase with depth from the pia (e.g., dendritic length and soma radius). The other features showed weak or no correlation with depth (e.g., dendritic diameter). The basal dendritic terminals in HL2/L3 PCs are particularly elongated, enabling multiple nonlinear processing units in these dendrites. Unlike the morphological features, the active biophysical features (e.g., spike shapes and rates) and passive/cable features (e.g., somatic input resistance, 47.68 ± 15.26 MΩ, membrane time constant, 12.03 ± 1.79 ms, average dendritic cable length, 0.99 ± 0.24) were depth-independent. A novel descriptor for apical dendritic topology yielded 2 distinct classes, termed hereby as “slim-tufted” and “profuse-tufted” HL2/L3 PCs; the latter class tends to fire at higher rates. Thus, our morpho-electrotonic analysis shows 2 distinct classes of HL2/L3 PCs. PMID:28968789

5-HT receptor subtypes as key targets in mediating pigment dispersion within melanophores of teleost, Oreochromis mossambicus.

PubMed

Salim, Saima; Ali, Ayesha S; Ali, Sharique A

2013-02-01

The presence of distinct class of 5-HT receptors in the melanophores of tilapia (Oreochromis mossambicus) is reported. The cellular responses to 5-HT (5-hydroxytryptamine), 5-HT(1), and 5-HT(2), agonists on isolated scale melanophores were observed with regard to pigment translocation within the cells. It was found that 5-HT exerted rapid and strong concentration dependent pigment granule dispersion within the melanophores. The threshold pharmacological dose of 5-HT that could elicit a measurable response was as low as 4.7×10(-12) M/L. Selective 5-HT(1) and 5-HT(2) agonists, sumatriptan and myristicin were investigated and resulted in dose-dependent pigment dispersion. The dispersing effects were effectively antagonized by receptor specific antagonists. It is suggested that 5-HT-induced physiological effects are mediated via distinct classes of receptors that possibly participate in modulation of pigmentary responses of the fish. Copyright © 2012 Elsevier Inc. All rights reserved.

Peripheral formalin injection induces unique spinal cord microglial phenotypic changes

PubMed Central

Fu, Kai-Yuan; Tan, Yong-Hui; Sung, Backil; Mao, Jianren

2014-01-01

Microglia are resident immune cells of brain and activated by peripheral tissue injury. In the present study, we investigated the possible induction of several microglial surface immunomolecules in the spinal cord, including leukocyte common antigen (LCA/CD45), MHC class I antigen, MHC class II antigen, Fc receptor, and CD11c following formalin injection into the rat’s hind paw. CD45 and MHC class I were upregulated in the activated microglia, which was evident on day 3 with the peak expression on day 7 following peripheral formalin injection. There was a very low basal expression of MHC class II, CD11c, and the Fc receptor, which did not change after the formalin injection. These results, for the first time, indicate that peripheral formalin injection can induce phenotypic changes of microglia with distinct upregulation of CD45 and MHC class I antigen. The data suggest that phenotypic changes of the activated microglia may be a unique pattern of central changes following peripheral tissue injury. PMID:19015000

p110α and p110β isoforms of PI3K signaling: are they two sides of the same coin?

PubMed

Singh, Paramjeet; Dar, Mohd Saleem; Dar, Mohd Jamal

2016-09-01

Class-1 phosphatidylinositol-3-kinases (PI3Ks) are activated by a variety of extracellular stimuli and have been implicated in a wide range of cellular processes. p110α and p110β are the two most studied isoforms of the class-1A PI3K signaling pathway. Although these two isoforms are ubiquitously expressed and play multiple redundant roles, they also have distinct functions within the cell. More recently, p110α and p110β isoforms have been shown to translocate into the nucleus and play a role in DNA replication and repair, and in cell cycle progression. In the following Review article, we discuss the overlapping and unique roles of p110α and p110β isoforms with a particular focus on their structure, expression analysis, subcellular localization, and signaling contributions in various cell types and model organisms. © 2016 Federation of European Biochemical Societies.

Recessive cardiac phenotypes in induced pluripotent stem cell models of Jervell and Lange-Nielsen syndrome: disease mechanisms and pharmacological rescue.

PubMed

Zhang, Miao; D'Aniello, Cristina; Verkerk, Arie O; Wrobel, Eva; Frank, Stefan; Ward-van Oostwaard, Dorien; Piccini, Ilaria; Freund, Christian; Rao, Jyoti; Seebohm, Guiscard; Atsma, Douwe E; Schulze-Bahr, Eric; Mummery, Christine L; Greber, Boris; Bellin, Milena

2014-12-16

Jervell and Lange-Nielsen syndrome (JLNS) is one of the most severe life-threatening cardiac arrhythmias. Patients display delayed cardiac repolarization, associated high risk of sudden death due to ventricular tachycardia, and congenital bilateral deafness. In contrast to the autosomal dominant forms of long QT syndrome, JLNS is a recessive trait, resulting from homozygous (or compound heterozygous) mutations in KCNQ1 or KCNE1. These genes encode the α and β subunits, respectively, of the ion channel conducting the slow component of the delayed rectifier K(+) current, IKs. We used complementary approaches, reprogramming patient cells and genetic engineering, to generate human induced pluripotent stem cell (hiPSC) models of JLNS, covering splice site (c.478-2A>T) and missense (c.1781G>A) mutations, the two major classes of JLNS-causing defects in KCNQ1. Electrophysiological comparison of hiPSC-derived cardiomyocytes (CMs) from homozygous JLNS, heterozygous, and wild-type lines recapitulated the typical and severe features of JLNS, including pronounced action and field potential prolongation and severe reduction or absence of IKs. We show that this phenotype had distinct underlying molecular mechanisms in the two sets of cell lines: the previously unidentified c.478-2A>T mutation was amorphic and gave rise to a strictly recessive phenotype in JLNS-CMs, whereas the missense c.1781G>A lesion caused a gene dosage-dependent channel reduction at the cell membrane. Moreover, adrenergic stimulation caused action potential prolongation specifically in JLNS-CMs. Furthermore, sensitivity to proarrhythmic drugs was strongly enhanced in JLNS-CMs but could be pharmacologically corrected. Our data provide mechanistic insight into distinct classes of JLNS-causing mutations and demonstrate the potential of hiPSC-CMs in drug evaluation.

Social space and cultural class divisions: the forms of capital and contemporary lifestyle differentiation.

PubMed

Flemmen, Magne; Jarness, Vegard; Rosenlund, Lennart

2018-03-01

In this article, we address whether and how contemporary social classes are marked by distinct lifestyles. We assess the model of the social space, a novel approach to class analysis pioneered by Bourdieu's Distinction. Although pivotal in Bourdieu's work, this model is too often overlooked in later research, making its contemporary relevance difficult to assess. We redress this by using the social space as a framework through which to study the cultural manifestation of class divisions in lifestyle differences in contemporary Norwegian society. Through a Multiple Correspondence Analysis (MCA) of unusually rich survey data, we reveal a structure strikingly similar to the model in Distinction, with a primary dimension of the volume of capital, and a secondary dimension of the composition of capital. While avoiding the substantialist fallacy of predefined notions of 'highbrow' and 'lowbrow' tastes, we explore how 168 lifestyle items map onto this social space. This reveals distinct classed lifestyles according to both dimensions of the social space. The lifestyles of the upper classes are distinctly demanding in terms of resources. Among those rich in economic capital, this manifests itself in a lifestyle which involves a quest for excitement, and which is bodily oriented and expensive. For their counterparts rich in cultural capital, a more ascetic and intellectually oriented lifestyle manifests itself, demanding of resources in the sense of requiring symbolic mastery, combining a taste for canonized, legitimate culture with more cosmopolitan and 'popular' items. In contrast to many studies' descriptions of the lower classes as 'disengaged' and 'inactive', we find evidence of distinct tastes on their part. Our analysis thus affirms the validity of Bourdieu's model of social class and the contention that classes tend to take the form of status groups. We challenge dominant positions in cultural stratification research, while questioning the aptness of the metaphor of the 'omnivore', as well as recent analyses of 'emerging cultural capital'. © London School of Economics and Political Science 2017.

Diversity amongst trigeminal neurons revealed by high throughput single cell sequencing

PubMed Central

Nguyen, Minh Q.; Wu, Youmei; Bonilla, Lauren S.; von Buchholtz, Lars J.

2017-01-01

The trigeminal ganglion contains somatosensory neurons that detect a range of thermal, mechanical and chemical cues and innervate unique sensory compartments in the head and neck including the eyes, nose, mouth, meninges and vibrissae. We used single-cell sequencing and in situ hybridization to examine the cellular diversity of the trigeminal ganglion in mice, defining thirteen clusters of neurons. We show that clusters are well conserved in dorsal root ganglia suggesting they represent distinct functional classes of somatosensory neurons and not specialization associated with their sensory targets. Notably, functionally important genes (e.g. the mechanosensory channel Piezo2 and the capsaicin gated ion channel Trpv1) segregate into multiple clusters and often are expressed in subsets of cells within a cluster. Therefore, the 13 genetically-defined classes are likely to be physiologically heterogeneous rather than highly parallel (i.e., redundant) lines of sensory input. Our analysis harnesses the power of single-cell sequencing to provide a unique platform for in silico expression profiling that complements other approaches linking gene-expression with function and exposes unexpected diversity in the somatosensory system. PMID:28957441

Highly parallel genome-wide expression profiling of individual cells using nanoliter droplets

PubMed Central

Macosko, Evan Z.; Basu, Anindita; Satija, Rahul; Nemesh, James; Shekhar, Karthik; Goldman, Melissa; Tirosh, Itay; Bialas, Allison R.; Kamitaki, Nolan; Martersteck, Emily M.; Trombetta, John J.; Weitz, David A.; Sanes, Joshua R.; Shalek, Alex K.; Regev, Aviv; McCarroll, Steven A.

2015-01-01

Summary Cells, the basic units of biological structure and function, vary broadly in type and state. Single-cell genomics can characterize cell identity and function, but limitations of ease and scale have prevented its broad application. Here we describe Drop-Seq, a strategy for quickly profiling thousands of individual cells by separating them into nanoliter-sized aqueous droplets, associating a different barcode with each cell’s RNAs, and sequencing them all together. Drop-Seq analyzes mRNA transcripts from thousands of individual cells simultaneously while remembering transcripts’ cell of origin. We analyzed transcriptomes from 44,808 mouse retinal cells and identified 39 transcriptionally distinct cell populations, creating a molecular atlas of gene expression for known retinal cell classes and novel candidate cell subtypes. Drop-Seq will accelerate biological discovery by enabling routine transcriptional profiling at single-cell resolution. PMID:26000488

The genetic origin of minor histocompatibility antigens.

PubMed

Roopenian, D C; Christianson, G J; Davis, A P; Zuberi, A R; Mobraaten, L E

1993-01-01

The purpose of this study was to elucidate the genetic origin of minor histocompatibility (H) antigens. Toward this end common inbred mouse strains, distinct subspecies, and species of the subgenus Mus were examined for expression of various minor H antigens. These antigens were encoded by the classical minor H loci H-3 and H-4 or by newly identified minor H antigens detected as a consequence of mutation. Both minor H antigens that stimulate MHC class I-restricted cytotoxic T cells (Tc) and antigens that stimulate MHC class II-restricted helper T cells (Th) were monitored. The results suggested that strains of distinct ancestry commonly express identical or cross-reactive antigens. Moreover, a correlation between the lack of expression of minor H antigens and ancestral heritage was observed. To address whether the antigens found on unrelated strains were allelic with the sensitizing minor H antigens or a consequence of antigen cross-reactivity, classical genetic segregation analysis was carried out. Even in distinct subspecies and species, the minor H antigens always mapped to the site of the appropriate minor H locus. Together the results suggest: 1) minor H antigen sequences are evolutionarily stable in that their pace of antigenic change is slow enough to predate subspeciation and speciation; 2) the minor H antigens originated in the inbred strains as a consequence of a rare polymorphism or loss mutation carried in a founder mouse stock that caused the mouse to perceive the wild-type protein as foreign; 3) there is a remarkable lack of antigenic cross-reactivity between the defined minor H antigens and other gene products.

The LIM and POU homeobox genes ttx-3 and unc-86 act as terminal selectors in distinct cholinergic and serotonergic neuron types.

PubMed

Zhang, Feifan; Bhattacharya, Abhishek; Nelson, Jessica C; Abe, Namiko; Gordon, Patricia; Lloret-Fernandez, Carla; Maicas, Miren; Flames, Nuria; Mann, Richard S; Colón-Ramos, Daniel A; Hobert, Oliver

2014-01-01

Transcription factors that drive neuron type-specific terminal differentiation programs in the developing nervous system are often expressed in several distinct neuronal cell types, but to what extent they have similar or distinct activities in individual neuronal cell types is generally not well explored. We investigate this problem using, as a starting point, the C. elegans LIM homeodomain transcription factor ttx-3, which acts as a terminal selector to drive the terminal differentiation program of the cholinergic AIY interneuron class. Using a panel of different terminal differentiation markers, including neurotransmitter synthesizing enzymes, neurotransmitter receptors and neuropeptides, we show that ttx-3 also controls the terminal differentiation program of two additional, distinct neuron types, namely the cholinergic AIA interneurons and the serotonergic NSM neurons. We show that the type of differentiation program that is controlled by ttx-3 in different neuron types is specified by a distinct set of collaborating transcription factors. One of the collaborating transcription factors is the POU homeobox gene unc-86, which collaborates with ttx-3 to determine the identity of the serotonergic NSM neurons. unc-86 in turn operates independently of ttx-3 in the anterior ganglion where it collaborates with the ARID-type transcription factor cfi-1 to determine the cholinergic identity of the IL2 sensory and URA motor neurons. In conclusion, transcription factors operate as terminal selectors in distinct combinations in different neuron types, defining neuron type-specific identity features.

Polymorphisms and Tissue Expression of the Feline Leukocyte Antigen Class I Loci FLAI-E, -H and -K

PubMed Central

Holmes, Jennifer C.; Holmer, Savannah G.; Ross, Peter; Buntzman, Adam S.; Frelinger, Jeffrey A.; Hess, Paul R.

2013-01-01

Cytotoxic CD8+ T-cell immunosurveillance for intracellular pathogens, such as viruses, is controlled by classical major histocompatibility complex (MHC) class Ia molecules, and ideally, these antiviral T-cell populations are defined by the specific peptide and restricting MHC allele. Surprisingly, despite the utility of the cat in modeling human viral immunity, little is known about the Feline Leukocyte Antigen class I complex (FLAI). Only a few coding sequences with uncertain locus origin and expression patterns have been reported. Of 19 class I genes, 3 loci - FLAI-E, -H and -K – are predicted to encode classical molecules, and our objective was to evaluate their status by analyzing polymorphisms and tissue expression. Using locus-specific, PCR-based genotyping, we amplified 33 FLAI-E, -H, and -K alleles from 12 cats of various breeds, identifying, for the first time, alleles across 3 distinct loci in a feline species. Alleles shared the expected polymorphic and invariant sites in the α1/α2 domains, and full-length cDNA clones possessed all characteristic class Ia exons. Alleles could be assigned to a specific locus with reasonable confidence, although there was evidence of potentially confounding interlocus recombination between FLAI-E and -K. Only FLAI-E, -H and -K-origin alleles were amplified from cDNAs of multiple tissue types. We also defined hypervariable regions across these genes, which permitted the assignment of names to both novel and established alleles. As predicted, FLAI-E, -H, and -K fulfill the major criteria of class Ia genes. These data represent a necessary prerequisite for studying epitope-specific antiviral CD8+ T-cell responses in cats. PMID:23812210

Analysis of class II (hydrolytic) and class I (beta-lyase) apurinic/apyrimidinic endonucleases with a synthetic DNA substrate.

PubMed Central

Levin, J D; Demple, B

1990-01-01

We have developed simple and sensitive assays that distinguish the main classes of apurinic/apyrimidinic (AP) endonucleases: Class I enzymes that cleave on the 3' side of AP sites by beta-elimination, and Class II enzymes that cleave by hydrolysis on the 5' side. The distinction of the two types depends on the use of a synthetic DNA polymer that contains AP sites with 5'-[32P]phosphate residues. Using this approach, we now show directly that Escherichia coli endonuclease IV and human AP endonuclease are Class II enzymes, as inferred previously on the basis of indirect assays. The assay method does not exhibit significant interference by nonspecific nucleases or primary amines, which allows the ready determination of different AP endonuclease activities in crude cell extracts. In this way, we show that virtually all of the Class II AP endonuclease activity in E. coli can be accounted for by two enzymes: exonuclease III and endonuclease IV. In the yeast Saccharomyces cerevisiae, the Class II AP endonuclease activity is totally dependent on a single enzyme, the Apn1 protein, but there are probably multiple Class I enzymes. The versatility and ease of our approach should be useful for characterizing this important class of DNA repair enzymes in diverse systems. PMID:1698278

Met receptor inhibitor SU11274 localizes in the endoplasmic reticulum.

PubMed

Wiest, Edwin J; Smith, Heather Jensen; Hollingsworth, Michael A

2018-07-02

We discovered that SU11274, a class I c-Met inhibitor, fluoresces when excited by 488 nm laser light and showed rapid specific accumulation in distinct subcellular compartments. Given that SU11274 reduces cancer cell viability, we exploited these newly identified spectral properties to determine SU11274 intracellular distribution and accumulation in human pancreatic cancer cells. The aim of the studies reported here was to identify organelle(s) to which SU11274 is trafficked. We conclude that SU11274 rapidly and predominantly accumulates in the endoplasmic reticulum. Copyright © 2018. Published by Elsevier Inc.

Local Circuits of V1 Layer 4B Neurons Projecting to V2 Thick Stripes Define Distinct Cell Classes and Avoid Cytochrome Oxidase Blobs

PubMed Central

Yarch, Jeff; Federer, Frederick

2017-01-01

Decades of anatomical studies on the primate primary visual cortex (V1) have led to a detailed diagram of V1 intrinsic circuitry, but this diagram lacks information about the output targets of V1 cells. Understanding how V1 local processing relates to downstream processing requires identification of neuronal populations defined by their output targets. In primates, V1 layers (L)2/3 and 4B send segregated projections to distinct cytochrome oxidase (CO) stripes in area V2: neurons in CO blob columns project to thin stripes while neurons outside blob columns project to thick and pale stripes, suggesting functional specialization of V1-to-V2 CO streams. However, the conventional diagram of V1 shows all L4B neurons, regardless of their soma location in blob or interblob columns, as projecting selectively to CO blobs in L2/3, suggesting convergence of blob/interblob information in L2/3 blobs and, possibly, some V2 stripes. However, it is unclear whether all L4B projection neurons show similar local circuitries. Using viral-mediated circuit tracing, we have identified the local circuits of L4B neurons projecting to V2 thick stripes in macaque. Consistent with previous studies, we found the somata of this L4B subpopulation to reside predominantly outside blob columns; however, unlike previous descriptions of local L4B circuits, these cells consistently projected outside CO blob columns in all layers. Thus, the local circuits of these L4B output neurons, just like their extrinsic projections to V2, preserve CO streams. Moreover, the intra-V1 laminar patterns of axonal projections identify two distinct neuron classes within this L4B subpopulation, including a rare novel neuron type, suggestive of two functionally specialized output channels. SIGNIFICANCE STATEMENT Conventional diagrams of primate primary visual cortex (V1) depict neuronal connections within and between different V1 layers, but lack information about the cells' downstream targets. This information is critical to understanding how local processing in V1 relates to downstream processing. We have identified the local circuits of a population of cells in V1 layer (L)4B that project to area V2. These cells' local circuits differ from classical descriptions of L4B circuits in both the laminar and functional compartments targeted by their axons, and identify two neuron classes. Our results demonstrate that both local intra-V1 and extrinsic V1-to-V2 connections of L4B neurons preserve CO-stream segregation, suggesting that across-stream integration occurs downstream of V1, and that output targets dictate local V1 circuitry. PMID:28077720

In Vitro Growth Inhibitory Activities of Natural Products from Irciniid Sponges against Cancer Cells: A Comparative Study

PubMed Central

BenRedjem Romdhane, Yosr; Elbour, Monia; Carbone, Marianna; Ciavatta, Maria Letizia; Gavagnin, Margherita; Mathieu, Véronique; Lefranc, Florence; Ktari, Leila; Ben Mustapha, Karim; Boudabous, Abdellatif; Kiss, Robert

2016-01-01

Marine sponges of the Irciniidae family contain both bioactive furanosesterterpene tetronic acids (FTAs) and prenylated hydroquinones (PHQs). Both classes of compounds are known for their anti-inflammatory, antioxidant, and antimicrobial properties and known to display growth inhibitory effects against various human tumor cell lines. However, the different experimental conditions of the reported in vitro bioassays, carried out on different cancer cell lines within separate studies, prevent realistic actual discrimination between the two classes of compounds from being carried out in terms of growth inhibitory effects. In the present work, a chemical investigation of irciniid sponges from Tunisian coasts led to the purification of three known FTAs and three known PHQs. The in vitro growth inhibitory properties of the six purified compounds have been evaluated in the same experiment in a panel of five human and one murine cancer cell lines displaying various levels of sensitivity to proapoptotic stimuli. Surprisingly, FTAs and PHQs elicited distinct profiles of growth inhibitory-responses, differing by one to two orders of magnitude in favor of the PHQs in all cell lines. The obtained comparative results are discussed in the light of a better selection of drug candidates from natural sources. PMID:27597966

Word Class Distinctions in Second Language Acquisition: An Experimental Study of L2 Spanish

ERIC Educational Resources Information Center

Zyzik, Eve; Azevedo, Clara

2009-01-01

Although the problem of word class has been explored in numerous first language studies, relatively little is known about this process in SLA. The present study measures second language (L2) learners' knowledge of word class distinctions (e.g., noun vs. adjective) in a variety of syntactic contexts. English-speaking learners of Spanish from…

Do recognizable lifetime eating disorder phenotypes naturally occur in a culturally asian population? A combined latent profile and taxometric approach.

PubMed

Thomas, Jennifer J; Eddy, Kamryn T; Ruscio, John; Ng, King Lam; Casale, Kristen E; Becker, Anne E; Lee, Sing

2015-05-01

We examined whether empirically derived eating disorder (ED) categories in Hong Kong Chinese patients (N = 454) would be consistent with recognizable lifetime ED phenotypes derived from latent structure models of European and American samples. We performed latent profile analysis (LPA) using indicator variables from data collected during routine assessment, and then applied taxometric analysis to determine whether latent classes were qualitatively versus quantitatively distinct. Latent profile analysis identified four classes: (i) binge/purge (47%); (ii) non-fat-phobic low-weight (34%); (iii) fat-phobic low-weight (12%); and (iv) overweight disordered eating (6%). Taxometric analysis identified qualitative (categorical) distinctions between the binge/purge and non-fat-phobic low-weight classes, and also between the fat-phobic and non-fat-phobic low-weight classes. Distinctions between the fat-phobic low-weight and binge/purge classes were indeterminate. Empirically derived categories in Hong Kong showed recognizable correspondence with recognizable lifetime ED phenotypes. Although taxometric findings support two distinct classes of low weight EDs, LPA findings also support heterogeneity among non-fat-phobic individuals. Copyright © 2015 John Wiley & Sons, Ltd and Eating Disorders Association.

Control of germline stem cell self-renewal and differentiation in the Drosophila ovary: concerted actions of niche signals and intrinsic factors.

PubMed

Xie, Ting

2013-01-01

In the Drosophila ovary, germline stem cells (GSCs) physically interact with their niche composed of terminal filament cells, cap cells, and possibly GSC-contacting escort cells (ECs). A GSC divides to generate a self-renewing stem cell that remains in the niche and a differentiating daughter that moves away from the niche. The GSC niche provides a bone morphogenetic protein (BMP) signal that maintains GSC self-renewal by preventing stem cell differentiation via repression of the differentiation-promoting gene bag of marbles (bam). In addition, it expresses E-cadherin, which mediates cell adhesion for anchoring GSCs in the niche, enabling continuous self-renewal. GSCs themselves also express different classes of intrinsic factors, including signal transducers, transcription factors, chromatin remodeling factors, translation regulators, and miRNAs, which control self-renewal by strengthening interactions with the niche and repressing various differentiation pathways. Differentiated GSC daughters, known as cystoblasts (CBs), also express distinct classes of intrinsic factors to inhibit self-renewal and promote germ cell differentiation. Surprisingly, GSC progeny are also dependent on their surrounding ECs for proper differentiation at least partly by preventing BMP from diffusing to the differentiated germ cell zone and by repressing ectopic BMP expression. Therefore, both GSC self-renewal and CB differentiation are controlled by collaborative actions of extrinsic signals and intrinsic factors. Copyright © 2012 Wiley Periodicals, Inc.

Anticancer Properties of Distinct Antimalarial Drug Classes

PubMed Central

Hooft van Huijsduijnen, Rob; Guy, R. Kiplin; Chibale, Kelly; Haynes, Richard K.; Peitz, Ingmar; Kelter, Gerhard; Phillips, Margaret A.; Vennerstrom, Jonathan L.; Yuthavong, Yongyuth; Wells, Timothy N. C.

2013-01-01

We have tested five distinct classes of established and experimental antimalarial drugs for their anticancer potential, using a panel of 91 human cancer lines. Three classes of drugs: artemisinins, synthetic peroxides and DHFR (dihydrofolate reductase) inhibitors effected potent inhibition of proliferation with IC50s in the nM- low µM range, whereas a DHODH (dihydroorotate dehydrogenase) and a putative kinase inhibitor displayed no activity. Furthermore, significant synergies were identified with erlotinib, imatinib, cisplatin, dasatinib and vincristine. Cluster analysis of the antimalarials based on their differential inhibition of the various cancer lines clearly segregated the synthetic peroxides OZ277 and OZ439 from the artemisinin cluster that included artesunate, dihydroartemisinin and artemisone, and from the DHFR inhibitors pyrimethamine and P218 (a parasite DHFR inhibitor), emphasizing their shared mode of action. In order to further understand the basis of the selectivity of these compounds against different cancers, microarray-based gene expression data for 85 of the used cell lines were generated. For each compound, distinct sets of genes were identified whose expression significantly correlated with compound sensitivity. Several of the antimalarials tested in this study have well-established and excellent safety profiles with a plasma exposure, when conservatively used in malaria, that is well above the IC50s that we identified in this study. Given their unique mode of action and potential for unique synergies with established anticancer drugs, our results provide a strong basis to further explore the potential application of these compounds in cancer in pre-clinical or and clinical settings. PMID:24391728

[BIOINFORMATIC SEARCH AND PHYLOGENETIC ANALYSIS OF THE CELLULOSE SYNTHASE GENES OF FLAX (LINUM USITATISSIMUM)].

PubMed

Pydiura, N A; Bayer, G Ya; Galinousky, D V; Yemets, A I; Pirko, Ya V; Podvitski, T A; Anisimova, N V; Khotyleva, L V; Kilchevsky, A V; Blume, Ya B

2015-01-01

A bioinformatic search of sequences encoding cellulose synthase genes in the flax genome, and their comparison to dicots orthologs was carried out. The analysis revealed 32 cellulose synthase gene candidates, 16 of which are highly likely to encode cellulose synthases, and the remaining 16--cellulose synthase-like proteins (Csl). Phylogenetic analysis of gene products of cellulose synthase genes allowed distinguishing 6 groups of cellulose synthase genes of different classes: CesA1/10, CesA3, CesA4, CesA5/6/2/9, CesA7 and CesA8. Paralogous sequences within classes CesA1/10 and CesA5/6/2/9 which are associated with the primary cell wall formation are characterized by a greater similarity within these classes than orthologous sequences. Whereas the genes controlling the biosynthesis of secondary cell wall cellulose form distinct clades: CesA4, CesA7, and CesA8. The analysis of 16 identified flax cellulose synthase gene candidates shows the presence of at least 12 different cellulose synthase gene variants in flax genome which are represented in all six clades of cellulose synthase genes. Thus, at this point genes of all ten known cellulose synthase classes are identify in flax genome, but their correct classification requires additional research.

Characterizing heterogeneous cellular responses to perturbations.

PubMed

Slack, Michael D; Martinez, Elisabeth D; Wu, Lani F; Altschuler, Steven J

2008-12-09

Cellular populations have been widely observed to respond heterogeneously to perturbation. However, interpreting the observed heterogeneity is an extremely challenging problem because of the complexity of possible cellular phenotypes, the large dimension of potential perturbations, and the lack of methods for separating meaningful biological information from noise. Here, we develop an image-based approach to characterize cellular phenotypes based on patterns of signaling marker colocalization. Heterogeneous cellular populations are characterized as mixtures of phenotypically distinct subpopulations, and responses to perturbations are summarized succinctly as probabilistic redistributions of these mixtures. We apply our method to characterize the heterogeneous responses of cancer cells to a panel of drugs. We find that cells treated with drugs of (dis-)similar mechanism exhibit (dis-)similar patterns of heterogeneity. Despite the observed phenotypic diversity of cells observed within our data, low-complexity models of heterogeneity were sufficient to distinguish most classes of drug mechanism. Our approach offers a computational framework for assessing the complexity of cellular heterogeneity, investigating the degree to which perturbations induce redistributions of a limited, but nontrivial, repertoire of underlying states and revealing functional significance contained within distinct patterns of heterogeneous responses.

NK cell activation: distinct stimulatory pathways counterbalancing inhibitory signals.

PubMed

Bakker, A B; Wu, J; Phillips, J H; Lanier, L L

2000-01-01

A delicate balance between positive and negative signals regulates NK cell effector function. Activation of NK cells may be initiated by the triggering of multiple adhesion or costimulatory molecules, and can be counterbalanced by inhibitory signals induced by receptors for MHC class I. A common pathway of inhibitory signaling is provided by immunoreceptor tyrosine-based inhibitory motifs (ITIMs) in the cytoplasmic domains of these receptors which mediate the recruitment of SH2 domain-bearing tyrosine phosphate-1 (SHP-1). In contrast to the extensive progress that has been made regarding the negative regulation of NK cell function, our knowledge of the signals that activate NK cells is still poor. Recent studies of the activating receptor complexes have shed new light on the induction of NK cell effector function. Several NK receptors using novel adaptors with immunoreceptor tyrosine-based activation motifs (ITAMs) and with PI 3-kinase recruiting motifs have been implicated in NK cell stimulation.

Zinc in innate and adaptive tumor immunity

PubMed Central

2010-01-01

Zinc is important. It is the second most abundant trace metal with 2-4 grams in humans. It is an essential trace element, critical for cell growth, development and differentiation, DNA synthesis, RNA transcription, cell division, and cell activation. Zinc deficiency has adverse consequences during embryogenesis and early childhood development, particularly on immune functioning. It is essential in members of all enzyme classes, including over 300 signaling molecules and transcription factors. Free zinc in immune and tumor cells is regulated by 14 distinct zinc importers (ZIP) and transporters (ZNT1-8). Zinc depletion induces cell death via apoptosis (or necrosis if apoptotic pathways are blocked) while sufficient zinc levels allows maintenance of autophagy. Cancer cells have upregulated zinc importers, and frequently increased zinc levels, which allow them to survive. Based on this novel synthesis, approaches which locally regulate zinc levels to promote survival of immune cells and/or induce tumor apoptosis are in order. PMID:21087493

Social class, power, and selfishness: when and why upper and lower class individuals behave unethically.

PubMed

Dubois, David; Rucker, Derek D; Galinsky, Adam D

2015-03-01

Are the rich more unethical than the poor? To answer this question, the current research introduces a key conceptual distinction between selfish and unethical behavior. Based on this distinction, the current article offers 2 novel findings that illuminate the relationship between social class and unethical behavior. First, the effects of social class on unethical behavior are not invariant; rather, the effects of social class are moderated by whether unethical behavior benefits the self or others. Replicating past work, social class positively predicted unethical behavior; however, this relationship was only observed when that behavior was self-beneficial. When unethical behavior was performed to benefit others, social class negatively predicted unethical behavior; lower class individuals were more likely than upper class individuals to engage in unethical behavior. Overall, social class predicts people's tendency to behave selfishly, rather than predicting unethical behavior per se. Second, individuals' sense of power drove the effects of social class on unethical behavior. Evidence for this relationship was provided in three forms. First, income, but not education level, predicted unethical behavior. Second, feelings of power mediated the effect of social class on unethical behavior, but feelings of status did not. Third, two distinct manipulations of power produced the same moderation by self-versus-other beneficiary as was found with social class. The current theoretical framework and data both synthesize and help to explain a range of findings in the social class and power literatures. PsycINFO Database Record (c) 2015 APA, all rights reserved.

High-content image informatics of the structural nuclear protein NuMA parses trajectories for stem/progenitor cell lineages and oncogenic transformation.

PubMed

Vega, Sebastián L; Liu, Er; Arvind, Varun; Bushman, Jared; Sung, Hak-Joon; Becker, Matthew L; Lelièvre, Sophie; Kohn, Joachim; Vidi, Pierre-Alexandre; Moghe, Prabhas V

2017-02-01

Stem and progenitor cells that exhibit significant regenerative potential and critical roles in cancer initiation and progression remain difficult to characterize. Cell fates are determined by reciprocal signaling between the cell microenvironment and the nucleus; hence parameters derived from nuclear remodeling are ideal candidates for stem/progenitor cell characterization. Here we applied high-content, single cell analysis of nuclear shape and organization to examine stem and progenitor cells destined to distinct differentiation endpoints, yet undistinguishable by conventional methods. Nuclear descriptors defined through image informatics classified mesenchymal stem cells poised to either adipogenic or osteogenic differentiation, and oligodendrocyte precursors isolated from different regions of the brain and destined to distinct astrocyte subtypes. Nuclear descriptors also revealed early changes in stem cells after chemical oncogenesis, allowing the identification of a class of cancer-mitigating biomaterials. To capture the metrology of nuclear changes, we developed a simple and quantitative "imaging-derived" parsing index, which reflects the dynamic evolution of the high-dimensional space of nuclear organizational features. A comparative analysis of parsing outcomes via either nuclear shape or textural metrics of the nuclear structural protein NuMA indicates the nuclear shape alone is a weak phenotypic predictor. In contrast, variations in the NuMA organization parsed emergent cell phenotypes and discerned emergent stages of stem cell transformation, supporting a prognosticating role for this protein in the outcomes of nuclear functions. Copyright © 2017 Elsevier Inc. All rights reserved.

Lipid rafts in T cell signalling and disease

PubMed Central

Jury, Elizabeth C.; Flores-Borja, Fabian; Kabouridis, Panagiotis S.

2007-01-01

Lipid rafts is a blanket term used to describe distinct areas in the plasma membrane rich in certain lipids and proteins and which are thought to perform diverse functions. A large number of studies report on lipid rafts having a key role in receptor signalling and activation of lymphocytes. In T cells, lipid raft involvement was demonstrated in the early steps during T cell receptor (TCR) stimulation. Interestingly, recent evidence has shown that signalling in these domains differs in T cells isolated from patients with autoimmune diseases such as systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA). Here, we discuss these findings and explore the potential of lipid rafts as targets for the development of a new class of agents to downmodulate immune responses and for the treatment of autoimmune diseases. PMID:17890113

Distinct Immunoglobulin Class and Immunoglobulin G Subclass Patterns against Ganglioside GQ1b in Miller Fisher Syndrome following Different Types of Infection

PubMed Central

Schwerer, Beatrix; Neisser, Andrea; Bernheimer, Hanno

1999-01-01

We studied serum antibodies against gangliosides GQ1b and GM1 in 13 patients with Miller Fisher syndrome (MFS) and in 18 patients with Guillain-Barré syndrome (GBS) with cranial nerve involvement. Anti-GQ1b titers were elevated in all patients with MFS cases (immunoglobulin G [IgG] > IgA, IgM), and in 8 of the 18 with GBS. Lower frequencies of increased anti-GM1 titers were observed in MFS patients (3 of 13), as well as in GBS patients (5 of 18). During the course of MFS, anti-GQ1b titers of all Ig classes decreased within 3 weeks after onset. By contrast, anti-GM1 titers (mainly IgM) transiently increased during the course of MFS in five of six patients, suggesting a nonspecific secondary immune response. In patients with MFS following respiratory infections, IgG was the major anti-GQ1b Ig class (six of six patients) and IgG3 was the major subclass (five of six). In contrast, four of five patients with MFS following gastrointestinal infections showed predominance of anti-GQ1b IgA or IgM over IgG and predominance of the IgG2 subclass; anti-GQ1b IgG (IgG3) prevailed in one patient only. These distinct Ig patterns strongly suggest that different infections may trigger different mechanisms of anti-GQ1b production, such as via T-cell-dependent as opposed to T-cell-independent pathways. Thus, the origin of antibodies against GQ1b in MFS may be determined by the type of infectious agent that precipitates the disease. PMID:10225903

Early recycling compartment trafficking of CD1a is essential for its intersection and presentation of lipid antigens.

PubMed

Cernadas, Manuela; Cavallari, Marco; Watts, Gerald; Mori, Lucia; De Libero, Gennaro; Brenner, Michael B

2010-02-01

A major step in understanding differences in the nature of Ag presentation was the realization that MHC class I samples peptides transported to the endoplasmic reticulum from the cytosol, whereas MHC class II samples peptides from lysosomes. In contrast to MHC class I and II molecules that present protein Ags, CD1 molecules present lipid Ags for recognition by specific T cells. Each of the five members of the CD1 family (CD1a-e) localizes to a distinct subcompartment of endosomes. Accordingly, it has been widely assumed that the distinct trafficking of CD1 isoforms must also have evolved to enable them to sample lipid Ags that traffic via different routes. Among the CD1 isoforms, CD1a is unusual because it does not have a tyrosine-based cytoplasmic sorting motif and uniquely localizes to the early endocytic recycling compartment. This led us to predict that CD1a might have evolved to focus on lipids that localize to early endocytic/recycling compartments. Strikingly, we found that the glycolipid Ag sulfatide also localized almost exclusively to early endocytic and recycling compartments. Consistent with colocalization of CD1a and sulfatide, wild-type CD1a molecules efficiently presented sulfatide to CD1a-restricted, sulfatide-specific T cells. In contrast, CD1a:CD1b tail chimeras, that retain the same Ag-binding capacity as CD1a but traffic based on the cytoplasmic tail of CD1b to lysosomes, failed to present sulfatide efficiently. Thus, the intracellular trafficking route of CD1a is essential for efficient presentation of lipid Ags that traffic through the early endocytic and recycling pathways.

PMS2 endonuclease activity has distinct biological functions and is essential for genome maintenance.

PubMed

van Oers, Johanna M M; Roa, Sergio; Werling, Uwe; Liu, Yiyong; Genschel, Jochen; Hou, Harry; Sellers, Rani S; Modrich, Paul; Scharff, Matthew D; Edelmann, Winfried

2010-07-27

The DNA mismatch repair protein PMS2 was recently found to encode a novel endonuclease activity. To determine the biological functions of this activity in mammals, we generated endonuclease-deficient Pms2E702K knock-in mice. Pms2EK/EK mice displayed increased genomic mutation rates and a strong cancer predisposition. In addition, class switch recombination, but not somatic hypermutation, was impaired in Pms2EK/EK B cells, indicating a specific role in Ig diversity. In contrast to Pms2-/- mice, Pms2EK/EK male mice were fertile, indicating that this activity is dispensable in spermatogenesis. Therefore, the PMS2 endonuclease activity has distinct biological functions and is essential for genome maintenance and tumor suppression.

Distinct mobilization of leukocytes and hematopoietic stem cells by CXCR4 peptide antagonist LY2510924 and monoclonal antibody LY2624587

PubMed Central

Peng, Sheng-Bin; Van Horn, Robert D.; Yin, Tinggui; Brown, Robin M.; Roell, William C.; Obungu, Victor H.; Ruegg, Charles; Wroblewski, Victor J.; Raddad, Eyas; Stille, John R.

2017-01-01

Stromal cell-derived factor-1 (SDF-1) and its receptor CXCR4 play a critical role in mobilization and redistribution of immune cells and hematopoietic stem cells (HSCs). We evaluated effects of two CXCR4-targeting agents, peptide antagonist LY2510924 and monoclonal antibody LY2624587, on mobilizing HSCs and white blood cells (WBCs) in humans, monkeys, and mice. Biochemical analysis showed LY2510924 peptide blocked SDF-1/CXCR4 binding in all three species; LY2624587 antibody blocked binding in human and monkey, with minimal activity in mouse. Cellular analysis showed LY2624587 antibody, but not LY2510924 peptide, down-regulated cell surface CXCR4 and induced hematological tumor cell death; both agents have been shown to inhibit SDF-1/CXCR4 interaction and downstream signaling. In animal models, LY2510924 peptide induced robust, prolonged, dose- and time-dependent WBC and HSC increases in mice and monkeys, whereas LY2624587 antibody induced only moderate, transient increases in monkeys. In clinical trials, similar pharmacodynamic effects were observed in patients with advanced cancer: LY2510924 peptide induced sustained WBC and HSC increases, while LY2624587 antibody induced only minimal, transient WBC changes. These distinct pharmacodynamic effects in two different classes of CXCR4 inhibitors are clinically important and should be carefully considered when designing combination studies with immune checkpoint inhibitors or other agents for cancer therapy. PMID:29212254

Super Memory Bros.: going from mirror patterns to concordant patterns via similarity enhancements.

PubMed

Ozubko, Jason D; Joordens, Steve

2008-12-01

When memory is contrasted for stimuli belonging to distinct stimulus classes, one of two patterns is observed: a mirror pattern, in which one stimulus gives rise to higher hits but lower false alarms (e.g., the frequency-based mirror effect) or a concordant pattern, in which one stimulus class gives rise both to higher hits and to higher false alarms (e.g., the pseudoword effect). On the basis of the dual-process account proposed by Joordens and Hockley (2000), we predict that mirror patterns occur when one stimulus class is more familiar and less distinctive than another, whereas concordant patterns occur when one stimulus class is more familiar than another. We tested these assumptions within a video game paradigm using novel stimuli that allow manipulations in terms of distinctiveness and familiarity (via similarity). When more distinctive, less familiar items are contrasted with less distinctive, more familiar items, a mirror pattern is observed. Systematically enhancing the familiarity of stimuli transforms the mirror pattern to a concordant pattern as predicted. Although our stimuli differ considerably from those used in examinations of the frequency-based mirror effect and the pseudoword effect, the implications of our findings with respect to those phenomena are also discussed.

CD8+ TCR repertoire formation is guided primarily by the peptide component of the antigenic complex.

PubMed

Koning, Dan; Costa, Ana I; Hoof, Ilka; Miles, John J; Nanlohy, Nening M; Ladell, Kristin; Matthews, Katherine K; Venturi, Vanessa; Schellens, Ingrid M M; Borghans, Jose A M; Kesmir, Can; Price, David A; van Baarle, Debbie

2013-02-01

CD8(+) T cells recognize infected or dysregulated cells via the clonotypically expressed αβ TCR, which engages Ag in the form of peptide bound to MHC class I (MHC I) on the target cell surface. Previous studies have indicated that a diverse Ag-specific TCR repertoire can be beneficial to the host, yet the determinants of clonotypic diversity are poorly defined. To better understand the factors that govern TCR repertoire formation, we conducted a comprehensive clonotypic analysis of CD8(+) T cell populations directed against epitopes derived from EBV and CMV. Neither pathogen source nor the restricting MHC I molecule were linked with TCR diversity; indeed, both HLA-A and HLA-B molecules were observed to interact with an overlapping repertoire of expressed TRBV genes. Peptide specificity, however, markedly impacted TCR diversity. In addition, distinct peptides sharing HLA restriction and viral origin mobilized TCR repertoires with distinct patterns of TRBV gene usage. Notably, no relationship was observed between immunodominance and TCR diversity. These findings provide new insights into the forces that shape the Ag-specific TCR repertoire in vivo and highlight a determinative role for the peptide component of the peptide-MHC I complex on the molecular frontline of CD8(+) T cell-mediated immune surveillance.

Maternal anaemia at delivery and haemoglobin evolution in children during their first 18 months of life using latent class analysis.

PubMed

Koura, Kobto G; Ouédraogo, Smaïla; Cottrell, Gilles; Le Port, Agnès; Massougbodji, Achille; Garcia, André

2012-01-01

Anaemia during pregnancy and at delivery is an important public health problem in low- and middle-income countries. Its association with the children's haemoglobin level over time remains unclear. Our goals were to identify distinct haemoglobin level trajectories using latent class analysis and to assess the association between these trajectories and maternal anaemia and other risk factors. A prospective study of children from birth to 18 months of life was conducted in a rural setting in Tori-Bossito, Benin. The main outcome measure was the haemoglobin levels repeatedly measured at 3, 6, 9, 12, 15 and 18 months. Variables were collected from the mothers at delivery and from their children at birth and during the follow-up. The analyses were performed by means of Latent Class Analysis which has never been used for this kind of data. All the analyses were performed with Stata software, version 11.0, using the generalized linear latent and mixed model (GLLAMM) framework. We showed that 33.7% of children followed a low haemoglobin trajectory and 66.3% a high trajectory during the first 18 months of life. Newborn anaemia, placental malaria, malaria attack, sickle cell trait and male gender were significantly associated with a lower children's haemoglobin level over time, whereas maternal age, children living in a polygamous family and with good feeding practices had a higher Hb level in the first18 months. We also showed that maternal anaemia was a predictor for 'low haemoglobin level trajectory' group membership but have no significant effect on children haemoglobin level over time. Latent Class Analyses framework seems well suited to analyse longitudinal data under the hypothesis that different subpopulations of subjects are present in the data, each with its own set of parameters, with distinctive evolutions that themselves may reflect distinctive aetiologies.

Maternal Anaemia at Delivery and Haemoglobin Evolution in Children during Their First 18 Months of Life Using Latent Class Analysis

PubMed Central

Koura, Kobto G.; Ouédraogo, Smaïla; Cottrell, Gilles; Le Port, Agnès; Massougbodji, Achille; Garcia, André

2012-01-01

Background Anaemia during pregnancy and at delivery is an important public health problem in low- and middle-income countries. Its association with the children’s haemoglobin level over time remains unclear. Our goals were to identify distinct haemoglobin level trajectories using latent class analysis and to assess the association between these trajectories and maternal anaemia and other risk factors. Method A prospective study of children from birth to 18 months of life was conducted in a rural setting in Tori-Bossito, Benin. The main outcome measure was the haemoglobin levels repeatedly measured at 3, 6, 9, 12, 15 and 18 months. Variables were collected from the mothers at delivery and from their children at birth and during the follow-up. The analyses were performed by means of Latent Class Analysis which has never been used for this kind of data. All the analyses were performed with Stata software, version 11.0, using the generalized linear latent and mixed model (GLLAMM) framework. Results We showed that 33.7% of children followed a low haemoglobin trajectory and 66.3% a high trajectory during the first 18 months of life. Newborn anaemia, placental malaria, malaria attack, sickle cell trait and male gender were significantly associated with a lower children’s haemoglobin level over time, whereas maternal age, children living in a polygamous family and with good feeding practices had a higher Hb level in the first18 months. We also showed that maternal anaemia was a predictor for ‘low haemoglobin level trajectory’ group membership but have no significant effect on children haemoglobin level over time. Conclusion Latent Class Analyses framework seems well suited to analyse longitudinal data under the hypothesis that different subpopulations of subjects are present in the data, each with its own set of parameters, with distinctive evolutions that themselves may reflect distinctive aetiologies. PMID:23185556

Type II Natural Killer T (NKT) Cells And Their Emerging Role In Health And Disease

PubMed Central

Dhodapkar, Madhav V.; Kumar, Vipin

2016-01-01

Natural killer T (NKT) cells recognize lipid antigens presented by a class I MHC-like molecule CD1d, a member of the CD1 family. While most of the initial studies on NKT cells focused on a subset with semi-invariant T cell receptor (TCR) termed iNKT cells, majority of CD1d-restricted lipid-reactive human T cells express diverse TCRs and are termed as type II NKT cells. These cells constitute a distinct population of circulating and tissue-resident effector T cells with immune-regulatory properties. They react to a growing list of self- as well as non-self lipid ligands, and share some properties with both iNKT as well as conventional T cells. Emerging body of evidence points to their role in the regulation of immunity to pathogens/tumors and in autoimmune/metabolic disorders. Improved understanding of the biology of these cells and the ability to manipulate their function may be of therapeutic benefit in diverse disease conditions. PMID:28115591

Morphology, classification, and distribution of the projection neurons in the dorsal lateral geniculate nucleus of the rat.

PubMed

Ling, Changying; Hendrickson, Michael L; Kalil, Ronald E

2012-01-01

The morphology of confirmed projection neurons in the dorsal lateral geniculate nucleus (dLGN) of the rat was examined by filling these cells retrogradely with biotinylated dextran amine (BDA) injected into the visual cortex. BDA-labeled projection neurons varied widely in the shape and size of their cell somas, with mean cross-sectional areas ranging from 60-340 µm(2). Labeled projection neurons supported 7-55 dendrites that spanned up to 300 µm in length and formed dendritic arbors with cross-sectional areas of up to 7.0 × 10(4) µm(2). Primary dendrites emerged from cell somas in three broad patterns. In some dLGN projection neurons, primary dendrites arise from the cell soma at two poles spaced approximately 180° apart. In other projection neurons, dendrites emerge principally from one side of the cell soma, while in a third group of projection neurons primary dendrites emerge from the entire perimeter of the cell soma. Based on these three distinct patterns in the distribution of primary dendrites from cell somas, we have grouped dLGN projection neurons into three classes: bipolar cells, basket cells and radial cells, respectively. The appendages seen on dendrites also can be grouped into three classes according to differences in their structure. Short "tufted" appendages arise mainly from the distal branches of dendrites; "spine-like" appendages, fine stalks with ovoid heads, typically are seen along the middle segments of dendrites; and "grape-like" appendages, short stalks that terminate in a cluster of ovoid bulbs, appear most often along the proximal segments of secondary dendrites of neurons with medium or large cell somas. While morphologically diverse dLGN projection neurons are intermingled uniformly throughout the nucleus, the caudal pole of the dLGN contains more small projection neurons of all classes than the rostral pole.

Different CHD chromatin remodelers are required for expression of distinct gene sets and specific stages during development of Dictyostelium discoideum

PubMed Central

Platt, James L.; Rogers, Benjamin J.; Rogers, Kelley C.; Harwood, Adrian J.; Kimmel, Alan R.

2013-01-01

Control of chromatin structure is crucial for multicellular development and regulation of cell differentiation. The CHD (chromodomain-helicase-DNA binding) protein family is one of the major ATP-dependent, chromatin remodeling factors that regulate nucleosome positioning and access of transcription factors and RNA polymerase to the eukaryotic genome. There are three mammalian CHD subfamilies and their impaired functions are associated with several human diseases. Here, we identify three CHD orthologs (ChdA, ChdB and ChdC) in Dictyostelium discoideum. These CHDs are expressed throughout development, but with unique patterns. Null mutants lacking each CHD have distinct phenotypes that reflect their expression patterns and suggest functional specificity. Accordingly, using genome-wide (RNA-seq) transcriptome profiling for each null strain, we show that the different CHDs regulate distinct gene sets during both growth and development. ChdC is an apparent ortholog of the mammalian Class III CHD group that is associated with the human CHARGE syndrome, and GO analyses of aberrant gene expression in chdC nulls suggest defects in both cell-autonomous and non-autonomous signaling, which have been confirmed through analyses of chdC nulls developed in pure populations or with low levels of wild-type cells. This study provides novel insight into the broad function of CHDs in the regulation development and disease, through chromatin-mediated changes in directed gene expression. PMID:24301467

Evolution and functional diversification of fructose bisphosphate aldolase genes in photosynthetic marine diatoms.

PubMed

Allen, Andrew E; Moustafa, Ahmed; Montsant, Anton; Eckert, Angelika; Kroth, Peter G; Bowler, Chris

2012-01-01

Diatoms and other chlorophyll-c containing, or chromalveolate, algae are among the most productive and diverse phytoplankton in the ocean. Evolutionarily, chlorophyll-c algae are linked through common, although not necessarily monophyletic, acquisition of plastid endosymbionts of red as well as most likely green algal origin. There is also strong evidence for a relatively high level of lineage-specific bacterial gene acquisition within chromalveolates. Therefore, analyses of gene content and derivation in chromalveolate taxa have indicated particularly diverse origins of their overall gene repertoire. As a single group of functionally related enzymes spanning two distinct gene families, fructose 1,6-bisphosphate aldolases (FBAs) illustrate the influence on core biochemical pathways of specific evolutionary associations among diatoms and other chromalveolates with various plastid-bearing and bacterial endosymbionts. Protein localization and activity, gene expression, and phylogenetic analyses indicate that the pennate diatom Phaeodactylum tricornutum contains five FBA genes with very little overall functional overlap. Three P. tricornutum FBAs, one class I and two class II, are plastid localized, and each appears to have a distinct evolutionary origin as well as function. Class I plastid FBA appears to have been acquired by chromalveolates from a red algal endosymbiont, whereas one copy of class II plastid FBA is likely to have originated from an ancient green algal endosymbiont. The other copy appears to be the result of a chromalveolate-specific gene duplication. Plastid FBA I and chromalveolate-specific class II plastid FBA are localized in the pyrenoid region of the chloroplast where they are associated with β-carbonic anhydrase, which is known to play a significant role in regulation of the diatom carbon concentrating mechanism. The two pyrenoid-associated FBAs are distinguished by contrasting gene expression profiles under nutrient limiting compared with optimal CO2 fixation conditions, suggestive of a distinct specialized function for each. Cytosolically localized FBAs in P. tricornutum likely play a role in glycolysis and cytoskeleton function and seem to have originated from the stramenopile host cell and from diatom-specific bacterial gene transfer, respectively.

Evolution and Functional Diversification of Fructose Bisphosphate Aldolase Genes in Photosynthetic Marine Diatoms

PubMed Central

Allen, Andrew E.; Moustafa, Ahmed; Montsant, Anton; Eckert, Angelika; Kroth, Peter G.; Bowler, Chris

2012-01-01

Diatoms and other chlorophyll-c containing, or chromalveolate, algae are among the most productive and diverse phytoplankton in the ocean. Evolutionarily, chlorophyll-c algae are linked through common, although not necessarily monophyletic, acquisition of plastid endosymbionts of red as well as most likely green algal origin. There is also strong evidence for a relatively high level of lineage-specific bacterial gene acquisition within chromalveolates. Therefore, analyses of gene content and derivation in chromalveolate taxa have indicated particularly diverse origins of their overall gene repertoire. As a single group of functionally related enzymes spanning two distinct gene families, fructose 1,6-bisphosphate aldolases (FBAs) illustrate the influence on core biochemical pathways of specific evolutionary associations among diatoms and other chromalveolates with various plastid-bearing and bacterial endosymbionts. Protein localization and activity, gene expression, and phylogenetic analyses indicate that the pennate diatom Phaeodactylum tricornutum contains five FBA genes with very little overall functional overlap. Three P. tricornutum FBAs, one class I and two class II, are plastid localized, and each appears to have a distinct evolutionary origin as well as function. Class I plastid FBA appears to have been acquired by chromalveolates from a red algal endosymbiont, whereas one copy of class II plastid FBA is likely to have originated from an ancient green algal endosymbiont. The other copy appears to be the result of a chromalveolate-specific gene duplication. Plastid FBA I and chromalveolate-specific class II plastid FBA are localized in the pyrenoid region of the chloroplast where they are associated with β-carbonic anhydrase, which is known to play a significant role in regulation of the diatom carbon concentrating mechanism. The two pyrenoid-associated FBAs are distinguished by contrasting gene expression profiles under nutrient limiting compared with optimal CO2 fixation conditions, suggestive of a distinct specialized function for each. Cytosolically localized FBAs in P. tricornutum likely play a role in glycolysis and cytoskeleton function and seem to have originated from the stramenopile host cell and from diatom-specific bacterial gene transfer, respectively. PMID:21903677

Comparative analyses of CTCF and BORIS occupancies uncover two distinct classes of CTCF binding genomic regions.

PubMed

Pugacheva, Elena M; Rivero-Hinojosa, Samuel; Espinoza, Celso A; Méndez-Catalá, Claudia Fabiola; Kang, Sungyun; Suzuki, Teruhiko; Kosaka-Suzuki, Natsuki; Robinson, Susan; Nagarajan, Vijayaraj; Ye, Zhen; Boukaba, Abdelhalim; Rasko, John E J; Strunnikov, Alexander V; Loukinov, Dmitri; Ren, Bing; Lobanenkov, Victor V

2015-08-14

CTCF and BORIS (CTCFL), two paralogous mammalian proteins sharing nearly identical DNA binding domains, are thought to function in a mutually exclusive manner in DNA binding and transcriptional regulation. Here we show that these two proteins co-occupy a specific subset of regulatory elements consisting of clustered CTCF binding motifs (termed 2xCTSes). BORIS occupancy at 2xCTSes is largely invariant in BORIS-positive cancer cells, with the genomic pattern recapitulating the germline-specific BORIS binding to chromatin. In contrast to the single-motif CTCF target sites (1xCTSes), the 2xCTS elements are preferentially found at active promoters and enhancers, both in cancer and germ cells. 2xCTSes are also enriched in genomic regions that escape histone to protamine replacement in human and mouse sperm. Depletion of the BORIS gene leads to altered transcription of a large number of genes and the differentiation of K562 cells, while the ectopic expression of this CTCF paralog leads to specific changes in transcription in MCF7 cells. We discover two functionally and structurally different classes of CTCF binding regions, 2xCTSes and 1xCTSes, revealed by their predisposition to bind BORIS. We propose that 2xCTSes play key roles in the transcriptional program of cancer and germ cells.

Visual projection neurons in the Drosophila lobula link feature detection to distinct behavioral programs

PubMed Central

Wu, Ming; Nern, Aljoscha; Williamson, W Ryan; Morimoto, Mai M; Reiser, Michael B; Card, Gwyneth M; Rubin, Gerald M

2016-01-01

Visual projection neurons (VPNs) provide an anatomical connection between early visual processing and higher brain regions. Here we characterize lobula columnar (LC) cells, a class of Drosophila VPNs that project to distinct central brain structures called optic glomeruli. We anatomically describe 22 different LC types and show that, for several types, optogenetic activation in freely moving flies evokes specific behaviors. The activation phenotypes of two LC types closely resemble natural avoidance behaviors triggered by a visual loom. In vivo two-photon calcium imaging reveals that these LC types respond to looming stimuli, while another type does not, but instead responds to the motion of a small object. Activation of LC neurons on only one side of the brain can result in attractive or aversive turning behaviors depending on the cell type. Our results indicate that LC neurons convey information on the presence and location of visual features relevant for specific behaviors. DOI: http://dx.doi.org/10.7554/eLife.21022.001 PMID:28029094

Single Sublattice Endotaxial Phase Separation Driven by Charge Frustration in a Complex Oxide

PubMed Central

2013-01-01

Complex transition-metal oxides are important functional materials in areas such as energy and information storage. The cubic ABO3 perovskite is an archetypal example of this class, formed by the occupation of small octahedral B-sites within an AO3 network defined by larger A cations. We show that introduction of chemically mismatched octahedral cations into a cubic perovskite oxide parent phase modifies structure and composition beyond the unit cell length scale on the B sublattice alone. This affords an endotaxial nanocomposite of two cubic perovskite phases with distinct properties. These locally B-site cation-ordered and -disordered phases share a single AO3 network and have enhanced stability against the formation of a competing hexagonal structure over the single-phase parent. Synergic integration of the distinct properties of these phases by the coherent interfaces of the composite produces solid oxide fuel cell cathode performance superior to that expected from the component phases in isolation. PMID:23750709

Immunostimulatory Properties of Lipid Modified CpG Oligonucleotides.

PubMed

Yu, Chunsong; An, Myunggi; Li, Meng; Liu, Haipeng

2017-08-07

Innate immune responses recognizing pathogen associated molecular patterns play important roles in adaptive immunity. As such, ligands which mimic the conserved products of microbial and activate innate immunity are widely used as adjuvants for vaccines. Synthetic single strand oligodeoxynucleotides (ODNs) containing unmethylated cytosine-guanine (CpG) motifs which bind Toll-like receptor 9 (TLR9) are powerful molecular adjuvants, potentiating both humoral and cellular responses. However, CpG ODN's in vitro potency has not been translated to in vivo settings primarily due to issues associated with delivery and toxicity. A major challenge in clinical application of CpG ODN is the efficient delivery to lymph nodes, the anatomic sites where all the immune responses are initiated. Targeting CpG to the key antigen presenting cells (APC) is essential for its application as a vaccine adjuvant, as it not only enhances CpG's efficacy, but also greatly reduces the systemic toxicity. We recently discovered an "albumin-hitchhiking" approach by which CpG ODNs were conjugated to a lipophilic lipid tail and follow subcutaneous injection, accumulated in lymph nodes by binding and transporting with endogenous albumin. This molecular approach targets CpG to antigen presenting cells in the draining lymph nodes via an endogenous albumin-mediated mechanism and simultaneously improves both the efficacy and safety of CpG as a vaccine adjuvant. Since CpG ODNs can be divided into structurally distinct classes, and each class of CpG ODN activates different types of immune cells and triggers different types of immunostimulatory activities, it is important to thoroughly evaluate the efficacy of this "albumin-hitchhiking" strategy in each class of CpG. Here we compare the immunostimulatory activities of three classes of lipid conjugated CpG ODNs in vitro and in vivo. Three representative sequences of lipid modified CpG ODNs were synthesized and their stimulatory effects as a vaccine adjuvant were evaluated. Our results showed that in vitro, lipid modified class A CpG exhibited enhanced stimulatory activities toward TLR transfected reporter cells or bone-marrow derived dendritic cells, whereas lipid-modification of class B or C CpG reduces the activation of TLR9 by 2-3 fold, as compared with unmodified class B and class C CpG, respectively. However, in vivo coadministration of ovalbumin (OVA) protein antigen mixed with lipid-conjugated class B or C CpG ODNs, but not class A CpGs induced dramatically increased OVA-specific humoral and cytotoxic CD8 + T cells responses compared with OVA mixed with unmodified CpGs. Further, lipid-modification greatly reduces the toxicity associated with CpG by minimizing the systemic dissemination. Taken together, these results demonstrated that amphiphilic modification of three classes of CpG motifs differentially affected and modulated the immunostimulatory activities in vitro and in vivo. Our study highlights the importance of in vivo lymph node targeting of CpG ODNs in fulfilling their use as vaccine adjuvants, providing implications for the rational design of molecular adjuvant for subunit vaccines.

The taccalonolides and paclitaxel cause distinct effects on microtubule dynamics and aster formation

PubMed Central

2014-01-01

Background Microtubule stabilizers suppress microtubule dynamics and, at the lowest antiproliferative concentrations, disrupt the function of mitotic spindles, leading to mitotic arrest and apoptosis. At slightly higher concentrations, these agents cause the formation of multiple mitotic asters with distinct morphologies elicited by different microtubule stabilizers. Results We tested the hypothesis that two classes of microtubule stabilizing drugs, the taxanes and the taccalonolides, cause the formation of distinct aster structures due, in part, to differential effects on microtubule dynamics. Paclitaxel and the taccalonolides suppressed the dynamics of microtubules formed from purified tubulin as well as in live cells. Both agents suppressed microtubule dynamic instability, with the taccalonolides having a more pronounced inhibition of microtubule catastrophe, suggesting that they stabilize the plus ends of microtubules more effectively than paclitaxel. Live cell microscopy was also used to evaluate the formation and resolution of asters after drug treatment. While each drug had similar effects on initial formation, substantial differences were observed in aster resolution. Paclitaxel-induced asters often coalesced over time resulting in fewer, larger asters whereas numerous compact asters persisted once they were formed in the presence of the taccalonolides. Conclusions We conclude that the increased resistance of microtubule plus ends to catastrophe may play a role in the observed inability of taccalonolide-induced asters to coalesce during mitosis, giving rise to the distinct morphologies observed after exposure to these agents. PMID:24576146

Exposure of a distinct PDCA-1+ (CD317) B cell population to agonistic anti-4-1BB (CD137) inhibits T and B cell responses both in vitro and in vivo.

PubMed

Vinay, Dass S; Lee, Seung J; Kim, Chang H; Oh, Ho Sik; Kwon, Byoung S

2012-01-01

4-1BB (CD137) is an important T cell activating molecule. Here we report that it also promotes development of a distinct B cell subpopulation co-expressing PDCA-1. 4-1BB is expressed constitutively, and its expression is increased when PDCA-1(+) B cells are activated. We found that despite a high level of surface expression of 4-1BB on PDCA-1(+) B cells, treatment of these cells with agonistic anti-4-1BB mAb stimulated the expression of only a few activation markers (B7-2, MHC II, PD-L2), cytokines (IL-12p40/p70), and chemokines (MCP-1, RANTES), as well as sTNFR1, and the immunosuppressive enzyme, IDO. Although the PDCA-1(+) B cells stimulated by anti-4-1BB expressed MHC II at high levels and took up antigens efficiently, Ig class switching was inhibited when they were pulsed with T-independent (TI) or T-dependent (TD) Ags and adoptively transferred into syngeneic recipients. Furthermore, when anti-4-1BB-treated PDCA-1(+) B cells were pulsed with OVA peptide and combined with Vα2(+)CD4(+) T cells, Ag-specific cell division was inhibited both in vitro and in vivo. Our findings suggest that the 4-1BB signal transforms PDCA-1(+) B cells into propagators of negative immune regulation, and establish an important role for 4-1BB in PDCA-1(+) B cell development and function.

CCR5 adopts three homodimeric conformations that control cell surface delivery.

PubMed

Jin, Jun; Momboisse, Fanny; Boncompain, Gaelle; Koensgen, Florian; Zhou, Zhicheng; Cordeiro, Nelia; Arenzana-Seisdedos, Fernando; Perez, Franck; Lagane, Bernard; Kellenberger, Esther; Brelot, Anne

2018-05-08

Biophysical methods and x-ray crystallography have revealed that class A G protein-coupled receptors (GPCRs) can form homodimers. We combined computational approaches with receptor cross-linking, energy transfer, and a newly developed functional export assay to characterize the residues involved in the dimerization interfaces of the chemokine receptor CCR5, the major co-receptor for HIV-1 entry into cells. We provide evidence of three distinct CCR5 dimeric organizations, involving residues of transmembrane helix 5. Two dimeric states corresponded to unliganded receptors, whereas the binding of the inverse agonist maraviroc stabilized a third state. We found that CCR5 dimerization was required for targeting the receptor to the plasma membrane. These data suggest that dimerization contributes to the conformational diversity of inactive class A GPCRs and may provide new opportunities to investigate the cellular entry of HIV-1 and mechanisms for its inhibition. Copyright © 2018 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

Signaling networks in joint development

PubMed Central

Salva, Joanna E.; Merrill, Amy E.

2016-01-01

Here we review studies identifying regulatory networks responsible for synovial, cartilaginous, and fibrous joint development. Synovial joints, characterized by the fluid-filled synovial space between the bones, are found in high-mobility regions and are the most common type of joint. Cartilaginous joints unite adjacent bones through either a hyaline cartilage or fibrocartilage intermediate. Fibrous joints, which include the cranial sutures, form a direct union between bones through fibrous connective tissue. We describe how the distinct morphologic and histogenic characteristics of these joint classes are established during embryonic development. Collectively, these studies reveal that despite the heterogeneity of joint strength and mobility, joint development throughout the skeleton utilizes common signaling networks via long-range morphogen gradients and direct cell-cell contact. This suggests that different joint types represent specialized variants of homologous developmental modules. Identifying the unifying aspects of the signaling networks between joint classes allows a more complete understanding of the signaling code for joint formation, which is critical to improving strategies for joint regeneration and repair. PMID:27859991

Effect of Alzheimer's disease risk and protective factors on cognitive trajectories in subjective memory complainers.

PubMed

Teipel, Stefan J; Cavedo, Enrica; Lista, Simone; Habert, Marie-Odile; Potier, Marie-Claude; Grothe, Michel J; Epelbaum, Stephane; Sambati, Luisa; Gagliardi, Geoffroy; Toschi, Nicola; Greicius, Michael; Dubois, Bruno; Hampel, Harald

2018-05-21

Cognitive change in people at risk of Alzheimer's disease (AD) such as subjective memory complainers is highly variable across individuals. We used latent class growth modeling to identify distinct classes of nonlinear trajectories of cognitive change over 2 years follow-up from 265 subjective memory complainers individuals (age 70 years and older) of the INSIGHT-preAD cohort. We determined the effect of cortical amyloid load, hippocampus and basal forebrain volumes, and education on the cognitive trajectory classes. Latent class growth modeling identified distinct nonlinear cognitive trajectories. Education was associated with higher performing trajectories, whereas global amyloid load and basal forebrain atrophy were associated with lower performing trajectories. Distinct classes of cognitive trajectories were associated with risk and protective factors of AD. These associations support the notion that the identified cognitive trajectories reflect different risk for AD that may be useful for selecting high-risk individuals for intervention trials. Copyright © 2018. Published by Elsevier Inc.

Natural HLA-B*2705 protein ligands with glutamine as anchor motif: implications for HLA-B27 association with spondyloarthropathy.

PubMed

Infantes, Susana; Lorente, Elena; Barnea, Eilon; Beer, Ilan; Barriga, Alejandro; Lasala, Fátima; Jiménez, Mercedes; Admon, Arie; López, Daniel

2013-04-12

The presentation of short viral peptide antigens by human leukocyte antigen (HLA) class I molecules on cell surfaces is a key step in the activation of cytotoxic T lymphocytes, which mediate the killing of pathogen-infected cells or initiate autoimmune tissue damage. HLA-B27 is a well known class I molecule that is used to study both facets of the cellular immune response. Using mass spectrometry analysis of complex HLA-bound peptide pools isolated from large amounts of HLA-B*2705(+) cells, we identified 200 naturally processed HLA-B*2705 ligands. Our analyses revealed that a change in the position (P) 2 anchor motif was detected in the 3% of HLA-B*2705 ligands identified. B*2705 class I molecules were able to bind these six GlnP2 peptides, which showed significant homology to pathogenic bacterial sequences, with a broad range of affinities. One of these ligands was able to bind with distinct conformations to HLA-B27 subtypes differentially associated with ankylosing spondylitis. These conformational differences could be sufficient to initiate autoimmune damage in patients with ankylosing spondylitis-associated subtypes. Therefore, these kinds of peptides (short, with GlnP2, and similar low affinity to all HLA-B27 subtypes tested but with unlike conformations in differentially ankylosing spondylitis-associated subtypes) must not be excluded from future researches involving potential arthritogenic peptides.

Natural HLA-B*2705 Protein Ligands with Glutamine as Anchor Motif

PubMed Central

Infantes, Susana; Lorente, Elena; Barnea, Eilon; Beer, Ilan; Barriga, Alejandro; Lasala, Fátima; Jiménez, Mercedes; Admon, Arie; López, Daniel

2013-01-01

The presentation of short viral peptide antigens by human leukocyte antigen (HLA) class I molecules on cell surfaces is a key step in the activation of cytotoxic T lymphocytes, which mediate the killing of pathogen-infected cells or initiate autoimmune tissue damage. HLA-B27 is a well known class I molecule that is used to study both facets of the cellular immune response. Using mass spectrometry analysis of complex HLA-bound peptide pools isolated from large amounts of HLA-B*2705+ cells, we identified 200 naturally processed HLA-B*2705 ligands. Our analyses revealed that a change in the position (P) 2 anchor motif was detected in the 3% of HLA-B*2705 ligands identified. B*2705 class I molecules were able to bind these six GlnP2 peptides, which showed significant homology to pathogenic bacterial sequences, with a broad range of affinities. One of these ligands was able to bind with distinct conformations to HLA-B27 subtypes differentially associated with ankylosing spondylitis. These conformational differences could be sufficient to initiate autoimmune damage in patients with ankylosing spondylitis-associated subtypes. Therefore, these kinds of peptides (short, with GlnP2, and similar low affinity to all HLA-B27 subtypes tested but with unlike conformations in differentially ankylosing spondylitis-associated subtypes) must not be excluded from future researches involving potential arthritogenic peptides. PMID:23430249

Screen of FDA-approved drug library reveals compounds that protect hair cells from aminoglycosides and cisplatin

PubMed Central

Vlasits, Anna L.; Simon, Julian A.; Raible, David W.; Rubel, Edwin W; Owens, Kelly N.

2012-01-01

Loss of mechanosensory hair cells in the inner ear accounts for many hearing loss and balance disorders. Several beneficial pharmaceutical drugs cause hair cell death as a side effect. These include aminoglycoside antibiotics, such as neomycin, kanamycin and gentamicin, and several cancer chemotherapy drugs, such as cisplatin. Discovering new compounds that protect mammalian hair cells from toxic insults is experimentally difficult because of the inaccessibility of the inner ear. We used the zebrafish lateral line sensory system as an in vivo screening platform to survey a library of FDA-approved pharmaceuticals for compounds that protect hair cells from neomycin, gentamicin, kanamycin and cisplatin. Ten compounds were identified that provide protection from at least two of the four toxins. The resulting compounds fall into several drug classes, including serotonin and dopamine-modulating drugs, adrenergic receptor ligands, and estrogen receptor modulators. The protective compounds show different effects against the different toxins, supporting the idea that each toxin causes hair cell death by distinct, but partially overlapping, mechanisms. Furthermore, some compounds from the same drug classes had different protective properties, suggesting that they might not prevent hair cell death by their known target mechanisms. Some protective compounds blocked gentamicin uptake into hair cells, suggesting that they may block mechanotransduction or other routes of entry. The protective compounds identified in our screen will provide a starting point for studies in mammals as well as further research discovering the cellular signaling pathways that trigger hair cell death. PMID:22967486

Investigating heterogeneity in violent offending liability among injection drug users from a developmental perspective.

PubMed

Torok, Michelle; Darke, Shane; Shand, Fiona; Kaye, Sharlene

2016-09-01

Violence is a major burden of harm among injecting drug users (IDU), however, the liability to violent offending is not well understood. The current study aimed to better understand differences in the liability to violence by determining whether IDU could be disaggregated into distinct violent offending classes, and determining the correlates of class membership. A total of 300 IDU from Sydney, Australia were administered a structured interview examining the prevalence and severity of drug use and violent offending histories, as well as early life risk factors (maltreatment, childhood mental disorder, trait personality). IDU were disaggregated into four distinct latent classes, comprising a non-violent class (24%), an adolescent-onset persistent class (33%), an adult-onset transient class (24%) and an early-onset, chronic class (19%). Pairwise and group comparisons of classes on predispositional and substance use risks showed that the EARLY class had the poorest psychosocial risk profile, while the NON class had the most favourable. Multinomial logistic regression revealed that higher trait impulsivity and aggression scores, having a history of conduct disorder, frequent childhood abuse, and more problematic alcohol use, were independently associated with more temporally stable and severe violent offending. The model explained 67% of variance in class membership (χ(2)=207.7, df=51, p<0.001). IDU can be meaningfully disaggregated into distinct violent offending classes using developmental criteria. The age of onset of violence was indicative of class membership insomuch as that the extent of early life risk exposure was differentially associated with greater long-term liability to violence and drug use. Copyright © 2016 Elsevier Ltd. All rights reserved.

The cells of cajal-retzius: still a mystery one century after.

PubMed

Soriano, Eduardo; Del Río, José Antonio

2005-05-05

Cajal-Retzius (CR) cells are an enigmatic class of neurons located at the surface of the cerebral cortex, playing a major role in cortical development. In this review, we discuss several distinct features of these neurons and the mechanisms by which they regulate cortical development. Many CR cells likely have extracortical origin and undergo cell death during development. Recent genetic studies report unique patterns of gene expression in CR cells, which may help to explain the developmental processes in which they participate. Moreover, a number of studies indicate that CR cells, and their secreted gene product, reelin, are involved in neuronal migration by acting on two key partners, migrating neurons and radial glial cells. Emerging data show that these neurons are a critical part of an early and complex network of neural activity in layer I, supporting the notion that CR cells modulate cortical maturation. Given these key and complex developmental properties, it is therefore conceivable for CR cells to be implicated in the pathogenesis of a variety of neurological disorders.

Identification and characterization of polyclonal αβ T cells with dendritic cell properties

PubMed Central

Kuka, Mirela; Munitic, Ivana; Ashwell, Jonathan D.

2012-01-01

An efficient immune response requires coordination between innate and adaptive immunity, which act through cells different in origin and function. Here we report the identification of thymus-derived αβ TCR+ cells that express CD11c and MHC class II, and require FLT3L for development (TDC). TDC express genes heretofore found uniquely in T cells or DC, as well as a distinctive signature of cytotoxicity-related genes. Unlike other innate T cell subsets, TDC have a polyclonal TCR repertoire andrespond to cognate antigens. However, they differ from conventional T cells in that they do not require help from antigen-presenting cells, respond to TLR-mediated stimulation by producing IL-12 and process and present antigen. The physiologic relevance of TDC, found in mice and humans, is still under investigation, but the fact that they combine key features of T and DC cells suggests that they provide a bridge between the innate and adaptive immune systems. PMID:23187623

New small-molecule inhibitor class targeting human immunodeficiency virus type 1 virion maturation.

PubMed

Blair, Wade S; Cao, Joan; Fok-Seang, Juin; Griffin, Paul; Isaacson, Jason; Jackson, R Lynn; Murray, Edward; Patick, Amy K; Peng, Qinghai; Perros, Manos; Pickford, Chris; Wu, Hua; Butler, Scott L

2009-12-01

A new small-molecule inhibitor class that targets virion maturation was identified from a human immunodeficiency virus type 1 (HIV-1) antiviral screen. PF-46396, a representative molecule, exhibits antiviral activity against HIV-1 laboratory strains and clinical isolates in T-cell lines and peripheral blood mononuclear cells (PBMCs). PF-46396 specifically inhibits the processing of capsid (CA)/spacer peptide 1 (SP1) (p25), resulting in the accumulation of CA/SP1 (p25) precursor proteins and blocked maturation of the viral core particle. Viral variants resistant to PF-46396 contain a single amino acid substitution in HIV-1 CA sequences (CAI201V), distal to the CA/SP1 cleavage site in the primary structure, which we demonstrate is sufficient to confer significant resistance to PF-46396 and 3-O-(3',3'-dimethylsuccinyl) betulinic acid (DSB), a previously described maturation inhibitor. Conversely, a single amino substitution in SP1 (SP1A1V), which was previously associated with DSB in vitro resistance, was sufficient to confer resistance to DSB and PF-46396. Further, the CAI201V substitution restored CA/SP1 processing in HIV-1-infected cells treated with PF-46396 or DSB. Our results demonstrate that PF-46396 acts through a mechanism that is similar to DSB to inhibit the maturation of HIV-1 virions. To our knowledge, PF-46396 represents the first small-molecule HIV-1 maturation inhibitor that is distinct in chemical class from betulinic acid-derived maturation inhibitors (e.g., DSB), demonstrating that molecules of diverse chemical classes can inhibit this mechanism.

Reverse crosstalk of TGFβ and PPARβ/δ signaling identified by transcriptional profiling

PubMed Central

Stockert, Josefine; Adhikary, Till; Kaddatz, Kerstin; Finkernagel, Florian; Meissner, Wolfgang; Müller-Brüsselbach, Sabine; Müller, Rolf

2011-01-01

Previous work has provided strong evidence for a role of peroxisome proliferator-activated receptor β/δ (PPARβ/δ) and transforming growth factor-β (TGFβ) in inflammation and tumor stroma function, raising the possibility that both signaling pathways are interconnected. We have addressed this hypothesis by microarray analyses of human diploid fibroblasts induced to myofibroblastic differentiation, which revealed a substantial, mostly reverse crosstalk of both pathways and identified distinct classes of genes. A major class encompasses classical PPAR target genes, including ANGPTL4, CPT1A, ADRP and PDK4. These genes are repressed by TGFβ, which is counteracted by PPARβ/δ activation. This is mediated, at least in part, by the TGFβ-induced recruitment of the corepressor SMRT to PPAR response elements, and its release by PPARβ/δ ligands, indicating that TGFβ and PPARβ/δ signals are integrated by chromatin-associated complexes. A second class represents TGFβ-induced genes that are downregulated by PPARβ/δ agonists, exemplified by CD274 and IL6, which is consistent with the anti-inflammatory properties of PPARβ/δ ligands. Finally, cooperative regulation by both ligands was observed for a minor group of genes, including several regulators of cell proliferation. These observations indicate that PPARβ/δ is able to influence the expression of distinct sets of both TGFβ-repressed and TGFβ-activated genes in both directions. PMID:20846954

Dissecting hematopoietic and renal cell heterogeneity in adult zebrafish at single-cell resolution using RNA sequencing.

PubMed

Tang, Qin; Iyer, Sowmya; Lobbardi, Riadh; Moore, John C; Chen, Huidong; Lareau, Caleb; Hebert, Christine; Shaw, McKenzie L; Neftel, Cyril; Suva, Mario L; Ceol, Craig J; Bernards, Andre; Aryee, Martin; Pinello, Luca; Drummond, Iain A; Langenau, David M

2017-10-02

Recent advances in single-cell, transcriptomic profiling have provided unprecedented access to investigate cell heterogeneity during tissue and organ development. In this study, we used massively parallel, single-cell RNA sequencing to define cell heterogeneity within the zebrafish kidney marrow, constructing a comprehensive molecular atlas of definitive hematopoiesis and functionally distinct renal cells found in adult zebrafish. Because our method analyzed blood and kidney cells in an unbiased manner, our approach was useful in characterizing immune-cell deficiencies within DNA-protein kinase catalytic subunit ( prkdc ), interleukin-2 receptor γ a ( il2rga ), and double-homozygous-mutant fish, identifying blood cell losses in T, B, and natural killer cells within specific genetic mutants. Our analysis also uncovered novel cell types, including two classes of natural killer immune cells, classically defined and erythroid-primed hematopoietic stem and progenitor cells, mucin-secreting kidney cells, and kidney stem/progenitor cells. In total, our work provides the first, comprehensive, single-cell, transcriptomic analysis of kidney and marrow cells in the adult zebrafish. © 2017 Tang et al.

Dissecting hematopoietic and renal cell heterogeneity in adult zebrafish at single-cell resolution using RNA sequencing

PubMed Central

Iyer, Sowmya; Lobbardi, Riadh; Chen, Huidong; Hebert, Christine; Shaw, McKenzie L.; Neftel, Cyril; Suva, Mario L.; Bernards, Andre; Aryee, Martin; Drummond, Iain A.

2017-01-01

Recent advances in single-cell, transcriptomic profiling have provided unprecedented access to investigate cell heterogeneity during tissue and organ development. In this study, we used massively parallel, single-cell RNA sequencing to define cell heterogeneity within the zebrafish kidney marrow, constructing a comprehensive molecular atlas of definitive hematopoiesis and functionally distinct renal cells found in adult zebrafish. Because our method analyzed blood and kidney cells in an unbiased manner, our approach was useful in characterizing immune-cell deficiencies within DNA–protein kinase catalytic subunit (prkdc), interleukin-2 receptor γ a (il2rga), and double-homozygous–mutant fish, identifying blood cell losses in T, B, and natural killer cells within specific genetic mutants. Our analysis also uncovered novel cell types, including two classes of natural killer immune cells, classically defined and erythroid-primed hematopoietic stem and progenitor cells, mucin-secreting kidney cells, and kidney stem/progenitor cells. In total, our work provides the first, comprehensive, single-cell, transcriptomic analysis of kidney and marrow cells in the adult zebrafish. PMID:28878000

Comprehensive benchmarking reveals H2BK20 acetylation as a distinctive signature of cell-state-specific enhancers and promoters.

PubMed

Kumar, Vibhor; Rayan, Nirmala Arul; Muratani, Masafumi; Lim, Stefan; Elanggovan, Bavani; Xin, Lixia; Lu, Tess; Makhija, Harshyaa; Poschmann, Jeremie; Lufkin, Thomas; Ng, Huck Hui; Prabhakar, Shyam

2016-05-01

Although over 35 different histone acetylation marks have been described, the overwhelming majority of regulatory genomics studies focus exclusively on H3K27ac and H3K9ac. In order to identify novel epigenomic traits of regulatory elements, we constructed a benchmark set of validated enhancers by performing 140 enhancer assays in human T cells. We tested 40 chromatin signatures on this unbiased enhancer set and identified H2BK20ac, a little-studied histone modification, as the most predictive mark of active enhancers. Notably, we detected a novel class of functionally distinct enhancers enriched in H2BK20ac but lacking H3K27ac, which was present in all examined cell lines and also in embryonic forebrain tissue. H2BK20ac was also unique in highlighting cell-type-specific promoters. In contrast, other acetylation marks were present in all active promoters, regardless of cell-type specificity. In stimulated microglial cells, H2BK20ac was more correlated with cell-state-specific expression changes than H3K27ac, with TGF-beta signaling decoupling the two acetylation marks at a subset of regulatory elements. In summary, our study reveals a previously unknown connection between histone acetylation and cell-type-specific gene regulation and indicates that H2BK20ac profiling can be used to uncover new dimensions of gene regulation. © 2016 Kumar et al.; Published by Cold Spring Harbor Laboratory Press.

Comprehensive benchmarking reveals H2BK20 acetylation as a distinctive signature of cell-state-specific enhancers and promoters

PubMed Central

Kumar, Vibhor; Rayan, Nirmala Arul; Muratani, Masafumi; Lim, Stefan; Elanggovan, Bavani; Xin, Lixia; Lu, Tess; Makhija, Harshyaa; Poschmann, Jeremie; Lufkin, Thomas; Ng, Huck Hui; Prabhakar, Shyam

2016-01-01

Although over 35 different histone acetylation marks have been described, the overwhelming majority of regulatory genomics studies focus exclusively on H3K27ac and H3K9ac. In order to identify novel epigenomic traits of regulatory elements, we constructed a benchmark set of validated enhancers by performing 140 enhancer assays in human T cells. We tested 40 chromatin signatures on this unbiased enhancer set and identified H2BK20ac, a little-studied histone modification, as the most predictive mark of active enhancers. Notably, we detected a novel class of functionally distinct enhancers enriched in H2BK20ac but lacking H3K27ac, which was present in all examined cell lines and also in embryonic forebrain tissue. H2BK20ac was also unique in highlighting cell-type-specific promoters. In contrast, other acetylation marks were present in all active promoters, regardless of cell-type specificity. In stimulated microglial cells, H2BK20ac was more correlated with cell-state-specific expression changes than H3K27ac, with TGF-beta signaling decoupling the two acetylation marks at a subset of regulatory elements. In summary, our study reveals a previously unknown connection between histone acetylation and cell-type-specific gene regulation and indicates that H2BK20ac profiling can be used to uncover new dimensions of gene regulation. PMID:26957309

A Novel Class of Somatic Small RNAs Similar to Germ Cell Pachytene PIWI-interacting Small RNAs*

PubMed Central

Ortogero, Nicole; Schuster, Andrew S.; Oliver, Daniel K.; Riordan, Connor R.; Hong, Annie S.; Hennig, Grant W.; Luong, Dickson; Bao, Jianqiang; Bhetwal, Bhupal P.; Ro, Seungil; McCarrey, John R.; Yan, Wei

2014-01-01

PIWI-interacting RNAs (piRNAs) are small noncoding RNAs that bind PIWI family proteins exclusively expressed in the germ cells of mammalian gonads. MIWI2-associated piRNAs are essential for silencing transposons during primordial germ cell development, and MIWI-bound piRNAs are required for normal spermatogenesis during adulthood in mice. Although piRNAs have long been regarded as germ cell-specific, increasing lines of evidence suggest that somatic cells also express piRNA-like RNAs (pilRNAs). Here, we report the detection of abundant pilRNAs in somatic cells, which are similar to MIWI-associated piRNAs mainly expressed in pachytene spermatocytes and round spermatids in the testis. Based on small RNA deep sequencing and quantitative PCR analyses, pilRNA expression is dynamic and displays tissue specificity. Although pilRNAs are similar to pachytene piRNAs in both size and genomic origins, they have a distinct ping-pong signature. Furthermore, pilRNA biogenesis appears to utilize a yet to be identified pathway, which is different from all currently known small RNA biogenetic pathways. In addition, pilRNAs appear to preferentially target the 3′-UTRs of mRNAs in a partially complementary manner. Our data suggest that pilRNAs, as an integral component of the small RNA transcriptome in somatic cell lineages, represent a distinct population of small RNAs that may have functions similar to germ cell piRNAs. PMID:25320077

The emergence of a new chlorophytan system, and Dr. Kornmann's contribution thereto

NASA Astrophysics Data System (ADS)

van den Hoek, C.; Stam, W. T.; Olsen, J. L.

1988-09-01

In traditional chlorophytan systems the organizational level was the primary character for the distinction of main groups (classes and orders). For instance, in Fott (1971), the flagellate level corresponds with the Volvocales, the coccoid level with the Chlorococcales, the filamentous level with the Ulotrichales, the siphonocladous level with the Siphonocladales, and the siphonous level with the Bryopsidales. The new system presented here is an elaboration and emendation of recently proposed taxonomies and their underlying phylogenetic hypotheses, and it is mainly based on ultrastructural features which have become available over the last 15 years. The following criteria are used for the distinction of classes and orders: (1) architecture of the flagellate cell (flagellate cells are considered as the depositories of primitive characters); (2) type of mitosis-cytokinesis; (3) place of meiosis in the life history and, consequently, the sexual life history type; (4) organizational level and thallus architecture; (5) habitat type (marine versus feshwater and terrestrial); (6) chloroplast type. The following classes are presented: Prasinophyceae, Chlamydophyceae, Ulvophyceae (orders Codiolales, Ulvales, Cladophorales, Bryopsidales, Dasycladales), Pleurastrophyceae (?), Chlorophyceae s.s. (orders Cylindrocapsales, Oedogoniales, Chaetophorales), Zygnematophyceae, Trentepohliophyceae, Charophyceae (orders Klebsormidiales, Coleochaetales, Charales). The new system no longer reflects the traditional hypothesis of a stepwise evolutionary progression of organizational levels in which the flagellate level represents the most primitive lineage, the coccoid and sarcinoid levels lineages of intermediate derivation, and the filamentous, siphonocladous and siphonous levels the most derived lineages. Instead, it is now hypothesized that these levels have arisen over and over again in different chlorophytan lineages which are primarily characterized by their type of flagellate cell. The flagellate green algal classes Prasinophyceae (with organic body scales) and Chlamydophyceae probably represent bundles of highly conservative lineages that diverged very long ago. Consequently, extant genera and species in these classes can be expected to have emerged long ago. Fossil evidence points to a minimum age of 600 Ma of certain extant Prasinophycean genera, and molecular evidence to a minimum age of 400 500 Ma of a few Chlamydomonas species. On the contrary, the most derived “green algal” lineage, the Angiosperms, can be expected to consist of, on average, much younger genera and species. Fossil evidence points to a minimum age of genera of 5 60 Ma. Lineages of intermediate evolutionary derivation (Ulvophyceae, Chlorophyceae, Charophyceae) can be expected to encompass genera and species of intermediate age. Fossil and (limited) molecular evidence point to a minimum age of 230 70 Ma of extant genera in Bryopsidales, Dasycladales and Cladophorales (Ulvophyceae) and of 250 80 Ma of extant genera in Charales (Charophyceae).

Classification of human natural killer cells based on migration behavior and cytotoxic response.

PubMed

Vanherberghen, Bruno; Olofsson, Per E; Forslund, Elin; Sternberg-Simon, Michal; Khorshidi, Mohammad Ali; Pacouret, Simon; Guldevall, Karolin; Enqvist, Monika; Malmberg, Karl-Johan; Mehr, Ramit; Önfelt, Björn

2013-02-21

Despite intense scrutiny of the molecular interactions between natural killer (NK) and target cells, few studies have been devoted to dissection of the basic functional heterogeneity in individual NK cell behavior. Using a microchip-based, time-lapse imaging approach allowing the entire contact history of each NK cell to be recorded, in the present study, we were able to quantify how the cytotoxic response varied between individual NK cells. Strikingly, approximately half of the NK cells did not kill any target cells at all, whereas a minority of NK cells was responsible for a majority of the target cell deaths. These dynamic cytotoxicity data allowed categorization of NK cells into 5 distinct classes. A small but particularly active subclass of NK cells killed several target cells in a consecutive fashion. These "serial killers" delivered their lytic hits faster and induced faster target cell death than other NK cells. Fast, necrotic target cell death was correlated with the amount of perforin released by the NK cells. Our data are consistent with a model in which a small fraction of NK cells drives tumor elimination and inflammation.

Exploiting genomics and natural genetic variation to decode macrophage enhancers

PubMed Central

Romanoski, Casey E.; Link, Verena M.; Heinz, Sven; Glass, Christopher K.

2015-01-01

The mammalian genome contains on the order of a million enhancer-like regions that are required to establish the identities and functions of specific cell types. Here, we review recent studies in immune cells that have provided insight into the mechanisms that selectively activate certain enhancers in response to cell lineage and environmental signals. We describe a working model wherein distinct classes of transcription factors define the repertoire of active enhancers in macrophages through collaborative and hierarchical interactions, and discuss important challenges to this model, specifically providing examples from T cells. We conclude by discussing the use of natural genetic variation as a powerful approach for decoding transcription factor combinations that play dominant roles in establishing the enhancer landscapes, and the potential that these insights have for advancing our understanding of the molecular causes of human disease. PMID:26298065

Perithecium morphogenesis in Sordaria macrospora.

PubMed

Lord, Kathryn M; Read, Nick D

2011-04-01

The perithecium of the self-fertile ascomycete Sordaria macrospora provides an excellent model in which to analyse fungal multicellular development. This study provides a detailed analysis of perithecium morphogenesis in the wild type and eight developmental mutants of S. macrospora, using a range of correlative microscopical techniques. Fundamentally, perithecia and other complex multicellular structures produced by fungi arise by hyphal aggregation and adhesion, and these processes are followed by specialization and septation of hyphal compartments within the aggregates. Perithecial morphogenesis can be divided into the ascogonial, protoperithecial, and perithecial stages of development. At least 13 specialized, morphologically distinct cell-types are involved in perithecium morphogenesis, and these fall into three basic classes: hyphae, conglutinate cells and spores. Conglutinate cells arise from hyphal adhesion and certain perithecial hyphae develop from conglutinate cells. Various hypha-conglutinate cell transitions play important roles during the development of the perithecial wall and neck. Copyright © 2010. Published by Elsevier Inc.

No prolongation of skin allograft survival by immunoproteasome inhibition in mice.

PubMed

Mundt, Sarah; Basler, Michael; Sawitzki, Birgit; Groettrup, Marcus

2017-08-01

The immunoproteasome, a distinct class of proteasomes, which is inducible under inflammatory conditions and constitutively expressed in monocytes and lymphocytes, is known to shape the antigenic repertoire presented on major histocompatibility complex (MHC) class I molecules. Moreover, inhibition of the immunoproteasome subunit LMP7 ameliorates clinical symptoms of autoimmune diseases in vivo and was shown to suppress the development of T helper cell (Th) 1 and Th17 cells and to promote regulatory T-cell (Treg) generation independently of its function in antigen processing. Since Th1 and Th17 cells are detrimental and Treg cells are critical for transplant acceptance, we investigated the influence of the LMP7-selective inhibitor ONX 0914 in a mixed lymphocyte reaction (MLR) in vitro as well as on allograft rejection in a MHC-disparate (C57BL/6 to BALB/c) and a multiple minor histocompatibility antigen (miHA)-disparate (B10.Br to C3H) model of skin transplantation in vivo. Although we observed reduced allo-specific IL-17 production of T cells in vitro, we found that selective inhibition of LMP7 had neither an influence on allograft survival in an MHC-mismatch model nor in a multiple minor mismatch skin transplantation model. We conclude that inhibition of the immunoproteasome is not effective in prolonging skin allograft survival in skin allotransplantation. Copyright © 2017 Elsevier Ltd. All rights reserved.

Phenotypic and genotypic expression of self-incompatibility haplotypes in Arabidopsis lyrata suggests unique origin of alleles in different dominance classes.

PubMed

Prigoda, Nadia L; Nassuth, Annette; Mable, Barbara K

2005-07-01

The highly divergent alleles of the SRK gene in outcrossing Arabidopsis lyrata have provided important insights into the evolutionary history of self-incompatibility (SI) alleles and serve as an ideal model for studies of the evolutionary and molecular interactions between alleles in cell-cell recognition systems in general. One tantalizing question is how new specificities arise in systems that require coordination between male and female components. Allelic recruitment via gene conversion has been proposed as one possibility, based on the division of DNA sequences at the SRK locus into two distinctive groups: (1) sequences whose relationships are not well resolved and display the long branch lengths expected for a gene under balancing selection (Class A); and (2) sequences falling into a well-supported group with shorter branch lengths (Class B) that are closely related to an unlinked paralogous locus. The purpose of this study was to determine if differences in phenotype (site of expression assayed using allele-specific reverse transcription-polymerase chain reaction) or function (dominance relationships assayed through controlled pollinations) accompany the sequence-based classification. Expression of Class A alleles was restricted to floral tissues, as predicted for genes involved in the SI response. In contrast, Class B alleles, despite being tightly linked to the SI phenotype, were unexpectedly expressed in both leaves and floral tissues; the same pattern found for a related unlinked paralogous sequence. Whereas Class A included haplotypes in three different dominance classes, all Class B haplotypes were found to be recessive to all except one Class A haplotype. In addition, mapping of expression and dominance patterns onto an S-domain-based genealogy suggested that allelic dominance may be determined more by evolutionary history than by frequency-dependent selection for lowered dominance as some theories suggest. The possibility that interlocus gene conversion might have contributed to allelic diversity is discussed.

Human scavenger receptor class B type I is expressed with cell-specific fashion in both initial and terminal site of reverse cholesterol transport.

PubMed

Nakagawa-Toyama, Yumiko; Hirano, Ken-ichi; Tsujii, Ken-ichi; Nishida, Makoto; Miyagawa, Jun-ichiro; Sakai, Naohiko; Yamashita, Shizuya

2005-11-01

The reverse cholesterol transport (RCT) is one of the major protective systems against atherosclerosis, in which high-density lipoprotein (HDL) removes cholesterol from lipid-laden cells and delivers it to the liver. Scavenger receptor class B type I (SR-BI) is a HDL receptor in the liver and adrenal glands and is involved in the selective uptake of cholesteryl ester from HDL, which has been extensively, analyzed using rodent models. However, the expression and regulation of the human homologue of this receptor are not known yet. We previously reported that this receptor is expressed in in vitro differentiated macrophages and its expression is up-regulated by the addition of modified lipoproteins into the medium [Hirano K, Yamashita S, Nakagawa Y, et al. Expression of human scavenger receptor class B type I in cultured human monocyte-derived macrophages and atherosclerotic lesions. Circ Res 1999;85:108-16]. In order to further investigate the physiological significance of this receptor in humans, we have performed extensive immunohistochemical analyses with specimens of the liver and adrenal glands as well as arteries with different stages of atherosclerotic lesions. In human liver and adrenal glands, a positive SR-BI immunoreactivity was detected in both hepatic and adrenal parenchymal cells as well as Kupffer cells. These parenchymal cells had a strong signal on the cell surface, whereas Kupffer cells showed a heterogeneous and punctate pattern. In human aorta and coronary arteries, SR-BI was highly expressed in atherosclerotic plaques, but not in non-atherosclerotic lesions. Double immunostaining revealed that SR-BI was expressed in a subpopulation of macrophages, of which staining pattern was similar to that observed in Kupffer cells. These data clearly demonstrated that SR-BI was expressed with cell-specific fashions in both the initial and terminal step of RCT in humans. Thus, SR-BI might be physiologically relevant and have distinct tissue-specific functions.

Detection of receptor-specific murine leukemia virus binding to cells by immunofluorescence analysis.

PubMed Central

Kadan, M J; Sturm, S; Anderson, W F; Eglitis, M A

1992-01-01

Four classes of murine leukemia virus (MuLV) which display distinct cellular tropisms and bind to different retrovirus receptors to initiate virus infection have been described. In the present study, we describe a rapid, sensitive immunofluorescence assay useful for characterizing the initial binding of MuLV to cells. By using the rat monoclonal antibody 83A25 (L. H. Evans, R. P. Morrison, F. G. Malik, J. Portis, and W. J. Britt, J. Virol. 64:6176-6183, 1990), which recognizes an epitope of the envelope gp70 molecule common to the different classes of MuLV, it is possible to analyse the binding of ecotropic, amphotropic, or xenotropic MuLV by using only a single combination of primary and secondary antibodies. The MuLV binding detected by this assay is envelope receptor specific and matches the susceptibility to infection determined for cells from a variety of species. The binding of amphotropic MuLV to NIH 3T3 cells was shown to be rapid, saturable, and temperature dependent. Chinese hamster ovary (CHO-K1) cells normally lack the ability to bind ecotropic virus and are not infectible by ecotropic vectors. Expression of the cloned ecotropic retrovirus receptor gene (Rec) in CHO-K1 cells confers high levels of ecotropic virus-specific binding and confers susceptibility to infection. Characterization of MuLV binding to primary cells may provide insight into the infectibility of cells by retroviruses and aid in the selection of appropriate vectors for gene transfer experiments. PMID:1312632

PMS2 endonuclease activity has distinct biological functions and is essential for genome maintenance

PubMed Central

van Oers, Johanna M. M.; Roa, Sergio; Werling, Uwe; Liu, Yiyong; Genschel, Jochen; Sellers, Rani S.; Modrich, Paul; Scharff, Matthew D.; Edelmann, Winfried

2010-01-01

The DNA mismatch repair protein PMS2 was recently found to encode a novel endonuclease activity. To determine the biological functions of this activity in mammals, we generated endonuclease-deficient Pms2E702K knock-in mice. Pms2EK/EK mice displayed increased genomic mutation rates and a strong cancer predisposition. In addition, class switch recombination, but not somatic hypermutation, was impaired in Pms2EK/EK B cells, indicating a specific role in Ig diversity. In contrast to Pms2−/− mice, Pms2EK/EK male mice were fertile, indicating that this activity is dispensable in spermatogenesis. Therefore, the PMS2 endonuclease activity has distinct biological functions and is essential for genome maintenance and tumor suppression. PMID:20624957

The Human NK Cell Response to Yellow Fever Virus 17D Is Primarily Governed by NK Cell Differentiation Independently of NK Cell Education.

PubMed

Marquardt, Nicole; Ivarsson, Martin A; Blom, Kim; Gonzalez, Veronica D; Braun, Monika; Falconer, Karolin; Gustafsson, Rasmus; Fogdell-Hahn, Anna; Sandberg, Johan K; Michaëlsson, Jakob

2015-10-01

NK cells play an important role in the defense against viral infections. However, little is known about the regulation of NK cell responses during the first days of acute viral infections in humans. In this study, we used the live attenuated yellow fever virus (YFV) vaccine 17D as a human in vivo model to study the temporal dynamics and regulation of NK cell responses in an acute viral infection. YFV induced a robust NK cell response in vivo, with an early activation and peak in NK cell function at day 6, followed by a delayed peak in Ki67 expression, which was indicative of proliferation, at day 10. The in vivo NK cell response correlated positively with plasma type I/III IFN levels at day 6, as well as with the viral load. YFV induced an increased functional responsiveness to IL-12 and IL-18, as well as to K562 cells, indicating that the NK cells were primed in vivo. The NK cell responses were associated primarily with the stage of differentiation, because the magnitude of induced Ki67 and CD69 expression was distinctly higher in CD57(-) NK cells. In contrast, NK cells expressing self- and nonself-HLA class I-binding inhibitory killer cell Ig-like receptors contributed, to a similar degree, to the response. Taken together, our results indicate that NK cells are primed by type I/III IFN in vivo early after YFV infection and that their response is governed primarily by the differentiation stage, independently of killer cell Ig-like receptor/HLA class I-mediated inhibition or education. Copyright © 2015 by The American Association of Immunologists, Inc.

Subgroups of Chemotherapy Patients With Distinct Morning and Evening Fatigue Trajectories

PubMed Central

Kober, Kord M.; Cooper, Bruce A.; Paul, Steven M.; Dunn, Laura B.; Levine, Jon D.; Wright, Fay; Hammer, Marilyn J.; Mastick, Judy; Venook, Alan; Aouizerat, Bradley E.; Miaskowski, Christine

2017-01-01

Purpose Purposes of this study were to: identify subgroups of patients with distinct trajectories for morning and evening fatigue; evaluate for differences in demographic and clinical characteristics among these subgroups; and compare and contrast the predictors of subgroup membership for morning and evening fatigue. Methods Outpatients with breast, gastrointestinal, gynecological, or lung cancer (n=582) completed questionnaires a total of six times over two cycles of CTX. Morning and evening fatigue severity were evaluated using the Lee Fatigue Scale. Latent profile analysis (LPA) was used to identify distinct subgroups. Results Three latent classes were identified for morning fatigue (i.e., Low (31.8%), High (51.4%), and Very High (16.8%)) and for evening evening fatigue (i.e., Moderate (20.0%), High (21.8%), and Very High (58.2%)). Most of the disease and treatment characteristics did not distinguish among the morning and evening fatigue classes. Compared to the Low class, patients in the High and Very High morning fatigue class were younger, had a lower functional status and higher level of comorbidity. Compared to the Moderate class, patients in the Very High evening fatigue class were younger, more likely to be female, had child care responsibilities, had a lower functional status, and a higher level of comorbidity. Conclusion LPA allows for the identification of risk factors for more severe fatigue. Since an overlap was not observed across the morning and evening fatigue classes and unique predictors for morning and evening fatigue were identified, these findings suggest that morning and evening fatigue may have distinct underlying mechanisms. PMID:26361758

Teachers in Class

ERIC Educational Resources Information Center

Van Galen, Jane

2008-01-01

In this article, I argue for a closer read of the daily "class work" of teachers, as posited by Reay, 1998. In developing exploratory class portraits of four teachers who occupy distinctive social positions (two from working-class homes now teaching upper-middle-class children and two from upper-middle-class homes now teaching poor children), I…

Genome-wide identification and expression analysis of the ClTCP transcription factors in Citrullus lanatus.

PubMed

Shi, Pibiao; Guy, Kateta Malangisha; Wu, Weifang; Fang, Bingsheng; Yang, Jinghua; Zhang, Mingfang; Hu, Zhongyuan

2016-04-12

The plant-specific TCP transcription factor family, which is involved in the regulation of cell growth and proliferation, performs diverse functions in multiple aspects of plant growth and development. However, no comprehensive analysis of the TCP family in watermelon (Citrullus lanatus) has been undertaken previously. A total of 27 watermelon TCP encoding genes distributed on nine chromosomes were identified. Phylogenetic analysis clustered the genes into 11 distinct subgroups. Furthermore, phylogenetic and structural analyses distinguished two homology classes within the ClTCP family, designated Class I and Class II. The Class II genes were differentiated into two subclasses, the CIN subclass and the CYC/TB1 subclass. The expression patterns of all members were determined by semi-quantitative PCR. The functions of two ClTCP genes, ClTCP14a and ClTCP15, in regulating plant height were confirmed by ectopic expression in Arabidopsis wild-type and ortholog mutants. This study represents the first genome-wide analysis of the watermelon TCP gene family, which provides valuable information for understanding the classification and functions of the TCP genes in watermelon.

Ovarian phagocyte subsets and their distinct tissue distribution patterns.

PubMed

Carlock, Colin; Wu, Jean; Zhou, Cindy; Ross, April; Adams, Henry; Lou, Yahuan

2013-01-01

Ovarian macrophages, which play critical roles in various ovarian events, are probably derived from multiple lineages. Thus, a systemic classification of their subsets is a necessary first step for determination of their functions. Utilizing antibodies to five phagocyte markers, i.e. IA/IE (major histocompatibility complex class II), F4/80, CD11b (Mac-1), CD11c, and CD68, this study investigated subsets of ovarian phagocytes in mice. Three-color immunofluorescence and flow cytometry, together with morphological observation on isolated ovarian cells, demonstrated complicated phenotypes of ovarian phagocytes. Four macrophage and one dendritic cell subset, in addition to many minor phagocyte subsets, were identified. A dendritic cell-like population with a unique phenotype of CD11c(high)IA/IE⁻F4/80⁻ was also frequently observed. A preliminary age-dependent study showed dramatic increases in IA/IE⁺ macrophages and IA/IE⁺ dendritic cells after puberty. Furthermore, immunofluorescences on ovarian sections showed that each subset displayed a distinct tissue distribution pattern. The pattern for each subset may hint to their role in an ovarian function. In addition, partial isolation of ovarian macrophage subset using CD11b antibodies was attempted. Establishment of this isolation method may have provided us a tool for more precise investigation of each subset's functions at the cellular and molecular levels.

Neurons Forming Optic Glomeruli Compute Figure–Ground Discriminations in Drosophila

PubMed Central

Aptekar, Jacob W.; Keleş, Mehmet F.; Lu, Patrick M.; Zolotova, Nadezhda M.

2015-01-01

Many animals rely on visual figure–ground discrimination to aid in navigation, and to draw attention to salient features like conspecifics or predators. Even figures that are similar in pattern and luminance to the visual surroundings can be distinguished by the optical disparity generated by their relative motion against the ground, and yet the neural mechanisms underlying these visual discriminations are not well understood. We show in flies that a diverse array of figure–ground stimuli containing a motion-defined edge elicit statistically similar behavioral responses to one another, and statistically distinct behavioral responses from ground motion alone. From studies in larger flies and other insect species, we hypothesized that the circuitry of the lobula—one of the four, primary neuropiles of the fly optic lobe—performs this visual discrimination. Using calcium imaging of input dendrites, we then show that information encoded in cells projecting from the lobula to discrete optic glomeruli in the central brain group these sets of figure–ground stimuli in a homologous manner to the behavior; “figure-like” stimuli are coded similar to one another and “ground-like” stimuli are encoded differently. One cell class responds to the leading edge of a figure and is suppressed by ground motion. Two other classes cluster any figure-like stimuli, including a figure moving opposite the ground, distinctly from ground alone. This evidence demonstrates that lobula outputs provide a diverse basis set encoding visual features necessary for figure detection. PMID:25972183

Neurons forming optic glomeruli compute figure-ground discriminations in Drosophila.

PubMed

Aptekar, Jacob W; Keleş, Mehmet F; Lu, Patrick M; Zolotova, Nadezhda M; Frye, Mark A

2015-05-13

Many animals rely on visual figure-ground discrimination to aid in navigation, and to draw attention to salient features like conspecifics or predators. Even figures that are similar in pattern and luminance to the visual surroundings can be distinguished by the optical disparity generated by their relative motion against the ground, and yet the neural mechanisms underlying these visual discriminations are not well understood. We show in flies that a diverse array of figure-ground stimuli containing a motion-defined edge elicit statistically similar behavioral responses to one another, and statistically distinct behavioral responses from ground motion alone. From studies in larger flies and other insect species, we hypothesized that the circuitry of the lobula--one of the four, primary neuropiles of the fly optic lobe--performs this visual discrimination. Using calcium imaging of input dendrites, we then show that information encoded in cells projecting from the lobula to discrete optic glomeruli in the central brain group these sets of figure-ground stimuli in a homologous manner to the behavior; "figure-like" stimuli are coded similar to one another and "ground-like" stimuli are encoded differently. One cell class responds to the leading edge of a figure and is suppressed by ground motion. Two other classes cluster any figure-like stimuli, including a figure moving opposite the ground, distinctly from ground alone. This evidence demonstrates that lobula outputs provide a diverse basis set encoding visual features necessary for figure detection. Copyright © 2015 the authors 0270-6474/15/357587-13$15.00/0.

β-cell-specific CD8 T cell phenotype in type 1 diabetes reflects chronic autoantigen exposure

PubMed Central

McLaren, James E.; Dolton, Garry; Matthews, Katherine K.; Gostick, Emma; Kronenberg-Versteeg, Deborah; Eichmann, Martin; Knight, Robin R.; Heck, Susanne; Powrie, Jake; Bingley, Polly J.; Dayan, Colin M.; Miles, John J.; Sewell, Andrew K.

2015-01-01

Autoreactive CD8 T cells play a central role in the destruction of pancreatic islet β-cells that leads to type 1 diabetes, yet the key features of this immune-mediated process remain poorly defined. In this study, we combined high definition polychromatic flow cytometry with ultrasensitive peptide-human leukocyte antigen class I (pHLAI) tetramer staining to quantify and characterize β-cell-specific CD8 T cell populations in patients with recent onset type 1 diabetes and healthy controls. Remarkably, we found that β-cell-specific CD8 T cell frequencies in peripheral blood were similar between subject groups. In contrast to healthy controls, however, patients with newly diagnosed type 1 diabetes displayed hallmarks of antigen-driven expansion uniquely within the β-cell-specific CD8 T cell compartment. Molecular analysis of selected β-cell-specific CD8 T cell populations further revealed highly skewed oligoclonal T cell receptor (TCR) repertoires comprising exclusively private clonotypes. Collectively, these data identify novel and distinctive features of disease-relevant CD8 T cells that inform the immunopathogenesis of type 1 diabetes. PMID:25249579

TETRASPANINs in Plants

PubMed Central

Reimann, Ronny; Kost, Benedikt; Dettmer, Jan

2017-01-01

Tetraspanins are small transmembrane proteins that laterally associate with each other and cluster with numerous partner proteins as well as lipids. These interactions result in the formation of a distinct class of membrane domains, the tetraspanin-enriched microdomains (TEMs), which influence numerous cellular processes such as cell adhesion and fusion, intracellular membrane trafficking, signaling, morphogenesis, motility as well as interaction with pathogens and cancer development. The majority of information available about tetraspanins is based on studies using animal models or cell lines, but tetraspanins are also present in fungi and plants. Recent studies indicate that tetraspanins have important functions in plant development, reproduction and stress responses. Here we provide a brief summary of the current state of tetraspanin research in plants. PMID:28458676

A Ferredoxin Disulfide Reductase Delivers Electrons to the Methanosarcina barkeri Class III Ribonucleotide Reductase

DOE PAGES

Wei, Yifeng; Li, Bin; Prakash, Divya; ...

2015-11-04

Two subtypes of class III anaerobic ribonucleotide reductases (RNRs) studied so far couple the reduction of ribonucleotides to the oxidation of formate, or the oxidation of NADPH via thioredoxin and thioredoxin reductase. Certain methanogenic archaea contain a phylogenetically distinct third subtype of class III RNR, with distinct active-site residues. Here we report the cloning and recombinant expression of the Methanosarcina barkeri class III RNR and show that the electrons required for ribonucleotide reduction can be delivered by a [4Fe-4S] protein ferredoxin disulfide reductase, and a conserved thioredoxin-like protein NrdH present in the RNR operon. The diversity of class III RNRsmore » reflects the diversity of electron carriers used in anaerobic metabolism« less

CD1-Restricted T Cells at the Crossroad of Innate and Adaptive Immunity.

PubMed

Pereira, Catia S; Macedo, M Fatima

2016-01-01

Lipid-specific T cells comprise a group of T cells that recognize lipids bound to the MHC class I-like CD1 molecules. There are four isoforms of CD1 that are expressed at the surface of antigen presenting cells and therefore capable of presenting lipid antigens: CD1a, CD1b, CD1c, and CD1d. Each one of these isoforms has distinct structural features and cellular localizations, which promotes binding to a broad range of different types of lipids. Lipid antigens originate from either self-tissues or foreign sources, such as bacteria, fungus, or plants and their recognition by CD1-restricted T cells has important implications in infection but also in cancer and autoimmunity. In this review, we describe the characteristics of CD1 molecules and CD1-restricted lipid-specific T cells, highlighting the innate-like and adaptive-like features of different CD1-restricted T cell subtypes.

TNF-induced target cell killing by CTL activated through cross-presentation.

PubMed

Wohlleber, Dirk; Kashkar, Hamid; Gärtner, Katja; Frings, Marianne K; Odenthal, Margarete; Hegenbarth, Silke; Börner, Carolin; Arnold, Bernd; Hämmerling, Günter; Nieswandt, Bernd; van Rooijen, Nico; Limmer, Andreas; Cederbrant, Karin; Heikenwalder, Mathias; Pasparakis, Manolis; Protzer, Ulrike; Dienes, Hans-Peter; Kurts, Christian; Krönke, Martin; Knolle, Percy A

2012-09-27

Viruses can escape cytotoxic T cell (CTL) immunity by avoiding presentation of viral components via endogenous MHC class I antigen presentation in infected cells. Cross-priming of viral antigens circumvents such immune escape by allowing noninfected dendritic cells to activate virus-specific CTLs, but they remain ineffective against infected cells in which immune escape is functional. Here, we show that cross-presentation of antigen released from adenovirus-infected hepatocytes by liver sinusoidal endothelial cells stimulated cross-primed effector CTLs to release tumor necrosis factor (TNF), which killed virus-infected hepatocytes through caspase activation. TNF receptor signaling specifically eliminated infected hepatocytes that showed impaired anti-apoptotic defense. Thus, CTL immune surveillance against infection relies on two similarly important but distinct effector functions that are both MHC restricted, requiring either direct antigen recognition on target cells and canonical CTL effector function or cross-presentation and a noncanonical effector function mediated by TNF. Copyright © 2012 The Authors. Published by Elsevier Inc. All rights reserved.

7 CFR 30.5 - Class.

Code of Federal Regulations, 2012 CFR

2012-01-01

... AND STANDARDS Classification of Leaf Tobacco Covering Classes, Types and Groups of Grades § 30.5 Class. One of the major divisions of leaf tobacco based on the distinct characteristics of the tobacco caused...

7 CFR 30.5 - Class.

Code of Federal Regulations, 2013 CFR

2013-01-01

... AND STANDARDS Classification of Leaf Tobacco Covering Classes, Types and Groups of Grades § 30.5 Class. One of the major divisions of leaf tobacco based on the distinct characteristics of the tobacco caused...

7 CFR 30.5 - Class.

Code of Federal Regulations, 2010 CFR

2010-01-01

... AND STANDARDS Classification of Leaf Tobacco Covering Classes, Types and Groups of Grades § 30.5 Class. One of the major divisions of leaf tobacco based on the distinct characteristics of the tobacco caused...

7 CFR 30.5 - Class.

Code of Federal Regulations, 2011 CFR

2011-01-01

... AND STANDARDS Classification of Leaf Tobacco Covering Classes, Types and Groups of Grades § 30.5 Class. One of the major divisions of leaf tobacco based on the distinct characteristics of the tobacco caused...

7 CFR 30.5 - Class.

Code of Federal Regulations, 2014 CFR

2014-01-01

... AND STANDARDS Classification of Leaf Tobacco Covering Classes, Types and Groups of Grades § 30.5 Class. One of the major divisions of leaf tobacco based on the distinct characteristics of the tobacco caused...

Forging T-Lymphocyte Identity: Intersecting Networks of Transcriptional Control

PubMed Central

Rothenberg, Ellen V.; Ungerbäck, Jonas; Champhekar, Ameya

2016-01-01

T lymphocyte development branches off from other lymphoid developmental programs through its requirement for sustained environmental signals through the Notch pathway. In the thymus, Notch signaling induces a succession of T-lineage regulatory factors that collectively create the T-cell identity through distinct steps. This process involves both the staged activation of T-cell identity genes and the staged repression of progenitor-cell-inherited regulatory genes once their roles in self-renewal and population expansion are no longer needed. With the recent characterization of Innate Lymphoid Cells (ILCs) that share transcriptional regulation programs extensively with T cell subsets, T-cell identity can increasingly be seen as defined in modular terms, as the processes selecting and actuating effector function are potentially detachable from the processes generating and selecting clonally unique T-cell receptor structures. The developmental pathways of different classes of T cells and ILCs are distinguished by the numbers of prerequisites of gene rearrangement, selection, and antigen contact before the cells gain access to nearly-common regulatory mechanisms for choosing effector function. Here, the major classes of transcription factors that interact with Notch signals during T-lineage specification are discussed in terms of their roles in these programs, the evidence for their spectra of target genes at different stages, and their cross-regulatory and cooperative actions with each other. Specific topics include Notch modulation of PU.1 and GATA-3, PU.1-Notch competition, the relationship between PU.1 and GATA-3, and the roles of E proteins, Bcl11b, and GATA-3 in guiding acquisition of T-cell identity while avoiding redirection to an ILC fate. PMID:26791859

A Dinitroaniline-Resistant Mutant of Eleusine indica Exhibits Cross-Resistance and Supersensitivity to Antimicrotubule Herbicides and Drugs 1

PubMed Central

Vaughn, Kevin C.; Marks, M. David; Weeks, Donald P.

1987-01-01

A dinitroaniline-resistant (R) biotype of Eleusine indica (L.) Gaertner. (goosegrass) is demonstrated to be cross-resistant to a structurally non-related herbicide, amiprophosmethyl, and supersensitive to two other classes of compounds which disrupt mitosis. These characteristics of the R biotype were discovered in a comparative test of the effects of 24 different antimitotic compounds on the R biotype and susceptible (S) wild-type Eleusine. The compounds tested could be classified into three groups based upon their effects on mitosis in root tips of the susceptible (S) biotype. Class I compounds induced effects like the well known mitotic disrupter colchicine: absence of cortical and spindle microtubules, mitosis arrested at prometaphase, and the formation of polymorphic nuclei after arrested mitosis. The R biotype was resistant to treatment with some class I inhibitors (all dinitroaniline herbicides and amiprophosmethyl) but not all (e.g. colchicine, podophyllotoxin, vinblastine, and pronamide). Roots of the R biotype, when treated with either dinitroaniline herbicides or amiprophosmethyl, exhibited no or only small increases in the mitotic index nor were the spindle and cortical microtubules affected. Compounds of class II (carbamate herbicides and griseofulvin) cause misorientation of microtubules which results in multinucleated cells. Compounds of class III (caffeine and structually related alkaloids) cause imcomplete cell walls to form at telophase. Each of these last two classes of compounds affected the R biotype more than the S biotype (supersensitivity). The cross-resistance and high levels of resistance of the R biotype of Eleusine to the dinitroaniline herbicides and the structurally distinct herbicide, amiprophosmethyl, indicate that a mechanism of resistance based upon metabolic modification, translocation, or compartmentation of the herbicides is probably not operative. Images Fig. 1 Fig. 2 Fig. 3 Fig. 5 Fig. 6 PMID:16665371

A Dinitroaniline-Resistant Mutant of Eleusine indica Exhibits Cross-Resistance and Supersensitivity to Antimicrotubule Herbicides and Drugs.

PubMed

Vaughn, K C; Marks, M D; Weeks, D P

1987-04-01

A dinitroaniline-resistant (R) biotype of Eleusine indica (L.) Gaertner. (goosegrass) is demonstrated to be cross-resistant to a structurally non-related herbicide, amiprophosmethyl, and supersensitive to two other classes of compounds which disrupt mitosis. These characteristics of the R biotype were discovered in a comparative test of the effects of 24 different antimitotic compounds on the R biotype and susceptible (S) wild-type Eleusine. The compounds tested could be classified into three groups based upon their effects on mitosis in root tips of the susceptible (S) biotype. Class I compounds induced effects like the well known mitotic disrupter colchicine: absence of cortical and spindle microtubules, mitosis arrested at prometaphase, and the formation of polymorphic nuclei after arrested mitosis. The R biotype was resistant to treatment with some class I inhibitors (all dinitroaniline herbicides and amiprophosmethyl) but not all (e.g. colchicine, podophyllotoxin, vinblastine, and pronamide). Roots of the R biotype, when treated with either dinitroaniline herbicides or amiprophosmethyl, exhibited no or only small increases in the mitotic index nor were the spindle and cortical microtubules affected. Compounds of class II (carbamate herbicides and griseofulvin) cause misorientation of microtubules which results in multinucleated cells. Compounds of class III (caffeine and structually related alkaloids) cause imcomplete cell walls to form at telophase. Each of these last two classes of compounds affected the R biotype more than the S biotype (supersensitivity). The cross-resistance and high levels of resistance of the R biotype of Eleusine to the dinitroaniline herbicides and the structurally distinct herbicide, amiprophosmethyl, indicate that a mechanism of resistance based upon metabolic modification, translocation, or compartmentation of the herbicides is probably not operative.

HLA-C expression pattern is spatially different between psoriasis and eczema skin lesions.

PubMed

Carlén, Lina; Sakuraba, Kazuko; Ståhle, Mona; Sánchez, Fabio

2007-02-01

Interactions between genetic and environmental factors underlie the immune dysregulation and keratinocyte abnormalities that characterize psoriasis. Among known psoriasis susceptibility loci (PSORS), PSORS1 on chromosome 6 has the strongest association to disease. Altered expression of some PSORS1 candidate genes has been reported but little is known about HLA-C expression in psoriasis. This study compared expression of major histocompatibility complex class Ia and HLA-C in psoriasis, allergic contact eczema, and normal skin. Although HLA-C was abundant in protein extracts from both eczema and psoriasis, a consistent and intriguing difference in the expression pattern was observed; strong immunoreactivity in the basal cell layer, polarized towards the basement membrane in psoriasis, whereas in eczema lesions HLA-C immunostaining was present mostly in suprabasal cells. Inflammatory cells in the dermis were strongly stained in both diseases. Normal skin epithelium showed less intense but similar HLA-C staining as eczema lesions. HLA class Ia expression overall resembled that of HLA-C in all samples. The distinct HLA-C expression patterns in psoriasis and eczema suggest a functional role in the specific psoriasis immune response and not only a general feature of inflammation.

Variants in members of the cadherin-catenin complex, CDH1 and CTNND1, cause blepharocheilodontic syndrome.

PubMed

Kievit, Anneke; Tessadori, Federico; Douben, Hannie; Jordens, Ingrid; Maurice, Madelon; Hoogeboom, Jeannette; Hennekam, Raoul; Nampoothiri, Sheela; Kayserili, Hülya; Castori, Marco; Whiteford, Margo; Motter, Connie; Melver, Catherine; Cunningham, Michael; Hing, Anne; Kokitsu-Nakata, Nancy M; Vendramini-Pittoli, Siulan; Richieri-Costa, Antonio; Baas, Annette F; Breugem, Corstiaan C; Duran, Karen; Massink, Maarten; Derksen, Patrick W B; van IJcken, Wilfred F J; van Unen, Leontine; Santos-Simarro, Fernando; Lapunzina, Pablo; Gil-da Silva Lopes, Vera L; Lustosa-Mendes, Elaine; Krall, Max; Slavotinek, Anne; Martinez-Glez, Victor; Bakkers, Jeroen; van Gassen, Koen L I; de Klein, Annelies; van den Boogaard, Marie-José H; van Haaften, Gijs

2018-02-01

Blepharocheilodontic syndrome (BCDS) consists of lagophthalmia, ectropion of the lower eyelids, distichiasis, euryblepharon, cleft lip/palate and dental anomalies and has autosomal dominant inheritance with variable expression. We identified heterozygous variants in two genes of the cadherin-catenin complex, CDH1, encoding E-cadherin, and CTNND1, encoding p120 catenin delta1 in 15 of 17 BCDS index patients, as was recently described in a different publication. CDH1 plays an essential role in epithelial cell adherence; CTNND1 binds to CDH1 and controls the stability of the complex. Functional experiments in zebrafish and human cells showed that the CDH1 variants impair the cell adhesion function of the cadherin-catenin complex in a dominant-negative manner. Variants in CDH1 have been linked to familial hereditary diffuse gastric cancer and invasive lobular breast cancer; however, no cases of gastric or breast cancer have been reported in our BCDS cases. Functional experiments reported here indicated the BCDS variants comprise a distinct class of CDH1 variants. Altogether, we identified the genetic cause of BCDS enabling DNA diagnostics and counseling, in addition we describe a novel class of dominant negative CDH1 variants.

Distinct T cell interactions with HLA class II tetramers characterize a spectrum of TCR affinities in the human antigen-specific T cell response.

PubMed

Reichstetter, S; Ettinger, R A; Liu, A W; Gebe, J A; Nepom, G T; Kwok, W W

2000-12-15

The polyclonal nature of T cells expanding in an ongoing immune response results in a range of disparate affinities and activation potential. Recently developed human class II tetramers provide a means to analyze this diversity by direct characterization of the trimolecular TCR-peptide-MHC interaction in live cells. Two HSV-2 VP16(369-379)-specific, DQA1*0102/DQB1*0602 (DQ0602)-restricted T cell clones were compared by means of T cell proliferation assay and HLA-DQ0602 tetramer staining. These two clones were obtained from the same subject, but show different TCR gene usage. Clone 48 was 10-fold more sensitive to VP16(369-379) peptide stimulation than clone 5 as assayed by proliferation assays, correlating with differences in MHC tetramer binding. Clone 48 gave positive staining with the DQ0602/VP16(369-379) tetramer at either 23 or 37 degrees C. Weak staining was also observed at 4 degrees C. Clone 5 showed weaker staining compared with clone 48 at 37 degrees C, and no staining was observed at 23 degrees C or on ice. Receptor internalization was not required for positive staining. Competitive binding indicates that the cell surface TCR of clone 48 has higher affinity for the DQ0602/VP16(369-379) complex than clone 5. The higher binding affinity of clone 48 for the peptide-MHC complex also correlates with a slower dissociation rate compared with clone 5.

Whole organism high content screening identifies stimulators of pancreatic beta-cell proliferation.

PubMed

Tsuji, Naoki; Ninov, Nikolay; Delawary, Mina; Osman, Sahar; Roh, Alex S; Gut, Philipp; Stainier, Didier Y R

2014-01-01

Inducing beta-cell mass expansion in diabetic patients with the aim to restore glucose homeostasis is a promising therapeutic strategy. Although several in vitro studies have been carried out to identify modulators of beta-cell mass expansion, restoring endogenous beta-cell mass in vivo has yet to be achieved. To identify potential stimulators of beta-cell replication in vivo, we established transgenic zebrafish lines that monitor and allow the quantification of cell proliferation by using the fluorescent ubiquitylation-based cell cycle indicator (FUCCI) technology. Using these new reagents, we performed an unbiased chemical screen, and identified 20 small molecules that markedly increased beta-cell proliferation in vivo. Importantly, these structurally distinct molecules, which include clinically-approved drugs, modulate three specific signaling pathways: serotonin, retinoic acid and glucocorticoids, showing the high sensitivity and robustness of our screen. Notably, two drug classes, retinoic acid and glucocorticoids, also promoted beta-cell regeneration after beta-cell ablation. Thus, this study establishes a proof of principle for a high-throughput small molecule-screen for beta-cell proliferation in vivo, and identified compounds that stimulate beta-cell proliferation and regeneration.

Multi-scale imaging and informatics pipeline for in situ pluripotent stem cell analysis.

PubMed

Gorman, Bryan R; Lu, Junjie; Baccei, Anna; Lowry, Nathan C; Purvis, Jeremy E; Mangoubi, Rami S; Lerou, Paul H

2014-01-01

Human pluripotent stem (hPS) cells are a potential source of cells for medical therapy and an ideal system to study fate decisions in early development. However, hPS cells cultured in vitro exhibit a high degree of heterogeneity, presenting an obstacle to clinical translation. hPS cells grow in spatially patterned colony structures, necessitating quantitative single-cell image analysis. We offer a tool for analyzing the spatial population context of hPS cells that integrates automated fluorescent microscopy with an analysis pipeline. It enables high-throughput detection of colonies at low resolution, with single-cellular and sub-cellular analysis at high resolutions, generating seamless in situ maps of single-cellular data organized by colony. We demonstrate the tool's utility by analyzing inter- and intra-colony heterogeneity of hPS cell cycle regulation and pluripotency marker expression. We measured the heterogeneity within individual colonies by analyzing cell cycle as a function of distance. Cells loosely associated with the outside of the colony are more likely to be in G1, reflecting a less pluripotent state, while cells within the first pluripotent layer are more likely to be in G2, possibly reflecting a G2/M block. Our multi-scale analysis tool groups colony regions into density classes, and cells belonging to those classes have distinct distributions of pluripotency markers and respond differently to DNA damage induction. Lastly, we demonstrate that our pipeline can robustly handle high-content, high-resolution single molecular mRNA FISH data by using novel image processing techniques. Overall, the imaging informatics pipeline presented offers a novel approach to the analysis of hPS cells that includes not only single cell features but also colony wide, and more generally, multi-scale spatial configuration.

Stimulus selectivity and response latency in putative inhibitory and excitatory neurons of the primate inferior temporal cortex

PubMed Central

Mruczek, Ryan E. B.

2012-01-01

The cerebral cortex is composed of many distinct classes of neurons. Numerous studies have demonstrated corresponding differences in neuronal properties across cell types, but these comparisons have largely been limited to conditions outside of awake, behaving animals. Thus the functional role of the various cell types is not well understood. Here, we investigate differences in the functional properties of two widespread and broad classes of cells in inferior temporal cortex of macaque monkeys: inhibitory interneurons and excitatory projection cells. Cells were classified as putative inhibitory or putative excitatory neurons on the basis of their extracellular waveform characteristics (e.g., spike duration). Consistent with previous intracellular recordings in cortical slices, putative inhibitory neurons had higher spontaneous firing rates and higher stimulus-evoked firing rates than putative excitatory neurons. Additionally, putative excitatory neurons were more susceptible to spike waveform adaptation following very short interspike intervals. Finally, we compared two functional properties of each neuron's stimulus-evoked response: stimulus selectivity and response latency. First, putative excitatory neurons showed stronger stimulus selectivity compared with putative inhibitory neurons. Second, putative inhibitory neurons had shorter response latencies compared with putative excitatory neurons. Selectivity differences were maintained and latency differences were enhanced during a visual search task emulating more natural viewing conditions. Our results suggest that short-latency inhibitory responses are likely to sculpt visual processing in excitatory neurons, yielding a sparser visual representation. PMID:22933717

Flow cytometry beads rather than the antihuman globulin method should be used to detect HLA Class I IgG antibody (PRA) in cadaveric renal regraft candidates.

PubMed

Bryan, Christopher F; McDonald, Scott B; Baier, Karen A; Luger, Alan M; Aeder, Mark I; Murillo, Daniel; Muruve, Nicolas A; Nelson, Paul W; Shield, Charles F; Warady, Bradley A

2002-01-01

HLA Class I antibody screening can be performed by flow cytometry using a mixture of 30 distinct bead populations each coated with the Class I antigen phenotype derived from different cell lines. In this study we compared the efficacy of Class I antibody screens done by flow cytometry beads with the antihuman globulin (AHG) method for patients awaiting cadaveric renal retransplantation. Class I panel reactive antibody (PRA) screening by flow cytometric beads of 21 regraft serum samples that had all been found to be negative by AHG DTT Class I PRA, revealed that 57.1% (12 of 21) had a flow Class I PRA of > or = 10%. Furthermore, when five regraft sera with an intermediate PRA were screened (mean AHG DTT PRA = 33.2 +/- 13%) the mean flow Class I PRA almost doubled (mean flow PRA = 72.4 +/- 10.2%) (p < 0.01). When active UNOS waiting list regraft candidates, after several months of screening the Class I PRA by flow beads, were divided into the three PRA categories based on their peak flow Class I PRA value (0-20%, 21-79% and > or = 80%), the incidence of a positive flow cross-match was 0%, 72% and 85% and the incidence of retransplantation was 60%, 22% and 10%, in each of these groups, respectively. These data provided our histocompatibility laboratory with the rationale to stop performing the AHG PRA and perform only the flow Class I PRA method for regraft candidates.

Impairment of the Ubiquitin-Proteasome Pathway by Methyl N-(6-Phenylsulfanyl-1H-benzimidazol-2-yl)carbamate Leads to a Potent Cytotoxic Effect in Tumor Cells

PubMed Central

Dogra, Nilambra; Mukhopadhyay, Tapas

2012-01-01

In recent years, there has been a great deal of interest in proteasome inhibitors as a novel class of anticancer drugs. We report that fenbendazole (FZ) (methyl N-(6-phenylsulfanyl-1H-benzimidazol-2-yl)carbamate) exhibits a potent growth-inhibitory activity against cancer cell lines but not normal cells. We show here, using fluorogenic substrates, that FZ treatment leads to the inhibition of proteasomal activity in the cells. Succinyl-Leu-Leu-Val-Tyr-methylcoumarinamide (MCA), benzyloxycarbonyl-Leu-Leu-Glu-7-amido-4-MCA, and t-butoxycarbonyl-Gln-Ala-Arg-7-amido-4-MCA fluorescent derivatives were used to assess chymotrypsin-like, post-glutamyl peptidyl-hydrolyzing, and trypsin-like protease activities, respectively. Non-small cell lung cancer cells transiently transfected with an expression plasmid encoding pd1EGFP and treated with FZ showed an accumulation of the green fluorescent protein in the cells due to an increase in its half-life. A number of apoptosis regulatory proteins that are normally degraded by the ubiquitin-proteasome pathway like cyclins, p53, and IκBα were found to be accumulated in FZ-treated cells. In addition, FZ induced distinct ER stress-associated genes like GRP78, GADD153, ATF3, IRE1α, and NOXA in these cells. Thus, treatment of human NSCLC cells with fenbendazole induced endoplasmic reticulum stress, reactive oxygen species production, decreased mitochondrial membrane potential, and cytochrome c release that eventually led to cancer cell death. This is the first report to demonstrate the inhibition of proteasome function and induction of endoplasmic reticulum stress/reactive oxygen species-dependent apoptosis in human lung cancer cell lines by fenbendazole, which may represent a new class of anticancer agents showing selective toxicity against cancer cells. PMID:22745125

Complicating the "Soccer Mom:" The Cultural Politics of Forming Class-Based Identity, Distinction, and Necessity

ERIC Educational Resources Information Center

Swanson, Lisa

2009-01-01

Using Pierre Bourdieu's theories of social class differentiation and class reproduction, this paper provides an analysis of class-based identity politics in contemporary suburban America. Through a critical ethnography of the emergent, American, upper-middle-class "soccer mom" phenomenon, this study contributes to a growing body of…

Substance use predictors of victimization profiles among homeless youth: a latent class analysis.

PubMed

Bender, Kimberly; Thompson, Sanna; Ferguson, Kristin; Langenderfer, Lisa

2014-02-01

Although a substantial body of literature demonstrates high prevalence of street victimization among homeless youth, few studies have investigated the existence of victimization classes that differ on the type and frequency of victimization experienced. Nor do we know how substance use patterns relate to victimization classes. Using latent class analysis (LCA), we examined the existence of victimization classes of homeless youth and investigated substance use predictors of class membership utilizing a large purposive sample (N=601) recruited from homeless youth-serving host agencies in three disparate regions of the U.S. Results of the LCA suggest the presence of three distinct victimization profiles - youth fit into a low-victimization class, a witness class, or a high-victimization class. These three victimization classes demonstrated differences in their substance use, including rates of substance abuse/dependence on alcohol and/or drugs. The presence of distinct victimization profiles suggests the need for screening and referral for differential services. Copyright © 2013 The Foundation for Professionals in Services for Adolescents. Published by Elsevier Ltd. All rights reserved.

Chemical imaging of a Symbiodinium sp. cell using synchrotron infrared microspectroscopy: a feasibility study.

PubMed

Gordon, B R; Martin, D E; Bambery, K R; Motti, C A

2018-04-01

The symbiotic relationship between corals and Symbiodinium spp. is the key to the success and survival of coral reef ecosystems the world over. Nutrient exchange and chemical communication between the two partners provides the foundation of this key relationship, yet we are far from a complete understanding of these processes. This is due, in part, to the difficulties associated with studying an intracellular symbiosis at the small spatial scales required to elucidate metabolic interactions between the two partners. This feasibility study, which accompanied a more extensive investigation of fixed Symbiodinium cells (data unpublished), examines the potential of using synchrotron radiation infrared microspectroscopy (SR-IRM) for exploring metabolite localisation within a single Symbiodinium cell. In doing so, three chemically distinct subcellular regions of a single Symbiodinium cell were established and correlated to cellular function based on assignment of diagnostic chemical classes. © 2017 The Authors Journal of Microscopy © 2017 Royal Microscopical Society.

Two distinct classes of functional α7-containing nicotinic receptor on rat superior cervical ganglion neurons

PubMed Central

Cuevas, Javier; Roth, Adelheid L; Berg, Darwin K

2000-01-01

Nicotinic acetylcholine receptors (nAChRs) that bind α-bungarotoxin (αBgt) were studied on isolated rat superior cervical ganglion (SCG) neurons using whole-cell patch clamp recording techniques.Rapid application of ACh onto the soma of voltage clamped neurons evoked a slowly desensitizing current that was reversibly blocked by αBgt (50 nm). The toxin-sensitive current constituted on average about half of the peak whole-cell response evoked by ACh.Nanomolar concentrations of methyllycaconitine blocked the αBgt-sensitive component of the ACh-evoked current as did intracellular dialysis with an anti-α7 monoclonal antibody. The results indicate that the slowly reversible toxin-sensitive response elicited by ACh arises from activation of an unusual class of α7-containing receptor (α7-nAChR) similar to that reported previously for rat intracardiac ganglion neurons.A second class of functional α7-nAChR was identified on some SCG neurons by using rapid application of choline to elicit responses. In these cases a biphasic response was obtained, which included a rapidly desensitizing component that was blocked by αBgt in a pseudo-irreversible manner. The pharmacology and kinetics of the responses resembled those previously attributed to α7-nAChRs in a number of other neuronal cell types.Experiments measuring the dissociation rate of 125I-labelled αBgt from SCG neurons revealed two classes of toxin-binding site. The times for toxin dissociation were consistent with those required to reverse blockade of the two kinds of αBgt-sensitive response.These results indicate that rat SCG neurons express two types of functional α7-nAChR, differing in pharmacology, desensitization and reversibility of αBgt blockade. PMID:10856125

Gα12 structural determinants of Hsp90 interaction are necessary for serum response element-mediated transcriptional activation.

PubMed

Montgomery, Ellyn R; Temple, Brenda R S; Peters, Kimberly A; Tolbert, Caitlin E; Booker, Brandon K; Martin, Joseph W; Hamilton, Tyler P; Tagliatela, Alicia C; Smolski, William C; Rogers, Stephen L; Jones, Alan M; Meigs, Thomas E

2014-04-01

The G12/13 class of heterotrimeric G proteins, comprising the α-subunits Gα12 and Gα13, regulates multiple aspects of cellular behavior, including proliferation and cytoskeletal rearrangements. Although guanine nucleotide exchange factors for the monomeric G protein Rho (RhoGEFs) are well characterized as effectors of this G protein class, a variety of other downstream targets has been reported. To identify Gα12 determinants that mediate specific protein interactions, we used a structural and evolutionary comparison between the G12/13, Gs, Gi, and Gq classes to identify "class-distinctive" residues in Gα12 and Gα13. Mutation of these residues in Gα12 to their deduced ancestral forms revealed a subset necessary for activation of serum response element (SRE)-mediated transcription, a G12/13-stimulated pathway implicated in cell proliferative signaling. Unexpectedly, this subset of Gα12 mutants showed impaired binding to heat-shock protein 90 (Hsp90) while retaining binding to RhoGEFs. Corresponding mutants of Gα13 exhibited robust SRE activation, suggesting a Gα12-specific mechanism, and inhibition of Hsp90 by geldanamycin or small interfering RNA-mediated lowering of Hsp90 levels resulted in greater downregulation of Gα12 than Gα13 signaling in SRE activation experiments. Furthermore, the Drosophila G12/13 homolog Concertina was unable to signal to SRE in mammalian cells, and Gα12:Concertina chimeras revealed Gα12-specific determinants of SRE activation within the switch regions and a C-terminal region. These findings identify Gα12 determinants of SRE activation, implicate Gα12:Hsp90 interaction in this signaling mechanism, and illuminate structural features that arose during evolution of Gα12 and Gα13 to allow bifurcated mechanisms of signaling to a common cell proliferative pathway.

Epitope predictions indicate the presence of two distinct types of epitope-antibody-reactivities determined by epitope profiling of intravenous immunoglobulins.

PubMed

Luštrek, Mitja; Lorenz, Peter; Kreutzer, Michael; Qian, Zilliang; Steinbeck, Felix; Wu, Di; Born, Nadine; Ziems, Bjoern; Hecker, Michael; Blank, Miri; Shoenfeld, Yehuda; Cao, Zhiwei; Glocker, Michael O; Li, Yixue; Fuellen, Georg; Thiesen, Hans-Jürgen

2013-01-01

Epitope-antibody-reactivities (EAR) of intravenous immunoglobulins (IVIGs) determined for 75,534 peptides by microarray analysis demonstrate that roughly 9% of peptides derived from 870 different human protein sequences react with antibodies present in IVIG. Computational prediction of linear B cell epitopes was conducted using machine learning with an ensemble of classifiers in combination with position weight matrix (PWM) analysis. Machine learning slightly outperformed PWM with area under the curve (AUC) of 0.884 vs. 0.849. Two different types of epitope-antibody recognition-modes (Type I EAR and Type II EAR) were found. Peptides of Type I EAR are high in tyrosine, tryptophan and phenylalanine, and low in asparagine, glutamine and glutamic acid residues, whereas for peptides of Type II EAR it is the other way around. Representative crystal structures present in the Protein Data Bank (PDB) of Type I EAR are PDB 1TZI and PDB 2DD8, while PDB 2FD6 and 2J4W are typical for Type II EAR. Type I EAR peptides share predicted propensities for being presented by MHC class I and class II complexes. The latter interaction possibly favors T cell-dependent antibody responses including IgG class switching. Peptides of Type II EAR are predicted not to be preferentially presented by MHC complexes, thus implying the involvement of T cell-independent IgG class switch mechanisms. The high extent of IgG immunoglobulin reactivity with human peptides implies that circulating IgG molecules are prone to bind to human protein/peptide structures under non-pathological, non-inflammatory conditions. A webserver for predicting EAR of peptide sequences is available at www.sysmed-immun.eu/EAR.

Dissecting local circuits in vivo: integrated optogenetic and electrophysiology approaches for exploring inhibitory regulation of cortical activity.

PubMed

Cardin, Jessica A

2012-01-01

Local cortical circuit activity in vivo comprises a complex and flexible series of interactions between excitatory and inhibitory neurons. Our understanding of the functional interactions between these different neural populations has been limited by the difficulty of identifying and selectively manipulating the diverse and sparsely represented inhibitory interneuron classes in the intact brain. The integration of recently developed optical tools with traditional electrophysiological techniques provides a powerful window into the role of inhibition in regulating the activity of excitatory neurons. In particular, optogenetic targeting of specific cell classes reveals the distinct impacts of local inhibitory populations on other neurons in the surrounding local network. In addition to providing the ability to activate or suppress spiking in target cells, optogenetic activation identifies extracellularly recorded neurons by class, even when naturally occurring spike rates are extremely low. However, there are several important limitations on the use of these tools and the interpretation of resulting data. The purpose of this article is to outline the uses and limitations of optogenetic tools, along with current methods for achieving cell type-specific expression, and to highlight the advantages of an experimental approach combining optogenetics and electrophysiology to explore the role of inhibition in active networks. To illustrate the efficacy of these combined approaches, I present data comparing targeted manipulations of cortical fast-spiking, parvalbumin-expressing and low threshold-spiking, somatostatin-expressing interneurons in vivo. Copyright © 2011 Elsevier Ltd. All rights reserved.

Glycogen content in hepatocytes is related with their size in normal rat liver but not in cirrhotic one.

PubMed

Bezborodkina, Natalia N; Chestnova, Anna Yu; Vorobev, Mikhail L; Kudryavtsev, Boris N

2016-04-01

Hepatocytes differ from one another by the degree of the ploidy, size, position in the liver lobule, and level of the DNA-synthetic processes. It is believed, that the cell size exerts substantial influence on the metabolism of the hepatocytes and the glycogen content in them. The aim of the present study was to test this hypothesis. Dry weight of hepatocytes, their ploidy and glycogen content were determined in the normal and the cirrhotic rat liver. Liver cirrhosis in rats was produced by chronic inhalation of CCl4 vapours in the course of 6 months. A combined cytophotometric method was used. Dry weight of the cell, its glycogen and DNA content were successively measured on a mapped preparation. Hepatocytes of each ploidy class in the normal and the cirrhotic rat liver accumulated glycogen at the same rate. In the normal liver, there was a distinct correlation between the size of hepatocytes and glycogen content in them. This correlation was observed in each ploidy class, and was especially pronounced in the class of mononucleate tetraploid hepatocytes. In the cirrhotic liver, there was no correlation between the size of the cells and their glycogen content. The impairment of liver lobular structure probably explains the observed lack of correlation between hepatocyte size and their glycogen content in the cirrhotic liver. © 2016 International Society for Advancement of Cytometry. © 2016 International Society for Advancement of Cytometry.

Molecular recognition of microbial lipid-based antigens by T cells.

PubMed

Gras, Stephanie; Van Rhijn, Ildiko; Shahine, Adam; Le Nours, Jérôme

2018-05-01

The immune system has evolved to protect hosts from pathogens. T cells represent a critical component of the immune system by their engagement in host defence mechanisms against microbial infections. Our knowledge of the molecular recognition by T cells of pathogen-derived peptidic antigens that are presented by the major histocompatibility complex glycoproteins is now well established. However, lipids represent an additional, distinct chemical class of molecules that when presented by the family of CD1 antigen-presenting molecules can serve as antigens, and be recognized by specialized subsets of T cells leading to antigen-specific activation. Over the past decades, numerous CD1-presented self- and bacterial lipid-based antigens have been isolated and characterized. However, our understanding at the molecular level of T cell immunity to CD1 molecules presenting microbial lipid-based antigens is still largely unexplored. Here, we review the insights and the molecular basis underpinning the recognition of microbial lipid-based antigens by T cells.

Optimizing Linked Perceptual Class Formation and Transfer of Function

ERIC Educational Resources Information Center

Fields, Lanny; Garruto, Michelle

2009-01-01

A linked perceptual class consists of two distinct perceptual classes, A' and B', the members of which have become related to each other. For example, a linked perceptual class might be composed of many pictures of a woman (one perceptual class) and the sounds of that woman's voice (the other perceptual class). In this case, any sound of the…

Maternal Milk T Cells Drive Development of Transgenerational Th1 Immunity in Offspring Thymus.

PubMed

Ghosh, Mrinal K; Nguyen, Virginia; Muller, H Konrad; Walker, Ameae M

2016-09-15

Using multiple murine foster-nursing protocols, thereby eliminating placental transfer and allowing a distinction between dam- and pup-derived cells, we show that foster nursing by an immunized dam results in development of CD8(+) T cells in nonimmunized foster pups that are specific for Ags against which the foster dam was immunized (Mycobacterium tuberculosis or Candida albicans). We have dubbed this process "maternal educational immunity" to distinguish it from passive cellular immunity. Of the variety of maternal immune cells present in milk, only T cells were detected in pup tissues. Maternal T cells, a substantial percentage of which were CD4(+)MHC class II(+), accumulated in the pup thymus and spleen during the nursing period. Further analysis of maternal cells in the pup thymus showed that a proportion was positive for maternal immunogen-specific MHC class II tetramers. To determine the outcome of Ag presentation in the thymus, the maternal or foster pup origin of immunogen-responding CD8(+) cells in foster pup spleens was assessed. Whereas ∼10% were maternally derived in the first few weeks after weaning, all immunogen-responding CD8(+) T cells were pup derived by 12 wk of age. Pup-derived immunogen-responsive CD8(+) cells persisted until at least 1 y of age. Passive cellular immunity is well accepted and has been demonstrated in the human population. In this study, we show an arguably more important role for transferred immune cells: the direction of offspring T cell development. Harnessing maternal educational immunity through prepregnancy immunization programs has potential for improvement of infant immunity. Copyright © 2016 by The American Association of Immunologists, Inc.

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

Wei, Yifeng; Li, Bin; Prakash, Divya

Two subtypes of class III anaerobic ribonucleotide reductases (RNRs) studied so far couple the reduction of ribonucleotides to the oxidation of formate, or the oxidation of NADPH via thioredoxin and thioredoxin reductase. Certain methanogenic archaea contain a phylogenetically distinct third subtype of class III RNR, with distinct active-site residues. Here we report the cloning and recombinant expression of the Methanosarcina barkeri class III RNR and show that the electrons required for ribonucleotide reduction can be delivered by a [4Fe-4S] protein ferredoxin disulfide reductase, and a conserved thioredoxin-like protein NrdH present in the RNR operon. The diversity of class III RNRsmore » reflects the diversity of electron carriers used in anaerobic metabolism« less

Parabrachial complex links pain transmission to descending pain modulation.

PubMed

Roeder, Zachary; Chen, QiLiang; Davis, Sophia; Carlson, Jonathan D; Tupone, Domenico; Heinricher, Mary M

2016-12-01

The rostral ventromedial medulla (RVM) has a well-documented role in pain modulation and exerts antinociceptive and pronociceptive influences mediated by 2 distinct classes of neurons, OFF-cells and ON-cells. OFF-cells are defined by a sudden pause in firing in response to nociceptive inputs, whereas ON-cells are characterized by a "burst" of activity. Although these reflex-related changes in ON- and OFF-cell firing are critical to their pain-modulating function, the pathways mediating these responses have not been identified. The present experiments were designed to test the hypothesis that nociceptive input to the RVM is relayed through the parabrachial complex (PB). In electrophysiological studies, ON- and OFF-cells were recorded in the RVM of lightly anesthetized male rats before and after an infusion of lidocaine or muscimol into PB. The ON-cell burst and OFF-cell pause evoked by noxious heat or mechanical probing were substantially attenuated by inactivation of the lateral, but not medial, parabrachial area. Retrograde tracing studies showed that neurons projecting to the RVM were scattered throughout PB. Few of these neurons expressed calcitonin gene-related peptide, suggesting that the RVM projection from PB is distinct from that to the amygdala. These data show that a substantial component of "bottom-up" nociceptive drive to RVM pain-modulating neurons is relayed through the PB. While the PB is well known as an important relay for ascending nociceptive information, its functional connection with the RVM allows the spinoparabrachial pathway to access descending control systems as part of a recurrent circuit.

Diverse environmental stresses elicit distinct responses at the level of pre-mRNA processing in yeast

PubMed Central

Bergkessel, Megan; Whitworth, Gregg B.; Guthrie, Christine

2011-01-01

Gene expression in eukaryotic cells is profoundly influenced by the post-transcriptional processing of mRNAs, including the splicing of introns in the nucleus and both nuclear and cytoplasmic degradation pathways. These processes have the potential to affect both the steady-state levels and the kinetics of changes to levels of intron-containing transcripts. Here we report the use of a splicing isoform-specific microarray platform to investigate the effects of diverse stress conditions on pre-mRNA processing. Interestingly, we find that diverse stresses cause distinct patterns of changes at this level. The responses we observed are most dramatic for the RPGs and can be categorized into three major classes. The first is characterized by accumulation of RPG pre-mRNA and is seen in multiple types of amino acid starvation regimes; the magnitude of splicing inhibition correlates with the severity of the stress. The second class is characterized by a rapid decrease in both pre- and mature RPG mRNA and is seen in many stresses that inactivate the TORC1 kinase complex. These decreases depend on nuclear turnover of the intron-containing pre-RNAs. The third class is characterized by a decrease in RPG pre-mRNA, with only a modest reduction in the mature species; this response is observed in hyperosmotic and cation-toxic stresses. We show that casein kinase 2 (CK2) makes important contributions to the changes in pre-mRNA processing, particularly for the first two classes of stress responses. In total, our data suggest that complex post-transcriptional programs cooperate to fine-tune expression of intron-containing transcripts in budding yeast. PMID:21697354

Diverse environmental stresses elicit distinct responses at the level of pre-mRNA processing in yeast.

PubMed

Bergkessel, Megan; Whitworth, Gregg B; Guthrie, Christine

2011-08-01

Gene expression in eukaryotic cells is profoundly influenced by the post-transcriptional processing of mRNAs, including the splicing of introns in the nucleus and both nuclear and cytoplasmic degradation pathways. These processes have the potential to affect both the steady-state levels and the kinetics of changes to levels of intron-containing transcripts. Here we report the use of a splicing isoform-specific microarray platform to investigate the effects of diverse stress conditions on pre-mRNA processing. Interestingly, we find that diverse stresses cause distinct patterns of changes at this level. The responses we observed are most dramatic for the RPGs and can be categorized into three major classes. The first is characterized by accumulation of RPG pre-mRNA and is seen in multiple types of amino acid starvation regimes; the magnitude of splicing inhibition correlates with the severity of the stress. The second class is characterized by a rapid decrease in both pre- and mature RPG mRNA and is seen in many stresses that inactivate the TORC1 kinase complex. These decreases depend on nuclear turnover of the intron-containing pre-RNAs. The third class is characterized by a decrease in RPG pre-mRNA, with only a modest reduction in the mature species; this response is observed in hyperosmotic and cation-toxic stresses. We show that casein kinase 2 (CK2) makes important contributions to the changes in pre-mRNA processing, particularly for the first two classes of stress responses. In total, our data suggest that complex post-transcriptional programs cooperate to fine-tune expression of intron-containing transcripts in budding yeast.

Transcriptional profiling at whole population and single cell levels reveals somatosensory neuron molecular diversity

PubMed Central

Chiu, Isaac M; Barrett, Lee B; Williams, Erika K; Strochlic, David E; Lee, Seungkyu; Weyer, Andy D; Lou, Shan; Bryman, Gregory S; Roberson, David P; Ghasemlou, Nader; Piccoli, Cara; Ahat, Ezgi; Wang, Victor; Cobos, Enrique J; Stucky, Cheryl L; Ma, Qiufu; Liberles, Stephen D; Woolf, Clifford J

2014-01-01

The somatosensory nervous system is critical for the organism's ability to respond to mechanical, thermal, and nociceptive stimuli. Somatosensory neurons are functionally and anatomically diverse but their molecular profiles are not well-defined. Here, we used transcriptional profiling to analyze the detailed molecular signatures of dorsal root ganglion (DRG) sensory neurons. We used two mouse reporter lines and surface IB4 labeling to purify three major non-overlapping classes of neurons: 1) IB4+SNS-Cre/TdTomato+, 2) IB4−SNS-Cre/TdTomato+, and 3) Parv-Cre/TdTomato+ cells, encompassing the majority of nociceptive, pruriceptive, and proprioceptive neurons. These neurons displayed distinct expression patterns of ion channels, transcription factors, and GPCRs. Highly parallel qRT-PCR analysis of 334 single neurons selected by membership of the three populations demonstrated further diversity, with unbiased clustering analysis identifying six distinct subgroups. These data significantly increase our knowledge of the molecular identities of known DRG populations and uncover potentially novel subsets, revealing the complexity and diversity of those neurons underlying somatosensation. DOI: http://dx.doi.org/10.7554/eLife.04660.001 PMID:25525749

A widespread class of reverse transcriptase-related cellular genes.

PubMed

Gladyshev, Eugene A; Arkhipova, Irina R

2011-12-20

Reverse transcriptases (RTs) polymerize DNA on RNA templates. They fall into several structurally related but distinct classes and form an assemblage of RT-like enzymes that, in addition to RTs, also includes certain viral RNA-dependent RNA polymerases (RdRP) synthesizing RNA on RNA templates. It is generally believed that most RT-like enzymes originate from retrotransposons or viruses and have no specific function in the host cell, with telomerases being the only notable exception. Here we report on the discovery and properties of a unique class of RT-related cellular genes collectively named rvt. We present evidence that rvts are not components of retrotransposons or viruses, but single-copy genes with a characteristic domain structure that may contain introns in evolutionarily conserved positions, occur in syntenic regions, and evolve under purifying selection. These genes can be found in all major taxonomic groups including protists, fungi, animals, plants, and even bacteria, although they exhibit patchy phylogenetic distribution in each kingdom. We also show that the RVT protein purified from one of its natural hosts, Neurospora crassa, exists in a multimeric form and has the ability to polymerize NTPs as well as dNTPs in vitro, with a strong preference for NTPs, using Mn(2+) as a cofactor. The existence of a previously unknown class of single-copy RT-related genes calls for reevaluation of the current views on evolution and functional roles of RNA-dependent polymerases in living cells.

Analyses of Dynein Heavy Chain Mutations Reveal Complex Interactions Between Dynein Motor Domains and Cellular Dynein Functions

PubMed Central

Sivagurunathan, Senthilkumar; Schnittker, Robert R.; Razafsky, David S.; Nandini, Swaran; Plamann, Michael D.; King, Stephen J.

2012-01-01

Cytoplasmic dynein transports cargoes for a variety of crucial cellular functions. However, since dynein is essential in most eukaryotic organisms, the in-depth study of the cellular function of dynein via genetic analysis of dynein mutations has not been practical. Here, we identify and characterize 34 different dynein heavy chain mutations using a genetic screen of the ascomycete fungus Neurospora crassa, in which dynein is nonessential. Interestingly, our studies show that these mutations segregate into five different classes based on the in vivo localization of the mutated dynein motors. Furthermore, we have determined that the different classes of dynein mutations alter vesicle trafficking, microtubule organization, and nuclear distribution in distinct ways and require dynactin to different extents. In addition, biochemical analyses of dynein from one mutant strain show a strong correlation between its in vitro biochemical properties and the aberrant intracellular function of that altered dynein. When the mutations were mapped to the published dynein crystal structure, we found that the three-dimensional structural locations of the heavy chain mutations were linked to particular classes of altered dynein functions observed in cells. Together, our data indicate that the five classes of dynein mutations represent the entrapment of dynein at five separate points in the dynein mechanochemical and transport cycles. We have developed N. crassa as a model system where we can dissect the complexities of dynein structure, function, and interaction with other proteins with genetic, biochemical, and cell biological studies. PMID:22649085

Interstitial Cells: Regulators of Smooth Muscle Function

PubMed Central

Sanders, Kenton M.; Ward, Sean M.; Koh, Sang Don

2014-01-01

Smooth muscles are complex tissues containing a variety of cells in addition to muscle cells. Interstitial cells of mesenchymal origin interact with and form electrical connectivity with smooth muscle cells in many organs, and these cells provide important regulatory functions. For example, in the gastrointestinal tract, interstitial cells of Cajal (ICC) and PDGFRα+ cells have been described, in detail, and represent distinct classes of cells with unique ultrastructure, molecular phenotypes, and functions. Smooth muscle cells are electrically coupled to ICC and PDGFRα+ cells, forming an integrated unit called the SIP syncytium. SIP cells express a variety of receptors and ion channels, and conductance changes in any type of SIP cell affect the excitability and responses of the syncytium. SIP cells are known to provide pacemaker activity, propagation pathways for slow waves, transduction of inputs from motor neurons, and mechanosensitivity. Loss of interstitial cells has been associated with motor disorders of the gut. Interstitial cells are also found in a variety of other smooth muscles; however, in most cases, the physiological and pathophysiological roles for these cells have not been clearly defined. This review describes structural, functional, and molecular features of interstitial cells and discusses their contributions in determining the behaviors of smooth muscle tissues. PMID:24987007

A pathway of costimulation that prevents anergy in CD28- T cells: B7- independent costimulation of CD1-restricted T cells

PubMed Central

1995-01-01

A class of molecules that is expressed on antigen presenting cells, exemplified by CD80 (B7), has been found to provide a necessary costimulatory signal for T cell activation and proliferation. CD28 and CTLA4 are the B7 counterreceptors and are expressed on the majority of human CD4+ T cells and many CD8+ T cells. The signal these molecules mediate is distinguished from other costimulatory signals by the finding that T cell recognition of antigen results in a prolonged state of T cell unresponsiveness or anergy, unless these costimulatory molecules are engaged. However, nearly half of the CD8+ and CD4-CD8- T cells lack CD28, and the costimulatory signals required for the activation of such cells are unknown. To understand the pathways of activation used by CD28- T cells, we have examined the costimulatory requirements of antigen-specific CD4-CD8- TCR(+)-alpha/beta circulating T cells that lack the expression of CD28. We have characterized two T cell lines, DN1 and DN6, that recognize a mycobacterial antigen, and are restricted not by major histocompatibility complex class I or II, but by CD1b or CD1c, two members of a family of major histocompatibility complex-related molecules that have been recently implicated in a distinct pathway for antigen presentation. Comparison of antigen-specific cytolytic responses of the DN1 and DN6 T cell lines against antigen-pulsed CD1+ monocytes or CD1+ B lymphoblastoid cell lines (B-LCL) demonstrated that these T cells recognized antigen presented by both types of cells. However, T cell proliferation occurred only when antigen was presented by CD1+ monocytes, indicating that the CD1+ monocytes expressed a costimulatory molecule that the B- LCL transfectants lacked. This hypothesis was confirmed by demonstrating that the T cells became anergic when incubated with the CD1(+)-transfected B-LCL in the presence of antigen, but not in the absence of antigen. The required costimulatory signal occurred by a CD28-independent mechanism since both the CD1+ monocytes and CD1+ B-LCL transfectants expressed B7-1 and B7-2, and DN1 and DN6 lacked surface expression of CD28. We propose that these data define a previously unrecognized pathway of costimulation for T cells distinct from that involving CD28 and its counterreceptors. We suggest that this B7- independent pathway plays a crucial role in the activation and maintenance of tolerance of at least a subset of CD28- T cells. PMID:7500046

Identification and staining of distinct populations of secretory organelles in astrocytes.

PubMed

Bezzi, Paola; Volterra, Andrea

2014-05-01

Increasing evidence indicates that astrocytes, the most abundant glial cell type in the brain, respond to an elevation in cytoplasmic calcium concentration ([Ca(2+)]i) by releasing chemical transmitters (also called gliotransmitters) via regulated exocytosis of heterogeneous classes of organelles. By this process, astrocytes exert modulatory influences on neighboring cells and are thought to participate in the control of synaptic circuits and cerebral blood flow. Studying the properties of exocytosis in astrocytes is a challenge, because the cell biological basis of this process is incompletely defined. Astrocytic exocytosis involves multiple populations of secretory vesicles, including synaptic-like microvesicles (SLMVs), dense-core granules (DCGs), and lysosomes. Here we summarize the available information for identifying individual populations of secretory organelles in astrocytes, including DCGs, SLMVs, and lysosomes, and present experimental procedures for specifically staining such populations.

Common and distinct DNA-binding and regulatory activities of the BEN-solo transcription factor family.

PubMed

Dai, Qi; Ren, Aiming; Westholm, Jakub O; Duan, Hong; Patel, Dinshaw J; Lai, Eric C

2015-01-01

Recently, the BEN (BANP, E5R, and NAC1) domain was recognized as a new class of conserved DNA-binding domain. The fly genome encodes three proteins that bear only a single BEN domain ("BEN-solo" factors); namely, Insensitive (Insv), Bsg25A (Elba1), and CG9883 (Elba2). Insv homodimers preferentially bind CCAATTGG palindromes throughout the genome to mediate transcriptional repression, whereas Bsg25A and Elba2 heterotrimerize with their obligate adaptor, Elba3 (i.e., the ELBA complex), to recognize a CCAATAAG motif in the Fab-7 insulator. While these data suggest distinct DNA-binding properties of BEN-solo proteins, we performed reporter assays that indicate that both Bsg25A and Elba2 can individually recognize Insv consensus sites efficiently. We confirmed this by solving the structure of Bsg25A complexed to the Insv site, which showed that key aspects of the BEN:DNA recognition strategy are similar between these proteins. We next show that both Insv and ELBA proteins are competent to mediate transcriptional repression via Insv consensus sequences but that the ELBA complex appears to be selective for the ELBA site. Reciprocally, genome-wide analysis reveals that Insv exhibits significant cobinding to class I insulator elements, indicating that it may also contribute to insulator function. Indeed, we observed abundant Insv binding within the Hox complexes with substantial overlaps with class I insulators, many of which bear Insv consensus sites. Moreover, Insv coimmunoprecipitates with the class I insulator factor CP190. Finally, we observed that Insv harbors exclusive activity among fly BEN-solo factors with respect to regulation of Notch-mediated cell fate choices in the peripheral nervous system. This in vivo activity is recapitulated by BEND6, a mammalian BEN-solo factor that conserves the Notch corepressor function of Insv but not its capacity to bind Insv consensus sites. Altogether, our data define an array of common and distinct biochemical and functional properties of this new family of transcription factors. © 2015 Dai et al.; Published by Cold Spring Harbor Laboratory Press.

Common and distinct DNA-binding and regulatory activities of the BEN-solo transcription factor family

PubMed Central

Dai, Qi; Ren, Aiming; Westholm, Jakub O.; Duan, Hong; Patel, Dinshaw J.

2015-01-01

Recently, the BEN (BANP, E5R, and NAC1) domain was recognized as a new class of conserved DNA-binding domain. The fly genome encodes three proteins that bear only a single BEN domain (“BEN-solo” factors); namely, Insensitive (Insv), Bsg25A (Elba1), and CG9883 (Elba2). Insv homodimers preferentially bind CCAATTGG palindromes throughout the genome to mediate transcriptional repression, whereas Bsg25A and Elba2 heterotrimerize with their obligate adaptor, Elba3 (i.e., the ELBA complex), to recognize a CCAATAAG motif in the Fab-7 insulator. While these data suggest distinct DNA-binding properties of BEN-solo proteins, we performed reporter assays that indicate that both Bsg25A and Elba2 can individually recognize Insv consensus sites efficiently. We confirmed this by solving the structure of Bsg25A complexed to the Insv site, which showed that key aspects of the BEN:DNA recognition strategy are similar between these proteins. We next show that both Insv and ELBA proteins are competent to mediate transcriptional repression via Insv consensus sequences but that the ELBA complex appears to be selective for the ELBA site. Reciprocally, genome-wide analysis reveals that Insv exhibits significant cobinding to class I insulator elements, indicating that it may also contribute to insulator function. Indeed, we observed abundant Insv binding within the Hox complexes with substantial overlaps with class I insulators, many of which bear Insv consensus sites. Moreover, Insv coimmunoprecipitates with the class I insulator factor CP190. Finally, we observed that Insv harbors exclusive activity among fly BEN-solo factors with respect to regulation of Notch-mediated cell fate choices in the peripheral nervous system. This in vivo activity is recapitulated by BEND6, a mammalian BEN-solo factor that conserves the Notch corepressor function of Insv but not its capacity to bind Insv consensus sites. Altogether, our data define an array of common and distinct biochemical and functional properties of this new family of transcription factors. PMID:25561495

Phase transitions in a model for the formation of herpes simplex ulcers

NASA Astrophysics Data System (ADS)

Ferreira, Claudia P.; Fontanari, J. F.; Zorzenon Dos Santos, Rita M.

2001-10-01

The critical properties of a cellular automaton model describing the spreading of infection of the herpes simplex virus in corneal tissue are investigated through the dynamic Monte Carlo method. The model takes into account different cell susceptibilities to the viral infection, as suggested by experimental findings. In a two-dimensional square lattice the sites are associated with two distinct types of cells, namely, permissive and resistant to the infection. While a permissive cell becomes infected in the presence of a single infected cell in its neighborhood, a resistant cell needs to be surrounded by at least R>1 infected or dead cells in order to become infected. The infection is followed by the death of the cells resulting in ulcers whose forms may be dendritic (self-limited clusters) or amoeboid (percolating clusters) depending on the degree of resistance R of the resistant cells as well as on the density of permissive cells in the healthy tissue. We show that a phase transition between these two regimes occurs only for R>=5 and, in addition, that the phase transition is in the universality class of the ordinary percolation.

A taxonomy of bacterial microcompartment loci constructed by a novel scoring method

DOE PAGES

Axen, Seth D.; Erbilgin, Onur; Kerfeld, Cheryl A.; ...

2014-10-23

Bacterial microcompartments (BMCs) are proteinaceous organelles involved in both autotrophic and heterotrophic metabolism. All BMCs share homologous shell proteins but differ in their complement of enzymes; these are typically encoded adjacent to shell protein genes in genetic loci, or operons. To enable the identification and prediction of functional (sub)types of BMCs, we developed LoClass, an algorithm that finds putative BMC loci and inventories, weights, and compares their constituent pfam domains to construct a locus similarity network and predict locus (sub)types. In addition to using LoClass to analyze sequences in the Non-redundant Protein Database, we compared predicted BMC loci found inmore » seven candidate bacterial phyla (six from single-cell genomic studies) to the LoClass taxonomy. Together, these analyses resulted in the identification of 23 different types of BMCs encoded in 30 distinct locus (sub)types found in 23 bacterial phyla. These include the two carboxysome types and a divergent set of metabolosomes, BMCs that share a common catalytic core and process distinct substrates via specific signature enzymes. Furthermore, many Candidate BMCs were found that lack one or more core metabolosome components, including one that is predicted to represent an entirely new paradigm for BMC-associated metabolism, joining the carboxysome and metabolosome. By placing these results in a phylogenetic context, we provide a framework for understanding the horizontal transfer of these loci, a starting point for studies aimed at understanding the evolution of BMCs. This comprehensive taxonomy of BMC loci, based on their constituent protein domains, foregrounds the functional diversity of BMCs and provides a reference for interpreting the role of BMC gene clusters encoded in isolate, single cell, and metagenomic data. Many loci encode ancillary functions such as transporters or genes for cofactor assembly; this expanded vocabulary of BMC-related functions should be useful for design of genetic modules for introducing BMCs in bioengineering applications.« less

A Taxonomy of Bacterial Microcompartment Loci Constructed by a Novel Scoring Method

PubMed Central

Kerfeld, Cheryl A.

2014-01-01

Bacterial microcompartments (BMCs) are proteinaceous organelles involved in both autotrophic and heterotrophic metabolism. All BMCs share homologous shell proteins but differ in their complement of enzymes; these are typically encoded adjacent to shell protein genes in genetic loci, or operons. To enable the identification and prediction of functional (sub)types of BMCs, we developed LoClass, an algorithm that finds putative BMC loci and inventories, weights, and compares their constituent pfam domains to construct a locus similarity network and predict locus (sub)types. In addition to using LoClass to analyze sequences in the Non-redundant Protein Database, we compared predicted BMC loci found in seven candidate bacterial phyla (six from single-cell genomic studies) to the LoClass taxonomy. Together, these analyses resulted in the identification of 23 different types of BMCs encoded in 30 distinct locus (sub)types found in 23 bacterial phyla. These include the two carboxysome types and a divergent set of metabolosomes, BMCs that share a common catalytic core and process distinct substrates via specific signature enzymes. Furthermore, many Candidate BMCs were found that lack one or more core metabolosome components, including one that is predicted to represent an entirely new paradigm for BMC-associated metabolism, joining the carboxysome and metabolosome. By placing these results in a phylogenetic context, we provide a framework for understanding the horizontal transfer of these loci, a starting point for studies aimed at understanding the evolution of BMCs. This comprehensive taxonomy of BMC loci, based on their constituent protein domains, foregrounds the functional diversity of BMCs and provides a reference for interpreting the role of BMC gene clusters encoded in isolate, single cell, and metagenomic data. Many loci encode ancillary functions such as transporters or genes for cofactor assembly; this expanded vocabulary of BMC-related functions should be useful for design of genetic modules for introducing BMCs in bioengineering applications. PMID:25340524

Adjuvants in the Driver’s Seat: How Magnitude, Type, Fine Specificity and Longevity of Immune Responses Are Driven by Distinct Classes of Immune Potentiators

PubMed Central

Bergmann-Leitner, Elke S.; Leitner, Wolfgang W.

2014-01-01

The mechanism by which vaccine adjuvants enhance immune responses has historically been considered to be the creation of an antigen depot. From here, the antigen is slowly released and provided to immune cells over an extended period of time. This “depot” was formed by associating the antigen with substances able to persist at the injection site, such as aluminum salts or emulsions. The identification of Pathogen-Associated Molecular Patterns (PAMPs) has greatly advanced our understanding of how adjuvants work beyond the simple concept of extended antigen release and has accelerated the development of novel adjuvants. This review focuses on the mode of action of different adjuvant classes in regards to the stimulation of specific immune cell subsets, the biasing of immune responses towards cellular or humoral immune response, the ability to mediate epitope spreading and the induction of persistent immunological memory. A better understanding of how particular adjuvants mediate their biological effects will eventually allow them to be selected for specific vaccines in a targeted and rational manner. PMID:26344620

Superparamagnetic iron oxide nanoparticles for MRI: contrast media pharmaceutical company R&D perspective.

PubMed

Corot, Claire; Warlin, David

2013-01-01

Superparamagnetic iron oxide (SPIO) nanoparticles are a relatively large class of contrast agents for magnetic resonance imaging. According to their biodistribution, distinct classes of SPIO nanoparticles have been investigated for clinical applications either as macrophage imaging agents or blood pool agents. Contrast agents which are pharmaceutics followed the same development rules as therapeutic drugs. Several drawbacks such as clinical development difficulties, organization of market access and imaging technological developments have limited the widespread use of these products. SPIO nanoparticles that are composed of thousands iron atoms providing large T2* effects are particularly suitable for theranostic. Stem cell migration and immune cell trafficking, as well as targeted SPIO nanoparticles for molecular imaging studies are mainly at the stage of proof of concept. A major economic challenge in the development of molecular imaging associated with a therapeutic treatment/procedure is to define innovative business models compatible with the needs of all players taking into account that theranostic solutions are promising to optimize resource allocation and ensure that expensive treatments are prescribed to responding patients. © 2013 Wiley Periodicals, Inc.

Genome-wide Analysis of the H3K4 Histone Demethylase RBP2 Reveals a Transcriptional Program Controlling Differentiation

PubMed Central

Lopez-Bigas, Nuria; Kisiel, Tomasz A.; DeWaal, Dannielle C.; Holmes, Katie B.; Volkert, Tom L.; Gupta, Sumeet; Love, Jennifer; Murray, Heather L.; Young, Richard A.; Benevolenskaya, Elizaveta V.

2010-01-01

SUMMARY Retinoblastoma protein (pRB) mediates cell-cycle withdrawal and differentiation by interacting with a variety of proteins. RB-Binding Protein 2 (RBP2) has been shown to be a key effector. We sought to determine transcriptional regulation by RBP2 genome-wide by using location analysis and gene expression profiling experiments. We describe that RBP2 shows high correlation with the presence of H3K4me3 and its target genes are separated into two functionally distinct classes: differentiation-independent and differentiation-dependent genes. The former class is enriched by genes that encode mitochondrial proteins, while the latter is represented by cell-cycle genes. We demonstrate the role of RBP2 in mitochondrial biogenesis, which involves regulation of H3K4me3-modified nucleosomes. Analysis of expression changes upon RBP2 depletion depicted genes with a signature of differentiation control, analogous to the changes seen upon reintroduction of pRB. We conclude that, during differentiation, RBP2 exerts inhibitory effects on multiple genes through direct interaction with their promoters. PMID:18722178

miR-1915 and miR-1225-5p Regulate the Expression of CD133, PAX2 and TLR2 in Adult Renal Progenitor Cells

PubMed Central

Costantino, Vincenzo; Curci, Claudia; Cox, Sharon N.; De Palma, Giuseppe; Schena, Francesco P.

2013-01-01

Adult renal progenitor cells (ARPCs) were recently identified in the cortex of the renal parenchyma and it was demonstrated that they were positive for PAX2, CD133, CD24 and exhibited multipotent differentiation ability. Recent studies on stem cells indicated that microRNAs (miRNAs), a class of noncoding small RNAs that participate in the regulation of gene expression, may play a key role in stem cell self-renewal and differentiation. Distinct sets of miRNAs are specifically expressed in pluripotent stem cells but not in adult tissues, suggesting a role for miRNAs in stem cell self-renewal. We compared miRNA expression profiles of ARPCs with that of mesenchymal stem cells (MSCs) and renal proximal tubular cells (RPTECs) finding distinct sets of miRNAs that were specifically expressed in ARPCs. In particular, miR-1915 and miR-1225-5p regulated the expression of important markers of renal progenitors, such as CD133 and PAX2, and important genes involved in the repair mechanisms of ARPCs, such as TLR2. We demonstrated that the expression of both the renal stem cell markers CD133 and PAX2 depends on lower miR-1915 levels and that the increase of miR-1915 levels improved capacity of ARPCs to differentiate into adipocyte-like and epithelial-like cells. Finally, we found that the low levels of miR-1225-5p were responsible for high TLR2 expression in ARPCs. Therefore, together, miR-1915 and miR-1225-5p seem to regulate important traits of renal progenitors: the stemness and the repair capacity. PMID:23861881

How unfinished business from S-phase affects mitosis and beyond

PubMed Central

Mankouri, Hocine W; Huttner, Diana; Hickson, Ian D

2013-01-01

The eukaryotic cell cycle is conventionally viewed as comprising several discrete steps, each of which must be completed before the next one is initiated. However, emerging evidence suggests that incompletely replicated, or unresolved, chromosomes from S-phase can persist into mitosis, where they present a potential threat to the faithful segregation of sister chromatids. In this review, we provide an overview of the different classes of loci where this ‘unfinished S-phase business' can lead to a variety of cytogenetically distinct DNA structures throughout the various steps of mitosis. Furthermore, we discuss the potential ways in which cells might not only tolerate this inevitable aspect of chromosome biology, but also exploit it to assist in the maintenance of genome stability. PMID:24065128

How unfinished business from S-phase affects mitosis and beyond.

PubMed

Mankouri, Hocine W; Huttner, Diana; Hickson, Ian D

2013-10-16

The eukaryotic cell cycle is conventionally viewed as comprising several discrete steps, each of which must be completed before the next one is initiated. However, emerging evidence suggests that incompletely replicated, or unresolved, chromosomes from S-phase can persist into mitosis, where they present a potential threat to the faithful segregation of sister chromatids. In this review, we provide an overview of the different classes of loci where this 'unfinished S-phase business' can lead to a variety of cytogenetically distinct DNA structures throughout the various steps of mitosis. Furthermore, we discuss the potential ways in which cells might not only tolerate this inevitable aspect of chromosome biology, but also exploit it to assist in the maintenance of genome stability.

Naive T-cell receptor transgenic T cells help memory B cells produce antibody

PubMed Central

Duffy, Darragh; Yang, Chun-Ping; Heath, Andrew; Garside, Paul; Bell, Eric B

2006-01-01

Injection of the same antigen following primary immunization induces a classic secondary response characterized by a large quantity of high-affinity antibody of an immunoglobulin G class produced more rapidly than in the initial response – the products of memory B cells are qualitatively distinct from that of the original naive B lymphocytes. Very little is known of the help provided by the CD4 T cells that stimulate memory B cells. Using antigen-specific T-cell receptor transgenic CD4 T cells (DO11.10) as a source of help, we found that naive transgenic T cells stimulated memory B cells almost as well (in terms of quantity and speed) as transgenic T cells that had been recently primed. There was a direct correlation between serum antibody levels and the number of naive transgenic T cells transferred. Using T cells from transgenic interleukin-2-deficient mice we showed that interleukin-2 was not required for a secondary response, although it was necessary for a primary response. The results suggested that the signals delivered by CD4 T cells and required by memory B cells for their activation were common to both antigen-primed and naive CD4 T cells. PMID:17067314

Fatty aldehyde dehydrogenases in Acinetobacter sp. strain HO1-N: role in hexadecanol metabolism.

PubMed Central

Singer, M E; Finnerty, W R

1985-01-01

The role of fatty aldehyde dehydrogenases (FALDHs) in hexadecane and hexadecanol metabolism was studied in Acinetobacter sp. strain HO1-N. Two distinct FALDHs were demonstrated in Acinetobacter sp. strain HO1-N: a membrane-bound, NADP-dependent FALDH activity induced 5-, 15-, and 9-fold by growth on hexadecanol, dodecyl aldehyde, and hexadecane, respectively, and a constitutive, NAD-dependent, membrane-localized FALDH. The NADP-dependent FALDH exhibited apparent Km and Vmax values for decyl aldehyde of 5.0, 13.0, 18.0, and 18.3 microM and 537.0, 500.0, 25.0, and 38.0 nmol/min in hexadecane-, hexadecanol-, ethanol-, palmitate-grown cells, respectively. FALDH isozymes ald-a, ald-b, and ald-c were demonstrated by gel electrophoresis in extracts of hexadecane- and hexadecanol-grown cells. ald-a, ald-b, and ald-d were present in dodecyl aldehyde-grown cells, while palmitate-grown control cells contained ald-b and ald-d. Dodecyl aldehyde-negative mutants were isolated and grouped into two phenotypic classes based on growth: class 1 mutants were hexadecane and hexadecanol negative and class 2 mutants were hexadecane and hexadecanol positive. Specific activity of NADP-dependent FALDH in Ald21 (class 1 mutant) was 85% lower than that of wild-type FALDH, while the specific activity of Ald24 (class 2 mutant) was 55% greater than that of wild-type FALDH. Ald21R, a dodecyl aldehyde-positive revertant able to grow on hexadecane, hexadecanol, and dodecyl aldehyde, exhibited a 100% increase in the specific activity of the NADP-dependent FALDH. The oxidation of [3H]hexadecane byAld21 yielded the accumulation of 61% more fatty aldehyde than the wild type, while Ald24 accumulated 27% more fatty aldehyde, 95% more fatty alcohol, and 65% more wax ester than the wild type. This study provides genetic and physiological evidence for the role of fatty aldehyde as an essential metabolic intermediate and NADP-dependent FALDH as a key enzyme in the dissimilation of hexadecane, hexadecanol, and dodecyl aldehyde in Acinetobactor sp. strain HO1-N. Images PMID:4066609

Changes in the Excitability of Neocortical Neurons in a Mouse Model of Amyotrophic Lateral Sclerosis Are Not Specific to Corticospinal Neurons and Are Modulated by Advancing Disease.

PubMed

Kim, Juhyun; Hughes, Ethan G; Shetty, Ashwin S; Arlotta, Paola; Goff, Loyal A; Bergles, Dwight E; Brown, Solange P

2017-09-13

Cell type-specific changes in neuronal excitability have been proposed to contribute to the selective degeneration of corticospinal neurons in amyotrophic lateral sclerosis (ALS) and to neocortical hyperexcitability, a prominent feature of both inherited and sporadic variants of the disease, but the mechanisms underlying selective loss of specific cell types in ALS are not known. We analyzed the physiological properties of distinct classes of cortical neurons in the motor cortex of hSOD1 G93A mice of both sexes and found that they all exhibit increases in intrinsic excitability that depend on disease stage. Targeted recordings and in vivo calcium imaging further revealed that neurons adapt their functional properties to normalize cortical excitability as the disease progresses. Although different neuron classes all exhibited increases in intrinsic excitability, transcriptional profiling indicated that the molecular mechanisms underlying these changes are cell type specific. The increases in excitability in both excitatory and inhibitory cortical neurons show that selective dysfunction of neuronal cell types cannot account for the specific vulnerability of corticospinal motor neurons in ALS. Furthermore, the stage-dependent alterations in neuronal function highlight the ability of cortical circuits to adapt as disease progresses. These findings show that both disease stage and cell type must be considered when developing therapeutic strategies for treating ALS. SIGNIFICANCE STATEMENT It is not known why certain classes of neurons preferentially die in different neurodegenerative diseases. It has been proposed that the enhanced excitability of affected neurons is a major contributor to their selective loss. We show using a mouse model of amyotrophic lateral sclerosis (ALS), a disease in which corticospinal neurons exhibit selective vulnerability, that changes in excitability are not restricted to this neuronal class and that excitability does not increase monotonically with disease progression. Moreover, although all neuronal cell types tested exhibited abnormal functional properties, analysis of their gene expression demonstrated cell type-specific responses to the ALS-causing mutation. These findings suggest that therapies for ALS may need to be tailored for different cell types and stages of disease. Copyright © 2017 the authors 0270-6474/17/379038-17$15.00/0.

Profiles of Community Violence Exposure Among African American Youth: An Examination of Desensitization to Violence Using Latent Class Analysis.

PubMed

Gaylord-Harden, Noni K; Dickson, Daniel; Pierre, Cynthia

2016-07-01

The current study employed latent class analysis (LCA) to identify distinct profiles of community violence exposure and their associations to desensitization outcomes in 241 African American early adolescents (M age = 12.86, SD = 1.28) in the sixth through eighth grade from under-resourced urban communities. Participants self-reported on their exposure to community violence, as well as on depressive and anxiety symptoms. The LCA revealed three distinct classes: a class exposed to low levels of violence (low exposure class), a class exposed to moderately high levels of victimization (victimization class), and a class exposed to high levels of all types of violence (high exposure class). Consistent with predictions, the high exposure class showed the lowest levels of depressive symptoms, suggesting a desensitization outcome. Gender and age were also examined in relation to the classes, and age was significantly associated with an increased risk of being a member of the high exposure class relative to the low exposure class. Using person-based analyses to examine desensitization outcomes provides useful information for prevention and intervention efforts, as it helps to identify a specific subgroup of youth that may be more likely to show desensitization outcomes in the context of community violence. © The Author(s) 2015.

Systematic Analysis of Primary Sequence Domain Segments for the Discrimination Between Class C GPCR Subtypes.

PubMed

König, Caroline; Alquézar, René; Vellido, Alfredo; Giraldo, Jesús

2018-03-01

G-protein-coupled receptors (GPCRs) are a large and diverse super-family of eukaryotic cell membrane proteins that play an important physiological role as transmitters of extracellular signal. In this paper, we investigate Class C, a member of this super-family that has attracted much attention in pharmacology. The limited knowledge about the complete 3D crystal structure of Class C receptors makes necessary the use of their primary amino acid sequences for analytical purposes. Here, we provide a systematic analysis of distinct receptor sequence segments with regard to their ability to differentiate between seven class C GPCR subtypes according to their topological location in the extracellular, transmembrane, or intracellular domains. We build on the results from the previous research that provided preliminary evidence of the potential use of separated domains of complete class C GPCR sequences as the basis for subtype classification. The use of the extracellular N-terminus domain alone was shown to result in a minor decrease in subtype discrimination in comparison with the complete sequence, despite discarding much of the sequence information. In this paper, we describe the use of Support Vector Machine-based classification models to evaluate the subtype-discriminating capacity of the specific topological sequence segments.

Crystal Structure of the PAC1R Extracellular Domain Unifies a Consensus Fold for Hormone Recognition by Class B G-Protein Coupled Receptors

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

Kumar, Shiva; Pioszak, Augen; Zhang, Chenghai

2012-02-21

Pituitary adenylate cyclase activating polypeptide (PACAP) is a member of the PACAP/glucagon family of peptide hormones, which controls many physiological functions in the immune, nervous, endocrine, and muscular systems. It activates adenylate cyclase by binding to its receptor, PAC1R, a member of class B G-protein coupled receptors (GPCR). Crystal structures of a number of Class B GPCR extracellular domains (ECD) bound to their respective peptide hormones have revealed a consensus mechanism of hormone binding. However, the mechanism of how PACAP binds to its receptor remains controversial as an NMR structure of the PAC1R ECD/PACAP complex reveals a different topology ofmore » the ECD and a distinct mode of ligand recognition. Here we report a 1.9 {angstrom} crystal structure of the PAC1R ECD, which adopts the same fold as commonly observed for other members of Class B GPCR. Binding studies and cell-based assays with alanine-scanned peptides and mutated receptor support a model that PAC1R uses the same conserved fold of Class B GPCR ECD for PACAP binding, thus unifying the consensus mechanism of hormone binding for this family of receptors.« less

Inhibitory Interneurons That Express GFP in the PrP-GFP Mouse Spinal Cord Are Morphologically Heterogeneous, Innervated by Several Classes of Primary Afferent and Include Lamina I Projection Neurons among Their Postsynaptic Targets

PubMed Central

Ganley, Robert P.; Iwagaki, Noboru; del Rio, Patricia; Baseer, Najma; Dickie, Allen C.; Boyle, Kieran A.; Polgár, Erika; Watanabe, Masahiko; Abraira, Victoria E; Zimmerman, Amanda

2015-01-01

The superficial dorsal horn of the spinal cord contains numerous inhibitory interneurons, which regulate the transmission of information perceived as touch, pain, or itch. Despite the importance of these cells, our understanding of their roles in the neuronal circuitry is limited by the difficulty in identifying functional populations. One group that has been identified and characterized consists of cells in the mouse that express green fluorescent protein (GFP) under control of the prion protein (PrP) promoter. Previous reports suggested that PrP-GFP cells belonged to a single morphological class (central cells), received inputs exclusively from unmyelinated primary afferents, and had axons that remained in lamina II. However, we recently reported that the PrP-GFP cells expressed neuronal nitric oxide synthase (nNOS) and/or galanin, and it has been shown that nNOS-expressing cells are more diverse in their morphology and synaptic connections. We therefore used a combined electrophysiological, pharmacological, and anatomical approach to reexamine the PrP-GFP cells. We provide evidence that they are morphologically diverse (corresponding to “unclassified” cells) and receive synaptic input from a variety of primary afferents, with convergence onto individual cells. We also show that their axons project into adjacent laminae and that they target putative projection neurons in lamina I. This indicates that the neuronal circuitry involving PrP-GFP cells is more complex than previously recognized, and suggests that they are likely to have several distinct roles in regulating the flow of somatosensory information through the dorsal horn. PMID:25972186

Forging T-Lymphocyte Identity: Intersecting Networks of Transcriptional Control.

PubMed

Rothenberg, Ellen V; Ungerbäck, Jonas; Champhekar, Ameya

2016-01-01

T-lymphocyte development branches off from other lymphoid developmental programs through its requirement for sustained environmental signals through the Notch pathway. In the thymus, Notch signaling induces a succession of T-lineage regulatory factors that collectively create the T-cell identity through distinct steps. This process involves both the staged activation of T-cell identity genes and the staged repression of progenitor-cell-inherited regulatory genes once their roles in self-renewal and population expansion are no longer needed. With the recent characterization of innate lymphoid cells (ILCs) that share transcriptional regulation programs extensively with T-cell subsets, T-cell identity can increasingly be seen as defined in modular terms, as the processes selecting and actuating effector function are potentially detachable from the processes generating and selecting clonally unique T-cell receptor structures. The developmental pathways of different classes of T cells and ILCs are distinguished by the numbers of prerequisites of gene rearrangement, selection, and antigen contact before the cells gain access to nearly common regulatory mechanisms for choosing effector function. Here, the major classes of transcription factors that interact with Notch signals during T-lineage specification are discussed in terms of their roles in these programs, the evidence for their spectra of target genes at different stages, and their cross-regulatory and cooperative actions with each other. Specific topics include Notch modulation of PU.1 and GATA-3, PU.1-Notch competition, the relationship between PU.1 and GATA-3, and the roles of E proteins, Bcl11b, and GATA-3 in guiding acquisition of T-cell identity while avoiding redirection to an ILC fate. © 2016 Elsevier Inc. All rights reserved.

Distinct Evening Fatigue Profiles in Oncology Outpatients Receiving Chemotherapy

PubMed Central

Wright, Fay; Cooper, Bruce A.; Conley, Yvette P.; Hammer, Marilyn J.; Chen, Lee-May; Paul, Steven M.; Levine, Jon D.; Miaskowski, Christine; Kober, Kord M.

2018-01-01

Background Fatigue is the most common and debilitating symptom experienced by oncology patients during chemotherapy (CTX). Fatigue severity demonstrates a large amount of inter-individual and diurnal variability. Purpose Study purposes were to evaluate for subgroups of patients with distinct evening fatigue profiles and evaluate how these subgroups differed on demographic, clinical, and symptom characteristics. Methods Outpatients with breast, gastrointestinal, gynecological, or lung cancer (n=1332) completed questionnaires six times over two cycles of CTX. Lee Fatigue Scale (LFS) evaluated evening fatigue severity. Latent profile analysis was used to identify distinct evening fatigue profiles. Results Four distinct evening fatigue classes (i.e., Low (14.0%), Moderate (17.2%), High (36.0%), Very High (32.8%)) were identified. Compared to the Low class, patients in the Very High evening fatigue class were: younger, female, had childcare responsibilities, had more years of education, had a lower functional status, had a higher comorbidity burden, and were diagnosed with breast cancer. Patients in the Very High class reported higher levels of depressive symptoms, sleep disturbance, and evening fatigue at enrollment. Conclusions Findings provide new insights into modifiable risk factors for higher levels of evening fatigue. Clinicians can use this information to identify higher risk patients and plan appropriate interventions. PMID:29725554

The structural and functional differentiation of hair cells in a lizard’s basilar papilla suggests an operational principle of amniote cochleas

PubMed Central

Chiappe, M. Eugenia; Kozlov, Andrei S.; Hudspeth, A. J.

2007-01-01

The hair cells in the mammalian cochlea are of two distinct types. Inner hair cells are responsible for transducing mechanical stimuli into electrical responses, which they forward to the brain through a copious afferent innervation. Outer hair cells, which are thought to mediate the active process that sensitizes and tunes the cochlea, possess a negligible afferent innervation. For every inner hair cell there are approximately three outer hair cells, so only a quarter of the hair cells directly deliver information to the central nervous system. Although this is a surprising feature for a sensory system, the occurrence of a similar innervation pattern in birds and crocodilians suggests that the arrangement has an adaptive value. Using a lizard with highly developed hearing, the tokay gecko, we demonstrate in the present study that the same principle operates in a third major group of terrestrial animals. We propose that the differentiation of hair cells into signaling and amplifying classes reflects incompatible strategies for the optimization of mechanoelectrical transduction and of an active process based on active hair-bundle motility. PMID:17978038

Potential antiproliferative activity of polyphenol metabolites against human breast cancer cells and their urine excretion pattern in healthy subjects following acute intake of a polyphenol-rich juice of grumixama (Eugenia brasiliensis Lam.).

PubMed

Teixeira, L L; Costa, G R; Dörr, F A; Ong, T P; Pinto, E; Lajolo, F M; Hassimotto, N M A

2017-06-21

The bioavailability and metabolism of anthocyanins and ellagitannins following acute intake of grumixama fruit, native Brazilian cherry, by humans, and its in vitro antiproliferative activity against breast cancer cells (MDA-MB-231) were investigated. A single dose of grumixama juice was administered to healthy women (n = 10) and polyphenol metabolites were analyzed in urine and plasma samples collected over 24 h. The majority of the metabolites circulating and excreted in urine were phenolic acids and urolithin conjugates, the gut microbiota catabolites of both classes of polyphenols, respectively. According to pharmacokinetic parameters, the subjects were divided into two distinct groups, high and low urinary metabolite excretors. The pool of polyphenol metabolites found in urine samples showed a significant inhibition of cell proliferation and G2/M cell cycle arrest in MDA-MB-231 cells. Our findings demonstrate the large interindividual variability concerning the polyphenol metabolism, which possibly could reflect in health promotion.

Herpesvirus Entry into Host Cells Mediated by Endosomal Low pH.

PubMed

Nicola, Anthony V

2016-09-01

Herpesviral pathogenesis stems from infection of multiple cell types including the site of latency and cells that support lytic replication. Herpesviruses utilize distinct cellular pathways, including low pH endocytic pathways, to enter different pathophysiologically relevant target cells. This review details the impact of the mildly acidic milieu of endosomes on the entry of herpesviruses, with particular emphasis on herpes simplex virus 1 (HSV-1). Epithelial cells, the portal of primary HSV-1 infection, support entry via low pH endocytosis mechanisms. Mildly acidic pH triggers reversible conformational changes in the HSV-1 class III fusion protein glycoprotein B (gB). In vitro treatment of herpes simplex virions with a similar pH range inactivates infectivity, likely by prematurely activating the viral entry machinery in the absence of a target membrane. How a given herpesvirus mediates both low pH and pH-independent entry events is a key unresolved question. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

The informational architecture of the cell.

PubMed

Walker, Sara Imari; Kim, Hyunju; Davies, Paul C W

2016-03-13

We compare the informational architecture of biological and random networks to identify informational features that may distinguish biological networks from random. The study presented here focuses on the Boolean network model for regulation of the cell cycle of the fission yeast Schizosaccharomyces pombe. We compare calculated values of local and global information measures for the fission yeast cell cycle to the same measures as applied to two different classes of random networks: Erdös-Rényi and scale-free. We report patterns in local information processing and storage that do indeed distinguish biological from random, associated with control nodes that regulate the function of the fission yeast cell-cycle network. Conversely, we find that integrated information, which serves as a global measure of 'emergent' information processing, does not differ from random for the case presented. We discuss implications for our understanding of the informational architecture of the fission yeast cell-cycle network in particular, and more generally for illuminating any distinctive physics that may be operative in life. © 2016 The Author(s).

Wealth, Poverty, and Happiness: Social Class Is Differentially Associated With Positive Emotions.

PubMed

Piff, Paul K; Moskowitz, Jake P

2017-12-18

Is higher social class associated with greater happiness? In a large nationally representative U.S. sample (N = 1,519), we examined the association between social class (household income) and self-reported tendencies to experience 7 distinct positive emotions that are core to happiness: amusement, awe, compassion, contentment, enthusiasm, love, and pride. Consistent with past research indicating that social class underlies differential patterns of attending to the self versus orienting to others, higher social class was associated with greater self-oriented feelings of contentment and pride, and with greater amusement. In contrast, lower social class was associated with more other-oriented feelings of compassion and love, and with greater awe. There were no class differences in enthusiasm. We discuss that individuals from different social class backgrounds may exhibit different patterns of emotional responding due to their distinct social concerns and priorities. Whereas self-oriented emotions may follow from, foster, and reinforce upper class individuals' desire for independence and self-sufficiency, greater other-oriented emotion may enable lower class individuals to form more interdependent bonds to cope with their more threatening environments. (PsycINFO Database Record (c) 2017 APA, all rights reserved).

KDM1 Class Flavin-Dependent Protein Lysine Demethylases

PubMed Central

Burg, Jonathan M.; Link, Jennifer E.; Morgan, Brittany S.; Heller, Frederick J.; Hargrove, Amanda E.; McCafferty, Dewey G.

2015-01-01

Flavin-dependent, lysine-specific protein demethylases (KDM1s) are a subfamily of amine oxidases that catalyze the selective posttranslational oxidative demethylation of methyllysine side chains within protein and peptide substrates. KDM1s participate in the widespread epigenetic regulation of both normal and disease state transcriptional programs. Their activities are central to various cellular functions, such as hematopoietic and neuronal differentiation, cancer proliferation and metastasis, and viral lytic replication and establishment of latency. Interestingly, KDM1s function as catalytic subunits within complexes with coregulatory molecules that modulate enzymatic activity of the demethylases and coordinate their access to specific substrates at distinct sites within the cell and chromatin. Although several classes of KDM1 -selective small molecule inhibitors have been recently developed, these pan-active site inhibition strategies lack the ability to selectively discriminate between KDM1 activity in specific, and occasionally opposing, functional contexts within these complexes. Here we review the discovery of this class of demethylases, their structures, chemical mechanisms, and specificity. Additionally, we review inhibition of this class of enzymes as well as emerging interactions with coregulatory molecules that regulate demethylase activity in highly specific functional contexts of biological and potential therapeutic importance. PMID:25787087

A novel strategy for the identification of antigens that are recognised by bovine MHC class I restricted cytotoxic T cells in a protozoan infection using reverse vaccinology.

PubMed

Graham, Simon P; Honda, Yoshikazu; Pellé, Roger; Mwangi, Duncan M; Glew, E Jane; de Villiers, Etienne P; Shah, Trushar; Bishop, Richard; van der Bruggen, Pierre; Nene, Vishvanath; Taracha, Evans L N

2007-02-09

Immunity against the bovine protozoan parasite Theileria parva has previously been shown to be mediated through lysis of parasite-infected cells by MHC class I restricted CD8+ cytotoxic T lymphocytes. It is hypothesized that identification of CTL target schizont antigens will aid the development of a sub-unit vaccine. We exploited the availability of the complete genome sequence data and bioinformatics tools to identify genes encoding secreted or membrane anchored proteins that may be processed and presented by the MHC class I molecules of infected cells to CTL. Of the 986 predicted open reading frames (ORFs) encoded by chromosome 1 of the T. parva genome, 55 were selected based on the presence of a signal peptide and/or a transmembrane helix domain. Thirty six selected ORFs were successfully cloned into a eukaryotic expression vector, transiently transfected into immortalized bovine skin fibroblasts and screened in vitro using T. parva-specific CTL. Recognition of gene products by CTL was assessed using an IFN-gamma ELISpot assay. A 525 base pair ORF encoding a 174 amino acid protein, designated Tp2, was identified by T. parva-specific CTL from 4 animals. These CTL recognized and lysed Tp2 transfected skin fibroblasts and recognized 4 distinct epitopes. Significantly, Tp2 specific CD8+ T cell responses were observed during the protective immune response against sporozoite challenge. The identification of an antigen containing multiple CTL epitopes and its apparent immunodominance during a protective anti-parasite response makes Tp2 an attractive candidate for evaluation of its vaccine potential.

The GPRC6A receptor displays constitutive internalization and sorting to the slow recycling pathway.

PubMed

Jacobsen, Stine Engesgaard; Ammendrup-Johnsen, Ina; Jansen, Anna Mai; Gether, Ulrik; Madsen, Kenneth Lindegaard; Bräuner-Osborne, Hans

2017-04-28

The class C G protein-coupled receptor GPRC6A is a putative nutrient-sensing receptor and represents a possible new drug target in metabolic disorders. However, the specific physiological role of this receptor has yet to be identified, and the mechanisms regulating its activity and cell surface availability also remain enigmatic. In the present study, we investigated the trafficking properties of GPRC6A by use of both a classical antibody feeding internalization assay in which cells were visualized using confocal microscopy and a novel internalization assay that is based on real-time measurements of fluorescence resonance energy transfer. Both assays revealed that GPRC6A predominantly undergoes constitutive internalization, whereas the agonist-induced effects were imperceptible. Moreover, postendocytic sorting was investigated by assessing the co-localization of internalized GPRC6A with selected Rab protein markers. Internalized GPRC6A was mainly co-localized with the early endosome marker Rab5 and the long loop recycling endosome marker Rab11 and to a much lesser extent with the late endosome marker Rab7. This suggests that upon agonist-independent internalization, GPRC6A is recycled via the Rab11-positive slow recycling pathway, which may be responsible for ensuring a persistent pool of GPRC6A receptors at the cell surface despite chronic agonist exposure. Distinct trafficking pathways have been reported for several of the class C receptors, and our results thus substantiate that non-canonical trafficking mechanisms are a common feature for the nutrient-sensing class C family that ensure functional receptors in the cell membrane despite prolonged agonist exposure. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

The GPRC6A receptor displays constitutive internalization and sorting to the slow recycling pathway

PubMed Central

Jacobsen, Stine Engesgaard; Ammendrup-Johnsen, Ina; Jansen, Anna Mai; Gether, Ulrik; Madsen, Kenneth Lindegaard; Bräuner-Osborne, Hans

2017-01-01

The class C G protein-coupled receptor GPRC6A is a putative nutrient-sensing receptor and represents a possible new drug target in metabolic disorders. However, the specific physiological role of this receptor has yet to be identified, and the mechanisms regulating its activity and cell surface availability also remain enigmatic. In the present study, we investigated the trafficking properties of GPRC6A by use of both a classical antibody feeding internalization assay in which cells were visualized using confocal microscopy and a novel internalization assay that is based on real-time measurements of fluorescence resonance energy transfer. Both assays revealed that GPRC6A predominantly undergoes constitutive internalization, whereas the agonist-induced effects were imperceptible. Moreover, postendocytic sorting was investigated by assessing the co-localization of internalized GPRC6A with selected Rab protein markers. Internalized GPRC6A was mainly co-localized with the early endosome marker Rab5 and the long loop recycling endosome marker Rab11 and to a much lesser extent with the late endosome marker Rab7. This suggests that upon agonist-independent internalization, GPRC6A is recycled via the Rab11-positive slow recycling pathway, which may be responsible for ensuring a persistent pool of GPRC6A receptors at the cell surface despite chronic agonist exposure. Distinct trafficking pathways have been reported for several of the class C receptors, and our results thus substantiate that non-canonical trafficking mechanisms are a common feature for the nutrient-sensing class C family that ensure functional receptors in the cell membrane despite prolonged agonist exposure. PMID:28280242

Brominated and organophosphate flame retardants target different neurodevelopmental stages, characterized with embryonic neural stem cells and neuronotypic PC12 cells.

PubMed

Slotkin, Theodore A; Skavicus, Samantha; Stapleton, Heather M; Seidler, Frederic J

2017-09-01

In addition to their activity as endocrine disruptors, brominated and organophosphate flame retardants are suspected to be developmental neurotoxicants, although identifying their specific mechanisms for that activity has been elusive. In the current study, we evaluated the effects of several flame retardants on neurodifferentiation using two in vitro models that assess distinct "decision nodes" in neural cell development: embryonic rat neural stem cells (NSCs), which evaluate the origination of neurons and glia from precursors, and rat neuronotypic PC12 cells, which characterize a later stage where cells committed to a neuronal phenotype undergo neurite outgrowth and neurotransmitter specification. In NSCs, both brominated and organophosphate flame retardants diverted the phenotype in favor of glia and away from formation of neurons, leading to an increased glia/neuron ratio, a common hallmark of the in vivo effects of neurotoxicants. For this early decision node, the brominated flame retardants were far more potent than the organophosphates. In PC12 cells, the brominated flame retardants were far less effective, whereas tris (1,3-dichloro-2-propyl) phosphate, an organophosphate, was more effective. Thus, the two classes of flame retardants differentially impact the two distinct vulnerable periods of neurodifferentiation. Furthermore, the effects on neurodifferentiation were separable from outright cytotoxicity, an important requirement in establishing a specific effect of these agents on neural cell development. These results reinforce the likelihood that flame retardants act as developmental neurotoxicants via direct effects on neural cell differentiation, over and above other activities that can impact nervous system development, such as endocrine disruption. Copyright © 2017 Elsevier B.V. All rights reserved.

Point process models for localization and interdependence of punctate cellular structures.

PubMed

Li, Ying; Majarian, Timothy D; Naik, Armaghan W; Johnson, Gregory R; Murphy, Robert F

2016-07-01

Accurate representations of cellular organization for multiple eukaryotic cell types are required for creating predictive models of dynamic cellular function. To this end, we have previously developed the CellOrganizer platform, an open source system for generative modeling of cellular components from microscopy images. CellOrganizer models capture the inherent heterogeneity in the spatial distribution, size, and quantity of different components among a cell population. Furthermore, CellOrganizer can generate quantitatively realistic synthetic images that reflect the underlying cell population. A current focus of the project is to model the complex, interdependent nature of organelle localization. We built upon previous work on developing multiple non-parametric models of organelles or structures that show punctate patterns. The previous models described the relationships between the subcellular localization of puncta and the positions of cell and nuclear membranes and microtubules. We extend these models to consider the relationship to the endoplasmic reticulum (ER), and to consider the relationship between the positions of different puncta of the same type. Our results do not suggest that the punctate patterns we examined are dependent on ER position or inter- and intra-class proximity. With these results, we built classifiers to update previous assignments of proteins to one of 11 patterns in three distinct cell lines. Our generative models demonstrate the ability to construct statistically accurate representations of puncta localization from simple cellular markers in distinct cell types, capturing the complex phenomena of cellular structure interaction with little human input. This protocol represents a novel approach to vesicular protein annotation, a field that is often neglected in high-throughput microscopy. These results suggest that spatial point process models provide useful insight with respect to the spatial dependence between cellular structures. © 2016 International Society for Advancement of Cytometry. © 2016 International Society for Advancement of Cytometry.

Use of Latent Class Analysis to define groups based on validity, cognition, and emotional functioning.

PubMed

Morin, Ruth T; Axelrod, Bradley N

Latent Class Analysis (LCA) was used to classify a heterogeneous sample of neuropsychology data. In particular, we used measures of performance validity, symptom validity, cognition, and emotional functioning to assess and describe latent groups of functioning in these areas. A data-set of 680 neuropsychological evaluation protocols was analyzed using a LCA. Data were collected from evaluations performed for clinical purposes at an urban medical center. A four-class model emerged as the best fitting model of latent classes. The resulting classes were distinct based on measures of performance validity and symptom validity. Class A performed poorly on both performance and symptom validity measures. Class B had intact performance validity and heightened symptom reporting. The remaining two Classes performed adequately on both performance and symptom validity measures, differing only in cognitive and emotional functioning. In general, performance invalidity was associated with worse cognitive performance, while symptom invalidity was associated with elevated emotional distress. LCA appears useful in identifying groups within a heterogeneous sample with distinct performance patterns. Further, the orthogonal nature of performance and symptom validities is supported.

Activation of mixed glia by Abeta-specific Th1 and Th17 cells and its regulation by Th2 cells.

PubMed

McQuillan, K; Lynch, Marina A; Mills, Kingston H G

2010-05-01

Microglia are innate immune cells of the CNS, that act as antigen-presenting cells (APC) for antigen-specific T cells and respond to inflammatory stimuli, such as amyloid-beta (Abeta), resulting in the release of neurotoxic factors and pro-inflammatory cytokines. Astrocytes can also act as APC and modulate the function of microglia. However, the role of distinct T cell subtypes, in particular Th17 cells, in glial activation and subsequent modulatory effects of Th2 cells are poorly understood. Here, we generated Abeta-specific Th1, Th2, and Th17 cells and examined their role in modulating Abeta-induced activation of microglia in a mixed glial culture, a preparation which mimics the complex APC types in the brain. We demonstrated that mixed glia acted as an effective APC for Abeta-specific Th1 and Th17 cells. Addition of Abeta-specific Th2 cells suppressed the Abeta-induced IFN-gamma production by Th1 cells and IL-17 production by Th17 cells with glia as the APC. Co-culture of Abeta-specific Th1 or Th17 cells with glia markedly enhanced Abeta-induced pro-inflammatory cytokine production and expression of MHC class II and co-stimulatory molecules on the microglia. Addition of Abeta-specific Th2 cells inhibited Th17 cell-induced IL-1beta and IL-6 production by mixed glia and attenuated Th1 cell-induced CD86 and CD40 expression on microglia. The modest enhancement of MHC class II and CD86 expression on astrocytes by Abeta-specific Th1 and Th17 was not attenuated by Th2 cells. These data indicate that Abeta-specific Th1 and Th17 cells induce inflammatory activation of glia, and that this is in part regulated by Th2 cells. Copyright 2010 Elsevier Inc. All rights reserved.

Quantitative two-dimensional gel electrophoresis analysis of human fibroblasts transformed by ras oncogenes.

PubMed

Miller, M J; Maher, V M; McCormick, J J

1992-11-01

Quantitative two-dimensional gel electrophoresis was used to compare the cellular protein patterns of a normal foreskin-derived human fibroblasts cell line (LG1) and three immortal derivatives of LG1. One derivative, designated MSU-1.1 VO, was selected for its ability to grow in the absence of serum and is non-tumorigenic in athymic mice. The other two strains were selected for focus-formation following transfection with either Ha-ras or N-ras oncogenes and form high grade malignant tumors. Correspondence and cluster analysis provided a nonbiased estimate of the relative similarity of the different two-dimensional patterns. These techniques separated the gel patterns into three distinct classes: LG1, MSU-1.1 VO, and the ras transformed cell strains. The MSU-1.1 VO cells were more closely related to the parental LG1 than to the ras-transformed cells. The differences between the three classes were primarily quantitative in nature: 16% of the spots demonstrated statistically significant changes (P < 0.01, T test, mean ratio of intensity > 2) in the rate of incorporation of radioactive amino acids. The patterns from the two ras-transformed cell strains were similar, and variations in the expression of proteins that occurred between the separate experiments obscured consistent differences between the Ha-ras and N-ras transformed cells. However, while only 9 out of 758 spots were classified as different (1%), correspondence analysis could consistently separate the two ras transformants. One of these spots was five times more intense in the Ha-ras transformed cells than the N-ras.(ABSTRACT TRUNCATED AT 250 WORDS)

Clear cell and endometrioid carcinomas: are their differences attributable to distinct cells of origin?

PubMed

Cochrane, Dawn R; Tessier-Cloutier, Basile; Lawrence, Katherine M; Nazeran, Tayyebeh; Karnezis, Anthony N; Salamanca, Clara; Cheng, Angela S; McAlpine, Jessica N; Hoang, Lien N; Gilks, C Blake; Huntsman, David G

2017-09-01

Endometrial epithelium is the presumed tissue of origin for both eutopic and endometriosis-derived clear cell and endometrioid carcinomas. We had previously hypothesized that the morphological, biological and clinical differences between these carcinomas are due to histotype-specific mutations. Although some mutations and genomic landscape features are more likely to be found in one of these histotypes, we were not able to identify a single class of mutations that was exclusively present in one histotype and not the other. This lack of genomic differences led us to an alternative hypothesis that these cancers could arise from distinct cells of origin within endometrial tissue, and that it is the cellular context that accounts for their differences. In a proteomic screen, we identified cystathionine γ-lyase (CTH) as a marker for clear cell carcinoma, as it is expressed at high levels in clear cell carcinomas of the ovary and endometrium. In the current study, we analysed normal Müllerian tissues, and found that CTH is expressed in ciliated cells of endometrium (both eutopic endometrium and endometriosis) and fallopian tubes. We then demonstrated that other ciliated cell markers are expressed in clear cell carcinomas, whereas endometrial secretory cell markers are expressed in endometrioid carcinomas. The same differential staining of secretory and ciliated cells was demonstrable in a three-dimensional organoid culture system, in which stem cells were stimulated to differentiate into an admixture of secretory and ciliated cells. These data suggest that endometrioid carcinomas are derived from cells of the secretory cell lineage, whereas clear cell carcinomas are derived from, or have similarities to, cells of the ciliated cell lineage. Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

T cell receptors used in cancer gene therapy cross-react with MART-1/Melan-A tumor antigens via distinct mechanisms1

PubMed Central

Borbulevych, Oleg Y.; Santhanagopolan, Sujatha M.; Hossain, Moushumi; Baker, Brian M.

2011-01-01

T cells engineered to express T cell receptors (TCRs) specific for tumor antigens can drive cancer regression. The first TCRs used in cancer gene therapy, DMF4 and DMF5, recognize two structurally distinct peptide epitopes of the melanoma-associated MART-1/Melan-A protein, both presented by the class I MHC protein HLA-A*0201. To help understand the mechanisms of TCR cross-reactivity and provide a foundation for the further development of immunotherapy, we determined the crystallographic structures of DMF4 and DMF5 in complex with both of the MART-1/Melan-A epitopes. The two TCRs use different mechanisms to accommodate the two ligands. Whereas DMF4 binds the two with a different orientation, altering its position over the peptide/MHC, DMF5 binds them both identically. The simpler mode of cross-reactivity by DMF5 is associated with higher affinity towards both ligands, consistent with the superior functional avidity of DMF5. More generally, the observation of two diverging mechanisms of cross-reactivity with the same antigens and the finding that TCR binding orientation can be determined by peptide alone extend our understanding of the mechanisms underlying TCR cross-reactivity. PMID:21795600

The late and dual origin of cerebrospinal fluid-contacting neurons in the mouse spinal cord

PubMed Central

Petracca, Yanina L.; Sartoretti, Maria Micaela; Di Bella, Daniela J.; Marin-Burgin, Antonia; Carcagno, Abel L.; Schinder, Alejandro F.; Lanuza, Guillermo M.

2016-01-01

Considerable progress has been made in understanding the mechanisms that control the production of specialized neuronal types. However, how the timing of differentiation contributes to neuronal diversity in the developing spinal cord is still a pending question. In this study, we show that cerebrospinal fluid-contacting neurons (CSF-cNs), an anatomically discrete cell type of the ependymal area, originate from surprisingly late neurogenic events in the ventral spinal cord. CSF-cNs are identified by the expression of the transcription factors Gata2 and Gata3, and the ionic channels Pkd2l1 and Pkd1l2. Contrasting with Gata2/3+ V2b interneurons, differentiation of CSF-cNs is independent of Foxn4 and takes place during advanced developmental stages previously assumed to be exclusively gliogenic. CSF-cNs are produced from two distinct dorsoventral regions of the mouse spinal cord. Most CSF-cNs derive from progenitors circumscribed to the late-p2 and the oligodendrogenic (pOL) domains, whereas a second subset of CSF-cNs arises from cells bordering the floor plate. The development of these two subgroups of CSF-cNs is differentially controlled by Pax6, they adopt separate locations around the postnatal central canal and they display electrophysiological differences. Our results highlight that spatiotemporal mechanisms are instrumental in creating neural cell diversity in the ventral spinal cord to produce distinct classes of interneurons, motoneurons, CSF-cNs, glial cells and ependymal cells. PMID:26839365

The soluble pattern recognition receptor PTX3 links humoral innate and adaptive immune responses by helping marginal zone B cells

PubMed Central

Sintes, Jordi; Polentarutti, Nadia; Walland, A. Cooper; Yeiser, John R.; Cunha, Cristina; Lacerda, João F.; Salvatori, Giovanni; Blander, J. Magarian

2016-01-01

Pentraxin 3 (PTX3) is a fluid-phase pattern recognition receptor of the humoral innate immune system with ancestral antibody-like properties but unknown antibody-inducing function. In this study, we found binding of PTX3 to splenic marginal zone (MZ) B cells, an innate-like subset of antibody-producing lymphocytes strategically positioned at the interface between the circulation and the adaptive immune system. PTX3 was released by a subset of neutrophils that surrounded the splenic MZ and expressed an immune activation–related gene signature distinct from that of circulating neutrophils. Binding of PTX3 promoted homeostatic production of IgM and class-switched IgG antibodies to microbial capsular polysaccharides, which decreased in PTX3-deficient mice and humans. In addition, PTX3 increased IgM and IgG production after infection with blood-borne encapsulated bacteria or immunization with bacterial carbohydrates. This immunogenic effect stemmed from the activation of MZ B cells through a neutrophil-regulated pathway that elicited class switching and plasmablast expansion via a combination of T cell–independent and T cell–dependent signals. Thus, PTX3 may bridge the humoral arms of the innate and adaptive immune systems by serving as an endogenous adjuvant for MZ B cells. This property could be harnessed to develop more effective vaccines against encapsulated pathogens. PMID:27621420

Homopiperazine Derivatives as a Novel Class of Proteasome Inhibitors with a Unique Mode of Proteasome Binding

PubMed Central

Kikuchi, Jiro; Shibayama, Naoya; Yamada, Satoshi; Wada, Taeko; Nobuyoshi, Masaharu; Izumi, Tohru; Akutsu, Miyuki; Kano, Yasuhiko; Sugiyama, Kanako; Ohki, Mio; Park, Sam-Yong; Furukawa, Yusuke

2013-01-01

The proteasome is a proteolytic machinery that executes the degradation of polyubiquitinated proteins to maintain cellular homeostasis. Proteasome inhibition is a unique and effective way to kill cancer cells because they are sensitive to proteotoxic stress. Indeed, the proteasome inhibitor bortezomib is now indispensable for the treatment of multiple myeloma and other intractable malignancies, but is associated with patient inconvenience due to intravenous injection and emerging drug resistance. To resolve these problems, we attempted to develop orally bioavailable proteasome inhibitors with distinct mechanisms of action and identified homopiperazine derivatives (HPDs) as promising candidates. Biochemical and crystallographic studies revealed that some HPDs inhibit all three catalytic subunits (ß 1, ß 2 and ß 5) of the proteasome by direct binding, whereas bortezomib and other proteasome inhibitors mainly act on the ß5 subunit. Proteasome-inhibitory HPDs exhibited cytotoxic effects on cell lines from various hematological malignancies including myeloma. Furthermore, K-7174, one of the HPDs, was able to inhibit the growth of bortezomib-resistant myeloma cells carrying a ß5-subunit mutation. Finally, K-7174 had additive effects with bortezomib on proteasome inhibition and apoptosis induction in myeloma cells. Taken together, HPDs could be a new class of proteasome inhibitors, which compensate for the weak points of conventional ones and overcome the resistance to bortezomib. PMID:23593271

Cell-Type and State-Dependent Synchronization among Rodent Somatosensory, Visual, Perirhinal Cortex, and Hippocampus CA1

PubMed Central

Vinck, Martin; Bos, Jeroen J.; Van Mourik-Donga, Laura A.; Oplaat, Krista T.; Klein, Gerbrand A.; Jackson, Jadin C.; Gentet, Luc J.; Pennartz, Cyriel M. A.

2016-01-01

Beta and gamma rhythms have been hypothesized to be involved in global and local coordination of neuronal activity, respectively. Here, we investigated how cells in rodent area S1BF are entrained by rhythmic fluctuations at various frequencies within the local area and in connected areas, and how this depends on behavioral state and cell type. We performed simultaneous extracellular field and unit recordings in four connected areas of the freely moving rat (S1BF, V1M, perirhinal cortex, CA1). S1BF spiking activity was strongly entrained by both beta and gamma S1BF oscillations, which were associated with deactivations and activations, respectively. We identified multiple classes of fast spiking and excitatory cells in S1BF, which showed prominent differences in rhythmic entrainment and in the extent to which phase locking was modulated by behavioral state. Using an additional dataset acquired by whole-cell recordings in head-fixed mice, these cell classes could be compared with identified phenotypes showing gamma rhythmicity in their membrane potential. We next examined how S1BF cells were entrained by rhythmic fluctuations in connected brain areas. Gamma-synchronization was detected in all four areas, however we did not detect significant gamma coherence among these areas. Instead, we only found long-range coherence in the theta-beta range among these areas. In contrast to local S1BF synchronization, we found long-range S1BF-spike to CA1–LFP synchronization to be homogeneous across inhibitory and excitatory cell types. These findings suggest distinct, cell-type contributions of low and high-frequency synchronization to intra- and inter-areal neuronal interactions. PMID:26834582

Trajectories of functional limitation in early rheumatoid arthritis and their association with mortality.

PubMed

Norton, Sam; Sacker, Amanda; Dixey, Josh; Done, John; Williams, Peter; Young, Adam

2013-11-01

This study aimed to identify subgroups with distinct trajectories of functional (HAQ) progression over 10 years following diagnosis of RA and identify baseline characteristics associated with the trajectories and their prognostic value for mortality. Between 1986 and 1998, 1460 patients with RA symptoms <2 years and prior to disease-modifying treatment (DMARDs) were recruited to an inception cohort (Early RA Study). Standard clinical, functional and laboratory assessments were performed at presentation and annually. Deaths were tracked by the National Health Service Central Register. Growth mixture modelling was used to identify distinct trajectories of HAQ score progression and survival analysis employed to compare all-cause mortality across the trajectory classes. Four HAQ score progression classes were identified: moderate increasing (46%), low stable (6%), moderate stable (28%) and high stable (20%). Only the moderate-increasing class exhibited an accelerated decline in function over normal ageing. Compared with the moderate-increasing class, individuals with high-stable HAQ scores were more likely to be female, have more severe disease and other coexistent conditions. Low-stable class patients were more likely to be male and report less pain. The high-stable class had increased risk of mortality compared with the moderate-increasing class after adjusting for potential confounding factors, whereas low-stable and moderate-stable classes were at reduced mortality risk. The effect of RA on function is set within the first few years and is affected by comorbidity. Identifying distinct groups of patients may help to target those at greater risk of poor functional outcome and mortality.

Qa-1/HLA-E-restricted regulatory CD8+ T cells and self-nonself discrimination: an essay on peripheral T-cell regulation.

PubMed

Jiang, Hong; Chess, Leonard

2008-11-01

By discriminating self from nonself and controlling the magnitude and class of immune responses, the immune system mounts effective immunity against virtually any foreign antigens but avoids harmful immune responses to self. These are two equally important and related but distinct processes, which function in concert to ensure an optimal function of the immune system. Immunologically relevant clinical problems often occur because of failure of either process, especially the former. Currently, there is no unified conceptual framework to characterize the precise relationship between thymic negative selection and peripheral immune regulation, which is the basis for understanding self-non-self discrimination versus control of magnitude and class of immune responses. In this article, we explore a novel hypothesis of how the immune system discriminates self from nonself in the periphery during adaptive immunity. This hypothesis permits rational analysis of various seemingly unrelated biomedical problems inherent in immunologic disorders that cannot be uniformly interpreted by any currently existing paradigms. The proposed hypothesis is based on a unified conceptual framework of the "avidity model of peripheral T-cell regulation" that we originally proposed and tested, in both basic and clinical immunology, to understand how the immune system achieves self-nonself discrimination in the periphery.

Functional dissociation in sweet taste receptor neurons between and within taste organs of Drosophila

PubMed Central

Thoma, Vladimiros; Knapek, Stephan; Arai, Shogo; Hartl, Marion; Kohsaka, Hiroshi; Sirigrivatanawong, Pudith; Abe, Ayako; Hashimoto, Koichi; Tanimoto, Hiromu

2016-01-01

Finding food sources is essential for survival. Insects detect nutrients with external taste receptor neurons. Drosophila possesses multiple taste organs that are distributed throughout its body. However, the role of different taste organs in feeding remains poorly understood. By blocking subsets of sweet taste receptor neurons, we show that receptor neurons in the legs are required for immediate sugar choice. Furthermore, we identify two anatomically distinct classes of sweet taste receptor neurons in the leg. The axonal projections of one class terminate in the thoracic ganglia, whereas the other projects directly to the brain. These two classes are functionally distinct: the brain-projecting neurons are involved in feeding initiation, whereas the thoracic ganglia-projecting neurons play a role in sugar-dependent suppression of locomotion. Distinct receptor neurons for the same taste quality may coordinate early appetitive responses, taking advantage of the legs as the first appendages to contact food. PMID:26893070

Targeted delivery of antigen to hamster nasal lymphoid tissue with M-cell-directed lectins.

PubMed Central

Giannasca, P J; Boden, J A; Monath, T P

1997-01-01

The nasal cavity of a rodent is lined by an epithelium organized into distinct regional domains responsible for specific physiological functions. Aggregates of nasal lymphoid tissue (NALT) located at the base of the nasal cavity are believed to be sites of induction of mucosal immune responses to airborne antigens. The epithelium overlying NALT contains M cells which are specialized for the transcytosis of immunogens, as demonstrated in other mucosal tissues. We hypothesized that NALT M cells are characterized by distinct glycoconjugate receptors which influence antigen uptake and immune responses to transcytosed antigens. To identify glycoconjugates that may distinguish NALT M cells from other cells of the respiratory epithelium (RE), we performed lectin histochemistry on sections of the hamster nasal cavity with a panel of lectins. Many classes of glycoconjugates were found on epithelial cells in this region. While most lectins bound to sites on both the RE and M cells, probes capable of recognizing alpha-linked galactose were found to label the follicle-associated epithelium (FAE) almost exclusively. By morphological criteria, the FAE contains >90% M cells. To determine if apical glycoconjugates on M cells were accessible from the nasal cavity, an M-cell-selective lectin and a control lectin in parallel were administered intranasally to hamsters. The M-cell-selective lectin was found to specifically target the FAE, while the control lectin did not. Lectin bound to M cells in vivo was efficiently endocytosed, consistent with the role of M cells in antigen transport. Intranasal immunization with lectin-test antigen conjugates without adjuvant stimulated induction of specific serum immunoglobulin G, whereas antigen alone or admixed with lectin did not. The selective recognition of NALT M cells by a lectin in vivo provides a model for microbial adhesin-host cell receptor interactions on M cells and the targeted delivery of immunogens to NALT following intranasal administration. PMID:9317039

Discovery and Characterization of Chromatin States for Systematic Annotation of the Human Genome

NASA Astrophysics Data System (ADS)

Ernst, Jason; Kellis, Manolis

A plethora of epigenetic modifications have been described in the human genome and shown to play diverse roles in gene regulation, cellular differentiation and the onset of disease. Although individual modifications have been linked to the activity levels of various genetic functional elements, their combinatorial patterns are still unresolved and their potential for systematic de novo genome annotation remains untapped. Here, we use a multivariate Hidden Markov Model to reveal chromatin states in human T cells, based on recurrent and spatially coherent combinations of chromatin marks.We define 51 distinct chromatin states, including promoter-associated, transcription-associated, active intergenic, largescale repressed and repeat-associated states. Each chromatin state shows specific enrichments in functional annotations, sequence motifs and specific experimentally observed characteristics, suggesting distinct biological roles. This approach provides a complementary functional annotation of the human genome that reveals the genome-wide locations of diverse classes of epigenetic function.

Parental attitudes towards measles vaccination in the canton of Aargau, Switzerland: a latent class analysis.

PubMed

Weiss, Carine; Schröpfer, Daniel; Merten, Sonja

2016-08-11

Despite the successes of routine national childhood vaccination programmes, measles remains a public health concern. The purpose of this paper is to investigate how patterns of parental attitudes are linked to the decision-making process for or against MMR vaccination. This exploratory study was designed to identify distinct patterns of attitudes towards or against measles vaccination through Latent Class Analysis (LCA) in a sub-sample of mothers living in the canton of Aargau in Switzerland. Parents of young children below 36 months of age were randomly selected through parents' counsellors' registries. Among other questions, respondents were asked to state their agreement in response to 14 belief statements regarding measles vaccination on a 5-point Likert scale. To identify groups of parents showing distinct patterns of attitudes and beliefs regarding measles vaccination, we used Latent Class Analysis (LCA). The LCA showed three classes of parents with different attitudes and believes towards measles vaccination: The biggest group (class 1) are those having positive attitudes towards immunisation, followed by the second biggest group (class 2) which is characterised by having fearful attitudes and by showing uncertainty about immunisation. The third group (class 3) shows distinct patterns of critical attitudes against immunisation. Within this group over 90 % agree or totally agree that immunisation is an artificial intrusion into the natural immune system and therefore want to vaccinate their children only if necessary. We find that parents in the Canton Aargau who hesitate to vaccinate their children against measles, mumps and rubella show distinct opinions and attitudes. Health professionals should be aware of these perceptions to tailor their messages accordingly and positively influence these parents to vaccinate their children. Special attention needs to be given to those parents who are planning to vaccinate their children but are not following the national guidelines.

Predicted structure of MIF/CD74 and RTL1000/CD74 complexes.

PubMed

Meza-Romero, Roberto; Benedek, Gil; Leng, Lin; Bucala, Richard; Vandenbark, Arthur A

2016-04-01

Macrophage migration inhibitory factor (MIF) is a key cytokine in autoimmune and inflammatory diseases that attracts and then retains activated immune cells from the periphery to the tissues. MIF exists as a homotrimer and its effects are mediated through its primary receptor, CD74 (the class II invariant chain that exhibits a highly structured trimerization domain), present on class II expressing cells. Although a number of binding residues have been identified between MIF and CD74 trimers, their spatial orientation has not been established. Using a docking program in silico, we have modeled binding interactions between CD74 and MIF as well as CD74 and a competitive MIF inhibitor, RTL1000, a partial MHC class II construct that is currently in clinical trials for multiple sclerosis. These analyses revealed 3 binding sites on the MIF trimer that each were predicted to bind one CD74 trimer through interactions with two distinct 5 amino acid determinants. Surprisingly, predicted binding of one CD74 trimer to a single RTL1000 antagonist utilized the same two 5 residue determinants, providing strong suggestive evidence in support of the MIF binding regions on CD74. Taken together, our structural modeling predicts a new MIF(CD74)3 dodecamer that may provide the basis for increased MIF potency and the requirement for ~3-fold excess RTL1000 to achieve full antagonism.

Lag-3, Tim-3, and TIGIT co-inhibitory receptors with specialized functions in immune regulation

PubMed Central

Anderson, Ana C.; Joller, Nicole; Kuchroo, Vijay K.

2016-01-01

Summary Co-inhibitory receptors, such as CTLA-4 and PD-1, have an important role in regulating T cell responses and have proven to be effective targets in the setting of chronic diseases where constitutive co-inhibitory receptor expression on T cells dampens effector T cell responses. Unfortunately, many patients still fail to respond to therapies that target CTLA-4 and PD-1. The next wave of co-inhibitory receptor targets that are being explored in clinical trials include Lag-3, Tim-3, and TIGIT. These receptors while belonging to the same class of receptors as PD-1 and CTLA-4 exhibit unique functions especially at tissue sites where they regulate distinct aspects of immunity. Increased understanding of the specialized functions of these receptors will inform the rational application of therapies that target these receptors to the clinic. PMID:27192565

New Human CD22/Siglec-2 Ligands with a Triazole Glycoside.

PubMed

Prescher, Horst; Schweizer, Astrid; Kuhfeldt, Elena; Nitschke, Lars; Brossmer, Reinhard

2017-07-04

CD22 is a member of the Siglec family. Considerable attention has been drawn to the design and synthesis of new Siglec ligands to explore target biology and innovative therapies. In particular, CD22-ligand-targeted nanoparticles with therapeutic functions have proved successful in preclinical settings for blood cancers, autoimmune diseases, and tolerance induction. Here we report the design, synthesis and affinity evaluation of a new class of Siglec ligands: namely sialic acid derivatives with a triazole moiety replacing the natural glycoside oxygen atom. In addition, we describe important and surprising differences in binding to CD22 expressed at the cell surface for compounds with distinct valences. The new class of compounds might serve as a template for the design of ligands for other members of the Siglec family and next-generation CD22-ligand-based targeted therapies. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

Murine endogenous retroviruses

PubMed Central

2016-01-01

Up to 10% of the mouse genome is comprised of endogenous retrovirus (ERV) sequences, and most represent the remains of ancient germ line infections. Our knowledge of the three distinct classes of ERVs is inversely correlated with their copy number, and their characterization has benefited from the availability of divergent wild mouse species and subspecies, and from ongoing analysis of the Mus genome sequence. In contrast to human ERVs, which are nearly all extinct, active mouse ERVs can still be found in all three ERV classes. The distribution and diversity of ERVs has been shaped by host-virus interactions over the course of evolution, but ERVs have also been pivotal in shaping the mouse genome by altering host genes through insertional mutagenesis, by adding novel regulatory and coding sequences, and by their co-option by host cells as retroviral resistance genes. We review mechanisms by which an adaptive coexistence has evolved. (Part of a Multi-author Review) PMID:18818872

Regulation of MHC class I expression by Foxp3 and its effect on Treg cell function

PubMed Central

Mu, Jie; Tai, Xuguang; Iyer, Shankar S.; Weissman, Jocelyn D.; Singer, Alfred; Singer, Dinah S.

2014-01-01

Expression of MHC class I molecules, which provide immune surveillance against intracellular pathogens, is higher on lymphoid cells than on any other cell types. In T cells, this is a result of activation of class I transcription by the T cell enhanceosome consisting of Runx1, CBFβ and LEF1. We now report that MHC class I transcription in T cells also is enhanced by Foxp3, resulting in higher levels of class I in CD4+CD25+ T regulatory cells than in conventional CD4+CD25− T cells. Interestingly, the effect of Foxp3 regulation of MHC class I transcription is cell-type specific: Foxp3 increases MHC class I expression in T cells but represses it in epithelial tumor cells. In both cell types, Foxp3 targets the upstream IRE and downstream core promoter of the class I gene. Importantly, expression of MHC class I contributes to the function of CD4+CD25+ T regulatory cells by enhancing immune suppression, both in in vitro and in vivo. These findings identify MHC class I genes as direct targets of Foxp3 whose expression augments regulatory T cell function. PMID:24523508

Regional distribution of calretinin and calbindin-D28k expression in the brain of the urodele amphibian Pleurodeles waltl during embryonic and larval development.

PubMed

Joven, Alberto; Morona, Ruth; Moreno, Nerea; González, Agustín

2013-07-01

The sequence of appearance of calretinin and calbindin-D28k immunoreactive (CRir and CBir, respectively) cells and fibers has been studied in the brain of the urodele amphibian Pleurodeles waltl. Embryonic, larval and juvenile stages were studied. The early expression and the dynamics of the distribution of CBir and CRir structures have been used as markers for developmental aspects of distinct neuronal populations, highlighting the accurate extent of many regions in the developing brain, not observed on the basis of cytoarchitecture alone. CR and, to a lesser extent, CB are expressed early in the central nervous system and show a progressively increasing expression from the embryonic stages throughout the larval life and, in general, the labeled structures in the developing brain retain their ability to express these proteins in the adult brain. The onset of CRir cells primarily served to follow the development of the olfactory bulbs, subpallium, thalamus, alar hypothalamus, mesencephalic tegmentum, and distinct cell populations in the rhombencephalic reticular formation. CBir cells highlighted the development of, among others, the pallidum, hypothalamus, dorsal habenula, midbrain tegmentum, cerebellum, and central gray of the rostral rhombencephalon. However, it was the relative and mostly segregated distribution of both proteins in distinct cell populations which evidenced the developing regionalization of the brain. The results have shown the usefulness in neuroanatomy of the analysis during development of the onset of CBir and CRir structures, but the comparison with previous data has shown extensive variability across vertebrate classes. Therefore, one should be cautious when comparing possible homologue structures across species only on the basis of the expression of these proteins, due to the variation of the content of calcium-binding proteins observed in well-established homologous regions in the brain of different vertebrates.

Mechanisms of Acquired Drug Resistance to the HDAC6 Selective Inhibitor Ricolinostat Reveals Rational Drug-Drug Combination with Ibrutinib.

PubMed

Amengual, Jennifer E; Prabhu, Sathyen A; Lombardo, Maximilian; Zullo, Kelly; Johannet, Paul M; Gonzalez, Yulissa; Scotto, Luigi; Serrano, Xavier Jirau; Wei, Ying; Duong, Jimmy; Nandakumar, Renu; Cremers, Serge; Verma, Akanksha; Elemento, Olivier; O'Connor, Owen A

2017-06-15

Purpose: Pan-class I/II histone deacetylase (HDAC) inhibitors are effective treatments for select lymphomas. Isoform-selective HDAC inhibitors are emerging as potentially more targeted agents. ACY-1215 (ricolinostat) is a first-in-class selective HDAC6 inhibitor. To better understand the discrete function of HDAC6 and its role in lymphoma, we developed a lymphoma cell line resistant to ACY-1215. Experimental Design: The diffuse large B-cell lymphoma cell line OCI-Ly10 was exposed to increasing concentrations of ACY-1215 over an extended period of time, leading to the development of a resistant cell line. Gene expression profiling (GEP) was performed to investigate differentially expressed genes. Combination studies of ACY-1215 and ibrutinib were performed in cell lines, primary human lymphoma tissue, and a xenograft mouse model. Results: Systematic incremental increases in drug exposure led to the development of distinct resistant cell lines with IC 50 values 10- to 20-fold greater than that for parental lines. GEP revealed upregulation of MAPK10, HELIOS, HDAC9, and FYN, as well as downregulation of SH3BP5 and LCK. Gene-set enrichment analysis (GSEA) revealed modulation of the BTK pathway. Ibrutinib was found to be synergistic with ACY-1215 in cell lines as well as in 3 primary patient samples of lymphoma. In vivo confirmation of antitumor synergy was demonstrated with a xenograft of DLBCL. Conclusions: The development of this ACY-1215-resistant cell line has provided valuable insights into the mechanistic role of HDAC6 in lymphoma and offered a novel method to identify rational synergistic drug combinations. Translation of these findings to the clinic is underway. Clin Cancer Res; 23(12); 3084-96. ©2016 AACR . ©2016 American Association for Cancer Research.

Spatial separation of the processing and MHC class I loading compartments for cross-presentation of the tumor-associated antigen HER2/neu by human dendritic cells

PubMed Central

Baleeiro, Renato B; Rietscher, René; Diedrich, Andrea; Czaplewska, Justyna A; Lehr, Claus-Michael; Scherließ, Regina; Hanefeld, Andrea; Gottschaldt, Michael; Walden, Peter

2015-01-01

Cross-presentation is the process by which professional antigen presenting cells (APCs) (B cells, dendritic cells (DCs) and macrophages) present endocytosed antigens (Ags) via MHC-I to CD8+ T cells. This process is crucial for induction of adaptive immune responses against tumors and infected cells. The pathways and cellular compartments involved in cross-presentation are unresolved and controversial. Among the cells with cross-presenting capacity, DCs are the most efficient, which was proposed to depend on prevention of endosomal acidification to block degradation of the epitopes. Contrary to this view, we show in this report that some cargoes induce strong endosomal acidification following uptake by human DCs, while others not. Moreover, processing of the tumor-associated antigen HER2/neu delivered in nanoparticles (NP) for cross-presentation of the epitope HER2/neu369–377 on HLA-A2 depended on endosomal acidification and cathepsin activity as well as proteasomes, and newly synthesized HLA class I. However, the HLA-A*0201/HER2/neu369–377 complexes were not found in the endoplasmic reticulum (ER) nor in endolysosomes but in hitherto not described vesicles. The data thus indicate spatial separation of antigen processing and loading of MHC-I for cross-presentation: antigen processing occurs in the uptake compartment and the cytosol whereas MHC-I loading with peptide takes place in a distinct subcellular compartment. The findings further elucidate the cellular pathways involved in the cross-presentation of a full-length, clinically relevant tumor-associated antigen by human DCs, and the impact of the vaccine formulation on antigen processing and CD8+ T cell induction. PMID:26985398

Evolution of bacterial-like phosphoprotein phosphatases in photosynthetic eukaryotes features ancestral mitochondrial or archaeal origin and possible lateral gene transfer.

PubMed

Uhrig, R Glen; Kerk, David; Moorhead, Greg B

2013-12-01

Protein phosphorylation is a reversible regulatory process catalyzed by the opposing reactions of protein kinases and phosphatases, which are central to the proper functioning of the cell. Dysfunction of members in either the protein kinase or phosphatase family can have wide-ranging deleterious effects in both metazoans and plants alike. Previously, three bacterial-like phosphoprotein phosphatase classes were uncovered in eukaryotes and named according to the bacterial sequences with which they have the greatest similarity: Shewanella-like (SLP), Rhizobiales-like (RLPH), and ApaH-like (ALPH) phosphatases. Utilizing the wealth of data resulting from recently sequenced complete eukaryotic genomes, we conducted database searching by hidden Markov models, multiple sequence alignment, and phylogenetic tree inference with Bayesian and maximum likelihood methods to elucidate the pattern of evolution of eukaryotic bacterial-like phosphoprotein phosphatase sequences, which are predominantly distributed in photosynthetic eukaryotes. We uncovered a pattern of ancestral mitochondrial (SLP and RLPH) or archaeal (ALPH) gene entry into eukaryotes, supplemented by possible instances of lateral gene transfer between bacteria and eukaryotes. In addition to the previously known green algal and plant SLP1 and SLP2 protein forms, a more ancestral third form (SLP3) was found in green algae. Data from in silico subcellular localization predictions revealed class-specific differences in plants likely to result in distinct functions, and for SLP sequences, distinctive and possibly functionally significant differences between plants and nonphotosynthetic eukaryotes. Conserved carboxyl-terminal sequence motifs with class-specific patterns of residue substitutions, most prominent in photosynthetic organisms, raise the possibility of complex interactions with regulatory proteins.

Opposing Development of Cytotoxic and Follicular Helper CD4 T Cells Controlled by the TCF-1-Bcl6 Nexus.

PubMed

Donnarumma, Tiziano; Young, George R; Merkenschlager, Julia; Eksmond, Urszula; Bongard, Nadine; Nutt, Stephen L; Boyer, Claude; Dittmer, Ulf; Le-Trilling, Vu Thuy Khanh; Trilling, Mirko; Bayer, Wibke; Kassiotis, George

2016-11-01

CD4 + T cells develop distinct and often contrasting helper, regulatory, or cytotoxic activities. Typically a property of CD8 + T cells, granzyme-mediated cytotoxic T cell (CTL) potential is also exerted by CD4 + T cells. However, the conditions that induce CD4 + CTLs are not entirely understood. Using single-cell transcriptional profiling, we uncover a unique signature of Granzyme B (GzmB) + CD4 + CTLs, which distinguishes them from other CD4 + T helper (Th) cells, including Th1 cells, and strongly contrasts with the follicular helper T (Tfh) cell signature. The balance between CD4 + CTL and Tfh differentiation heavily depends on the class of infecting virus and is jointly regulated by the Tfh-related transcription factors Bcl6 and Tcf7 (encoding TCF-1) and by the expression of the inhibitory receptors PD-1 and LAG3. This unique profile of CD4 + CTLs offers targets for their study, and its antagonism by the Tfh program separates CD4 + T cells with either helper or killer functions. Copyright © 2016 The Author(s). Published by Elsevier Inc. All rights reserved.

How neurosecretory vesicles release their cargo.

PubMed

Scalettar, Bethe A

2006-04-01

Neurons and related cell types often contain two major classes of neurosecretory vesicles, synaptic vesicles (SVs) and dense-core granules (DCGs), which store and release distinct cargo. SVs store and release classic neurotransmitters, which facilitate propagation of action potentials across the synaptic cleft, whereas DCGs transport, store, and release hormones, proteins, and neuropeptides, which facilitate neuronal survival, synaptic transmission, and learning. Over the past few years, there has been a major surge in our understanding of many of the key molecular mechanisms underlying cargo release from SVs and DCGs. This surge has been driven largely by the use of fluorescence microscopy (especially total internal reflection fluorescence microscopy) to visualize SVs or DCGs in living cells. This review highlights some of the recent insights into cargo release from neurosecretory vesicles provided by fluorescence microscopy, with emphasis on DCGs.

Human Corneal Limbal-Epithelial Cell Response to Varying Silk Film Geometric Topography In Vitro

PubMed Central

Lawrence, Brian D.; Pan, Zhi; Liu, Aihong; Kaplan, David L.; Rosenblatt, Mark I.

2012-01-01

Silk fibroin films are a promising class of biomaterials that have a number of advantages for use in ophthalmic applications due to their transparent nature, mechanical properties and minimal inflammatory response upon implantation. Freestanding silk films with parallel line and concentric ring topographies were generated for in vitro characterization of human corneal limbal-epithelial (HCLE) cell response upon differing geometric patterned surfaces. Results indicated that silk film topography significantly affected initial HCLE culture substrate attachment, cellular alignment, cell-to-cell contact formation, actin cytoskeleton alignment, and focal adhesion (FA) localization. Most notably, parallel line patterned surfaces displayed a 36%–54% increase on average in initial cell attachment, which corresponded to an over 2-fold increase in FA localization when compared to other silk film surfaces and controls. In addition, distinct localization of FA formation was observed along the edges for all patterned silk film topographies. In conclusion, silk film feature topography appears to help direct corneal epithelial cell response and cytoskeleton development, especially in regards to FA distribution, in vitro. PMID:22705042

Intra-adrenal murine TH-MYCN neuroblastoma tumors grow more aggressive and exhibit a distinct tumor microenvironment relative to their subcutaneous equivalents.

PubMed

Kroesen, Michiel; Brok, Ingrid C; Reijnen, Daphne; van Hout-Kuijer, Maaike A; Zeelenberg, Ingrid S; Den Brok, Martijn H; Hoogerbrugge, Peter M; Adema, Gosse J

2015-05-01

In around half of the patients with neuroblastoma (NBL), the primary tumor is located in one of the adrenal glands. We have previously reported on a transplantable TH-MYCN model of subcutaneous (SC) growing NBL in C57Bl/6 mice for immunological studies. In this report, we describe an orthotopic TH-MYCN transplantable model where the tumor cells were injected intra-adrenally (IA) by microsurgery. Strikingly, 9464D cells grew out much faster in IA tumors compared to the subcutis. Tumors were infiltrated by equal numbers of lymphocytes and myeloid cells. Within the myeloid cell population, however, tumor-infiltrating macrophages were more abundant in IA tumors compared to SC tumors and expressed lower levels of MHC class II, indicative of a more immunosuppressive phenotype. Using 9464D cells stably expressing firefly luciferase, enhanced IA tumor growth could be confirmed using bioluminescence. Collectively, these data show that the orthotopic IA localization of TH-MYCN cells impacts the NBL tumor microenvironment, resulting in a more stringent NBL model to study novel immunotherapeutic approaches for NBL.

The Enigma of Tripeptidyl-Peptidase II: Dual Roles in Housekeeping and Stress

PubMed Central

Preta, Giulio; de Klark, Rainier; Gavioli, Riccardo; Glas, Rickard

2010-01-01

The tripeptidyl-peptidase II complex consists of repeated 138 kDa subunits, assembled into two twisted strands that form a high molecular weight complex (>5 MDa). TPPII, like many other cytosolic peptidases, plays a role in the ubiquitin-proteasome pathway downstream of the proteasome as well as in the production and destruction of MHC class I antigens and degradation of neuropeptides. Tripeptidyl-peptidase II activity is increased in cells with an increased demand for protein degradation, but whether degradation of cytosolic peptides is the only cell biological role for TPPII has remained unclear. Recent data indicated that TPPII translocates into the nucleus to control DNA damage responses in malignant cells, supporting that cytosolic “housekeeping peptidases” may have additional roles in cell biology, besides their contribution to protein turnover. Overall, TPPII has an emerging importance in several cancer-related fields, such as metabolism, cell death control, and control of genome integrity; roles that are not understood in detail. The present paper reviews the cell biology of TPPII and discusses distinct roles for TPPII in the nucleus and cytosol. PMID:20847939

Small Modifications to Network Topology Can Induce Stochastic Bistable Spiking Dynamics in a Balanced Cortical Model

PubMed Central

McDonnell, Mark D.; Ward, Lawrence M.

2014-01-01

Abstract Directed random graph models frequently are used successfully in modeling the population dynamics of networks of cortical neurons connected by chemical synapses. Experimental results consistently reveal that neuronal network topology is complex, however, in the sense that it differs statistically from a random network, and differs for classes of neurons that are physiologically different. This suggests that complex network models whose subnetworks have distinct topological structure may be a useful, and more biologically realistic, alternative to random networks. Here we demonstrate that the balanced excitation and inhibition frequently observed in small cortical regions can transiently disappear in otherwise standard neuronal-scale models of fluctuation-driven dynamics, solely because the random network topology was replaced by a complex clustered one, whilst not changing the in-degree of any neurons. In this network, a small subset of cells whose inhibition comes only from outside their local cluster are the cause of bistable population dynamics, where different clusters of these cells irregularly switch back and forth from a sparsely firing state to a highly active state. Transitions to the highly active state occur when a cluster of these cells spikes sufficiently often to cause strong unbalanced positive feedback to each other. Transitions back to the sparsely firing state rely on occasional large fluctuations in the amount of non-local inhibition received. Neurons in the model are homogeneous in their intrinsic dynamics and in-degrees, but differ in the abundance of various directed feedback motifs in which they participate. Our findings suggest that (i) models and simulations should take into account complex structure that varies for neuron and synapse classes; (ii) differences in the dynamics of neurons with similar intrinsic properties may be caused by their membership in distinctive local networks; (iii) it is important to identify neurons that share physiological properties and location, but differ in their connectivity. PMID:24743633

Hantaviruses induce cell type- and viral species-specific host microRNA expression signatures

PubMed Central

Shin, Ok Sarah; Kumar, Mukesh; Yanagihara, Richard; Song, Jin-Won

2014-01-01

The mechanisms of hantavirus-induced modulation of host cellular immunity remain poorly understood. Recently, microRNAs (miRNAs) have emerged as a class of essential regulators of host immune response genes. To ascertain if differential host miRNA expression toward representative hantavirus species correlated with immune response genes, miRNA expression profiles were analyzed in human endothelial cells, macrophages and epithelial cells infected with pathogenic and nonpathogenic rodent- and shrew-borne hantaviruses. Distinct miRNA expression profiles were observed in a cell type- and viral species-specific pattern. A subset of miRNAs, including miR-151-5p and miR-1973, were differentially expressed between Hantaan virus and Prospect Hill virus. Pathway analyses confirmed that the targets of selected miRNAs were associated with inflammatory responses and innate immune receptor-mediated signaling pathways. Our data suggest that differential immune responses following hantavirus infection may be regulated in part by cellular miRNA through dysregulation of genes critical to the inflammatory process. PMID:24074584

Reprogramming cell fate with a genome-scale library of artificial transcription factors.

PubMed

Eguchi, Asuka; Wleklinski, Matthew J; Spurgat, Mackenzie C; Heiderscheit, Evan A; Kropornicka, Anna S; Vu, Catherine K; Bhimsaria, Devesh; Swanson, Scott A; Stewart, Ron; Ramanathan, Parameswaran; Kamp, Timothy J; Slukvin, Igor; Thomson, James A; Dutton, James R; Ansari, Aseem Z

2016-12-20

Artificial transcription factors (ATFs) are precision-tailored molecules designed to bind DNA and regulate transcription in a preprogrammed manner. Libraries of ATFs enable the high-throughput screening of gene networks that trigger cell fate decisions or phenotypic changes. We developed a genome-scale library of ATFs that display an engineered interaction domain (ID) to enable cooperative assembly and synergistic gene expression at targeted sites. We used this ATF library to screen for key regulators of the pluripotency network and discovered three combinations of ATFs capable of inducing pluripotency without exogenous expression of Oct4 (POU domain, class 5, TF 1). Cognate site identification, global transcriptional profiling, and identification of ATF binding sites reveal that the ATFs do not directly target Oct4; instead, they target distinct nodes that converge to stimulate the endogenous pluripotency network. This forward genetic approach enables cell type conversions without a priori knowledge of potential key regulators and reveals unanticipated gene network dynamics that drive cell fate choices.

Some Lipid Droplets Are More Equal Than Others: Different Metabolic Lipid Droplet Pools in Hepatic Stellate Cells.

PubMed

Molenaar, Martijn R; Vaandrager, Arie B; Helms, J Bernd

2017-01-01

Hepatic stellate cells (HSCs) are professional lipid-storing cells and are unique in their property to store most of the retinol (vitamin A) as retinyl esters in large-sized lipid droplets. Hepatic stellate cell activation is a critical step in the development of chronic liver disease, as activated HSCs cause fibrosis. During activation, HSCs lose their lipid droplets containing triacylglycerols, cholesteryl esters, and retinyl esters. Lipidomic analysis revealed that the dynamics of disappearance of these different classes of neutral lipids are, however, very different from each other. Although retinyl esters steadily decrease during HSC activation, triacylglycerols have multiple pools one of which becomes transiently enriched in polyunsaturated fatty acids before disappearing. These observations are consistent with the existence of preexisting "original" lipid droplets with relatively slow turnover and rapidly recycling lipid droplets that transiently appear during activation of HSCs. Elucidation of the molecular machinery involved in the regulation of these distinct lipid droplet pools may open new avenues for the treatment of liver fibrosis.

Cancer vulnerabilities unveiled by genomic loss

PubMed Central

Nijhawan, Deepak; Zack, Travis I.; Ren, Yin; Strickland, Matthew R.; Lamothe, Rebecca; Schumacher, Steven E.; Tsherniak, Aviad; Besche, Henrike C.; Rosenbluh, Joseph; Shehata, Shyemaa; Cowley, Glenn S.; Weir, Barbara A.; Goldberg, Alfred L.; Mesirov, Jill P.; Root, David E.; Bhatia, Sangeeta N.; Beroukhim, Rameen; Hahn, William C.

2012-01-01

Summary Due to genome instability, most cancers exhibit loss of regions containing tumor suppressor genes and collateral loss of other genes. To identify cancer-specific vulnerabilities that are the result of copy-number losses, we performed integrated analyses of genome-wide copy-number and RNAi profiles and identified 56 genes for which gene suppression specifically inhibited the proliferation of cells harboring partial copy-number loss of that gene. These CYCLOPS (Copy-number alterations Yielding Cancer Liabilities Owing to Partial losS) genes are enriched for spliceosome, proteasome and ribosome components. One CYCLOPS gene, PSMC2, encodes an essential member of the 19S proteasome. Normal cells express excess PSMC2, which resides in a complex with PSMC1, PSMD2, and PSMD5 and acts as a reservoir protecting cells from PSMC2 suppression. Cells harboring partial PSMC2 copy-number loss lack this complex and die after PSMC2 suppression. These observations define a distinct class of cancer-specific liabilities resulting from genome instability. PMID:22901813

Reprogramming cell fate with a genome-scale library of artificial transcription factors

PubMed Central

Eguchi, Asuka; Wleklinski, Matthew J.; Spurgat, Mackenzie C.; Heiderscheit, Evan A.; Kropornicka, Anna S.; Vu, Catherine K.; Bhimsaria, Devesh; Swanson, Scott A.; Stewart, Ron; Ramanathan, Parameswaran; Kamp, Timothy J.; Slukvin, Igor; Thomson, James A.; Dutton, James R.; Ansari, Aseem Z.

2016-01-01

Artificial transcription factors (ATFs) are precision-tailored molecules designed to bind DNA and regulate transcription in a preprogrammed manner. Libraries of ATFs enable the high-throughput screening of gene networks that trigger cell fate decisions or phenotypic changes. We developed a genome-scale library of ATFs that display an engineered interaction domain (ID) to enable cooperative assembly and synergistic gene expression at targeted sites. We used this ATF library to screen for key regulators of the pluripotency network and discovered three combinations of ATFs capable of inducing pluripotency without exogenous expression of Oct4 (POU domain, class 5, TF 1). Cognate site identification, global transcriptional profiling, and identification of ATF binding sites reveal that the ATFs do not directly target Oct4; instead, they target distinct nodes that converge to stimulate the endogenous pluripotency network. This forward genetic approach enables cell type conversions without a priori knowledge of potential key regulators and reveals unanticipated gene network dynamics that drive cell fate choices. PMID:27930301

41 CFR 301-10.121 - What classes of airline accommodations are available?

Code of Federal Regulations, 2011 CFR

2011-07-01

... airlines as “tourist class,” “economy class,” or as “single class” when the airline offers only one class... available, i.e., two “cabins”, with two distinctly different seating types (such as girth and pitch) and the... airline flight has only two cabins available but equips both with one type of seating, (i.e., seating...

Clustering of adversity in young adults on disability pension due to mental disorders: a latent class analysis.

PubMed

Joensuu, Matti; Mattila-Holappa, Pauliina; Ahola, Kirsi; Ervasti, Jenni; Kivimäki, Mika; Kivekäs, Teija; Koskinen, Aki; Vahtera, Jussi; Virtanen, Marianna

2016-02-01

Mental disorders are the leading cause of work disability among young adults. This study examined whether distinct classes could be identified among young adults on the basis of medical history before receiving a disability pension due to a mental disorder. Medical history was obtained from pension applications and attached medical certificates for 1163 individuals aged 18-34 years who, in 2008, received a disability pension due to a mental disorder. Using latent class analysis, 10 clinical and individual adversities and their associations with sex, age and diagnostic category were examined. Three classes were identified: childhood adversity (prevalence, 33%), comorbidity (23%), and undefined (44%). The childhood adversity class was characterized by adverse events and symptoms reported during childhood and it associated with depressive disorders. The comorbidity class was characterized by comorbid mental disorders, suicide attempts and substance abuse and associated with younger age and bipolar disorder. The undefined class formed no distinct profile; individuals in this class had the lowest number of adversities and it associated with psychotic disorders. The identification of subgroups characterized by childhood circumstances and comorbidity may help planning of prevention and support practices for young adults with mental disorders and risk of work disability.

The Educational Strategies of Danish University Students from Professional and Working-Class Backgrounds

ERIC Educational Resources Information Center

Thomsen, Jens Peter; Munk, Martin D.; Eiberg-Madsen, Misja; Hansen, Gro Inge

2013-01-01

This article studies the educational strategies adopted by university students from different class backgrounds in a Scandinavian welfare regime. Studies show distinct differences among classes relating to economic considerations, risk-averse behavior, and patterns of socialization among university students. We investigate these differences…

Children's Understanding of Social Class Differences.

ERIC Educational Resources Information Center

Naimark, Hedwin

Investigated were school children's recognition of social class distinctions, cues they used, and the relationship of their ideas about social class to other beliefs and attitudes about the social world. An ethnically mixed group of 201 male and female subjects participated in the study. Five age groups were represented: second, fifth, and sixth…

Is there Complex Trauma Experience typology for Australian's experiencing extreme social disadvantage and low housing stability?

PubMed

Keane, Carol A; Magee, Christopher A; Kelly, Peter J

2016-11-01

Traumatic childhood experiences predict many adverse outcomes in adulthood including Complex-PTSD. Understanding complex trauma within socially disadvantaged populations has important implications for policy development and intervention implementation. This paper examined the nature of complex trauma experienced by disadvantaged individuals using a latent class analysis (LCA) approach. Data were collected through the large-scale Journeys Home Study (N=1682), utilising a representative sample of individuals experiencing low housing stability. Data on adverse childhood experiences, adulthood interpersonal trauma and relevant covariates were collected through interviews at baseline (Wave 1). Latent class analysis (LCA) was conducted to identify distinct classes of childhood trauma history, which included physical assault, neglect, and sexual abuse. Multinomial logistic regression investigated childhood relevant factors associated with class membership such as biological relationship of primary carer at age 14 years and number of times in foster care. Of the total sample (N=1682), 99% reported traumatic adverse childhood experiences. The most common included witnessing of violence, threat/experience of physical abuse, and sexual assault. LCA identified six distinct childhood trauma history classes including high violence and multiple traumas. Significant covariate differences between classes included: gender, biological relationship of primary carer at age 14 years, and time in foster care. Identification of six distinct childhood trauma history profiles suggests there might be unique treatment implications for individuals living in extreme social disadvantage. Further research is required to examine the relationship between these classes of experience, consequent impact on adulthood engagement, and future transitions though homelessness. Copyright © 2016 Elsevier Ltd. All rights reserved.

Cytotoxic activity against human neuroblastoma and melanoma cells mediated by IgM antibodies derived from peripheral blood of healthy donors.

PubMed

Devarapu, Satish Kumar; Mamidi, Srinivas; Plöger, Frank; Dill, Othmar; Blixt, Ola; Kirschfink, Michael; Schwartz-Albiez, Reinhard

2016-06-15

A small percentage of healthy donors identified in the Western population carry antibodies in their peripheral blood which convey cytotoxic activity against certain human melanoma and neuroblastoma cell lines. We measured the cytotoxic activity of sera and plasmas from healthy donors on the human neuroblastoma cell line Kelly and various melanoma cell lines. Antibodies of IgM isotype, presumably belonging to the class of naturally occurring antibodies, exerted cytotoxic activity in a complement-dependent fashion. Apart from complement-dependent tumor cell lysis, we observed C3 opsonization in all tumor cell lines upon treatment with cytotoxic plasmas. Cell lines tested primarily expressed membrane complement regulatory proteins (mCRP) CD46, CD55 and CD59 to various extents. Blocking of mCRPs by monoclonal antibodies enhanced cell lysis and opsonization, though some melanoma cells remained resistant to complement attack. Epitopes recognized by cytotoxic antibodies were represented by gangliosides such as GD2 and GD3, as evidenced by cellular sialidase pretreatment and enhanced expression of distinct gangliosides. It remains to be clarified why only a small fraction of healthy persons carry these antitumor cytotoxic antibodies. © 2016 UICC.

The Split Personality of Glutamate Transporters: A Chloride Channel and a Transporter.

PubMed

Cater, Rosemary J; Ryan, Renae M; Vandenberg, Robert J

2016-03-01

Transporters and ion channels are conventionally categorised into distinct classes of membrane proteins. However, some membrane proteins have a split personality and can function as both transporters and ion channels. The excitatory amino acid transporters (EAATs) in particular, function as both glutamate transporters and chloride (Cl(-)) channels. The EAATs couple the transport of glutamate to the co-transport of three Na(+) ions and one H(+) ion into the cell, and the counter-transport of one K(+) ion out of the cell. The EAAT Cl(-) channel is activated by the binding of glutamate and Na(+), but is thermodynamically uncoupled from glutamate transport and involves molecular determinants distinct from those responsible for glutamate transport. Several crystal structures of an EAAT archaeal homologue, GltPh, at different stages of the transport cycle, alongside numerous functional studies and molecular dynamics simulations, have provided extensive insights into the mechanism of substrate transport via these transporters. However, the molecular determinants involved in Cl(-) permeation, and the mechanism by which this channel is activated are not entirely understood. Here we will discuss what is currently known about the molecular determinants involved in EAAT-mediated Cl(-) permeation and the mechanisms that underlie their split personality.

NetMHCstab – predicting stability of peptide–MHC-I complexes; impacts for cytotoxic T lymphocyte epitope discovery

PubMed Central

Jørgensen, Kasper W; Rasmussen, Michael; Buus, Søren; Nielsen, Morten

2014-01-01

Major histocompatibility complex class I (MHC-I) molecules play an essential role in the cellular immune response, presenting peptides to cytotoxic T lymphocytes (CTLs) allowing the immune system to scrutinize ongoing intracellular production of proteins. In the early 1990s, immunogenicity and stability of the peptide–MHC-I (pMHC-I) complex were shown to be correlated. At that time, measuring stability was cumbersome and time consuming and only small data sets were analysed. Here, we investigate this fairly unexplored area on a large scale compared with earlier studies. A recent small-scale study demonstrated that pMHC-I complex stability was a better correlate of CTL immunogenicity than peptide–MHC-I affinity. We here extended this study and analysed a total of 5509 distinct peptide stability measurements covering 10 different HLA class I molecules. Artificial neural networks were used to construct stability predictors capable of predicting the half-life of the pMHC-I complex. These predictors were shown to predict T-cell epitopes and MHC ligands from SYFPEITHI and IEDB to form significantly more stable MHC-I complexes compared with affinity-matched non-epitopes. Combining the stability predictions with a state-of-the-art affinity predictions NetMHCcons significantly improved the performance for identification of T-cell epitopes and ligands. For the HLA alleles included in the study, we could identify distinct sub-motifs that differentiate between stable and unstable peptide binders and demonstrate that anchor positions in the N-terminal of the binding motif (primarily P2 and P3) play a critical role for the formation of stable pMHC-I complexes. A webserver implementing the method is available at http://www.cbs.dtu.dk/services/NetMHCstab. PMID:23927693

Broad spectrum activity of a lectin-like bacterial serine protease family on human leukocytes.

PubMed

Ayala-Lujan, Jorge Luis; Vijayakumar, Vidhya; Gong, Mei; Smith, Rachel; Santiago, Araceli E; Ruiz-Perez, Fernando

2014-01-01

The serine protease autotransporter from Enterobacteriaceae (SPATE) family, which number more than 25 proteases with apparent diverse functions, have been phylogenetically divided into two distinct classes, designated 1 and 2. We recently demonstrated that Pic and Tsh, two members of the class-2 SPATE family produced by intestinal and extraintestinal pathogenic E. coli, were able to cleave a number of O-glycosylated proteins on neutrophils and lymphocytes resulting in impaired leukocyte functions. Here we show that most members of the class-2 SPATE family have lectin-like properties and exhibit differential protease activity reliant on glycoprotein type and cell lineage. Protease activity was seen in virtually all tested O-glycosylated proteins including CD34, CD55, CD164, TIM1, TIM3, TIM4 and C1-INH. We also show that although SPATE proteins bound and cleaved glycoproteins more efficiently on granulocytes and monocytes, they also targeted glycoproteins on B, T and natural killer lymphocytes. Finally, we found that the characteristic domain-2 of class-2 SPATEs is not required for glycoprotease activity, but single amino acid mutations in Pic domain-1 to those residues naturally occurring in domain-1 of SepA, were sufficient to hamper Pic glycoprotease activity. This study shows that most class-2 SPATEs have redundant activities and suggest that they may function as immunomodulators at several levels of the immune system.

Broad Spectrum Activity of a Lectin-Like Bacterial Serine Protease Family on Human Leukocytes

PubMed Central

Ayala-Lujan, Jorge Luis; Vijayakumar, Vidhya; Gong, Mei; Smith, Rachel; Santiago, Araceli E.; Ruiz-Perez, Fernando

2014-01-01

The serine protease autotransporter from Enterobacteriaceae (SPATE) family, which number more than 25 proteases with apparent diverse functions, have been phylogenetically divided into two distinct classes, designated 1 and 2. We recently demonstrated that Pic and Tsh, two members of the class-2 SPATE family produced by intestinal and extraintestinal pathogenic E. coli, were able to cleave a number of O-glycosylated proteins on neutrophils and lymphocytes resulting in impaired leukocyte functions. Here we show that most members of the class-2 SPATE family have lectin-like properties and exhibit differential protease activity reliant on glycoprotein type and cell lineage. Protease activity was seen in virtually all tested O-glycosylated proteins including CD34, CD55, CD164, TIM1, TIM3, TIM4 and C1-INH. We also show that although SPATE proteins bound and cleaved glycoproteins more efficiently on granulocytes and monocytes, they also targeted glycoproteins on B, T and natural killer lymphocytes. Finally, we found that the characteristic domain-2 of class-2 SPATEs is not required for glycoprotease activity, but single amino acid mutations in Pic domain-1 to those residues naturally occurring in domain-1 of SepA, were sufficient to hamper Pic glycoprotease activity. This study shows that most class-2 SPATEs have redundant activities and suggest that they may function as immunomodulators at several levels of the immune system. PMID:25251283

Cell-to-cell stimulation of movement in nonmotile mutants of Myxococcus

PubMed Central

Hodgkin, Jonathan; Kaiser, Dale

1977-01-01

A large number of nonmotile mutants of the gliding bacterium Myxococcus xanthus have been isolated and partly characterized. About [unk] of these mutants are conditional mutants of a novel kind: mutant cells become transiently motile after contact with nonmutant cells or with cells of a different mutant type. These “stimulatable” mutants fall into five phenotypic classes (types B, C, D, E, and F). Most mutants are nonstimulatable (type A) and never become motile, but type A cells (and wild-type cells) can stimulate cells of any of the other five types. Stimulatable mutants of different types are capable of stimulating each other. For example, in a mixture of B and C cells, both become motile. Linkage analysis using a generalized transducing phage has shown that each of types B, C, D, E, and F corresponds to a single distinct genetic locus. Type A mutants, by contrast, belong to at least 17 different loci. Stimulation depends on close apposition of interacting cells, because stimulation does not occur when contact between cells is prevented. It is possible that the stimulatable mutants are defective in components of the gliding mechanism that can be exchanged between cells. Alternatively, they may be defective in a system of cell communication controlling the coordinated cell movements observed in Myxococcus. Images PMID:16592422

Neurons innervating the lamina in the butterfly, Papilio xuthus.

PubMed

Hamanaka, Yoshitaka; Shibasaki, Hiromichi; Kinoshita, Michiyo; Arikawa, Kentaro

2013-05-01

The butterfly Papilio xuthus has compound eyes with three types of ommatidia. Each type houses nine spectrally heterogeneous photoreceptors (R1-R9) that are divided into six spectral classes: ultraviolet, violet, blue, green, red, and broad-band. Analysis of color discrimination has shown that P. xuthus uses the ultraviolet, blue, green, and red receptors for foraging. The ultraviolet and blue receptors are long visual fibers terminating in the medulla, whereas the green and red receptors are short visual fibers terminating in the lamina. This suggests that processing of wavelength information begins in the lamina in P. xuthus, unlike in flies. To establish the anatomical basis of color discrimination mechanisms, we examined neurons innervating the lamina by injecting neurobiotin into this neuropil. We found that in addition to photoreceptors and lamina monopolar cells, three distinct groups of cells project fibers into the lamina. Their cell bodies are located (1) at the anterior rim of the medulla, (2) between the proximal surface of the medulla and lobula plate, and (3) in the medulla cell body rind. Neurobiotin injection also labeled distinct terminals in medulla layers 1, 2, 3, 4 and 5. Terminals in layer 4 belong to the long visual fibers (R1, 2 and 9), while arbors in layers 1, 2 and 3 probably correspond to terminals of three subtypes of lamina monopolar cells, respectively. Immunocytochemistry coupled with neurobiotin injection revealed their transmitter candidates; neurons in (1) and a subset of neurons in (2) are immunoreactive to anti-serotonin and anti-γ-aminobutyric acid, respectively.

The late and dual origin of cerebrospinal fluid-contacting neurons in the mouse spinal cord.

PubMed

Petracca, Yanina L; Sartoretti, Maria Micaela; Di Bella, Daniela J; Marin-Burgin, Antonia; Carcagno, Abel L; Schinder, Alejandro F; Lanuza, Guillermo M

2016-03-01

Considerable progress has been made in understanding the mechanisms that control the production of specialized neuronal types. However, how the timing of differentiation contributes to neuronal diversity in the developing spinal cord is still a pending question. In this study, we show that cerebrospinal fluid-contacting neurons (CSF-cNs), an anatomically discrete cell type of the ependymal area, originate from surprisingly late neurogenic events in the ventral spinal cord. CSF-cNs are identified by the expression of the transcription factors Gata2 and Gata3, and the ionic channels Pkd2l1 and Pkd1l2. Contrasting with Gata2/3(+) V2b interneurons, differentiation of CSF-cNs is independent of Foxn4 and takes place during advanced developmental stages previously assumed to be exclusively gliogenic. CSF-cNs are produced from two distinct dorsoventral regions of the mouse spinal cord. Most CSF-cNs derive from progenitors circumscribed to the late-p2 and the oligodendrogenic (pOL) domains, whereas a second subset of CSF-cNs arises from cells bordering the floor plate. The development of these two subgroups of CSF-cNs is differentially controlled by Pax6, they adopt separate locations around the postnatal central canal and they display electrophysiological differences. Our results highlight that spatiotemporal mechanisms are instrumental in creating neural cell diversity in the ventral spinal cord to produce distinct classes of interneurons, motoneurons, CSF-cNs, glial cells and ependymal cells. © 2016. Published by The Company of Biologists Ltd.

Histone deacetylase inhibitors prevent p53-dependent and p53-independent Bax-mediated neuronal apoptosis through two distinct mechanisms.

PubMed

Uo, Takuma; Veenstra, Timothy D; Morrison, Richard S

2009-03-04

Pharmacological manipulation of protein acetylation levels by histone deacetylase (HDAC) inhibitors represents a novel therapeutic strategy to treat neurodegeneration as well as cancer. However, the molecular mechanisms that determine how HDAC inhibition exerts a protective effect in neurons as opposed to a cytotoxic action in tumor cells has not been elucidated. We addressed this issue in cultured postnatal mouse cortical neurons whose p53-dependent and p53-independent intrinsic apoptotic programs require the proapoptotic multidomain protein, Bax. Despite promoting nuclear p53 accumulation, Class I/II HDAC inhibitors (HDACIs) protected neurons from p53-dependent cell death induced by camptothecin, etoposide, heterologous p53 expression or the MDM2 inhibitor, nutlin-3a. HDACIs suppressed p53-dependent PUMA expression, a critical signaling intermediate linking p53 to Bax activation, thus preventing postmitochondrial events including cleavage of caspase-9 and caspase-3. In human SH-SY5Y neuroblastoma cells, however, HDACIs were not able to prevent p53-dependent cell death. Moreover, HDACIs also prevented caspase-3 cleavage in postnatal cortical neurons treated with staurosporine, 3-nitropropionic acid and a Bcl-2 inhibitor, all of which require the presence of Bax but not p53 to promote apoptosis. Although these three toxic agents displayed a requirement for Bax, they did not promote PUMA induction. These results demonstrate that HDACIs block Bax-dependent cell death by two distinct mechanisms to prevent neuronal apoptosis, thus identifying for the first time a defined molecular target for their neuroprotective actions.

Histone Deacetylase Inhibitors Prevent p53-dependent and Independent Bax-Mediated Neuronal Apoptosis Through Two Distinct Mechanisms

PubMed Central

Uo, Takuma; Veenstra, Timothy D.; Morrison, Richard S.

2009-01-01

Pharmacological manipulation of protein acetylation levels by histone deacetylase (HDAC) inhibitors represents a novel therapeutic strategy to treat neurodegeneration as well as cancer. However, the molecular mechanisms that determine how HDAC inhibition exerts a protective effect in neurons as opposed to a cytotoxic action in tumor cells has not been elucidated. We addressed this issue in cultured postnatal mouse cortical neurons whose p53-dependent and —independent intrinsic apoptotic programs require the pro-apoptotic multidomain protein, Bax. Despite promoting nuclear p53 accumulation, Class I/II HDAC inhibitors (HDACIs) protected neurons from p53-dependent cell death induced by camptothecin, etoposide, heterologous p53 expression or the MDM2 inhibitor, nutlin-3a. HDACIs suppressed p53-dependent PUMA expression, a critical signaling intermediate linking p53 to Bax activation, thus preventing post-mitochondrial events including cleavage of caspase-9 and -3. In human SH-SY5Y neuroblastoma cells, however, HDACIs were not able to prevent p53-dependent cell death. Moreover, HDACIs also prevented caspase-3 cleavage in postnatal cortical neurons treated with staurosporine, 3-nitropropionic acid and a Bcl-2 inhibitor, all of which require the presence of Bax but not p53 to promote apoptosis. Although these three toxic agents displayed a requirement for Bax, they did not promote PUMA induction. These results demonstrate that HDACIs block Bax-dependent cell death by two distinct mechanisms to prevent neuronal apoptosis, thus identifying for the first time a defined molecular target for their neuroprotective actions. PMID:19261878

Response of γδ T cells to plant-derived tannins

PubMed Central

Holderness, Jeff; Hedges, Jodi F.; Daughenbaugh, Katie; Kimmel, Emily; Graff, Jill; Freedman, Brett; Jutila, Mark A.

2008-01-01

Many pharmaceutical drugs are isolated from plants used in traditional medicines. Through screening plant extracts, both traditional medicines and compound libraries, new pharmaceutical drugs continue to be identified. Currently, two plant-derived agonists for γδ T cells are described. These plant-derived agonists impart innate effector functions upon distinct γδ T cell subsets. Plant tannins represent one class of γδ T cell agonist and preferentially activate the mucosal population. Mucosal γδ T cells function to modulate tissue immune responses and induce epithelium repair. Select tannins, isolated from apple peel, rapidly induce immune gene transcription in γδ T cells, leading to cytokine production and increased responsiveness to secondary signals. Activity of these tannin preparations tracks to the procyanidin fraction, with the procyanidin trimer (C1) having the most robust activity defined to date. The response to the procyanidins is evolutionarily conserved in that responses are seen with human, bovine, and murine γδ T cells. Procyanidin-induced responses described in this review likely account for the expansion of mucosal γδ T cells seen in mice and rats fed soluble extracts of tannins. Procyanidins may represent a novel approach for treatment of tissue damage, chronic infection, and autoimmune therpies. PMID:19166386

Structural basis for Epstein–Barr virus host cell tropism mediated by gp42 and gHgL entry glycoproteins

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

Sathiyamoorthy, Karthik; Hu, Yao Xiong; Möhl, Britta S.

Herpesvirus entry into host cells is mediated by multiple virally encoded receptor binding and membrane fusion glycoproteins. Despite their importance in host cell tropism and associated disease pathology, the underlying and essential interactions between these viral glycoproteins remain poorly understood. For Epstein–Barr virus (EBV), gHgL/gp42 complexes bind HLA class II to activate membrane fusion with B cells, but gp42 inhibits fusion and entry into epithelial cells. To clarify the mechanism by which gp42 controls the cell specificity of EBV infection, in this paper we determined the structure of gHgL/gp42 complex bound to an anti-gHgL antibody (E1D1). The critical regulator ofmore » EBV tropism is the gp42 N-terminal domain, which tethers the HLA-binding domain to gHgL by wrapping around the exterior of three gH domains. Both the gp42 N-terminal domain and E1D1 selectively inhibit epithelial-cell fusion; however, they engage distinct surfaces of gHgL. Finally, these observations clarify key determinants of EBV host cell tropism.« less

Structural basis for Epstein–Barr virus host cell tropism mediated by gp42 and gHgL entry glycoproteins

DOE PAGES

Sathiyamoorthy, Karthik; Hu, Yao Xiong; Möhl, Britta S.; ...

2016-12-08

Herpesvirus entry into host cells is mediated by multiple virally encoded receptor binding and membrane fusion glycoproteins. Despite their importance in host cell tropism and associated disease pathology, the underlying and essential interactions between these viral glycoproteins remain poorly understood. For Epstein–Barr virus (EBV), gHgL/gp42 complexes bind HLA class II to activate membrane fusion with B cells, but gp42 inhibits fusion and entry into epithelial cells. To clarify the mechanism by which gp42 controls the cell specificity of EBV infection, in this paper we determined the structure of gHgL/gp42 complex bound to an anti-gHgL antibody (E1D1). The critical regulator ofmore » EBV tropism is the gp42 N-terminal domain, which tethers the HLA-binding domain to gHgL by wrapping around the exterior of three gH domains. Both the gp42 N-terminal domain and E1D1 selectively inhibit epithelial-cell fusion; however, they engage distinct surfaces of gHgL. Finally, these observations clarify key determinants of EBV host cell tropism.« less

Longevity of T-cell memory following acute viral infection.

PubMed

Walker, Joshua M; Slifka, Mark K

2010-01-01

Investigation of T-cell-mediated immunity following acute viral infection represents an area of research with broad implications for both fundamental immunology research as well as vaccine development. Here, we review techniques that are used to assess T-cell memory including limiting dilution analysis, enzyme-linked immunospot (ELISPOT) assays, intracellular cytokine staining (ICCS) and peptide-MHC Class I tetramer staining. The durability of T-cell memory is explored in the context of several acute viral infections including vaccinia virus (VV), measles virus (MV) and yellow fever virus (YFV). Following acute infection, different virus-specific T-cell subpopulations exhibit distinct cytokine profiles and these profiles change over the course of infection. Differential regulation of the cytotoxic proteins, granzyme A, granzyme B and perforin are also observed in virus-specific T cells following infection. As a result of this work, we have gained a broader understanding of the kinetics and magnitude of antiviral T-cell immunity as well as new insight into the patterns of immunodominance and differential regulation of cytokines and cytotoxicity-associated molecules. This information may eventually lead to the generation of more effective vaccines that elicit T-cell memory with the optimal combination of functional characteristics required for providing protective immunity against infectious disease.

MHC class II expression in lung cancer.

PubMed

He, Yayi; Rozeboom, Leslie; Rivard, Christopher J; Ellison, Kim; Dziadziuszko, Rafal; Yu, Hui; Zhou, Caicun; Hirsch, Fred R

2017-10-01

Immunotherapy is an exciting development in lung cancer research. In this study we described major histocompatibility complex (MHC) Class II protein expression in lung cancer cell lines and patient tissues. We studied MHC Class II (DP, DQ, DR) (CR3/43, Abcam) protein expression in 55 non-small cell lung cancer (NSCLC) cell lines, 42 small cell lung cancer (SCLC) cell lines and 278 lung cancer patient tissues by immunohistochemistry (IHC). Seven (12.7%) NSCLC cell lines were positive for MHC Class II. No SCLC cell lines were found to be MHC Class II positive. We assessed 139 lung cancer samples available in the Hirsch Lab for MHC Class II. There was no positive MHC Class II staining on SCLC tumor cells. MHC Class II expression on TILs in SCLC was significantly lower than that on TILs in NSCLC (P＜0.001). MHC Class II was also assessed in an additional 139 NSCLC tumor tissues from Medical University of Gdansk, Poland. Patients with positive staining of MHC Class II on TILs had longer regression-free survival (RFS) and overall survival (OS) than those whose TILs were MHC Class II negative (2.980 years, 95% CI 1.628-4.332 vs. 1.050 years, 95% CI 0.556-1.554, P=0.028) (3.230 years, 95% CI 2.617-3.843 vs. 1.390 years, 95% CI 0.629-2.151, P=0.014). MHC Class II was expressed both in NSCLC cell lines and tissues. However, MHC Class II was not detected in SCLC cell lines or tissue tumor cells. MHC Class II expression was lower on SCLC TILs than on NSCLC TILs. Loss of expression of MHC Class II on SCLC tumor cells and reduced expression on SCLC TILs may be a means of escaping anti-cancer immunity. Higher MHC Class II expression on TILs was correlated with better prognosis in patients with NSCLC. Copyright © 2017. Published by Elsevier B.V.

Modular control of glutamatergic neuronal identity in C.elegans by distinct homeodomain proteins

PubMed Central

Serrano-Saiz, Esther; Poole, Richard J.; Felton, Terry; Zhang, Feifan; De La Cruz, Estanisla Daniel; Hobert, Oliver

2013-01-01

The choice of using one of many possible neurotransmitter systems is a critical step in defining the identity of an individual neuron type. We show here that the key defining feature of glutamatergic neurons, the vesicular glutamate transporter EAT-4/VGLUT is expressed in 38 of the 118 anatomically defined neuron classes of the C.elegans nervous system. We show that eat-4/VGLUT expression is controlled in a modular manner, with distinct cis-regulatory modules driving expression in distinct glutamatergic neuron classes. We identify 13 different transcription factors, 11 of them homeodomain proteins, that act in specific combinations in 25 different glutamatergic neuron classes to initiate and maintain eat-4/VGLUT expression. We show that the adoption of a glutamatergic phenotype is linked to the adoption of other terminal identity features of a neuron, including cotransmitter phenotypes. Examination of mouse orthologs of these homeodomain proteins resulted in the identification of mouse LHX1 as a regulator of glutamatergic neurons in the brainstem. PMID:24243022

The difference between emotion and affect. Comment on "The quartet theory of human emotions: An integrative and neurofunctional model" by S. Koelsch et al.

NASA Astrophysics Data System (ADS)

Cochrane, Tom

2015-06-01

Koelsch and colleagues have produced a fascinating model that convincingly argues for the existence of four neuroanatomically distinct systems involved in our emotions. As I understood it, the claim was that these systems build upon each other, each one introducing a new sophistication that allows a distinct class of emotional states to emerge. For instance, the hippocampus enables the generation of emotions related to long-term attachment as opposed to simple reward and punishment [1, Section 2.3.2]. However, it was uncertain why we should stop at four affect systems. Many different brain structures contribute an additional sophistication to emotional function; could there not be a class of emotions associated with each of these sophistications? How distinctive does the new function, or the new class of emotions have to be to qualify? Should we be on the lookout for neural structures associated with meta-emotions [2] or epistemic emotions [3] as well?

Detecting Math Anxiety with a Mixture Partial Credit Model

ERIC Educational Resources Information Center

Ölmez, Ibrahim Burak; Cohen, Allan S.

2017-01-01

The purpose of this study was to investigate a new methodology for detection of differences in middle grades students' math anxiety. A mixture partial credit model analysis revealed two distinct latent classes based on homogeneities in response patterns within each latent class. Students in Class 1 had less anxiety about apprehension of math…

Thoughts of Death and Suicide in Early Adolescence

ERIC Educational Resources Information Center

Vander Stoep, Ann; McCauley, Elizabeth; Flynn, Cynthia; Stone, Andrea

2009-01-01

The prevalence and persistence of thoughts of death and suicide during early adolescence were estimated in a community-based cohort. A latent class approach was used to identify distinct subgroups based on endorsements to depression items administered repeatedly over 24 months. Two classes emerged, with 75% in a low ideation class across four…

Preschool Children Use Linguistic Form Class and Pragmatic Cues To Interpret Generics.

ERIC Educational Resources Information Center

Gelman, Susan A.; Raman, Lakshmi

2003-01-01

Five studies examined preschoolers' understanding of linguistic form class and pragmatic context in presence of a single exemplar or multiexemplars. Data indicated that by 2 years, children use linguistic form class, and by age 3, use pragmatic context. Young children have begun to understand the distinction between generic and nongeneric noun…

Classes in the Balance: Latent Class Analysis and the Balance Scale Task

ERIC Educational Resources Information Center

Boom, Jan; ter Laak, Jan

2007-01-01

Latent class analysis (LCA) has been successfully applied to tasks measuring higher cognitive functioning, suggesting the existence of distinct strategies used in such tasks. With LCA it became possible to classify post hoc. This important step forward in modeling and analyzing cognitive strategies is relevant to the overlapping waves model for…

Gender and Social Class Differences in Japanese Mothers' Beliefs about Children's Education and Socialisation

ERIC Educational Resources Information Center

Yamamoto, Yoko

2016-01-01

Despite increasing rates of university attendance among women, a significant gender gap remains in socialisation and educational processes in Japan. To understand why and how gender-distinctive socialisation processes persist, this study aimed to examine both middle-class and working-class mothers' beliefs about gender, education, and children's…

Attentional Requirements for the Selection of Words from Different Grammatical Categories

ERIC Educational Resources Information Center

Ayora, Pauline; Janssen, Niels; Dell'Acqua, Roberto; Alario, F.-Xavier

2009-01-01

Two grammatical classes are commonly distinguished in psycholinguistic research. The open-class includes content words such as nouns, whereas the closed-class includes function words such as determiners. A standing issue is to identify whether these words are retrieved through similar or distinct selection mechanisms. We report a comparative…

Acquiring Word Class Distinctions in American Sign Language: Evidence from Handshape

ERIC Educational Resources Information Center

Brentari, Diane; Coppola, Marie; Jung, Ashley; Goldin-Meadow, Susan

2013-01-01

Handshape works differently in nouns versus a class of verbs in American Sign Language (ASL) and thus can serve as a cue to distinguish between these two word classes. Handshapes representing characteristics of the object itself ("object" handshapes) and handshapes representing how the object is handled ("handling" handshapes)…

Latent Class Analysis of Peer Conformity: Who Is Yielding to Pressure and Why?

ERIC Educational Resources Information Center

Kosten, Paul A.; Scheier, Lawrence M.; Grenard, Jerry L.

2013-01-01

This study used latent class analysis to examine typologies of peer conformity in a community sample of middle school students. Students responded to 31 items assessing diverse facets of conformity dispositions. The most parsimonious model produced three qualitatively distinct classes that differed on the basis of conformity to recreational…

Studying Psychosocial Barriers to Drug Treatment Among Chinese Methamphetamine Users Using A 3-Step Latent Class Analysis.

PubMed

Wang, Jichuan; Kelly, Brian C; Liu, Tieqiao; Hao, Wei

2016-03-01

Given the growth in methamphetamine use in China during the 21st century, we assessed perceived psychosocial barriers to drug treatment among this population. Using a sample of 303 methamphetamine users recruited via Respondent Driven Sampling, we use Latent Class Analysis (LCA) to identify possible distinct latent groups among Chinese methamphetamine users on the basis of their perceptions of psychosocial barriers to drug treatment. After covariates were included to predict latent class membership, the 3-step modeling approach was applied. Our findings indicate that the Chinese methamphetamine using population was heterogeneous on perceptions of drug treatment barriers; four distinct latent classes (subpopulations) were identified--Unsupported Deniers, Deniers, Privacy Anxious, and Low Barriers--and individual characteristics shaped the probability of class membership. Efforts to link Chinese methamphetamine users to treatment may require a multi-faceted approach that attends to differing perceptions about impediments to drug treatment. Copyright © 2015. Published by Elsevier Inc.

Binding and activation of major histocompatibility complex class II-deficient macrophages by staphylococcal exotoxins

NASA Technical Reports Server (NTRS)

Beharka, A. A.; Armstrong, J. W.; Iandolo, J. J.; Chapes, S. K.; Spooner, B. S. (Principal Investigator)

1994-01-01

Macrophages from C2D transgenic mice deficient in the expression of major histocompatibility complex (MHC) class II proteins were used to identify binding sites for superantigens distinct from the MHC class II molecule. Iodinated staphylococcal enterotoxins A and B (SEA and SEB) and exfoliative toxins A and B (ETA and ETB) bound to C2D macrophages in a concentration-dependent and competitive manner. All four toxins increased F-actin concentration within 30 s of their addition to C2D macrophages, indicating that signal transduction occurred in response to toxin in the absence of class II MHC. Furthermore, ETA, ETB, SEA, and, to a lesser extent, SEB induced C2D macrophages to produce interleukin 6. Several molecular species on C2D macrophages with molecular masses of 140, 97, 61, 52, 43, and 37 kDa bound SEA in immunoprecipitation experiments. These data indicate the presence of novel, functionally active toxin binding sites on murine macrophages distinct from MHC class II molecules.

Distinct mutations in yeast TAF(II)25 differentially affect the composition of TFIID and SAGA complexes as well as global gene expression patterns.

PubMed

Kirschner, Doris B; vom Baur, Elmar; Thibault, Christelle; Sanders, Steven L; Gangloff, Yann-Gaël; Davidson, Irwin; Weil, P Anthony; Tora, Làszlò

2002-05-01

The RNA polymerase II transcription factor TFIID, composed of the TATA-binding protein (TBP) and TBP-associated factors (TAF(II)s), nucleates preinitiation complex formation at protein-coding gene promoters. SAGA, a second TAF(II)-containing multiprotein complex, is involved in transcription regulation in Saccharomyces cerevisiae. One of the essential protein components common to SAGA and TFIID is yTAF(II)25. We define a minimal evolutionarily conserved 91-amino-acid region of TAF(II)25 containing a histone fold domain that is necessary and sufficient for growth in vivo. Different temperature-sensitive mutations of yTAF(II)25 or chimeras with the human homologue TAF(II)30 arrested cell growth at either the G(1) or G(2)/M cell cycle phase and displayed distinct phenotypic changes and gene expression patterns. Immunoprecipitation studies revealed that TAF(II)25 mutation-dependent gene expression and phenotypic changes correlated at least partially with the integrity of SAGA and TFIID. Genome-wide expression analysis revealed that the five TAF(II)25 temperature-sensitive mutant alleles individually affect the expression of between 18 and 33% of genes, whereas taken together they affect 64% of all class II genes. Thus, different yTAF(II)25 mutations induce distinct phenotypes and affect the regulation of different subsets of genes, demonstrating that no individual TAF(II) mutant allele reflects the full range of its normal functions.

Tri-city study of Ecstasy use problems: a latent class analysis.

PubMed

Scheier, Lawrence M; Ben Abdallah, Arbi; Inciardi, James A; Copeland, Jan; Cottler, Linda B

2008-12-01

This study used latent class analysis to examine distinctive subtypes of Ecstasy users based on 24 abuse and dependence symptoms underlying standard DSM-IV criteria. Data came from a three site, population-based, epidemiological study to examine diagnostic nosology for Ecstasy use. Subject inclusion criteria included lifetime Ecstasy use exceeding five times and once in the past year, with participants ranging in age between 16 and 47 years of age from St. Louis, Miami, U.S. and Sydney, Australia. A satisfactory model typified four latent classes representing clearly differentiated diagnostic clusters including: (1) a group of sub-threshold users endorsing few abuse and dependence symptoms (negatives), (2) a group of 'diagnostic orphans' who had characteristic features of dependence for a select group of symptoms (mild dependent), (3) a 'transitional group' mimicking the orphans with regard to their profile of dependence also but reporting some abuse symptoms (moderate dependent), and (4) a 'severe dependent' group with a distinct profile of abuse and dependence symptoms. A multinomial logistic regression model indicated that certain latent classes showed unique associations with external non-diagnostic markers. Controlling for demographic characteristics and lifetime quantity of Ecstasy pill use, criminal behavior and motivational cues for Ecstasy use were the most efficient predictors of cluster membership. This study reinforces the heuristic utility of DSM-IV criteria applied to Ecstasy but with a different collage of symptoms that produced four distinct classes of Ecstasy users.

VE-cadherin expression allows identification of a new class of hematopoietic stem cells within human embryonic liver.

PubMed

Oberlin, Estelle; Fleury, Maud; Clay, Denis; Petit-Cocault, Laurence; Candelier, Jean-Jacques; Mennesson, Benoît; Jaffredo, Thierry; Souyri, Michèle

2010-11-25

Edification of the human hematopoietic system during development is characterized by the production of waves of hematopoietic cells separated in time, formed in distinct embryonic sites (ie, yolk sac, truncal arteries including the aorta, and placenta). The embryonic liver is a major hematopoietic organ wherein hematopoietic stem cells (HSCs) expand, and the future, adult-type, hematopoietic cell hierarchy becomes established. We report herein the identification of a new, transient, and rare cell population in the human embryonic liver, which coexpresses VE-cadherin, an endothelial marker, CD45, a pan-hematopoietic marker, and CD34, a common endothelial and hematopoietic marker. This population displays an outstanding self-renewal, proliferation, and differentiation potential, as detected by in vitro and in vivo hematopoietic assays compared with its VE-cadherin negative counterpart. Based on VE-cadherin expression, our data demonstrate the existence of 2 phenotypically and functionally separable populations of multipotent HSCs in the human embryo, the VE-cadherin(+) one being more primitive than the VE-cadherin(-) one, and shed a new light on the hierarchical organization of the embryonic liver HSC compartment.

Requirements for efficient cell-type proportioning: regulatory timescales, stochasticity and lateral inhibition

NASA Astrophysics Data System (ADS)

Pfeuty, B.; Kaneko, K.

2016-04-01

The proper functioning of multicellular organisms requires the robust establishment of precise proportions between distinct cell types. This developmental differentiation process typically involves intracellular regulatory and stochastic mechanisms to generate cell-fate diversity as well as intercellular signaling mechanisms to coordinate cell-fate decisions at tissue level. We thus surmise that key insights about the developmental regulation of cell-type proportion can be captured by the modeling study of clustering dynamics in population of inhibitory-coupled noisy bistable systems. This general class of dynamical system is shown to exhibit a very stable two-cluster state, but also metastability, collective oscillations or noise-induced state hopping, which can prevent from timely and reliably reaching a robust and well-proportioned clustered state. To circumvent these obstacles or to avoid fine-tuning, we highlight a general strategy based on dual-time positive feedback loops, such as mediated through transcriptional versus epigenetic mechanisms, which improves proportion regulation by coordinating early and flexible lineage priming with late and firm commitment. This result sheds new light on the respective and cooperative roles of multiple regulatory feedback, stochasticity and lateral inhibition in developmental dynamics.

Specification of anteroposterior cell fates in Caenorhabditis elegans by Drosophila Hox proteins.

PubMed

Hunter, C P; Kenyon, C

1995-09-21

Antennapedia class homeobox (Hox) genes specify cell fates in successive anteroposterior body domains in vertebrates, insects and nematodes. The DNA-binding homeodomain sequences are very similar between vertebrate and Drosophila Hox proteins, and this similarity allows vertebrate Hox proteins to function in Drosophila. In contrast, the Caenorhabditis elegans homeodomains are substantially divergent. Further, C. elegans differs from both insects and vertebrates in having a non-segmented body as well as a distinctive mode of development that involves asymmetric early cleavages and invariant cell lineages. Here we report that, despite these differences, Drosophila Hox proteins expressed in C. elegans can substitute for C. elegans Hox proteins in the control of three different cell-fate decisions: the regulation of cell migration, the specification of serotonergic neurons, and the specification of a sensory structure. We also show that the specificity of one C. elegans Hox protein is partly determined by two amino acids that have been implicated in sequence-specific DNA binding. Together these findings suggest that factors important for target recognition by specific Hox proteins have been conserved throughout much of the animal kingdom.

Functional organization of the transcriptome in human brain

PubMed Central

Oldham, Michael C; Konopka, Genevieve; Iwamoto, Kazuya; Langfelder, Peter; Kato, Tadafumi; Horvath, Steve; Geschwind, Daniel H

2009-01-01

The enormous complexity of the human brain ultimately derives from a finite set of molecular instructions encoded in the human genome. These instructions can be directly studied by exploring the organization of the brain’s transcriptome through systematic analysis of gene coexpression relationships. We analyzed gene coexpression relationships in microarray data generated from specific human brain regions and identified modules of coexpressed genes that correspond to neurons, oligodendrocytes, astrocytes and microglia. These modules provide an initial description of the transcriptional programs that distinguish the major cell classes of the human brain and indicate that cell type–specific information can be obtained from whole brain tissue without isolating homogeneous populations of cells. Other modules corresponded to additional cell types, organelles, synaptic function, gender differences and the subventricular neurogenic niche. We found that subventricular zone astrocytes, which are thought to function as neural stem cells in adults, have a distinct gene expression pattern relative to protoplasmic astrocytes. Our findings provide a new foundation for neurogenetic inquiries by revealing a robust and previously unrecognized organization to the human brain transcriptome. PMID:18849986

Parsimony and complexity: Cell fate assignment in the developing Drosophila eye.

PubMed

Mavromatakis, Yannis Emmanuel; Tomlinson, Andrew

2017-07-03

The specification of the R7 photoreceptor in the Drosophila eye has become a classic model for understanding how cell fates are assigned in developing systems. R7 is derived from a group of cells that also gives rise to the R1/6 photoreceptor class and the non-photoreceptor cone cells. Our studies examine the signals and cellular information that direct each of these cell types. The cell fates are directed by the combined actions of the Receptor Tyrosine Kinase (RTK) and Notch (N) signaling pathways. The RTK pathway acts to remove the transcription factor Tramtrack (Ttk) which represses the photoreceptor fate. If a cell receives an RTK signal sufficient to remove Ttk then the photoreceptor fate is specified; if not, the cone cell fate results. If Ttk is removed from a cell and its N activity is high then it is specified as an R7, but if its N activity is low then it becomes an R1/6 class photoreceptor. Thus, a remarkably simple molecular code underlies the specification of the fates: 1. Ttk degraded or not: 2. N activity high or low. In the R1/6 and cone cell precursors the molecular codes are achieved with relative simplicity but in the R7 precursor, manifold interactions occur between the RTK and N pathways, and to-date we have identified 4 distinct roles played by N in R7 fate specification. In this review we detail this molecular complexity, and describe how the RTK/N pathway crosstalk eventually leads to the simple molecular code of Tramtrack removed and N activity high. Furthermore, we describe the role played by the transcription factor Lozenge (Lz) in directing retinal precursor fates, and how the RTK/N signals specify different retinal cell types depending on the presence or absence of Lz.

Differential regulation of microtubule severing by APC underlies distinct patterns of projection neuron and interneuron migration

PubMed Central

Eom, Tae-Yeon; Stanco, Amelia; Guo, Jiami; Wilkins, Gary; Deslauriers, Danielle; Yan, Jessica; Monckton, Chase; Blair, Josh; Oon, Eesim; Perez, Abby; Salas, Eduardo; Oh, Adrianna; Ghukasyan, Vladimir; Snider, William D.; Rubenstein, John L. R.; Anton, E. S.

2014-01-01

Coordinated migration of distinct classes of neurons to appropriate positions leads to the formation of functional neuronal circuitry in the cerebral cortex. Two major classes of cortical neurons, interneurons and projection neurons, utilize distinctly different modes (radial vs. tangential) and routes of migration to arrive at their final positions in the cerebral cortex. Here, we show that adenomatous polyposis coli (APC) modulates microtubule (MT) severing in interneurons to facilitate tangential mode of interneuron migration, but not the glial-guided, radial migration of projection neurons. APC regulates the stability and activity of the MT severing protein p60-katanin in interneurons to promote the rapid remodeling of neuronal processes necessary for interneuron migration. These findings reveal how severing and restructuring of MTs facilitate distinct modes of neuronal migration necessary for laminar organization of neurons in the developing cerebral cortex. PMID:25535916

Non-equivalence of Wnt and R-spondin ligands during Lgr5+ intestinal stem-cell self-renewal.

PubMed

Yan, Kelley S; Janda, Claudia Y; Chang, Junlei; Zheng, Grace X Y; Larkin, Kathryn A; Luca, Vincent C; Chia, Luis A; Mah, Amanda T; Han, Arnold; Terry, Jessica M; Ootani, Akifumi; Roelf, Kelly; Lee, Mark; Yuan, Jenny; Li, Xiao; Bolen, Christopher R; Wilhelmy, Julie; Davies, Paige S; Ueno, Hiroo; von Furstenberg, Richard J; Belgrader, Phillip; Ziraldo, Solongo B; Ordonez, Heather; Henning, Susan J; Wong, Melissa H; Snyder, Michael P; Weissman, Irving L; Hsueh, Aaron J; Mikkelsen, Tarjei S; Garcia, K Christopher; Kuo, Calvin J

2017-05-11

The canonical Wnt/β-catenin signalling pathway governs diverse developmental, homeostatic and pathological processes. Palmitoylated Wnt ligands engage cell-surface frizzled (FZD) receptors and LRP5 and LRP6 co-receptors, enabling β-catenin nuclear translocation and TCF/LEF-dependent gene transactivation. Mutations in Wnt downstream signalling components have revealed diverse functions thought to be carried out by Wnt ligands themselves. However, redundancy between the 19 mammalian Wnt proteins and 10 FZD receptors and Wnt hydrophobicity have made it difficult to attribute these functions directly to Wnt ligands. For example, individual mutations in Wnt ligands have not revealed homeostatic phenotypes in the intestinal epithelium-an archetypal canonical, Wnt pathway-dependent, rapidly self-renewing tissue, the regeneration of which is fueled by proliferative crypt Lgr5 + intestinal stem cells (ISCs). R-spondin ligands (RSPO1-RSPO4) engage distinct LGR4-LGR6, RNF43 and ZNRF3 receptor classes, markedly potentiate canonical Wnt/β-catenin signalling, and induce intestinal organoid growth in vitro and Lgr5 + ISCs in vivo. However, the interchangeability, functional cooperation and relative contributions of Wnt versus RSPO ligands to in vivo canonical Wnt signalling and ISC biology remain unknown. Here we identify the functional roles of Wnt and RSPO ligands in the intestinal crypt stem-cell niche. We show that the default fate of Lgr5 + ISCs is to differentiate, unless both RSPO and Wnt ligands are present. However, gain-of-function studies using RSPO ligands and a new non-lipidated Wnt analogue reveal that these ligands have qualitatively distinct, non-interchangeable roles in ISCs. Wnt proteins are unable to induce Lgr5 + ISC self-renewal, but instead confer a basal competency by maintaining RSPO receptor expression that enables RSPO ligands to actively drive and specify the extent of stem-cell expansion. This functionally non-equivalent yet cooperative interaction between Wnt and RSPO ligands establishes a molecular precedent for regulation of mammalian stem cells by distinct priming and self-renewal factors, with broad implications for precise control of tissue regeneration.

The D0 immunoglobulin-like domain plays a central role for the stronger binding of KIR3DL2 to B27 free heavy chain dimers

PubMed Central

Hatano, Hiroko; Shaw, Jacqueline; Marquardt, Kaitlin; Zhang, Zhiyong; Gauthier, Laurent; Chanteux, Stephanie; Rossi, Benjamin; Li, Demin; Mitchell, Julie; Kollnberger, Simon

2015-01-01

We have proposed that the killer cell immunoglobulin-like receptor KIR3DL2 binding more strongly to HLA-B27 (B27) β2m-free heavy chain (FHC) dimers regulates lymphocyte function in arthritis and infection. We compared the function of B27 FHC dimers with other class I heavy chains and identified contact residues in KIR3DL2. B27 FHC dimers interacted functionally with KIR3DL2 on NK and reporter cells more strongly than other class I FHC. Mutagenesis identified key residues in the D0 and other immunoglobulin-like domains which were shared and distinct from KIR3DL1, for KIR3DL2 binding to B27 and other class I FHC. We modeled B27 dimer binding to KIR3DL2 and compared experimental mutagenesis data with computational “hot spot” predictions. Modelling predicts the stronger binding of B27 dimers to KIR3DL2 is mediated by non-symmetrical complementary contacts of the D0 and D1 domains with the α1, α2 and α3 domains of both B27 heavy chains. By contrast, the D2 domain primarily contacts residues in the α2 domain of one B27 heavy chain. These findings both provide novel insights about the molecular basis of KIR3DL2 binding to HLA-B27 and other ligands and suggest an important role for KIR3DL2 HLA-B27 interactions in controlling the function of NK cells in HLA-B27+ individuals. PMID:25582852

GWAS of Follicular Lymphoma Reveals Allelic Heterogeneity at 6p21.32 and Suggests Shared Genetic Susceptibility with Diffuse Large B-cell Lymphoma

PubMed Central

Skibola, Christine F.; Darabi, Hatef; Conde, Lucia; Hjalgrim, Henrik; Kumar, Vikrant; Chang, Ellen T.; Rothman, Nathaniel; Cerhan, James R.; Brooks-Wilson, Angela R.; Rehnberg, Emil; Irwan, Ishak D.; Ryder, Lars P.; Brown, Peter N.; Bracci, Paige M.; Agana, Luz; Riby, Jacques; Cozen, Wendy; Davis, Scott; Hartge, Patricia; Morton, Lindsay M.; Severson, Richard K.; Wang, Sophia S.; Slager, Susan L.; Fredericksen, Zachary S.; Novak, Anne J.; Kay, Neil E.; Habermann, Thomas M.; Armstrong, Bruce; Kricker, Anne; Milliken, Sam; Purdue, Mark P.; Vajdic, Claire M.; Boyle, Peter; Lan, Qing; Zahm, Shelia H.; Zhang, Yawei; Zheng, Tongzhang; Leach, Stephen; Spinelli, John J.; Smith, Martyn T.; Chanock, Stephen J.; Padyukov, Leonid; Alfredsson, Lars; Klareskog, Lars; Glimelius, Bengt; Melbye, Mads; Liu, Edison T.; Adami, Hans-Olov; Humphreys, Keith; Liu, Jianjun

2011-01-01

Non-Hodgkin lymphoma (NHL) represents a diverse group of hematological malignancies, of which follicular lymphoma (FL) is a prevalent subtype. A previous genome-wide association study has established a marker, rs10484561 in the human leukocyte antigen (HLA) class II region on 6p21.32 associated with increased FL risk. Here, in a three-stage genome-wide association study, starting with a genome-wide scan of 379 FL cases and 791 controls followed by validation in 1,049 cases and 5,790 controls, we identified a second independent FL–associated locus on 6p21.32, rs2647012 (ORcombined = 0.64, Pcombined = 2×10−21) located 962 bp away from rs10484561 (r2<0.1 in controls). After mutual adjustment, the associations at the two SNPs remained genome-wide significant (rs2647012:ORadjusted = 0.70, Padjusted = 4×10−12; rs10484561:ORadjusted = 1.64, Padjusted = 5×10−15). Haplotype and coalescence analyses indicated that rs2647012 arose on an evolutionarily distinct haplotype from that of rs10484561 and tags a novel allele with an opposite (protective) effect on FL risk. Moreover, in a follow-up analysis of the top 6 FL–associated SNPs in 4,449 cases of other NHL subtypes, rs10484561 was associated with risk of diffuse large B-cell lymphoma (ORcombined = 1.36, Pcombined = 1.4×10−7). Our results reveal the presence of allelic heterogeneity within the HLA class II region influencing FL susceptibility and indicate a possible shared genetic etiology with diffuse large B-cell lymphoma. These findings suggest that the HLA class II region plays a complex yet important role in NHL. PMID:21533074

2-(4-aminophenyl) benzothiazole: a potent and selective pharmacophore with novel mechanistic action towards various tumour cell lines.

PubMed

Dubey, Raghvendra; Shrivastava, Prabhat K; Basniwal, Pawan K; Bhattacharya, Snehendu; Moorthy, Narayana S Hari Narayana

2006-06-01

2-(4-aminophenyl) benzothiazole (CJM -126) (Table 1 (1) and its analogues represent a potent and highly selective class of antitumor agents. These compounds in nanomolar range elicit potent growth inhibition in human-derived breast, colon, ovarian and renal tumour cell lines. Metabolism of benzothiazole plays a central role in its mode of action. Cytocrome P450 isoform, CYP1A1, biotransforms benzothiazoles, to active, as well as inactive metabolites. In vitro studies had confirmed that N-oxidation and N-acetylation (only 3' halogen congener) as main active metabolic transformation (generating cytotoxic electrophilic species), while C-6 oxidation and N-acetylation (except 3' halogen congener) as inactive metabolic transformation pathway. Generation of an inactive metabolite 2-(4-aminophenyl)-6-hydoxybenzothiazole [6-OH 126, (Table 1) (10)] is blocked by fluorinated analogue, substituted around benzothiazole nucleus, especially at 5-position. National Cancer Institute (NCI), USA, confirms this series as a unique mechanistic class distinct from clinically used chemotherapeutic agents. Benzothiazoles are potent aryl hydrocarbon receptor (AhR) agonists, binding to AhR results in induction of CYP1A1, causes generation of electrophilic reactive species which forms DNA adduct, ultimately resulting in cell death by activation of apoptotic machinery. To overcome the poor physiochemical and pharmaceutical properties (bioavailability problem) of this compounds, prodrug of benzothiazole derivatives were synthesized, which are introduced in clinical trails.

The class III PI(3)K Vps34 promotes autophagy and endocytosis but not TOR signaling in Drosophila.

PubMed

Juhász, Gábor; Hill, Jahda H; Yan, Ying; Sass, Miklós; Baehrecke, Eric H; Backer, Jonathan M; Neufeld, Thomas P

2008-05-19

Degradation of cytoplasmic components by autophagy requires the class III phosphatidylinositol 3 (PI(3))-kinase Vps34, but the mechanisms by which this kinase and its lipid product PI(3) phosphate (PI(3)P) promote autophagy are unclear. In mammalian cells, Vps34, with the proautophagic tumor suppressors Beclin1/Atg6, Bif-1, and UVRAG, forms a multiprotein complex that initiates autophagosome formation. Distinct Vps34 complexes also regulate endocytic processes that are critical for late-stage autophagosome-lysosome fusion. In contrast, Vps34 may also transduce activating nutrient signals to mammalian target of rapamycin (TOR), a negative regulator of autophagy. To determine potential in vivo functions of Vps34, we generated mutations in the single Drosophila melanogaster Vps34 orthologue, causing cell-autonomous disruption of autophagosome/autolysosome formation in larval fat body cells. Endocytosis is also disrupted in Vps34(-/-) animals, but we demonstrate that this does not account for their autophagy defect. Unexpectedly, TOR signaling is unaffected in Vps34 mutants, indicating that Vps34 does not act upstream of TOR in this system. Instead, we show that TOR/Atg1 signaling regulates the starvation-induced recruitment of PI(3)P to nascent autophagosomes. Our results suggest that Vps34 is regulated by TOR-dependent nutrient signals directly at sites of autophagosome formation.

The Phragmoplast-Orienting Kinesin-12 Class Proteins Translate the Positional Information of the Preprophase Band to Establish the Cortical Division Zone in Arabidopsis thaliana[C][W

PubMed Central

Lipka, Elisabeth; Gadeyne, Astrid; Stöckle, Dorothee; Zimmermann, Steffi; De Jaeger, Geert; Ehrhardt, David W.; Kirik, Viktor; Van Damme, Daniel; Müller, Sabine

2014-01-01

The preprophase band (PPB) is a faithful but transient predictor of the division plane in somatic cell divisions. Throughout mitosis the PPBs positional information is preserved by factors that continuously mark the division plane at the cell cortex, the cortical division zone, by their distinct spatio-temporal localization patterns. However, the mechanism maintaining these identity factors at the plasma membrane after PPB disassembly remains obscure. The pair of kinesin-12 class proteins PHRAGMOPLAST ORIENTING KINESIN1 (POK1) and POK2 are key players in division plane maintenance. Here, we show that POK1 is continuously present at the cell cortex, providing a spatial reference for the site formerly occupied by the PPB. Fluorescence recovery after photobleaching analysis combined with microtubule destabilization revealed dynamic microtubule-dependent recruitment of POK1 to the PPB during prophase, while POK1 retention at the cortical division zone in the absence of cortical microtubules appeared static. POK function is strictly required to maintain the division plane identity factor TANGLED (TAN) after PPB disassembly, although POK1 and TAN recruitment to the PPB occur independently during prophase. Together, our data suggest that POKs represent fundamental early anchoring components of the cortical division zone, translating and preserving the positional information of the PPB by maintaining downstream identity markers. PMID:24972597

The role of eigenvalues in linear feature selection theory

NASA Technical Reports Server (NTRS)

Brown, D. R.; Omalley, M. J.

1976-01-01

A particular measure of pattern class distinction called the average interclass divergence, or more simply, divergence, is considered. Here divergence will be the pairwise average of the expected interclass divergence derived from Hajek's two-class divergence.

RNA splicing regulates the temporal order of TNF-induced gene expression.

PubMed

Hao, Shengli; Baltimore, David

2013-07-16

When cells are induced to express inflammatory genes by treatment with TNF, the mRNAs for the induced genes appear in three distinct waves, defining gene groups I, II, and III, or early, intermediate, and late genes. To examine the basis for these different kinetic classes, we have developed a PCR-based procedure to distinguish pre-mRNAs from mRNAs. It shows that the three groups initiate transcription virtually simultaneously but that delays in splicing characterize groups II and III. We also examined the elongation times, concluding that pre-mRNA synthesis is coordinate but splicing differences directly regulate the timing of mRNA production.

Structural basis of viral invasion: lessons from paramyxovirus F

PubMed Central

Lamb, Robert A.; Jardetzky, Theodore S.

2007-01-01

Summary The structures of glycoproteins that mediate enveloped virus entry into cells have revealed dramatic structural changes that accompany membrane fusion and provided mechanistic insights into this process. The group of class I viral fusion proteins includes the influenza hemagglutinin, paramyxovirus F, HIV env and other mechanistically related fusogens, but these proteins are unrelated in sequence and exhibit clearly distinct structural features. Recently determined crystal structures of the paramyxovirus F protein in two conformations, representing prefusion and postfusion states, reveal a novel protein architecture that undergoes large-scale, irreversible refolding during membrane fusion, extending our understanding of this diverse group of membrane fusion machines. PMID:17870467

Lay Americans' views of why scientists disagree with each other.

PubMed

Johnson, Branden B; Dieckmann, Nathan F

2017-10-01

A survey experiment assessed response to five explanations of scientific disputes: problem complexity, self-interest, values, competence, and process choices (e.g. theories and methods). A US lay sample ( n = 453) did not distinguish interests from values, nor competence from process, as explanations of disputes. Process/competence was rated most likely and interests/values least; all, on average, were deemed likely to explain scientific disputes. Latent class analysis revealed distinct subgroups varying in their explanation preferences, with a more complex latent class structure for participants who had heard of scientific disputes in the past. Scientific positivism and judgments of science's credibility were the strongest predictors of latent class membership, controlling for scientific reasoning, political ideology, confidence in choice, scenario, education, gender, age, and ethnicity. The lack of distinction observed overall between different explanations, as well as within classes, raises challenges for further research on explanations of scientific disputes people find credible and why.

Evaluating lipid mediator structural complexity using ion mobility spectrometry combined with mass spectrometry

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

Kyle, Jennifer E.; Aly, Noor; Zheng, Xueyun

Lipid mediators (LMs) are broadly defined as a class of bioactive lipophilic molecules that regulate cell-to-cell communication events with many having a strong correlation with various human diseases and conditions. LMs are usually analyzed with liquid chromatography and mass spectrometry (LC-MS), but their numerous isomers greatly complicate the measurements with essentially identical fragmentation spectra and LC separations not always sufficient for distinguishing the features. In this work, we characterized LMs having specific categories using ion mobility spectrometry coupled with mass spectrometry (IMS-MS). The IMS collision cross sections and MS m/z values displayed distinct trends for each LM category studied. LC-IMS-MSmore » analyses on flu infected mouse tissue samples also illustrated the presence of additional LM species not in our databases.« less

Clinical potential of meningioma genomic insights: a practical review for neurosurgeons.

PubMed

Karsy, Michael; Azab, Mohammed A; Abou-Al-Shaar, Hussam; Guan, Jian; Eli, Ilyas; Jensen, Randy L; Ormond, D Ryan

2018-06-01

Meningiomas are among the most common intracranial pathological conditions, accounting for 36% of intracranial lesions treated by neurosurgeons. Although the majority of these lesions are benign, the classical categorization of tumors by histological type or World Health Organization (WHO) grade has not fully captured the potential for meningioma progression and recurrence. Many targeted treatments have failed to generate a long-lasting effect on these tumors. Recently, several seminal studies evaluating the genomics of intracranial meningiomas have rapidly changed the understanding of the disease. The importance of NF2 (neurofibromin 2), TRAF7 (tumor necrosis factor [TNF] receptor-associated factor 7), KLF4 (Kruppel-like factor 4), AKT1, SMO (smoothened), PIK3CA (phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit alpha), and POLR2 (RNA polymerase II subunit A) demonstrates that there are at least 6 distinct mutational classes of meningiomas. In addition, 6 methylation classes of meningioma have been appreciated, enabling improved prediction of prognosis compared with traditional WHO grades. Genomic studies have shed light on the nature of recurrent meningioma, distinct intracranial locations and mutational patterns, and a potential embryonic cancer stem cell-like origin. However, despite these exciting findings, the clinical relevance of these findings remains elusive. The authors review the key findings from recent genomic studies in meningiomas, specifically focusing on how these findings relate to clinical insights for the practicing neurosurgeon.

Combined Patterns of Risk for Problem and Obesogenic Behaviors in Adolescents: A Latent Class Analysis Approach

ERIC Educational Resources Information Center

Fleary, Sasha A.

2017-01-01

Background: Several studies have used latent class analyses to explore obesogenic behaviors and substance use in adolescents independently. We explored a variety of health risks jointly to identify distinct patterns of risk behaviors among adolescents. Methods: Latent class models were estimated using Youth Risk Behavior Surveillance System…

Complicating the "soccer mom:"the cultural politics of forming class-based identity, distinction, and necessity.

PubMed

Swanson, Lisa

2009-06-01

Using Pierre Bourdieu's theories of social class differentiation and class reproduction, this paper provides an analysis of class-based identity politics in contemporary suburban America. Through a critical ethnography of the emergent, American, upper-middle-class "soccer mom" phenomenon, this study contributes to a growing body of research that interrogates class-based, cultural practices of status differentiation. As part of a larger; longitudinal ethnographic study, this paper specifically focuses on the ways in which women, who are driven by upper-middle-class habitus, contest and construct their identity as mothers of young, soccer-playing children.

Rocks of the Columbia Hills

USGS Publications Warehouse

Squyres, S. W.; Arvidson, R. E.; Blaney, D.L.; Clark, B. C.; Crumpler, L.; Farrand, W. H.; Gorevan, S.; Herkenhoff, K. E.; Hurowitz, J.; Kusack, A.; McSween, H.Y.; Ming, D. W.; Morris, R.V.; Ruff, S.W.; Wang, A.; Yen, A.

2006-01-01

The Mars Exploration Rover Spirit has identified five distinct rock types in the Columbia Hills of Gusev crater. Clovis Class rock is a poorly sorted clastic rock that has undergone substantial aqueous alteration. We interpret it to be aqueously altered ejecta deposits formed by impacts into basaltic materials. Wishstone Class rock is also a poorly sorted clastic rock that has a distinctive chemical composition that is high in Ti and P and low in Cr. Wishstone Class rock may be pyroclastic or impact in origin. Peace Class rock is a sedimentary material composed of ultramafic sand grains cemented by significant quantities of Mg- and Ca-sulfates. Peace Class rock may have formed when water briefly saturated the ultramafic sands and evaporated to allow precipitation of the sulfates. Watchtower Class rocks are similar chemically to Wishstone Class rocks and have undergone widely varying degrees of near-isochemical aqueous alteration. They may also be ejecta deposits, formed by impacts into Wishstone-rich materials and altered by small amounts of water. Backstay Class rocks are basalt/trachybasalt lavas that were emplaced in the Columbia Hills after the other rock classes were, either as impact ejecta or by localized volcanic activity. The geologic record preserved in the rocks of the Columbia Hills reveals a period very early in Martian history in which volcanic materials were widespread, impact was a dominant process, and water was commonly present. Copyright 2006 by the American Geophysical Union.

The Rocks of the Columbia Hills

NASA Technical Reports Server (NTRS)

Squyres, Steven W.; Arvidson, Raymond E.; Blaney, Diana L.; Clark, Benton C.; Crumpler, Larry; Farrand, William H.; Gorevan, Stephen; Herkenhoff, Kenneth; Hurowitz, Joel; Kusack, Alastair;

Evidence of Associations between Cytokine Genes and Subjective Reports of Sleep Disturbance in Oncology Patients and Their Family Caregivers

PubMed Central

Miaskowski, Christine; Cooper, Bruce A.; Dhruva, Anand; Dunn, Laura B.; Langford, Dale J.; Cataldo, Janine K.; Baggott, Christina R.; Merriman, John D.; Dodd, Marylin; Lee, Kathryn; West, Claudia; Paul, Steven M.; Aouizerat, Bradley E.

2012-01-01

The purposes of this study were to identify distinct latent classes of individuals based on subjective reports of sleep disturbance; to examine differences in demographic, clinical, and symptom characteristics between the latent classes; and to evaluate for variations in pro- and anti-inflammatory cytokine genes between the latent classes. Among 167 oncology outpatients with breast, prostate, lung, or brain cancer and 85 of their FCs, growth mixture modeling (GMM) was used to identify latent classes of individuals based on General Sleep Disturbance Scale (GSDS) obtained prior to, during, and for four months following completion of radiation therapy. Single nucleotide polymorphisms (SNPs) and haplotypes in candidate cytokine genes were interrogated for differences between the two latent classes. Multiple logistic regression was used to assess the effect of phenotypic and genotypic characteristics on GSDS group membership. Two latent classes were identified: lower sleep disturbance (88.5%) and higher sleep disturbance (11.5%). Participants who were younger and had a lower Karnofsky Performance status score were more likely to be in the higher sleep disturbance class. Variation in two cytokine genes (i.e., IL6, NFKB) predicted latent class membership. Evidence was found for latent classes with distinct sleep disturbance trajectories. Unique genetic markers in cytokine genes may partially explain the interindividual heterogeneity characterizing these trajectories. PMID:22844404

Role of senescence and mitotic catastrophe in cancer therapy

PubMed Central

2010-01-01

Senescence and mitotic catastrophe (MC) are two distinct crucial non-apoptotic mechanisms, often triggered in cancer cells and tissues in response to anti-cancer drugs. Chemotherapeuticals and myriad other factors induce cell eradication via these routes. While senescence drives the cells to a state of quiescence, MC drives the cells towards death during the course of mitosis. The senescent phenotype distinguishes tumor cells that survived drug exposure but lost the ability to form colonies from those that recover and proliferate after treatment. Although senescent cells do not proliferate, they are metabolically active and may secrete proteins with potential tumor-promoting activities. The other anti-proliferative response of tumor cells is MC that is a form of cell death that results from abnormal mitosis and leads to the formation of interphase cells with multiple micronuclei. Different classes of cytotoxic agents induce MC, but the pathways of abnormal mitosis differ depending on the nature of the inducer and the status of cell-cycle checkpoints. In this review, we compare the two pathways and mention that they are activated to curb the growth of tumors. Altogether, we have highlighted the possibilities of the use of senescence targeting drugs, mitotic kinases and anti-mitotic agents in fabricating novel strategies in cancer control. PMID:20205872

Scarcity of autoreactive human blood IgA+ memory B cells

PubMed Central

Prigent, Julie; Lorin, Valérie; Kök, Ayrin; Hieu, Thierry; Bourgeau, Salomé

2016-01-01

Class‐switched memory B cells are key components of the “reactive” humoral immunity, which ensures a fast and massive secretion of high‐affinity antigen‐specific antibodies upon antigenic challenge. In humans, IgA class‐switched (IgA+) memory B cells and IgA antibodies are abundant in the blood. Although circulating IgA+ memory B cells and their corresponding secreted immunoglobulins likely possess major protective and/or regulatory immune roles, little is known about their specificity and function. Here, we show that IgA+ and IgG+ memory B‐cell antibodies cloned from the same healthy humans share common immunoglobulin gene features. IgA and IgG memory antibodies have comparable lack of reactivity to vaccines, common mucosa‐tropic viruses and commensal bacteria. However, the IgA+ memory B‐cell compartment contains fewer polyreactive clones and importantly, only rare self‐reactive clones compared to IgG+ memory B cells. Self‐reactivity of IgAs is acquired following B‐cell affinity maturation but not antibody class switching. Together, our data suggest the existence of different regulatory mechanisms for removing autoreactive clones from the IgG+ and IgA+ memory B‐cell repertoires, and/or different maturation pathways potentially reflecting the distinct nature and localization of the cognate antigens recognized by individual B‐cell populations. PMID:27469325

Mechanisms regulating enhanced HLA class II-mediated CD4+ T cell recognition of human B-cell lymphoma by resveratrol

PubMed Central

RADWAN, FAISAL F. Y.; ZHANG, LIXIA; HOSSAIN, AZIM; DOONAN, BENTLY P.; GOD, JASON; HAQUE, AZIZUL

2015-01-01

Malignant B-cells express measurable levels of HLA class II proteins, but often escape immune recognition by CD4+ T cells. Resveratrol (Resv) has been the focus of numerous investigations due to its potential chemopreventive and anti-cancer effects, but it has never been tested in the regulation of immune components in B-cell tumors. Here, we show for the first time that Resv treatment enhances HLA class II-mediated immune detection of B-cell lymphomas by altering immune components and class II presentation in tumor cells. Resv treatment induced an upregulation of both classical and non-classical HLA class II proteins (DR and DM) in B-lymphoma cells. Resv also altered endolysosomal cathepsins (Cat S, B and D) and a thiol reductase (GILT), increasing HLA class II-mediated antigen (Ag) processing in B-cell lymphomas and their subsequent recognition by CD4+ T cells. Mechanistic study demonstrated that Resv treatment activated the recycling class II pathway of Ag presentation through upregulation of Rab 4B protein expression in B-lymphoma cells. These findings suggest that HLA class II-mediated immune recognition of malignant B-cells can be improved by Resv treatment, thus encouraging its potential use in chemoimmunotherapy of B-cell lymphoma. PMID:21854084

Antigen-B Cell Receptor Complexes Associate with Intracellular major histocompatibility complex (MHC) Class II Molecules*

PubMed Central

Barroso, Margarida; Tucker, Heidi; Drake, Lisa; Nichol, Kathleen; Drake, James R.

2015-01-01

Antigen processing and MHC class II-restricted antigen presentation by antigen-presenting cells such as dendritic cells and B cells allows the activation of naïve CD4+ T cells and cognate interactions between B cells and effector CD4+ T cells, respectively. B cells are unique among class II-restricted antigen-presenting cells in that they have a clonally restricted antigen-specific receptor, the B cell receptor (BCR), which allows the cell to recognize and respond to trace amounts of foreign antigen present in a sea of self-antigens. Moreover, engagement of peptide-class II complexes formed via BCR-mediated processing of cognate antigen has been shown to result in a unique pattern of B cell activation. Using a combined biochemical and imaging/FRET approach, we establish that internalized antigen-BCR complexes associate with intracellular class II molecules. We demonstrate that the M1-paired MHC class II conformer, shown previously to be critical for CD4 T cell activation, is incorporated selectively into these complexes and loaded selectively with peptide derived from BCR-internalized cognate antigen. These results demonstrate that, in B cells, internalized antigen-BCR complexes associate with intracellular MHC class II molecules, potentially defining a site of class II peptide acquisition, and reveal a selective role for the M1-paired class II conformer in the presentation of cognate antigen. These findings provide key insights into the molecular mechanisms used by B cells to control the source of peptides charged onto class II molecules, allowing the immune system to mount an antibody response focused on BCR-reactive cognate antigen. PMID:26400081

A Heterodimer of a VHH (Variable Domains of Camelid Heavy Chain-only) Antibody That Inhibits Anthrax Toxin Cell Binding Linked to a VHH Antibody That Blocks Oligomer Formation Is Highly Protective in an Anthrax Spore Challenge Model*

PubMed Central

Moayeri, Mahtab; Leysath, Clinton E.; Tremblay, Jacqueline M.; Vrentas, Catherine; Crown, Devorah; Leppla, Stephen H.; Shoemaker, Charles B.

2015-01-01

Anthrax disease is caused by a toxin consisting of protective antigen (PA), lethal factor, and edema factor. Antibodies against PA have been shown to be protective against the disease. Variable domains of camelid heavy chain-only antibodies (VHHs) with affinity for PA were obtained from immunized alpacas and screened for anthrax neutralizing activity in macrophage toxicity assays. Two classes of neutralizing VHHs were identified recognizing distinct, non-overlapping epitopes. One class recognizes domain 4 of PA at a well characterized neutralizing site through which PA binds to its cellular receptor. A second neutralizing VHH (JKH-C7) recognizes a novel epitope. This antibody inhibits conversion of the PA oligomer from “pre-pore” to its SDS and heat-resistant “pore” conformation while not preventing cleavage of full-length 83-kDa PA (PA83) by cell surface proteases to its oligomer-competent 63-kDa form (PA63). The antibody prevents endocytosis of the cell surface-generated PA63 subunit but not preformed PA63 oligomers formed in solution. JKH-C7 and the receptor-blocking VHH class (JIK-B8) were expressed as a heterodimeric VHH-based neutralizing agent (VNA2-PA). This VNA displayed improved neutralizing potency in cell assays and protected mice from anthrax toxin challenge with much better efficacy than the separate component VHHs. The VNA protected virtually all mice when separately administered at a 1:1 ratio to toxin and protected mice against Bacillus anthracis spore infection. Thus, our studies show the potential of VNAs as anthrax therapeutics. Due to their simple and stable nature, VNAs should be amenable to genetic delivery or administration via respiratory routes. PMID:25564615

A heterodimer of a VHH (variable domains of camelid heavy chain-only) antibody that inhibits anthrax toxin cell binding linked to a VHH antibody that blocks oligomer formation is highly protective in an anthrax spore challenge model.

PubMed

Moayeri, Mahtab; Leysath, Clinton E; Tremblay, Jacqueline M; Vrentas, Catherine; Crown, Devorah; Leppla, Stephen H; Shoemaker, Charles B

2015-03-06

Anthrax disease is caused by a toxin consisting of protective antigen (PA), lethal factor, and edema factor. Antibodies against PA have been shown to be protective against the disease. Variable domains of camelid heavy chain-only antibodies (VHHs) with affinity for PA were obtained from immunized alpacas and screened for anthrax neutralizing activity in macrophage toxicity assays. Two classes of neutralizing VHHs were identified recognizing distinct, non-overlapping epitopes. One class recognizes domain 4 of PA at a well characterized neutralizing site through which PA binds to its cellular receptor. A second neutralizing VHH (JKH-C7) recognizes a novel epitope. This antibody inhibits conversion of the PA oligomer from "pre-pore" to its SDS and heat-resistant "pore" conformation while not preventing cleavage of full-length 83-kDa PA (PA83) by cell surface proteases to its oligomer-competent 63-kDa form (PA63). The antibody prevents endocytosis of the cell surface-generated PA63 subunit but not preformed PA63 oligomers formed in solution. JKH-C7 and the receptor-blocking VHH class (JIK-B8) were expressed as a heterodimeric VHH-based neutralizing agent (VNA2-PA). This VNA displayed improved neutralizing potency in cell assays and protected mice from anthrax toxin challenge with much better efficacy than the separate component VHHs. The VNA protected virtually all mice when separately administered at a 1:1 ratio to toxin and protected mice against Bacillus anthracis spore infection. Thus, our studies show the potential of VNAs as anthrax therapeutics. Due to their simple and stable nature, VNAs should be amenable to genetic delivery or administration via respiratory routes. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Direct binding to antigen-coated beads refines the specificity and cross-reactivity of four monoclonal antibodies that recognize polymorphic epitopes of HLA class I molecules

PubMed Central

Hilton, Hugo G; Parham, Peter

2013-01-01

Monoclonal antibodies with specificity for HLA class I determinants of HLA were originally characterized using serological assays in which the targets were cells expressing 3-6 HLA class I variants. Because of this complexity, the specificities of the antibodies were defined indirectly by correlation. Here we use a direct binding assay, in which the targets are synthetic beads coated with one of 111 HLA class I variants, representing the full range of HLA-A, -B and -C variation. We studied one monoclonal antibody with monomorphic specificity (W6/32) and four with polymorphic specificity (MA2.1, PA2.1, BB7.2 and BB7.1) and compared the results with those obtained previously. W6/32 reacted with all HLA class I variants. MA2.1 exhibits high specificity for HLA-A*02, -B*57 and -B*58, but also exhibited cross-reactivity with HLA-A*11 and -B*15:16. At low concentration (1μg/ml) PA2.1 and BB7.2 were both specific for HLA-A*02 and -A*69, and at high concentration (50μg/ml) exhibited significant cross-reactions with HLA-A*68, -A*23, and -A*24. BB7.1 exhibits specificity for HLA-B*07 and -B*42, as previously described, but reacts equally well with HLA-B*81, a rare allotype defined some 16 years after the description of BB7.1. The results obtained with cell-based and bead-based assays are consistent and, in combination with amino acid sequence comparison, increase understanding of the polymorphic epitopes recognized by the MA2.1, PA2.1, BB7.2 and BB7.1 antibodies. Comparison of two overlapping but distinctive bead sets from two sources gave similar results, but the overall levels of binding were significantly different. Several weaker reactions were observed with only one of the bead sets. PMID:23510417

Definition of Proteasomal Peptide Splicing Rules for High-Efficiency Spliced Peptide Presentation by MHC Class I Molecules

PubMed Central

Berkers, Celia R.; de Jong, Annemieke; Schuurman, Karianne G.; Linnemann, Carsten; Meiring, Hugo D.; Janssen, Lennert; Neefjes, Jacques J.; Schumacher, Ton N. M.; Rodenko, Boris

2015-01-01

Peptide splicing, in which two distant parts of a protein are excised and then ligated to form a novel peptide, can generate unique MHC class I–restricted responses. Because these peptides are not genetically encoded and the rules behind proteasomal splicing are unknown, it is difficult to predict these spliced Ags. In the current study, small libraries of short peptides were used to identify amino acid sequences that affect the efficiency of this transpeptidation process. We observed that splicing does not occur at random, neither in terms of the amino acid sequences nor through random splicing of peptides from different sources. In contrast, splicing followed distinct rules that we deduced and validated both in vitro and in cells. Peptide ligation was quantified using a model peptide and demonstrated to occur with up to 30% ligation efficiency in vitro, provided that optimal structural requirements for ligation were met by both ligating partners. In addition, many splicing products could be formed from a single protein. Our splicing rules will facilitate prediction and detection of new spliced Ags to expand the peptidome presented by MHC class I Ags. PMID:26401003

Notch Decoys that Selectively Block Dll/Notch or Jagged/Notch Disrupt Angiogenesis by Unique Mechanisms to Inhibit Tumor Growth

PubMed Central

Kangsamaksin, Thaned; Murtomaki, Aino; Kofler, Natalie M.; Cuervo, Henar; Chaudhri, Reyhaan A.; Tattersall, Ian W.; Rosenstiel, Paul E.; Shawber, Carrie J.; Kitajewski, Jan

2015-01-01

A pro-angiogenic role for Jagged-dependent activation of Notch signaling in the endothelium has yet to be described. Using proteins that encoded different NOTCH1 EGF-like repeats, we identified unique regions of DLL-class and JAG-class ligand/receptor interactions, and developed Notch decoys that function as ligand-specific Notch inhibitors. N110-24 decoy blocked JAG1/JAG2-mediated NOTCH1 signaling, angiogenic sprouting in vitro and retinal angiogenesis, demonstrating JAG-dependent Notch signal activation promotes angiogenesis. In tumors, N110-24 decoy reduced angiogenic sprouting, vessel perfusion, pericyte coverage, and tumor growth. JAG/NOTCH signaling uniquely inhibited expression of anti-angiogenic sVEFGFR-1/sFlt-1. N11-13 decoy interfered with DLL1/DLL4-mediated NOTCH1 signaling and caused endothelial hypersprouting in vitro, in retinal angiogenesis and in tumors. Thus, blockade of JAG- or DLL-mediated Notch signaling inhibits angiogenesis by distinct mechanisms. JAG/Notch signaling positively regulates angiogenesis by suppressing sVEGFR-1/sFlt-1 and promoting mural/endothelial cell interactions. Blockade of JAG-class ligands represents a novel, viable therapeutic approach to block tumor angiogenesis and growth. PMID:25387766

Interferon-gamma: biologic functions and HCV therapy (type I/II) (1 of 2 parts).

PubMed

Gattoni, A; Parlato, A; Vangieri, B; Bresciani, M; Derna, R

2006-01-01

This review is aimed at exhaustively presenting and discussing the interferon-gamma (IFN-gamma), a cytokine that plays an important role in inducing and modulating an array of immune responses. A review of the most significant and recent clinical trials was performed. Although IFN-gamma has some antiviral activity, it is much less active in this regard than type I IFNs. IFN-gamma is involved in the regulation of nearly all phases of the immune and inflammatory responses, including the activation and differentiation of T cells, B cells, NK cells, macrophages, and others. It is therefore best regarded as a distint immunoregulatory cytokine. IFN-gamma secretion is a hallmark of Th1 lymphocytes. It is also secreted by nearly all CD8 T cells, by some Th0 cells, and by NK cells. Each of these cell types secretes IFN-gamma only when activated, usually as part of immune response and especially in response to IL-2 and IL-12. IFN-gamma production is inhibited by IL-4, IL-10, TGFbeta, glucocorticoids, cyclosporin A and FK506. Nearly all cell types express the heterodimeric receptor for IFN-beta and respond to this cytokine by increasing the surface expression of class I MHC proteins. As a result, virtually any cell in the vicinity of an IFN-beta-secreting cell becomes more efficient at presenting endogenous antigens and hence a better target for cytotoxic killing if it harbors an intracellular pathogen. Unlike the type I IFNs, IFN-gamma also increases the expression of class II MHC proteins on professional APCs, and so promotes antigen presentation to helper T cells as well. It also induces de novo expression of class II MHC proteins on venular endothelial cells and on some other epithelial and connective tissue cells that do not otherwise express them, thus enabling these cell types to function as temporary APCs at sites of intense immune reactions. The effector functions of NK cells are to lyse virus-infected cells and to secrete IFN-gamma, which activates macrofages to destroy phagocytosed microbes. The mechanism of NK cell-mediates cytolysis is essentially the same as that of cytolysis by CTLS. NK cells lyse virally infected cells before antigen specific CTLS came become fully active, that is, during the first few days after viral infection. NK cells are expanded and activated by cytokines of innate immunity, such as IL-12 and IL-15, and they kill infected cells, especially those that display reduced levels of class I molecoles. Some tumors, especially those of hematopoietic origin, are targets of NK cells, perlevels or types of class I MHC molecules. Therefore, IFN-gamma serves critical functions in innate immunity and in specific cell-mediated immunity (in addition, IFN activates neutrophilis and stimulates the cytolitic activity of NK cells). Many IFNs-gamma induced effects result in heigtened immune surveillance. IFN-gamma is a remarkable cytokine that orchestrates many distinct cellular programs through transcriptional control over large numbers of genes. Many IFNs-gamma-induced effects resulting in heightend immune surveillance and immune system function during infection have been discussed in this review. As the pathogens (microorganism with the potential to cause tissue injury or disease) augment local IFN-gamma production, and IFN-gamma augments the immune system response, an important function of IFN-gamma during in vivo infection is suggested. IFN-gamma is primarily secreted by activated T cells and natural killer cells, and can promote macrophage activation, mediate antiviral e antibacterial immunity, enhance antigen presentation, orchestrate activation of the innate immune system, coordinate lymphocyte-endothelium interaction, regulate Th1/Th2 balance, and control cellular proliferation and apoptosis.

Structural characterization of encapsulated ferritin provides insight into iron storage in bacterial nanocompartments

PubMed Central

He, Didi; Hughes, Sam; Vanden-Hehir, Sally; Georgiev, Atanas; Altenbach, Kirsten; Tarrant, Emma; Mackay, C Logan; Waldron, Kevin J; Clarke, David J; Marles-Wright, Jon

2016-01-01

Ferritins are ubiquitous proteins that oxidise and store iron within a protein shell to protect cells from oxidative damage. We have characterized the structure and function of a new member of the ferritin superfamily that is sequestered within an encapsulin capsid. We show that this encapsulated ferritin (EncFtn) has two main alpha helices, which assemble in a metal dependent manner to form a ferroxidase center at a dimer interface. EncFtn adopts an open decameric structure that is topologically distinct from other ferritins. While EncFtn acts as a ferroxidase, it cannot mineralize iron. Conversely, the encapsulin shell associates with iron, but is not enzymatically active, and we demonstrate that EncFtn must be housed within the encapsulin for iron storage. This encapsulin nanocompartment is widely distributed in bacteria and archaea and represents a distinct class of iron storage system, where the oxidation and mineralization of iron are distributed between two proteins. DOI: http://dx.doi.org/10.7554/eLife.18972.001 PMID:27529188

Neuronal cell fate specification by the molecular convergence of different spatio-temporal cues on a common initiator terminal selector gene

PubMed Central

Stratmann, Johannes

2017-01-01

The extensive genetic regulatory flows underlying specification of different neuronal subtypes are not well understood at the molecular level. The Nplp1 neuropeptide neurons in the developing Drosophila nerve cord belong to two sub-classes; Tv1 and dAp neurons, generated by two distinct progenitors. Nplp1 neurons are specified by spatial cues; the Hox homeotic network and GATA factor grn, and temporal cues; the hb -> Kr -> Pdm -> cas -> grh temporal cascade. These spatio-temporal cues combine into two distinct codes; one for Tv1 and one for dAp neurons that activate a common terminal selector feedforward cascade of col -> ap/eya -> dimm -> Nplp1. Here, we molecularly decode the specification of Nplp1 neurons, and find that the cis-regulatory organization of col functions as an integratory node for the different spatio-temporal combinatorial codes. These findings may provide a logical framework for addressing spatio-temporal control of neuronal sub-type specification in other systems. PMID:28414802

Coordinated temporal and spatial control of motor neuron and serotonergic neuron generation from a common pool of CNS progenitors.

PubMed

Pattyn, Alexandre; Vallstedt, Anna; Dias, José M; Samad, Omar Abdel; Krumlauf, Robb; Rijli, Filippo M; Brunet, Jean-Francois; Ericson, Johan

2003-03-15

Neural progenitor cells often produce distinct types of neurons in a specific order, but the determinants that control the sequential generation of distinct neuronal subclasses in the vertebrate CNS remain poorly defined. We examined the sequential generation of visceral motor neurons and serotonergic neurons from a common pool of neural progenitors located in the ventral hindbrain. We found that the temporal specification of these neurons varies along the anterior-posterior axis of the hindbrain, and that the timing of their generation critically depends on the integrated activities of Nkx- and Hox-class homeodomain proteins. A primary function of these proteins is to coordinate the spatial and temporal activation of the homeodomain protein Phox2b, which in turn acts as a binary switch in the selection of motor neuron or serotonergic neuronal fate. These findings assign new roles for Nkx, Hox, and Phox2 proteins in the control of temporal neuronal fate determination, and link spatial and temporal patterning of CNS neuronal fates.

Regulatory logic of pan-neuronal gene expression in C. elegans

PubMed Central

Stefanakis, Nikolaos; Carrera, Ines; Hobert, Oliver

2015-01-01

While neuronal cell types display an astounding degree of phenotypic diversity, most if not all neuron types share a core panel of terminal features. However, little is known about how pan-neuronal expression patterns are genetically programmed. Through an extensive analysis of the cis-regulatory control regions of a battery of pan-neuronal C.elegans genes, including genes involved in synaptic vesicle biology and neuropeptide signaling, we define a common organizational principle in the regulation of pan-neuronal genes in the form of a surprisingly complex array of seemingly redundant, parallel-acting cis-regulatory modules that direct expression to broad, overlapping domains throughout the nervous system. These parallel-acting cis-regulatory modules are responsive to a multitude of distinct trans-acting factors. Neuronal gene expression programs therefore fall into two fundamentally distinct classes. Neuron type-specific genes are generally controlled by discrete and non-redundantly acting regulatory inputs, while pan-neuronal gene expression is controlled by diverse, coincident and seemingly redundant regulatory inputs. PMID:26291158

Trajectories of depression in adults with newly diagnosed type 1 diabetes: results from the German Multicenter Diabetes Cohort Study.

PubMed

Kampling, Hanna; Petrak, Frank; Farin, Erik; Kulzer, Bernd; Herpertz, Stephan; Mittag, Oskar

2017-01-01

There is a paucity of longitudinal data on type 1 diabetes and depression, especially in adults. The present study prospectively analysed trajectories of depressive symptoms in adults during the first 5 years of living with type 1 diabetes. We aimed to identify distinct trajectories of depressive symptoms and to examine how they affect diabetes outcome. We reanalysed data from a prospective multicentre observational cohort study including 313 adults with newly diagnosed type 1 diabetes. At baseline and in annual postal surveys over 5 consecutive years, we gathered patient characteristics and behavioural and psychosocial data (e.g. Symptom Checklist-90-R [SCL-90-R]). Medical data (e.g. HbA 1c levels) was obtained from the treating physicians. We applied growth mixture modelling (GMM) to identify distinct trajectories of depression over time. Five years after diagnosis, 7.8% (n = 20) of patients were moderately depressed and 10.2% (n = 26) were severely depressed. GMM statistics identified three possible models of trajectories (class 1, 'no depressive symptoms'; class 2, 'worsening depressive symptoms that improve after 2 years'; class 3, 'worsening depressive symptoms'). Severity of depression symptoms at baseline (subscale of the SCL-90-R questionnaire) significantly predicted membership of classes 2 and 3 vs class 1. After 5 years, higher HbA 1c values were detected in class 3 patients (mean = 8.2%, 66 mmol/mol) compared with class 1 and class 2 (both: mean = 7.2%, 55 mmol/mol). We identified distinct trajectories of depressive symptoms that are also relevant for diabetes outcome. Patients with worsening depressive symptoms over time exhibited poor glycaemic control after the first 5 years of living with diabetes. They also exhibited a reduced quality of life and increased diabetes-related distress.

Differential incorporation of docosahexaenoic acid into distinct cholesterol-rich membrane raft domains.

PubMed

Duraisamy, Yasotha; Lambert, Daniel; O'Neill, Catherine A; Padfield, Philip J

2007-09-07

We investigated the influence of docosahexaenoic acid (DHA) on the fatty acid and protein compositions of two populations of membrane rafts present in Caco-2 cells. DHA (100 microM) had no significant influence on the fatty acid or protein compositions of tight junction-associated, Lubrol insoluble, membrane rafts. However, DHA did significantly alter the fatty acid and protein compositions of "archetypal" Triton X-100 insoluble membrane rafts. The DHA content of the raft lipids increased 25-fold and was accompanied by a redistribution of src and fyn out of the rafts. DHA also increased Caco-2 cell monolayer permeability producing a 95% drop in transepithelial electrical resistance and a 8.56-fold increase in the flux of dextran. In conclusion, the data demonstrate that DHA does not increase permeability through modifying the TJ-associated rafts. The data do, however, show that DHA is differentially incorporated into different classes of membrane rafts, which has significant implications to our understanding of how omega-3 PUFAs modulate plasma membrane organization and cell function.

Lineage Tracking for Probing Heritable Phenotypes at Single-Cell Resolution

PubMed Central

Cottinet, Denis; Condamine, Florence; Bremond, Nicolas; Griffiths, Andrew D.; Rainey, Paul B.; de Visser, J. Arjan G. M.; Baudry, Jean; Bibette, Jérôme

2016-01-01

Determining the phenotype and genotype of single cells is central to understand microbial evolution. DNA sequencing technologies allow the detection of mutants at high resolution, but similar approaches for phenotypic analyses are still lacking. We show that a drop-based millifluidic system enables the detection of heritable phenotypic changes in evolving bacterial populations. At time intervals, cells were sampled and individually compartmentalized in 100 nL drops. Growth through 15 generations was monitored using a fluorescent protein reporter. Amplification of heritable changes–via growth–over multiple generations yields phenotypically distinct clusters reflecting variation relevant for evolution. To demonstrate the utility of this approach, we follow the evolution of Escherichia coli populations during 30 days of starvation. Phenotypic diversity was observed to rapidly increase upon starvation with the emergence of heritable phenotypes. Mutations corresponding to each phenotypic class were identified by DNA sequencing. This scalable lineage-tracking technology opens the door to large-scale phenotyping methods with special utility for microbiology and microbial population biology. PMID:27077662

Lineage Tracking for Probing Heritable Phenotypes at Single-Cell Resolution.

PubMed

Cottinet, Denis; Condamine, Florence; Bremond, Nicolas; Griffiths, Andrew D; Rainey, Paul B; de Visser, J Arjan G M; Baudry, Jean; Bibette, Jérôme

2016-01-01

Determining the phenotype and genotype of single cells is central to understand microbial evolution. DNA sequencing technologies allow the detection of mutants at high resolution, but similar approaches for phenotypic analyses are still lacking. We show that a drop-based millifluidic system enables the detection of heritable phenotypic changes in evolving bacterial populations. At time intervals, cells were sampled and individually compartmentalized in 100 nL drops. Growth through 15 generations was monitored using a fluorescent protein reporter. Amplification of heritable changes-via growth-over multiple generations yields phenotypically distinct clusters reflecting variation relevant for evolution. To demonstrate the utility of this approach, we follow the evolution of Escherichia coli populations during 30 days of starvation. Phenotypic diversity was observed to rapidly increase upon starvation with the emergence of heritable phenotypes. Mutations corresponding to each phenotypic class were identified by DNA sequencing. This scalable lineage-tracking technology opens the door to large-scale phenotyping methods with special utility for microbiology and microbial population biology.

Systematic Identification of Molecular Subtype-Selective Vulnerabilities in Non Small Cell Lung Cancer

PubMed Central

Kim, Hyun Seok; Mendiratta, Saurabh; Kim, Jiyeon; Pecot, Chad Victor; Larsen, Jill E.; Zubovych, Iryna; Seo, Bo Yeun; Kim, Jimi; Eskiocak, Banu; Chung, Hannah; McMillan, Elizabeth; Wu, Sherry; De Brabander, Jef; Komurov, Kakajan; Toombs, Jason E.; Wei, Shuguang; Peyton, Michael; Williams, Noelle; Gazdar, Adi F.; Posner, Bruce A.; Brekken, Rolf; Sood, Anil K.; Deberardinis, Ralph J.; Roth, Michael G.; Minna, John D.; White, Michael A.

2013-01-01

SUMMARY Context-specific molecular vulnerabilities that arise during tumor evolution represent an attractive intervention target class. However, the frequency and diversity of somatic lesions detected among lung tumors can confound efforts to identify these targets. To confront this challenge, we have applied parallel screening of chemical and genetic perturbations within a panel of molecularly annotated NSCLC lines to identify intervention opportunities tightly linked to molecular response indicators predictive of target sensitivity. Anchoring this analysis on a matched tumor/normal cell model from a lung adenocarcinoma patient identified three distinct target/response-indicator pairings that are represented with significant frequencies (6–16%) in the patient population. These include NLRP3 mutation/inflammasome activation-dependent FLIP addiction, co-occuring KRAS and LKB1 mutation-driven COPI addiction, and selective sensitivity to a synthetic indolotriazine that is specified by a 7-gene expression signature. Target efficacies were validated in vivo, and mechanism of action studies uncovered new cancer cell biology. PMID:24243015

Systematic identification of molecular subtype-selective vulnerabilities in non-small-cell lung cancer.

PubMed

Kim, Hyun Seok; Mendiratta, Saurabh; Kim, Jiyeon; Pecot, Chad Victor; Larsen, Jill E; Zubovych, Iryna; Seo, Bo Yeun; Kim, Jimi; Eskiocak, Banu; Chung, Hannah; McMillan, Elizabeth; Wu, Sherry; De Brabander, Jef; Komurov, Kakajan; Toombs, Jason E; Wei, Shuguang; Peyton, Michael; Williams, Noelle; Gazdar, Adi F; Posner, Bruce A; Brekken, Rolf A; Sood, Anil K; Deberardinis, Ralph J; Roth, Michael G; Minna, John D; White, Michael A

2013-10-24

Context-specific molecular vulnerabilities that arise during tumor evolution represent an attractive intervention target class. However, the frequency and diversity of somatic lesions detected among lung tumors can confound efforts to identify these targets. To confront this challenge, we have applied parallel screening of chemical and genetic perturbations within a panel of molecularly annotated NSCLC lines to identify intervention opportunities tightly linked to molecular response indicators predictive of target sensitivity. Anchoring this analysis on a matched tumor/normal cell model from a lung adenocarcinoma patient identified three distinct target/response-indicator pairings that are represented with significant frequencies (6%-16%) in the patient population. These include NLRP3 mutation/inflammasome activation-dependent FLIP addiction, co-occurring KRAS and LKB1 mutation-driven COPI addiction, and selective sensitivity to a synthetic indolotriazine that is specified by a seven-gene expression signature. Target efficacies were validated in vivo, and mechanism-of-action studies informed generalizable principles underpinning cancer cell biology. Copyright © 2013 Elsevier Inc. All rights reserved.

Diminished nuclear RNA decay upon Salmonella infection upregulates antibacterial noncoding RNAs.

PubMed

Imamura, Katsutoshi; Takaya, Akiko; Ishida, Yo-Ichi; Fukuoka, Yayoi; Taya, Toshiki; Nakaki, Ryo; Kakeda, Miho; Imamachi, Naoto; Sato, Aiko; Yamada, Toshimichi; Onoguchi-Mizutani, Rena; Akizuki, Gen; Tanu, Tanzina; Tao, Kazuyuki; Miyao, Sotaro; Suzuki, Yutaka; Nagahama, Masami; Yamamoto, Tomoko; Jensen, Torben Heick; Akimitsu, Nobuyoshi

2018-06-07

Cytoplasmic mRNA degradation controls gene expression to help eliminate pathogens during infection. However, it has remained unclear whether such regulation also extends to nuclear RNA decay. Here, we show that 145 unstable nuclear RNAs, including enhancer RNAs (eRNAs) and long noncoding RNAs (lncRNAs) such as NEAT1v2, are stabilized upon Salmonella infection in HeLa cells. In uninfected cells, the RNA exosome, aided by the Nuclear EXosome Targeting (NEXT) complex, degrades these labile transcripts. Upon infection, the levels of the exosome/NEXT components, RRP6 and MTR4, dramatically decrease, resulting in transcript stabilization. Depletion of lncRNAs, NEAT1v2, or eRNA07573 in HeLa cells triggers increased susceptibility to Salmonella infection concomitant with the deregulated expression of a distinct class of immunity-related genes, indicating that the accumulation of unstable nuclear RNAs contributes to antibacterial defense. Our results highlight a fundamental role for regulated degradation of nuclear RNA in the response to pathogenic infection. © 2018 The Authors.

Neurogliaform cortical interneurons derive from cells in the preoptic area

PubMed Central

Cadilhac, Christelle; Prados, Julien; Holtmaat, Anthony

2018-01-01

Delineating the basic cellular components of cortical inhibitory circuits remains a fundamental issue in order to understand their specific contributions to microcircuit function. It is still unclear how current classifications of cortical interneuron subtypes relate to biological processes such as their developmental specification. Here we identified the developmental trajectory of neurogliaform cells (NGCs), the main effectors of a powerful inhibitory motif recruited by long-range connections. Using in vivo genetic lineage-tracing in mice, we report that NGCs originate from a specific pool of 5-HT3AR-expressing Hmx3+ cells located in the preoptic area (POA). Hmx3-derived 5-HT3AR+ cortical interneurons (INs) expressed the transcription factors PROX1, NR2F2, the marker reelin but not VIP and exhibited the molecular, morphological and electrophysiological profile of NGCs. Overall, these results indicate that NGCs are a distinct class of INs with a unique developmental trajectory and open the possibility to study their specific functional contribution to cortical inhibitory microcircuit motifs. PMID:29557780

Ubiquitinated Sirtuin 1 (SIRT1) Function Is Modulated during DNA Damage-induced Cell Death and Survival*

PubMed Central

Peng, Lirong; Yuan, Zhigang; Li, Yixuan; Ling, Hongbo; Izumi, Victoria; Fang, Bin; Fukasawa, Kenji; Koomen, John; Chen, Jiandong; Seto, Edward

2015-01-01

Downstream signaling of physiological and pathological cell responses depends on post-translational modification such as ubiquitination. The mechanisms regulating downstream DNA damage response (DDR) signaling are not completely elucidated. Sirtuin 1 (SIRT1), the founding member of Class III histone deacetylases, regulates multiple steps in DDR and is closely associated with many physiological and pathological processes. However, the role of post-translational modification or ubiquitination of SIRT1 during DDR is unclear. We show that SIRT1 is dynamically and distinctly ubiquitinated in response to DNA damage. SIRT1 was ubiquitinated by the MDM2 E3 ligase in vitro and in vivo. SIRT1 ubiquitination under normal conditions had no effect on its enzymatic activity or rate of degradation; hypo-ubiquitination, however, reduced SIRT1 nuclear localization. Ubiquitination of SIRT1 affected its function in cell death and survival in response to DNA damage. Our results suggest that ubiquitination is required for SIRT1 function during DDR. PMID:25670865

Human IgG repertoire of malaria antigen-immunized human immune system (HIS) mice.

PubMed

Nogueira, Raquel Tayar; Sahi, Vincent; Huang, Jing; Tsuji, Moriya

2017-08-01

Humanized mouse models present an important tool for preclinical evaluation of new vaccines and therapeutics. Here we show the human variable repertoire of antibody sequences cloned from a previously described human immune system (HIS) mouse model that possesses functional human CD4+ T cells and B cells, namely HIS-CD4/B mice. We sequenced variable IgG genes from single memory B-cell and plasma-cell sorted from splenocytes or whole blood lymphocytes of HIS-CD4/B mice that were vaccinated with a human plasmodial antigen, a recombinant Plasmodium falciparum circumsporozoite protein (rPfCSP). We demonstrate that rPfCSP immunization triggers a diverse B-cell IgG repertoire composed of various human VH family genes and distinct V(D)J recombinations that constitute diverse CDR3 sequences similar to humans, although low hypermutated sequences were generated. These results demonstrate the substantial genetic diversity of responding human B cells of HIS-CD4/B mice and their capacity to mount human IgG class-switched antibody response upon vaccination. Copyright © 2017 European Federation of Immunological Societies. Published by Elsevier B.V. All rights reserved.

Bacterial Biofilms as Complex Communities

NASA Astrophysics Data System (ADS)

Vlamakis, Hera

2010-03-01

Many microbial populations form surface-associated multicellular communities known as biofilms. These multicellular communities are encased in a self-produced extracellular matrix composed of polysaccharides and proteins. Division of labor is a key feature of these communities and different cells serve distinct functions. We have found that in biofilms of the bacterium Bacillus subtilis, different cell types including matrix-producing and sporulating cells coexist and localize to distinct regions within the structured community. We were interested in understanding how these different cell types arise. Using fluorescence reporters under the control of promoters that are specific for distinct cell types we were able to follow the dynamics of differentiation throughout biofilm development. We found that a series of extracellular signals leads to differentiation of distinct cell types during biofilm formation. In addition, we found that extracellular matrix functions as a differentiation signal for timely sporulation within a biofilm and mutants unable to produce matrix were delayed in sporulation. Our results indicate that within a biofilm, cell-cell signaling is directional in that one cell type produces a signal that is sensed by another distinct cell type. Furthermore, once differentiated, cells become resistant to the action of other signaling molecules making it possible to maintain distinct cell populations over prolonged periods.

Disruption of HLA class II antigen presentation in Burkitt lymphoma: implication of a 47 000 MW acid labile protein in CD4+ T-cell recognition

PubMed Central

God, Jason M; Zhao, Dan; Cameron, Christine A; Amria, Shereen; Bethard, Jennifer R; Haque, Azizul

2014-01-01

While Burkitt lymphoma (BL) has a well-known defect in HLA class I-mediated antigen presentation, the exact role of BL-associated HLA class II in generating a poor CD4+ T-cell response remains unresolved. Here, we found that BL cells are deficient in their ability to optimally stimulate CD4+ T cells via the HLA class II pathway. This defect in CD4+ T-cell recognition was not associated with low levels of co-stimulatory molecules on BL cells, as addition of external co-stimulation failed to elicit CD4+ T-cell activation by BL. Further, the defect was not caused by faulty antigen/class II interaction, because antigenic peptides bound with measurable affinity to BL-associated class II molecules. Interestingly, functional class II–peptide complexes were formed at acidic pH 5·5, which restored immune recognition. Acidic buffer (pH 5·5) eluate from BL cells contained molecules that impaired class II-mediated antigen presentation and CD4+ T-cell recognition. Biochemical analysis showed that these molecules were greater than 30 000 molecular weight in size, and proteinaceous in nature. In addition, BL was found to have decreased expression of a 47 000 molecular weight enolase-like molecule that enhances class II-mediated antigen presentation in B cells, macrophages and dendritic cells, but not in BL cells. These findings demonstrate that BL likely has multiple defects in HLA class II-mediated antigen presentation and immune recognition, which may be exploited for future immunotherapies. PMID:24628049

Languages Discourses in Australian Middle-Class Schools: Parent and Student Perspectives

ERIC Educational Resources Information Center

Wright, Jan; Cruickshank, Ken; Black, Stephen

2018-01-01

Much of the literature on social class and language study in schools argues that for middle-class parents and their children, languages are chosen for their capacity to offer forms of distinction that provide an edge in the global labour market. In this paper, we draw on data collected from interviews with parents and children in middle-class…

Measuring Type 2 Diabetes Mellitus Knowledge and Perceptions of Risk in Middle-Class African Americans

ERIC Educational Resources Information Center

Spears, Erica C.; Guidry, Jeffrey J.; Harvey, Idethia S.

2018-01-01

There is a paucity in the literature examining the African American middle-class. Most studies of African Americans and Type 2 Diabetes Mellitus (T2DM) have concentrated on lower-SES individuals, or make no distinction between African Americans of varying socio-economic positions. Middle-class African Americans are vulnerable in ways often…

What do somatic hypermutation and class switch recombination teach us about chronic lymphocytic leukaemia pathogenesis?

PubMed

Oppezzo, P; Dighiero, G

2005-01-01

B-CLL cells express CD5 and IgM/IgD and thus have a mantle zone-like phenotype of naive cells, which, in normal conditions express unmutated Ig genes. However, recent studies have shown that 50%-70% of CLL harbour somatic mutations of VH genes, as if they had matured in a lymphoid follicle. Interestingly, the presence or absence of somatic hypermutation (SHM) process is associated with the use of particular VH genes. Particular alleles of the VH1-69 gene and the VH4-39 gene are preferentially expressed in an unmutated form, while VH4-34 or the majority of VH3 family genes frequently contain somatic mutations. The fact that some genes like VH1-69 and VH3-07 recombine this VH segment to particular JH segments and the restricted use of CDR3 sequences by CLLs expressing the VH4-39 gene suggest that the observed differences in BCR structure in B-CLL could result from selection by distinct antigenic epitopes. It is currently unclear whether this putative antigen-driven process could occur prior to leukaemic transformation and/or that the precursors were transformed into leukaemic cells at distinct maturational stages. The mutational profile of Ig genes has been shown to be associated with disease prognosis. These results could favour the idea that CLL could correspond to two different diseases that look alike in morphologic and phenotypic terms. In CLL with mutated Ig genes, the proliferating B cell may have transited through germinal centres, the physiologic site of hypermutation, whereas in CLL with unmutated Ig genes the malignant B cell may derive from a pre-germinal centre naïve B cell. Despite these clinical and molecular differences, recent studies on gene expression profiling of B-CLL cells showed that CLL is characterized by a common gene expression signature that is irrespective of Ig mutational status and differs from other lymphoid cancers and normal lymphoid subpopulations, suggesting that CLL cases share a common mechanism of transformation and/or cell of origin. Activation induced cytidine deaminase (AID) plays a key role in SHM and class switch recombination (CSR). However, the mechanisms accounting for AID action and control of its expression remain unclear. In a recent work we have shown that in contrast to normal circulating B-cells, AID transcripts are expressed constitutively in CLL patients undergoing active CSR, but interestingly this expression occurs predominately in unmutated CLL B-cells. These data favour the view that AID protein may act differentially on CSR and SHM pathways, but the role-played by AID in both processes remains to be elucidated. Recent work indicates that AID is expressed in a small fraction of tumoral cells, which could suggest that this small fraction of cells may correspond to B-CLL cells that would have recently experienced an AID-inducing stimulus occurring in a specific microenvironment.

Prostitution

PubMed Central

Sawa, R.J.

1987-01-01

A review of the literature discloses that prostitutes are distinguishable into distinct classes, each with distinct clinical implications. The spectre of AIDS suggests that we review the implications of the health risks associated with this profession. This article discusses the potential causes, health problems, and treatment of prostitutes. PMID:21263810

Probes of Ubiquitin E3 ligases distinguish different stages of Parkin activation

PubMed Central

Pao, Kuan-Chuan; Stanley, Mathew; Han, Cong; Lai, Yu-Chiang; Murphy, Paul; Balk, Kristin; Wood, Nicola T.; Corti, Olga; Corvol, Jean-Christophe; Muqit, Miratul M.K.; Virdee, Satpal

2016-01-01

E3 ligases represent an important class of enzymes, yet there are currently no chemical probes to profile their activity. We develop a new class of activity-based probe by reengineering of a ubiquitin-charged E2 conjugating enzyme and demonstrate their utility by profiling the transthiolation activity of the RING-in-between-RING (RBR) E3 ligase Parkin in vitro and in cellular extracts. Our study provides valuable insight into the roles, and cellular hierarchy, of distinct phosphorylation events in Parkin activation. We also profile Parkin patient disease-associated mutations and strikingly demonstrate that they largely mediate their effect by altering transthiolation activity. Furthermore, our probes enable direct and quantitative measurement of endogenous Parkin activity revealing that endogenous Parkin is activated in neuronal cell lines (≥75 %) in response to mitochondrial depolarization. This new technology also holds promise as a novel biomarker of PINK1-Parkin signalling as demonstrated by compatibility with Parkinson’s disease patient-derived samples. PMID:26928937

Investigation of antibacterial mode of action for traditional and amphiphilic aminoglycosides.

PubMed

Udumula, Venkatareddy; Ham, Young Wan; Fosso, Marina Y; Chan, Ka Yee; Rai, Ravi; Zhang, Jianjun; Li, Jie; Chang, Cheng-Wei Tom

2013-03-15

Aminoglycoside represents a class of versatile and broad spectrum antibacterial agents. In an effort to revive the antibacterial activity against aminoglycoside resistant bacteria, our laboratory has developed two new classes of aminoglycoside, pyranmycin and amphiphilic neomycin (NEOF004). The former resembles the traditional aminoglycoside, neomycin. The latter, albeit derived from neomycin, appears to exert antibacterial action via a different mode of action. In order to discern that these aminoglycoside derivatives have distinct antibacterial mode of action, RNA-binding affinity and fluorogenic dye were employed. These studies, together with our previous investigation, confirm that pyranmycin exhibit the traditional antibacterial mode of action of aminoglycosides by binding toward the bacterial rRNA. On the other hand, the amphiphilic neomycin, NEOF004 disrupts the bacterial cell wall. In a broader perspective, it verifies that structurally modified neomycin can exert different antibacterial mode of action leading to the revival of activity against aminoglycoside resistant bacteria. Copyright © 2013 Elsevier Ltd. All rights reserved.

Computational and Statistical Analyses of Amino Acid Usage and Physico-Chemical Properties of the Twelve Late Embryogenesis Abundant Protein Classes

PubMed Central

Jaspard, Emmanuel; Macherel, David; Hunault, Gilles

2012-01-01

Late Embryogenesis Abundant Proteins (LEAPs) are ubiquitous proteins expected to play major roles in desiccation tolerance. Little is known about their structure - function relationships because of the scarcity of 3-D structures for LEAPs. The previous building of LEAPdb, a database dedicated to LEAPs from plants and other organisms, led to the classification of 710 LEAPs into 12 non-overlapping classes with distinct properties. Using this resource, numerous physico-chemical properties of LEAPs and amino acid usage by LEAPs have been computed and statistically analyzed, revealing distinctive features for each class. This unprecedented analysis allowed a rigorous characterization of the 12 LEAP classes, which differed also in multiple structural and physico-chemical features. Although most LEAPs can be predicted as intrinsically disordered proteins, the analysis indicates that LEAP class 7 (PF03168) and probably LEAP class 11 (PF04927) are natively folded proteins. This study thus provides a detailed description of the structural properties of this protein family opening the path toward further LEAP structure - function analysis. Finally, since each LEAP class can be clearly characterized by a unique set of physico-chemical properties, this will allow development of software to predict proteins as LEAPs. PMID:22615859

Distinct cargo-specific response landscapes underpin the complex and nuanced role of galectin-glycan interactions in clathrin-independent endocytosis.

PubMed

Mathew, Mohit P; Donaldson, Julie G

2018-05-11

Clathrin-independent endocytosis (CIE) is a form of endocytosis that lacks a defined cytoplasmic machinery. Here, we asked whether glycan interactions, acting from the outside, could be a part of that endocytic machinery. We show that the perturbation of global cellular patterns of protein glycosylation by modulation of metabolic flux affects CIE. Interestingly, these changes in glycosylation had cargo-specific effects. For example, in HeLa cells, GlcNAc treatment, which increases glycan branching, increased major histocompatibility complex class I (MHCI) internalization but inhibited CIE of the glycoprotein CD59 molecule (CD59). The effects of knocking down the expression of galectin 3, a carbohydrate-binding protein and an important player in galectin-glycan interactions, were also cargo-specific and stimulated CD59 uptake. By contrast, inhibition of all galectin-glycan interactions by lactose inhibited CIE of both MHCI and CD59. None of these treatments affected clathrin-mediated endocytosis, implying that glycosylation changes specifically affect CIE. We also found that the galectin lattice tailors membrane fluidity and cell spreading. Furthermore, changes in membrane dynamics mediated by the galectin lattice affected macropinocytosis, an altered form of CIE, in HT1080 cells. Our results suggest that glycans play an important and nuanced role in CIE, with each cargo being affected uniquely by alterations in galectin and glycan profiles and their interactions. We conclude that galectin-driven effects exist on a continuum from stimulatory to inhibitory, with distinct CIE cargo proteins having unique response landscapes and with different cell types starting at different positions on these conceptual landscapes.

DNA methylation analysis reveals distinct methylation signatures in pediatric germ cell tumors.

PubMed

Amatruda, James F; Ross, Julie A; Christensen, Brock; Fustino, Nicholas J; Chen, Kenneth S; Hooten, Anthony J; Nelson, Heather; Kuriger, Jacquelyn K; Rakheja, Dinesh; Frazier, A Lindsay; Poynter, Jenny N

2013-06-27

Aberrant DNA methylation is a prominent feature of many cancers, and may be especially relevant in germ cell tumors (GCTs) due to the extensive epigenetic reprogramming that occurs in the germ line during normal development. We used the Illumina GoldenGate Cancer Methylation Panel to compare DNA methylation in the three main histologic subtypes of pediatric GCTs (germinoma, teratoma and yolk sac tumor (YST); N = 51) and used recursively partitioned mixture models (RPMM) to test associations between methylation pattern and tumor and demographic characteristics. We identified genes and pathways that were differentially methylated using generalized linear models and Ingenuity Pathway Analysis. We also measured global DNA methylation at LINE1 elements and evaluated methylation at selected imprinted loci using pyrosequencing. Methylation patterns differed by tumor histology, with 18/19 YSTs forming a distinct methylation class. Four pathways showed significant enrichment for YSTs, including a human embryonic stem cell pluripotency pathway. We identified 190 CpG loci with significant methylation differences in mature and immature teratomas (q < 0.05), including a number of CpGs in stem cell and pluripotency-related pathways. Both YST and germinoma showed significantly lower methylation at LINE1 elements compared with normal adjacent tissue while there was no difference between teratoma (mature and immature) and normal tissue. DNA methylation at imprinted loci differed significantly by tumor histology and location. Understanding methylation patterns may identify the developmental stage at which the GCT arose and the at-risk period when environmental exposures could be most harmful. Further, identification of relevant genetic pathways could lead to the development of new targets for therapy.

The class politics of prejudice: Brexit and the land of no-hope and glory.

PubMed

Mckenzie, Lisa

2017-11-01

The debates relating to social class and whether it is still a useful concept in describing a lived reality of the British population has never been far away from media, political and academic dispute. Thatcher's Britain throughout the 1980s attempted to dilute class meaning with what was called 'a home owning democracy' and thus end class collective politics through easily available credit for the working class while simultaneously attacking trade union organization, recruitment and political action. During the late 1990s and into the noughties a 'New Labour' administration attempted to exacerbate the end of class politics through an agenda of a 'cultural distinction' to class identity. Class struggle, class politics and class identity is embedded deep within the cultural norms practices, and history of British democracy. Consequently it is difficult if not impossible to prise class inequality in the UK away from and out of national, local and personal politics (Savage et al. 2015: 393-8). This paper focuses upon the sense that class politics, and cultural class distinction, within the UK had the biggest influence in determining a working-class 'Leave Vote' in the 2016 referendum within the UK. This paper explores accounts and narratives from working-class 'leave' voters though an ethnographic study of the political and social viewpoints of working-class communities of East London, and of ex-mining towns of Nottinghamshire. Framing into fuller context the anger and apathy of being 'left out', arguing that being 'left out' has been part of working-class political narratives for over 30 years. Going beyond frustration and apathy, a significant part of the narrative of working people was of 'not existing', suggesting certain important linkages with ongoing debates about new ways of conceptualizing class differences and class structures. © London School of Economics and Political Science 2017.

ROOT HAIR DEFECTIVE SIX-LIKE Class I Genes Promote Root Hair Development in the Grass Brachypodium distachyon

PubMed Central

Kim, Chul Min

2016-01-01

Genes encoding ROOT HAIR DEFECTIVE SIX-LIKE (RSL) class I basic helix loop helix proteins are expressed in future root hair cells of the Arabidopsis thaliana root meristem where they positively regulate root hair cell development. Here we show that there are three RSL class I protein coding genes in the Brachypodium distachyon genome, BdRSL1, BdRSL2 and BdRSL3, and each is expressed in developing root hair cells after the asymmetric cell division that forms root hair cells and hairless epidermal cells. Expression of BdRSL class I genes is sufficient for root hair cell development: ectopic overexpression of any of the three RSL class I genes induces the development of root hairs in every cell of the root epidermis. Expression of BdRSL class I genes in root hairless Arabidopsis thaliana root hair defective 6 (Atrhd6) Atrsl1 double mutants, devoid of RSL class I function, restores root hair development indicating that the function of these proteins has been conserved. However, neither AtRSL nor BdRSL class I genes is sufficient for root hair development in A. thaliana. These data demonstrate that the spatial pattern of class I RSL activity can account for the pattern of root hair cell differentiation in B. distachyon. However, the spatial pattern of class I RSL activity cannot account for the spatial pattern of root hair cells in A. thaliana. Taken together these data indicate that that the functions of RSL class I proteins have been conserved among most angiosperms—monocots and eudicots—despite the dramatically different patterns of root hair cell development. PMID:27494519

Essential role of Stat6 in IL-4 signalling.

PubMed

Takeda, K; Tanaka, T; Shi, W; Matsumoto, M; Minami, M; Kashiwamura, S; Nakanishi, K; Yoshida, N; Kishimoto, T; Akira, S

1996-04-18

Interleukin-4 (IL-4) is a pleiotropic lymphokine which plays an important role in the immune system. IL-4 activates two distinct signalling pathways through tyrosine phosphorylation of Stat6, a signal transducer and activator of transcription, and of a 170K protein called 4PS. To investigate the functional role of Stat6 in IL-4 signalling, we generated mice deficient in Stat6 by gene targeting. We report here that in the mutant mice, expression of CD23 and major histocompatibility complex (MHC) class II in resting B cells was not enhanced in response to IL-4. IL-4 induced B-cell proliferation costimulated by anti-IgM antibody was abolished. The T-cell proliferative response was also notably reduced. Furthermore, production of Th2 cytokines from T cells as well as IgE and IgG1 responses after nematode infection were profoundly reduced. These findings agreed with those obtained in IL-4 deficient mice or using antibodies to IL-4 and the IL-4 receptor. We conclude that Stat6 plays a central role in exerting IL-4 mediated biological responses.

Transdifferentiation and Proliferation in Two Distinct Hemocyte Lineages in Drosophila melanogaster Larvae after Wasp Infection

PubMed Central

Ihalainen, Teemu O.; Vanha-aho, Leena-Maija; Andó, István; Rämet, Mika

2016-01-01

Cellular immune responses require the generation and recruitment of diverse blood cell types that recognize and kill pathogens. In Drosophila melanogaster larvae, immune-inducible lamellocytes participate in recognizing and killing parasitoid wasp eggs. However, the sequence of events required for lamellocyte generation remains controversial. To study the cellular immune system, we developed a flow cytometry approach using in vivo reporters for lamellocytes as well as for plasmatocytes, the main hemocyte type in healthy larvae. We found that two different blood cell lineages, the plasmatocyte and lamellocyte lineages, contribute to the generation of lamellocytes in a demand-adapted hematopoietic process. Plasmatocytes transdifferentiate into lamellocyte-like cells in situ directly on the wasp egg. In parallel, a novel population of infection-induced cells, which we named lamelloblasts, appears in the circulation. Lamelloblasts proliferate vigorously and develop into the major class of circulating lamellocytes. Our data indicate that lamellocyte differentiation upon wasp parasitism is a plastic and dynamic process. Flow cytometry with in vivo hemocyte reporters can be used to study this phenomenon in detail. PMID:27414410

Transdifferentiation and Proliferation in Two Distinct Hemocyte Lineages in Drosophila melanogaster Larvae after Wasp Infection.

PubMed

Anderl, Ines; Vesala, Laura; Ihalainen, Teemu O; Vanha-Aho, Leena-Maija; Andó, István; Rämet, Mika; Hultmark, Dan

2016-07-01

Cellular immune responses require the generation and recruitment of diverse blood cell types that recognize and kill pathogens. In Drosophila melanogaster larvae, immune-inducible lamellocytes participate in recognizing and killing parasitoid wasp eggs. However, the sequence of events required for lamellocyte generation remains controversial. To study the cellular immune system, we developed a flow cytometry approach using in vivo reporters for lamellocytes as well as for plasmatocytes, the main hemocyte type in healthy larvae. We found that two different blood cell lineages, the plasmatocyte and lamellocyte lineages, contribute to the generation of lamellocytes in a demand-adapted hematopoietic process. Plasmatocytes transdifferentiate into lamellocyte-like cells in situ directly on the wasp egg. In parallel, a novel population of infection-induced cells, which we named lamelloblasts, appears in the circulation. Lamelloblasts proliferate vigorously and develop into the major class of circulating lamellocytes. Our data indicate that lamellocyte differentiation upon wasp parasitism is a plastic and dynamic process. Flow cytometry with in vivo hemocyte reporters can be used to study this phenomenon in detail.

Large oncosomes contain distinct protein cargo and represent a separate functional class of tumor-derived extracellular vesicles

PubMed Central

Minciacchi, Valentina R.; You, Sungyong; Spinelli, Cristiana; Morley, Samantha; Zandian, Mandana; Aspuria, Paul-Joseph; Cavallini, Lorenzo; Ciardiello, Chiara; Sobreiro, Mariana Reis; Morello, Matteo; Kharmate, Geetanjali; Jang, Su Chul; Kim, Dae-Kyum; Hosseini-Beheshti, Elham; Guns, Emma Tomlinson; Gleave, Martin; Gho, Yong Song; Mathivanan, Suresh; Yang, Wei; Freeman, Michael R.; Di Vizio, Dolores

2015-01-01

Large oncosomes (LO) are atypically large (1-10μm diameter) cancer-derived extracellular vesicles (EVs), originating from the shedding of membrane blebs and associated with advanced disease. We report that 25% of the proteins, identified by a quantitative proteomics analysis, are differentially represented in large and nano-sized EVs from prostate cancer cells. Proteins enriched in large EVs included enzymes involved in glucose, glutamine and amino acid metabolism, all metabolic processes relevant to cancer. Glutamine metabolism was altered in cancer cells exposed to large EVs, an effect that was not observed upon treatment with exosomes. Large EVs exhibited discrete buoyant densities in iodixanol (OptiPrepTM) gradients. Fluorescent microscopy of large EVs revealed an appearance consistent with LO morphology, indicating that these structures can be categorized as LO. Among the proteins enriched in LO, cytokeratin 18 (CK18) was one of the most abundant (within the top 5th percentile) and was used to develop an assay to detect LO in the circulation and tissues of mice and patients with prostate cancer. These observations indicate that LO represent a discrete EV type that may play a distinct role in tumor progression and that may be a source of cancer-specific markers. PMID:25857301

Subjective reactions to cocaine and marijuana are associated with abuse and dependence.

PubMed

Grant, Julia D; Scherrer, Jeffrey F; Lyons, Michael J; Tsuang, Ming; True, William R; Bucholz, Kathleen K

2005-09-01

Subjective effects of marijuana and cocaine use are associated with amount of drug use and potentially with risk of abuse and dependence. We used Latent Class Analyses (LCA) to examine subjective responses to two categories of drugs and link these to abuse and dependence. In 1992, subjective responses were queried of 2506 marijuana and 661 cocaine lifetime users who were members of the Vietnam Era Twin Registry. LCA was used to identify classes of subjective effects. Multinomial logistic regression models were computed to test for an association between classes and marijuana and cocaine abuse or dependence. The best LCA solution for marijuana identified 6 distinct classes characterized as positive, relaxed, reactive, adverse, low and very reactive. The best LCA solution for cocaine identified 5 distinct classes characterized as positive, alert, adverse, low and very reactive. Marijuana abuse and dependence were significantly associated with each latent class. Cocaine abuse was associated with the reactive class (OR=3.9; 95% CI: 1.6-9.5). Cocaine dependence was associated with reactive (OR=15.3; 95% CI: 7.1-32.6), adverse (OR=9.7; 95% CI: 4.5-21.0) and very reactive (OR=18.7; 95% CI: 5.6-62.6) classes. We found evidence for both qualitative and quantitative subjective effect profiles. Subjective effects, both positive and adverse are associated with lifetime risk for marijuana and cocaine dependence.

Novel fusion proteins for the antigen-specific staining and elimination of B cell receptor-positive cell populations demonstrated by a tetanus toxoid fragment C (TTC) model antigen.

PubMed

Klose, Diana; Saunders, Ute; Barth, Stefan; Fischer, Rainer; Jacobi, Annett Marita; Nachreiner, Thomas

2016-02-17

In an earlier study we developed a unique strategy allowing us to specifically eliminate antigen-specific murine B cells via their distinct B cell receptors using a new class of fusion proteins. In the present work we elaborated our idea to demonstrate the feasibility of specifically addressing and eliminating human memory B cells. The present study reveals efficient adaptation of the general approach to selectively target and eradicate human memory B cells. In order to demonstrate the feasibility we engineered a fusion protein following the principle of recombinant immunotoxins by combining a model antigen (tetanus toxoid fragment C, TTC) for B cell receptor targeting and a truncated version of Pseudomonas aeruginosa exotoxin A (ETA') to induce apoptosis after cellular uptake. The TTC-ETA' fusion protein not only selectively bound to a TTC-reactive murine B cell hybridoma cell line in vitro but also to freshly isolated human memory B cells from immunized donors ex vivo. Specific toxicity was confirmed on an antigen-specific population of human CD27(+) memory B cells. This protein engineering strategy can be used as a generalized platform approach for the construction of therapeutic fusion proteins with disease-relevant antigens as B cell receptor-binding domains, offering a promising approach for the specific depletion of autoreactive B-lymphocytes in B cell-driven autoimmune diseases.

IgG1 memory B cells keep the memory of IgE responses.

PubMed

He, Jin-Shu; Subramaniam, Sharrada; Narang, Vipin; Srinivasan, Kandhadayar; Saunders, Sean P; Carbajo, Daniel; Wen-Shan, Tsao; Hidayah Hamadee, Nur; Lum, Josephine; Lee, Andrea; Chen, Jinmiao; Poidinger, Michael; Zolezzi, Francesca; Lafaille, Juan J; Curotto de Lafaille, Maria A

2017-09-21

The unique differentiation of IgE cells suggests unconventional mechanisms of IgE memory. IgE germinal centre cells are transient, most IgE cells are plasma cells, and high affinity IgE is produced by the switching of IgG1 cells to IgE. Here we investigate the function of subsets of IgG1 memory B cells in IgE production and find that two subsets of IgG1 memory B cells, CD80 + CD73 + and CD80 - CD73 - , contribute distinctively to the repertoires of high affinity pathogenic IgE and low affinity non-pathogenic IgE. Furthermore, repertoire analysis indicates that high affinity IgE and IgG1 plasma cells differentiate from rare CD80 + CD73 + high affinity memory clones without undergoing further mutagenesis. By identifying the cellular origin of high affinity IgE and the clonal selection of high affinity memory B cells into the plasma cell fate, our findings provide fundamental insights into the pathogenesis of allergies, and on the mechanisms of antibody production in memory B cell responses.IgE is an important mediator of protective immunity as well as allergic reaction, but how high affinity IgE antibodies are produced in memory responses is not clear. Here the authors show that IgE can be generated via class-switch recombination in IgG1 memory B cells without additional somatic hypermutation.

A novel examination of exposure patterns and posttraumatic stress after a university mass murder.

PubMed

Liu, Sabrina R; Kia-Keating, Maryam

2018-03-05

Occurring at an alarming rate in the United States, mass violence has been linked to posttraumatic stress symptoms (PTSS) in both direct victims and community members who are indirectly exposed. Identifying what distinct exposure patterns exist and their relation to later PTSS has important clinical implications. The present study determined classes of exposure to an event of mass violence, and if PTSS differed across classes. First- and second-year college students (N = 1,189) participated in a confidential online survey following a mass murder at their university, which assessed event exposure and PTSS 3 months later. Latent class analysis (LCA) was used to empirically determine distinct classes of exposure patterns and links between class membership and PTSS. The final model yielded 4 classes: minimal exposure (55.5% of sample), auditory exposure (29.4% of sample), visual exposure (10% of sample), and interpersonal exposure (5% of sample). More severe direct exposure (i.e., the visual exposure class) was associated with significantly higher levels of PTSS than the auditory exposure or minimal exposure classes, as was the interpersonal exposure class. There were no significant differences in PTSS between the auditory exposure and minimal exposure classes or the visual exposure and interpersonal exposure classes. Results point to the differential impact of exposure categories, and provide empirical evidence for distinguishing among auditory, visual, and interpersonal exposures to events of mass violence on college campuses. Clinical implications suggest that visual and interpersonal exposure may warrant targeted efforts following mass violence. (PsycINFO Database Record (c) 2018 APA, all rights reserved).

Social power and social class: conceptualization, consequences, and current challenges.

PubMed

Rucker, Derek D; Galinsky, Adam D

2017-12-01

This article offers a primer on social power and social class with respect to their theoretical importance, conceptual distinction, and empirical relationship. We introduce and define the constructs of social power, social class, and one's psychological sense of power. We next explore the complex relationship between social power and social class. Because social class can produce a sense of power within an individual, studies on social power can inform theory and research on social class. We conclude with a discussion of the current challenges and future opportunities for the study of social power and social class. Copyright © 2017 Elsevier Ltd. All rights reserved.

Designed Proteins Induce the Formation of Nanocage-containing Extracellular Vesicles

PubMed Central

Votteler, Jörg; Ogohara, Cassandra; Yi, Sue; Hsia, Yang; Nattermann, Una; Belnap, David M.; King, Neil P.; Sundquist, Wesley I.

2017-01-01

Complex biological processes are often performed by self-organizing nanostructures comprising multiple classes of macromolecules, such as ribosomes (proteins and RNA) or enveloped viruses (proteins, nucleic acids, and lipids). Approaches have been developed for designing self-assembling structures consisting of either nucleic acids1,2 or proteins3–5, but strategies for engineering hybrid biological materials are only beginning to emerge6,7. Here, we describe the design of self-assembling protein nanocages that direct their own release from human cells inside small vesicles in a manner that resembles some viruses. We refer to these hybrid biomaterials as Enveloped Protein Nanocages (EPNs). Robust EPN biogenesis required protein sequence elements that encode three distinct functions: membrane binding, self-assembly, and recruitment of the Endosomal Sorting Complexes Required for Transport (ESCRT) machinery8. A variety of synthetic proteins with these functional elements induced EPN biogenesis, highlighting the modularity and generality of the design strategy. Biochemical and electron cryomicroscopic (cryo-EM) analyses revealed that one design, EPN-01, comprised small (~100 nm) vesicles containing multiple protein nanocages that closely matched the structure of the designed 60-subunit self-assembling scaffold9. EPNs that incorporated the vesicular stomatitis viral glycoprotein (VSV-G) could fuse with target cells and deliver their contents, thereby transferring cargoes from one cell to another. These studies show how proteins can be programmed to direct the formation of hybrid biological materials that perform complex tasks, and establish EPNs as a novel class of designed, modular, genetically-encoded nanomaterials that can transfer molecules between cells. PMID:27919066

Structural basis of semaphorin–plexin signalling

PubMed Central

Janssen, Bert J. C.; Robinson, Ross A.; Pérez-Brangulí, Francesc; Bell, Christian H.; Mitchell, Kevin J.; Siebold, Christian; Jones, E. Yvonne

2013-01-01

Cell-cell signalling of semaphorin ligands through interaction with plexin receptors is important for the homeostasis and morphogenesis of many tissues and is widely studied for its role in neural connectivity, cancer, cell migration and immune responses1. SEMA4D and Sema6A exemplify two diverse vertebrate, membrane-spanning semaphorin classes (4 and 6) that are capable of direct signalling through members of the two largest plexin classes, B and A, respectively2,3. In the absence of any structural information on the plexin ectodomain or its interaction with semaphorins the extracellular specificity and mechanism controlling plexin signalling has remained unresolved. Here we present crystal structures of cognate complexes of the semaphorin-binding regions of plexins B1 and A2 with semaphorin ectodomains (human PLXNB11–2–SEMA4Decto and murine PlxnA21–4–Sema6Aecto), plus unliganded structures of PlxnA21–4 and Sema6Aecto. These structures, together with biophysical and cellular assays of wild-type and mutant proteins, reveal that semaphorin dimers independently bind two plexin molecules and that signalling is critically dependent on the avidity of the resulting bivalent 2:2 complex (monomeric semaphorin binds plexin but fails to trigger signalling). In combination, our data favour a cell-cell signalling mechanism involving semaphorin-stabilized plexin dimerization, possibly followed by clustering, which is consistent with previous functional data. Furthermore, the shared generic architecture of the complexes, formed through conserved contacts of the amino-terminal seven-bladed β-propeller (sema) domains of both semaphorin and plexin, suggests that a common mode of interaction triggers all semaphorin–plexin based signalling, while distinct insertions within or between blades of the sema domains determine binding specificity. PMID:20877282

From The Cover: Induction of antiviral immunity requires Toll-like receptor signaling in both stromal and dendritic cell compartments

NASA Astrophysics Data System (ADS)

Sato, Ayuko; Iwasaki, Akiko

2004-11-01

Pattern recognition by Toll-like receptors (TLRs) is known to be important for the induction of dendritic cell (DC) maturation. DCs, in turn, are critically important in the initiation of T cell responses. However, most viruses do not infect DCs. This recognition system poses a biological problem in ensuring that most viral infections be detected by pattern recognition receptors. Furthermore, it is unknown what, if any, is the contribution of TLRs expressed by cells that are infected by a virus, versus TLRs expressed by DCs, in the initiation of antiviral adaptive immunity. Here we address these issues using a physiologically relevant model of mucosal infection with herpes simplex virus type 2. We demonstrate that innate immune recognition of viral infection occurs in two distinct stages, one at the level of the infected epithelial cells and the other at the level of the noninfected DCs. Importantly, both TLR-mediated recognition events are required for the induction of effector T cells. Our results demonstrate that virally infected tissues instruct DCs to initiate the appropriate class of effector T cell responses and reveal the critical importance of the stromal cells in detecting infectious agents through their own pattern recognition receptors. mucosal immunity | pattern recognition | viral infection

Human immunodeficiency virus infection of helper T cell clones. Early proliferative defects despite intact antigen-specific recognition and interleukin 4 secretion.

PubMed Central

Laurence, J; Friedman, S M; Chartash, E K; Crow, M K; Posnett, D N

1989-01-01

HIV selectively inhibited the proliferative response of clonal CD4+ T lymphocytes to alloantigen while other alloantigen-dependent responses were unperturbed. Specifically, impaired blastogenesis could be dissociated from alloantigen-specific induction of the B cell activation molecule CD23, IL-4 release, and inositol lipid hydrolysis. In addition, membrane expression of pertinent T cell receptor molecules, including CD2, CD3, and T cell antigen receptor (Ti), remained intact. Using two MHC class II-specific human CD4+ helper T cell clones, the proliferative defect was shown to be an early consequence of HIV infection, occurring within 4 d of viral inoculation and preceding increases in mature virion production. It was generalizable to three distinct methods of T cell activation, all independent of antigen-presenting cells: anti-CD3 mediated cross-linking of the CD3/Ti complex; anti-CD2 and phorbol 12-myristic 13-acetate (PMA); and anti-CD28 plus PMA. These abnormalities were not mitigated by addition of exogenous IL-2, even though expression of the IL-2 receptor (CD25) was unaltered. These studies define a selective blockade in T cell function early after HIV exposure that could serve as a model for certain in vivo manifestations of AIDS. PMID:2470786

Lineage tracing of human B cells reveals the in vivo landscape of human antibody class switching

PubMed Central

Horns, Felix; Vollmers, Christopher; Croote, Derek; Mackey, Sally F; Swan, Gary E; Dekker, Cornelia L; Davis, Mark M; Quake, Stephen R

2016-01-01

Antibody class switching is a feature of the adaptive immune system which enables diversification of the effector properties of antibodies. Even though class switching is essential for mounting a protective response to pathogens, the in vivo patterns and lineage characteristics of antibody class switching have remained uncharacterized in living humans. Here we comprehensively measured the landscape of antibody class switching in human adult twins using antibody repertoire sequencing. The map identifies how antibodies of every class are created and delineates a two-tiered hierarchy of class switch pathways. Using somatic hypermutations as a molecular clock, we discovered that closely related B cells often switch to the same class, but lose coherence as somatic mutations accumulate. Such correlations between closely related cells exist when purified B cells class switch in vitro, suggesting that class switch recombination is directed toward specific isotypes by a cell-autonomous imprinted state. DOI: http://dx.doi.org/10.7554/eLife.16578.001 PMID:27481325

Differences in antigen presentation to MHC class I-and class II- restricted influenza virus-specific cytolytic T lymphocyte clones

PubMed Central

1986-01-01

We have examined requirements for antigen presentation to a panel of MHC class I-and class II-restricted, influenza virus-specific CTL clones by controlling the form of virus presented on the target cell surface. Both H-2K/D- and I region-restricted CTL recognize target cells exposed to infectious virus, but only the I region-restricted clones efficiently lysed histocompatible target cells pulsed with inactivated virus preparations. The isolated influenza hemagglutinin (HA) polypeptide also could sensitize target cells for recognition by class II-restricted, HA-specific CTL, but not by class I-restricted, HA- specific CTL. Inhibition of nascent viral protein synthesis abrogated the ability of target cells to present viral antigen relevant for class I-restricted CTL recognition. Significantly, presentation for class II- restricted recognition was unaffected in target cells exposed to preparations of either inactivated or infectious virus. This differential sensitivity suggested that these H-2I region-restricted CTL recognized viral polypeptides derived from the exogenously introduced virions, rather than viral polypeptides newly synthesized in the infected cell. In support of this contention, treatment of the target cells with the lysosomotropic agent chloroquine abolished recognition of infected target cells by class II-restricted CTL without diminishing class I-restricted recognition of infected target cells. Furthermore, when the influenza HA gene was introduced into target cells without exogenous HA polypeptide, the target cells that expressed the newly synthesized protein product of the HA gene were recognized only by H-2K/D-restricted CTL. These observations suggest that important differences may exist in requirements for antigen presentation between H-2K/D and H-2I region-restricted CTL. These differences may reflect the nature of the antigenic epitopes recognized by these two CTL subsets. PMID:3485173

Functional photoreceptor loss revealed with adaptive optics: an alternate cause of color blindness.

PubMed

Carroll, Joseph; Neitz, Maureen; Hofer, Heidi; Neitz, Jay; Williams, David R

2004-06-01

There is enormous variation in the X-linked L/M (long/middle wavelength sensitive) gene array underlying "normal" color vision in humans. This variability has been shown to underlie individual variation in color matching behavior. Recently, red-green color blindness has also been shown to be associated with distinctly different genotypes. This has opened the possibility that there may be important phenotypic differences within classically defined groups of color blind individuals. Here, adaptive optics retinal imaging has revealed a mechanism for producing dichromatic color vision in which the expression of a mutant cone photopigment gene leads to the loss of the entire corresponding class of cone photoreceptor cells. Previously, the theory that common forms of inherited color blindness could be caused by the loss of photoreceptor cells had been discounted. We confirm that remarkably, this loss of one-third of the cones does not impair any aspect of vision other than color.

Two distinct arginine methyltransferases are required for biogenesis of Sm-class ribonucleoproteins.

PubMed

Gonsalvez, Graydon B; Tian, Liping; Ospina, Jason K; Boisvert, François-Michel; Lamond, Angus I; Matera, A Gregory

2007-08-27

Small nuclear ribonucleoproteins (snRNPs) are core components of the spliceosome. The U1, U2, U4, and U5 snRNPs each contain a common set of seven Sm proteins. Three of these Sm proteins are posttranslationally modified to contain symmetric dimethylarginine (sDMA) residues within their C-terminal tails. However, the precise function of this modification in the snRNP biogenesis pathway is unclear. Several lines of evidence suggest that the methyltransferase protein arginine methyltransferase 5 (PRMT5) is responsible for sDMA modification of Sm proteins. We found that in human cells, PRMT5 and a newly discovered type II methyltransferase, PRMT7, are each required for Sm protein sDMA modification. Furthermore, we show that the two enzymes function nonredundantly in Sm protein methylation. Lastly, we provide in vivo evidence demonstrating that Sm protein sDMA modification is required for snRNP biogenesis in human cells.

Two distinct arginine methyltransferases are required for biogenesis of Sm-class ribonucleoproteins

PubMed Central

Gonsalvez, Graydon B.; Tian, Liping; Ospina, Jason K.; Boisvert, François-Michel; Lamond, Angus I.; Matera, A. Gregory

2007-01-01

Small nuclear ribonucleoproteins (snRNPs) are core components of the spliceosome. The U1, U2, U4, and U5 snRNPs each contain a common set of seven Sm proteins. Three of these Sm proteins are posttranslationally modified to contain symmetric dimethylarginine (sDMA) residues within their C-terminal tails. However, the precise function of this modification in the snRNP biogenesis pathway is unclear. Several lines of evidence suggest that the methyltransferase protein arginine methyltransferase 5 (PRMT5) is responsible for sDMA modification of Sm proteins. We found that in human cells, PRMT5 and a newly discovered type II methyltransferase, PRMT7, are each required for Sm protein sDMA modification. Furthermore, we show that the two enzymes function nonredundantly in Sm protein methylation. Lastly, we provide in vivo evidence demonstrating that Sm protein sDMA modification is required for snRNP biogenesis in human cells. PMID:17709427

DNA repair mechanisms in cancer development and therapy.

PubMed

Torgovnick, Alessandro; Schumacher, Björn

2015-01-01

DNA damage has been long recognized as causal factor for cancer development. When erroneous DNA repair leads to mutations or chromosomal aberrations affecting oncogenes and tumor suppressor genes, cells undergo malignant transformation resulting in cancerous growth. Genetic defects can predispose to cancer: mutations in distinct DNA repair systems elevate the susceptibility to various cancer types. However, DNA damage not only comprises a root cause for cancer development but also continues to provide an important avenue for chemo- and radiotherapy. Since the beginning of cancer therapy, genotoxic agents that trigger DNA damage checkpoints have been applied to halt the growth and trigger the apoptotic demise of cancer cells. We provide an overview about the involvement of DNA repair systems in cancer prevention and the classes of genotoxins that are commonly used for the treatment of cancer. A better understanding of the roles and interactions of the highly complex DNA repair machineries will lead to important improvements in cancer therapy.

Community structure of planktonic methane-oxidizing bacteria in a subtropical reservoir characterized by dominance of phylotype closely related to nitrite reducer

NASA Astrophysics Data System (ADS)

Kojima, Hisaya; Tokizawa, Riho; Kogure, Kouhei; Kobayashi, Yuki; Itoh, Masayuki; Shiah, Fuh-Kwo; Okuda, Noboru; Fukui, Manabu

2014-07-01

Methane-oxidizing bacteria (MOB) gain energy from the oxidation of methane and may play important roles in freshwater ecosystems. In this study, the community structure of planktonic MOB was investigated in a subtropical reservoir. Bacterial community structure was investigated through the analysis of the 16S rRNA gene. Three groups of phylogenetically distinct MOB were detected in the clone libraries of polymerase chain reaction products obtained with universal primers. The groups belonged to the class Gammaproteobacteria, the class Alphaproteobacteria, and the candidate phylum NC10. The last group, which consists of close relatives of the nitrite reducer `Candidatus Methylomirabilis oxyfera', was frequently detected in the clone libraries of deep-water environments. The presence of 3 groups of MOB in deep water was also shown by a cloning analysis of the pmoA gene encoding particulate methane monooxygenase. The dominance of `M. oxyfera'-like organisms in deep water was confirmed by catalyzed reporter deposition-fluorescence in situ hybridization, in which cells stained with a specific probe accounted for 16% of total microbial cells. This is the first study to demonstrate that close relatives of the nitrite reducer can be major component of planktonic MOB community which may affect carbon flow in aquatic ecosystems.

Trajectories of Quality of Life after Hematopoietic Cell Transplantation: Secondary Analysis of BMT CTN 0902 Data

PubMed Central

Jim, Heather S.L.; Sutton, Steven K.; Small, Brent J.; Jacobsen, Paul B.; Wood, William A.; Knight, Jennifer M.; Majhail, Navneet S.; Syrjala, Karen L.; Lee, Stephanie J.

2016-01-01

Summary Quality of life is increasingly recognized as an important secondary endpoint of hematopoietic cell transplantation (HCT). The current study examined the extent to which attrition results in biased estimates of patient quality of life. The study also examined whether patients differ in terms of trajectories of quality of life in the first six months post-transplant. A secondary data analysis was conducted of 701 participants who enrolled in the Blood and Marrow Transplantation Clinical Trials Network (BMT CTN) 0902 trial. Participants completed the SF-36, a measure of quality of life, prior to transplant and 100 and 180 days post-transplant. Results indicated that attrition resulted in slightly biased overestimates of quality of life but the amount of overestimation remained stable over time. Patients could be grouped into three distinct classes based on physical quality of life: 1) low and stable; 2) average and declining, then stable; and 3) average and stable. Four classes of patients emerged for mental quality of life: 1) low and stable; 2) average, improving, then stable; 3) higher than average (by almost 1 SD) and stable; and 4) average and stable. Taken together, these data provide a more comprehensive understanding of quality of life that can be used to educate HCT recipients and their caregivers. PMID:27538374

Phytochrome regulation of cellulose synthesis in Arabidopsis.

PubMed

Bischoff, Volker; Desprez, Thierry; Mouille, Gregory; Vernhettes, Samantha; Gonneau, Martine; Höfte, Herman

2011-11-08

Plant development is highly plastic and dependent on light quantity and quality monitored by specific photoreceptors. Although we have a detailed knowledge of light signaling pathways, little is known about downstream targets involved in growth control. Cell size and shape are in part controlled by cellulose microfibrils extruded from large cellulose synthase complexes (CSCs) that migrate in the plasma membrane along cortical microtubules. Here we show a role for the red/far-red light photoreceptor PHYTOCHROME B (PHYB) in the regulation of cellulose synthesis in the growing Arabidopsis hypocotyl. In this organ, CSCs contains three distinct cellulose synthase (CESA) isoform classes: nonredundant CESA1 and CESA3 and a third class represented by partially redundant CESA2, CESA5, and CESA6. Interestingly, in the dark, depending on which CESA subunits occupy the third position, CSC velocity is more or less inhibited through an interaction with microtubules. Activation of PHYB overrules this inhibition. The analysis of cesa5 mutants shows a role for phosphorylation in the control of CSC velocity. These results, combined with the cesa5 mutant phenotype, suggest that cellulose synthesis is fine tuned through the regulated interaction of CSCs with microtubules and that PHYB signaling impinges on this process to maintain cell wall strength and growth in changing environments. Copyright © 2011 Elsevier Ltd. All rights reserved.

Structurally distinct membrane nanotubes between human macrophages support long-distance vesicular traffic or surfing of bacteria.

PubMed

Onfelt, Björn; Nedvetzki, Shlomo; Benninger, Richard K P; Purbhoo, Marco A; Sowinski, Stefanie; Hume, Alistair N; Seabra, Miguel C; Neil, Mark A A; French, Paul M W; Davis, Daniel M

2006-12-15

We report that two classes of membrane nanotubes between human monocyte-derived macrophages can be distinguished by their cytoskeletal structure and their functional properties. Thin membrane nanotubes contained only F-actin, whereas thicker nanotubes, i.e., those > approximately 0.7 microm in diameter, contained both F-actin and microtubules. Bacteria could be trapped and surf along thin, but not thick, membrane nanotubes toward connected macrophage cell bodies. Once at the cell body, bacteria could then be phagocytosed. The movement of bacteria is aided by a constitutive flow of the nanotube surface because streptavidin-coated beads were similarly able to traffic along nanotubes between surface-biotinylated macrophages. Mitochondria and intracellular vesicles, including late endosomes and lysosomes, could be detected within thick, but not thin, membrane nanotubes. Analysis from kymographs demonstrated that vesicles moved in a stepwise, bidirectional manner at approximately 1 microm/s, consistent with their traffic being mediated by the microtubules found only in thick nanotubes. Vesicular traffic in thick nanotubes and surfing of beads along thin nanotubes were both stopped upon the addition of azide, demonstrating that both processes require ATP. However, microtubule destabilizing agents colchicine or nocodazole abrogated vesicular transport but not the flow of the nanotube surface, confirming that distinct cytoskeletal structures of nanotubes give rise to different functional properties. Thus, membrane nanotubes between macrophages are more complex than unvarying ubiquitous membrane tethers and facilitate several means for distal interactions between immune cells.

Profiling of wheat class III peroxidase genes derived from powdery mildew-attacked epidermis reveals distinct sequence-associated expression patterns.

PubMed

Liu, Guosheng; Sheng, Xiaoyan; Greenshields, David L; Ogieglo, Adam; Kaminskyj, Susan; Selvaraj, Gopalan; Wei, Yangdou

2005-07-01

A cDNA library was constructed from leaf epidermis of diploid wheat (Triticum monococcum) infected with the powdery mildew fungus (Blumeria graminis f. sp. tritici) and was screened for genes encoding peroxidases. From 2,500 expressed sequence tags (ESTs), 36 cDNAs representing 10 peroxidase genes (designated TmPRX1 to TmPRX10) were isolated and further characterized. Alignment of the deduced amino acid sequences and phylogenetic clustering with peroxidases from other plant species demonstrated that these peroxidases fall into four distinct groups. Differential expression and tissue-specific localization among the members were observed during the B. graminis f. sp. tritici attack using Northern blots and reverse-transcriptase polymerase chain reaction analyses. Consistent with its abundance in the EST collection, TmPRX1 expression showed the highest induction during pathogen attack and fluctuated in response to the fungal parasitic stages. TmPRX1 to TmPRX6 were expressed predominantly in mesophyll cells, whereas TmPRX7 to TmPRX10, which feature a putative C-terminal propeptide, were detectable mainly in epidermal cells. Using TmPRX8 as a representative, we demonstrated that its C-terminal propeptide was sufficient to target a green fluorescent protein fusion protein to the vacuoles in onion cells. Finally, differential expression profiles of the TmPRXs after abiotic stresses and signal molecule treatments were used to dissect the potential role of these peroxidases in multiple stress and defense pathways.

Tangential migration of glutamatergic neurons and cortical patterning during development: Lessons from Cajal-Retzius cells.

PubMed

Barber, Melissa; Pierani, Alessandra

2016-08-01

Tangential migration is a mode of cell movement, which in the developing cerebral cortex, is defined by displacement parallel to the ventricular surface and orthogonal to the radial glial fibers. This mode of long-range migration is a strategy by which distinct neuronal classes generated from spatially and molecularly distinct origins can integrate to form appropriate neural circuits within the cortical plate. While it was previously believed that only GABAergic cortical interneurons migrate tangentially from their origins in the subpallial ganglionic eminences to integrate in the cortical plate, it is now known that transient populations of glutamatergic neurons also adopt this mode of migration. These include Cajal-Retzius cells (CRs), subplate neurons (SPs), and cortical plate transient neurons (CPTs), which have crucial roles in orchestrating the radial and tangential development of the embryonic cerebral cortex in a noncell-autonomous manner. While CRs have been extensively studied, it is only in the last decade that the molecular mechanisms governing their tangential migration have begun to be elucidated. To date, the mechanisms of SPs and CPTs tangential migration remain unknown. We therefore review the known signaling pathways, which regulate parameters of CRs migration including their motility, contact-redistribution and adhesion to the pial surface, and discuss this in the context of how CR migration may regulate their signaling activity in a spatial and temporal manner. © 2015 Wiley Periodicals, Inc. Develop Neurobiol 76: 847-881, 2016. © 2015 Wiley Periodicals, Inc.

Update on "What" and "Where" in Spatial Language: A New Division of Labor for Spatial Terms.

PubMed

Landau, Barbara

2017-03-01

In this article, I revisit Landau and Jackendoff's () paper, "What and where in spatial language and spatial cognition," proposing a friendly amendment and reformulation. The original paper emphasized the distinct geometries that are engaged when objects are represented as members of object kinds (named by count nouns), versus when they are represented as figure and ground in spatial expressions (i.e., play the role of arguments of spatial prepositions). We provided empirical and theoretical arguments for the link between these distinct representations in spatial language and their accompanying nonlinguistic neural representations, emphasizing the "what" and "where" systems of the visual system. In the present paper, I propose a second division of labor between two classes of spatial prepositions in English that appear to be quite distinct. One class includes prepositions such as in and on, whose core meanings engage force-dynamic, functional relationships between objects, with geometry only a marginal player. The second class includes prepositions such as above/below and right/left, whose core meanings engage geometry, with force-dynamic relationships a passing or irrelevant variable. The insight that objects' force-dynamic relationships matter to spatial terms' uses is not new; but thinking of these terms as a distinct set within spatial language has theoretical and empirical consequences that are new. I propose three such consequences, rooted in the fact that geometric knowledge is highly constrained and early-emerging in life, while force-dynamic knowledge of objects and their interactions is relatively unconstrained and needs to be learned piecemeal over a lengthy timeline. First, the two classes will engage different learning problems, with different developmental trajectories for both first and second language learners; second, the classes will naturally lead to different degrees of cross-linguistic variation; and third, they may be rooted in different neural representations. Copyright © 2016 Cognitive Science Society, Inc.

Haemophilus ducreyi Cutaneous Ulcer Strains Diverged from Both Class I and Class II Genital Ulcer Strains: Implications for Epidemiological Studies.

PubMed

Gangaiah, Dharanesh; Spinola, Stanley M

2016-12-01

Haemophilus ducreyi has emerged as a major cause of cutaneous ulcers (CU) in yaws-endemic regions of the tropics in the South Pacific, South East Asia and Africa. H. ducreyi was once thought only to cause the genital ulcer (GU) disease chancroid; GU strains belong to 2 distinct classes, class I and class II. Using whole-genome sequencing of 4 CU strains from Samoa, 1 from Vanuatu and 1 from Papua New Guinea, we showed that CU strains diverged from the class I strain 35000HP and that one CU strain expressed β-lactamase. Recently, the Center for Disease Control and Prevention released the genomes of 11 additional CU strains from Vanuatu and Ghana; however, the evolutionary relationship of these CU strains to previously-characterized CU and GU strains is unknown. We performed phylogenetic analysis of 17 CU and 10 GU strains. Class I and class II GU strains formed two distinct clades. The class I strains formed two subclades, one containing 35000HP and HD183 and the other containing the remainder of the class I strains. Twelve of the CU strains formed a subclone under the class I 35000HP subclade, while 2 CU strains formed a subclone under the other class I subclade. Unexpectedly, 3 of the CU strains formed a subclone under the class II clade. Phylogenetic analysis of dsrA-hgbA-ncaA sequences yielded a tree similar to that of whole-genome phylogenetic tree. CU strains diverged from multiple lineages within both class I and class II GU strains. Multilocus sequence typing of dsrA-hgbA-ncaA could be reliably used for epidemiological investigation of CU and GU strains. As class II strains grow relatively poorly and are relatively more susceptible to vancomycin than class I strains, these findings have implications for methods to recover CU strains. Comparison of contemporary CU and GU isolates would help clarify the relationship between these entities.

Trajectories of Social Withdrawal from Middle Childhood to Early Adolescence

PubMed Central

Oh, Wonjung; Bowker, Julie C.; Booth-LaForce, Cathryn; Rose-Krasnor, Linda; Laursen, Brett

2013-01-01

Heterogeneity and individual differences in the developmental course of social withdrawal were examined longitudinally in a community sample (N=392). General Growth Mixture Modeling (GGMM) was used to identify distinct pathways of social withdrawal, differentiate valid subgroup trajectories, and examine factors that predicted change in trajectories within subgroups. Assessments of individual (social withdrawal), interactive (prosocial behavior), relationship (friendship involvement, stability and quality, best friend’s withdrawal and exclusion/victimization) and group- (exclusion/victimization) level characteristics were used to define growth trajectories from the final year of elementary school, across the transition to middle school, and then to the final year of middle school (fifth-to-eighth grades). Three distinct trajectory classes were identified: low stable, increasing, and decreasing. Peer exclusion, prosocial behavior, and mutual friendship involvement differentiated class membership. Friendlessness, friendship instability, and exclusion were significant predictors of social withdrawal for the increasing class, whereas lower levels of peer exclusion predicted a decrease in social withdrawal for the decreasing class. PMID:18193479

Search for Type Ia supernova NUV-optical subclasses

NASA Astrophysics Data System (ADS)

Cinabro, David; Scolnic, Daniel; Kessler, Richard; Li, Ashley; Miller, Jake

2017-04-01

In response to a recently reported observation of evidence for two classes of Type Ia supernovae (SNe Ia) distinguished by their brightness in the rest-frame near-ultraviolet (NUV), we search for the phenomenon in publicly available light-curve data. We use the SNANA supernova analysis package to simulate SN Ia light curves in the Sloan Digital Sky Survey (SDSS) Supernova Search and the Supernova Legacy Survey (SNLS) with a model of two distinct ultraviolet classes of SNe Ia and a conventional model with a single broad distribution of SN-Ia ultraviolet brightnesses. We compare simulated distributions of rest-frame colours with these two models to those observed in 158 SNe Ia in the SDSS and SNLS data. The SNLS sample of 99 SNe Ia is in clearly better agreement with a model with one class of SN Ia light curves and shows no evidence for distinct NUV sub-classes. The SDSS sample of 59 SNe Ia with poorer colour resolution does not distinguish between the two models.

Trajectories of social withdrawal from middle childhood to early adolescence.

PubMed

Oh, Wonjung; Rubin, Kenneth H; Bowker, Julie C; Booth-LaForce, Cathryn; Rose-Krasnor, Linda; Laursen, Brett

2008-05-01

Heterogeneity and individual differences in the developmental course of social withdrawal were examined longitudinally in a community sample (N = 392). General Growth Mixture Modeling (GGMM) was used to identify distinct pathways of social withdrawal, differentiate valid subgroup trajectories, and examine factors that predicted change in trajectories within subgroups. Assessments of individual (social withdrawal), interactive (prosocial behavior), relationship (friendship involvement, stability and quality, best friend's withdrawal and exclusion/victimization) and group- (exclusion/victimization) level characteristics were used to define growth trajectories from the final year of elementary school, across the transition to middle school, and then to the final year of middle school (fifth-to-eighth grades). Three distinct trajectory classes were identified: low stable, increasing, and decreasing. Peer exclusion, prosocial behavior, and mutual friendship involvement differentiated class membership. Friendlessness, friendship instability, and exclusion were significant predictors of social withdrawal for the increasing class, whereas lower levels of peer exclusion predicted a decrease in social withdrawal for the decreasing class.

The DSIF subunits Spt4 and Spt5 have distinct roles at various phases of immunoglobulin class switch recombination.

PubMed

Stanlie, Andre; Begum, Nasim A; Akiyama, Hideo; Honjo, Tasuku

2012-01-01

Class-switch recombination (CSR), induced by activation-induced cytidine deaminase (AID), can be divided into two phases: DNA cleavage of the switch (S) regions and the joining of the cleaved ends of the different S regions. Here, we show that the DSIF complex (Spt4 and Spt5), a transcription elongation factor, is required for CSR in a switch-proficient B cell line CH12F3-2A cells, and Spt4 and Spt5 carry out independent functions in CSR. While neither Spt4 nor Spt5 is required for transcription of S regions and AID, expression array analysis suggests that Spt4 and Spt5 regulate a distinct subset of transcripts in CH12F3-2A cells. Curiously, Spt4 is critically important in suppressing cryptic transcription initiating from the intronic Sμ region. Depletion of Spt5 reduced the H3K4me3 level and DNA cleavage at the Sα region, whereas Spt4 knockdown did not perturb the H3K4me3 status and S region cleavage. H3K4me3 modification level thus correlated well with the DNA breakage efficiency. Therefore we conclude that Spt5 plays a role similar to the histone chaperone FACT complex that regulates H3K4me3 modification and DNA cleavage in CSR. Since Spt4 is not involved in the DNA cleavage step, we suspected that Spt4 might be required for DNA repair in CSR. We examined whether Spt4 or Spt5 is essential in non-homologous end joining (NHEJ) and homologous recombination (HR) as CSR utilizes general repair pathways. Both Spt4 and Spt5 are required for NHEJ and HR as determined by assay systems using synthetic repair substrates that are actively transcribed even in the absence of Spt4 and Spt5. Taken together, Spt4 and Spt5 can function independently in multiple transcription-coupled steps of CSR.

Antitumor Lipids--Structure, Functions, and Medical Applications.

PubMed

Kostadinova, Aneliya; Topouzova-Hristova, Tanya; Momchilova, Albena; Tzoneva, Rumiana; Berger, Martin R

2015-01-01

Cell proliferation and metastasis are considered hallmarks of tumor progression. Therefore, efforts have been made to develop novel anticancer drugs that inhibit both the proliferation and the motility of tumor cells. Synthetic antitumor lipids (ATLs), which are chemically divided into two main classes, comprise (i) alkylphospholipids (APLs) and (ii) alkylphosphocholines (APCs). They represent a new entity of drugs with distinct antiproliferative properties in tumor cells. These compounds do not interfere with the DNA or mitotic spindle apparatus of the cell, instead, they incorporate into cell membranes, where they accumulate and interfere with lipid metabolism and lipid-dependent signaling pathways. Recently, it has been shown that the most commonly studied APLs inhibit proliferation by inducing apoptosis in malignant cells while leaving normal cells unaffected and are potent sensitizers of conventional chemo- and radiotherapy, as well as of electrical field therapy. APLs resist catabolic degradation to a large extent, therefore accumulate in the cell and interfere with lipid-dependent survival signaling pathways, notably PI3K-Akt and Raf-Erk1/2, and de novo phospholipid biosynthesis. They are internalized in the cell membrane via raft domains and cause downstream reactions as inhibition of cell growth and migration, cell cycle arrest, actin stress fibers collapse, and apoptosis. This review summarizes the in vitro, in vivo, and clinical trials of most common ATLs and their mode of action at molecular and biochemical levels. © 2015 Elsevier Inc. All rights reserved.

Four Distinct Subgroups of Self-Injurious Behavior among Chinese Adolescents: Findings from a Latent Class Analysis

PubMed Central

Xin, Xiuhong; Ming, Qingsen; Zhang, Jibiao; Wang, Yuping; Liu, Mingli; Yao, Shuqiao

2016-01-01

Self-injurious behavior (SIB) among adolescents is an important public health issue worldwide. It is still uncertain whether homogeneous subgroups of SIB can be identified and whether constellations of SIBs can co-occur due to the high heterogeneity of these behaviors. In this study, a cross-sectional study was conducted on a large school-based sample and latent class analysis was performed (n = 10,069, mean age = 15 years) to identify SIB classes based on 11 indicators falling under direct SIB (DSIB), indirect SIB (ISIB), and suicide attempts (SAs). Social and psychological characteristics of each subgroup were examined after controlling for age and gender. Results showed that a four-class model best fit the data and each class had a distinct pattern of co-occurrence of SIBs and external measures. Class 4 (the baseline/normative group, 65.3%) had a low probability of SIB. Class 3 (severe SIB group, 3.9%) had a high probability of SIB and the poorest social and psychological status. Class 1 (DSIB+SA group, 14.2%) had similar scores for external variables compared to class 3, and included a majority of girls [odds ratio (OR) = 1.94]. Class 2 (ISIB group, 16.6%) displayed moderate endorsement of ISIB items, and had a majority of boys and older adolescents (OR = 1.51). These findings suggest that SIB is a heterogeneous entity, but it may be best explained by four homogenous subgroups that display quantitative and qualitative differences. Findings in this study will improve our understanding on SIB and may facilitate the prevention and treatment of SIB. PMID:27392132

Characterization of Non-coding DNA Satellites Associated with Sweepoviruses (Genus Begomovirus, Geminiviridae) – Definition of a Distinct Class of Begomovirus-Associated Satellites

PubMed Central

Lozano, Gloria; Trenado, Helena P.; Fiallo-Olivé, Elvira; Chirinos, Dorys; Geraud-Pouey, Francis; Briddon, Rob W.; Navas-Castillo, Jesús

2016-01-01

Begomoviruses (family Geminiviridae) are whitefly-transmitted, plant-infecting single-stranded DNA viruses that cause crop losses throughout the warmer parts of the World. Sweepoviruses are a phylogenetically distinct group of begomoviruses that infect plants of the family Convolvulaceae, including sweet potato (Ipomoea batatas). Two classes of subviral molecules are often associated with begomoviruses, particularly in the Old World; the betasatellites and the alphasatellites. An analysis of sweet potato and Ipomoea indica samples from Spain and Merremia dissecta samples from Venezuela identified small non-coding subviral molecules in association with several distinct sweepoviruses. The sequences of 18 clones were obtained and found to be structurally similar to tomato leaf curl virus-satellite (ToLCV-sat, the first DNA satellite identified in association with a begomovirus), with a region with significant sequence identity to the conserved region of betasatellites, an A-rich sequence, a predicted stem–loop structure containing the nonanucleotide TAATATTAC, and a second predicted stem–loop. These sweepovirus-associated satellites join an increasing number of ToLCV-sat-like non-coding satellites identified recently. Although sharing some features with betasatellites, evidence is provided to suggest that the ToLCV-sat-like satellites are distinct from betasatellites and should be considered a separate class of satellites, for which the collective name deltasatellites is proposed. PMID:26925037

Shavenbaby Couples Patterning to Epidermal Cell Shape Control

PubMed Central

Fernandes, Isabelle; Roch, Fernando; Payre, François

2006-01-01

It is well established that developmental programs act during embryogenesis to determine animal morphogenesis. How these developmental cues produce specific cell shape during morphogenesis, however, has remained elusive. We addressed this question by studying the morphological differentiation of the Drosophila epidermis, governed by a well-known circuit of regulators leading to a stereotyped pattern of smooth cells and cells forming actin-rich extensions (trichomes). It was shown that the transcription factor Shavenbaby plays a pivotal role in the formation of trichomes and underlies all examined cases of the evolutionary diversification of their pattern. To gain insight into the mechanisms of morphological differentiation, we sought to identify shavenbaby's downstream targets. We show here that Shavenbaby controls epidermal cell shape, through the transcriptional activation of different classes of cellular effectors, directly contributing to the organization of actin filaments, regulation of the extracellular matrix, and modification of the cuticle. Individual inactivation of shavenbaby's targets produces distinct trichome defects and only their simultaneous inactivation prevent trichome formation. Our data show that shavenbaby governs an evolutionarily conserved developmental module consisting of a set of genes collectively responsible for trichome formation, shedding new light on molecular mechanisms acting during morphogenesis and the way they can influence evolution of animal forms. PMID:16933974

Different receptors binding to distinct interfaces on herpes simplex virus gD can trigger events leading to cell fusion and viral entry

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

Spear, Patricia G.; Manoj, Sharmila; Yoon, Miri

2006-01-05

One of the herpes simplex virus envelope glycoproteins, designated gD, is the principal determinant of cell recognition for viral entry. Other viral glycoproteins, gB, gH and gL, cooperate with gD to mediate the membrane fusion that is required for viral entry and cell fusion. Membrane fusion is triggered by the binding of gD to one of its receptors. These receptors belong to three different classes of cell surface molecules. This review summarizes recent findings on the structure and function of gD. The results presented indicate that gD may assume more than one conformation, one in the absence of receptor, anothermore » when gD is bound to the herpesvirus entry mediator, a member of the TNF receptor family, and a third when gD is bound to nectin-1, a cell adhesion molecule in the immunoglobulin superfamily. Finally, information and ideas are presented about a membrane-proximal region of gD that is required for membrane fusion, but not for receptor binding, and that may have a role in activating the fusogenic activity of gB, gH and gL.« less

Transformation to continuous growth of primary human T lymphocytes by human T-cell leukemia virus type I X-region genes transduced by a herpesvirus saimiri vector

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

Grassmann, R.; Dengler, C.; Mueller-Fleckenstein, I.

1989-05-01

The role of the X region of the genome of the human T-cell leukemia virus type I (HTLV-I) in the immortalization of lymphocytes has been difficult to distinguish from its role in viral replication as this region encodes at least two genes, tax and rex, required for replication and the expression of viral proteins. To determine whether the X region does encode immortalizing functions, a fragment of the HTLV-I provirus capable of expressing known X-region proteins was inserted into the genome of a transformation-defective, replication-competent Herpesvirus saimiri. Infection of fresh mitogen-activated human cord blood and thymocytes yielded immortal T-cell linesmore » that had the same phenotype (CD4{sup +}, Cd5{sup +}, HLA class II{sup +}, interleukin 2 receptor {alpha}-chain +) as lymphocytes transformed by cocultivation with HTLV-I. These experiments demonstrate that the X region encodes the functions of HTLV-I that immortalize a distinct subpopulation of human T cells. The experiments also demonstrate the utility of the H. saimiri vector for the transduction of heterologous genes into human T cells.« less

Distinct roles of KAP1, HP1 and G9a/GLP in silencing of the two-cell-specific retrotransposon MERVL in mouse ES cells

PubMed Central

2013-01-01

Background In mouse embryonic stem cells (mESCs), transcriptional silencing of numerous class I and II endogenous retroviruses (ERVs), including IAP, ETn and MMERVK10C, is dependent upon the H3K9 methyltransferase (KMTase) SETDB1/ESET and its binding partner KAP1/TRIM28. In contrast, the H3K9 KMTases G9a and GLP and HP1 proteins are dispensable for this process. Intriguingly, MERVL retroelements are actively transcribed exclusively in the two-cell (2C) embryo, but the molecular basis of silencing of these class III ERVs at later developmental stages has not been systematically addressed. Results Here, we characterized the roles of these chromatin factors in MERVL silencing in mESCs. While MMERVK10C and IAP ERVs are bound by SETDB1 and KAP1 and are induced following their deletion, MERVL ERVs show relatively low levels of SETDB1 and KAP1 binding and are upregulated exclusively following KAP1 depletion, indicating that KAP1 influences MERVL expression independent of SETDB1. In contrast to class I and class II ERVs, MERVL and MERVL LTR-driven genic transcripts are also upregulated following depletion of G9a or GLP, and G9a binds directly to these ERVs. Consistent with a direct role for H3K9me2 in MERVL repression, these elements are highly enriched for G9a-dependent H3K9me2, and catalytically active G9a is required for silencing of MERVL LTR-driven transcripts. MERVL is also derepressed in HP1α and HP1β KO ESCs. However, like KAP1, HP1α and HP1β are only modestly enriched at MERVL relative to IAP LTRs. Intriguingly, as recently shown for KAP1, RYBP, LSD1 and G9a-deficient mESCs, many genes normally expressed in the 2C embryo are also induced in HP1 KO mESCs, revealing that aberrant expression of a subset of 2C-specific genes is a common feature in each of these KO lines. Conclusions Our results indicate that G9a and GLP, which are not required for silencing of class I and II ERVs, are recruited to MERVL elements and play a direct role in silencing of these class III ERVs, dependent upon G9a catalytic activity. In contrast, induction of MERVL expression in KAP1, HP1α and HP1β KO ESCs may occur predominantly as a consequence of indirect effects, in association with activation of a subset of 2C-specific genes. PMID:23735015

Skeletal muscle satellite cells

NASA Technical Reports Server (NTRS)

Schultz, E.; McCormick, K. M.

1994-01-01

Evidence now suggests that satellite cells constitute a class of myogenic cells that differ distinctly from other embryonic myoblasts. Satellite cells arise from somites and first appear as a distinct myoblast type well before birth. Satellite cells from different muscles cannot be functionally distinguished from one another and are able to provide nuclei to all fibers without regard to phenotype. Thus, it is difficult to ascribe any significant function to establishing or stabilizing fiber type, even during regeneration. Within a muscle, satellite cells exhibit marked heterogeneity with respect to their proliferative behavior. The satellite cell population on a fiber can be partitioned into those that function as stem cells and those which are readily available for fusion. Recent studies have shown that the cells are not simply spindle shaped, but are very diverse in their morphology and have multiple branches emanating from the poles of the cells. This finding is consistent with other studies indicating that the cells have the capacity for extensive migration within, and perhaps between, muscles. Complexity of cell shape usually reflects increased cytoplasmic volume and organelles including a well developed Golgi, and is usually associated with growing postnatal muscle or muscles undergoing some form of induced adaptive change or repair. The appearance of activated satellite cells suggests some function of the cells in the adaptive process through elaboration and secretion of a product. Significant advances have been made in determining the potential secretion products that satellite cells make. The manner in which satellite cell proliferative and fusion behavior is controlled has also been studied. There seems to be little doubt that cellcell coupling is not how satellite cells and myofibers communicate. Rather satellite cell regulation is through a number of potential growth factors that arise from a number of sources. Critical to the understanding of this form of control is to determine which of the many growth factors that can alter satellite cell behavior in vitro are at work in vivo. Little work has been done to determine what controls are at work after a regeneration response has been initiated. It seems likely that, after injury, growth factors are liberated through proteolytic activity and initiate an activation process whereby cells enter into a proliferative phase. After myofibers are formed, it also seems likely that satellite cell behavior is regulated through diffusible factors arising from the fibers rather than continuous control by circulating factors.(ABSTRACT TRUNCATED AT 400 WORDS).

The Specter of Divorce: Views from Working- and Middle-Class Cohabitors

ERIC Educational Resources Information Center

Miller, Amanda J.; Sassler, Sharon; Kusi-Appouh, Dela

2011-01-01

Young Americans increasingly express apprehension about their ability to successfully manage intimate relationships. Partially in response, cohabitation has become normative over the past few decades. Little research, however, examines social class distinctions in how emerging adults perceive challenges to sustaining intimate unions. We examine…

Experimental Support for the Ecoimmunity Theory: Distinct Phenotypes of Nonlymphocytic Cells in SCID and Wild-Type Mice.

PubMed

Ochayon, David E; Baranovski, Boris M; Malkin, Peter; Schuster, Ronen; Kalay, Noa; Ben-Hamo, Rotem; Sloma, Ido; Levinson, Justin; Brazg, Jared; Efroni, Sol; Lewis, Eli C; Nevo, Uri

2016-01-01

Immune tolerance toward "self" is critical in multiple immune disorders. While there are several mechanisms to describe the involvement of immune cells in the process, the role of peripheral tissue cells in that context is not yet clear. The theory of ecoimmunity postulates that interactions between immune and tissue cells represent a predator-prey relationship. A lifelong interaction, shaped mainly during early ontogeny, leads to selection of nonimmune cell phenotypes. Normally, therefore, nonimmune cells that evolve alongside an intact immune system would be phenotypically capable of evading immune responses, and cells whose phenotype falls short of satisfying this steady state would expire under hostile immune responses. This view was supported until recently by experimental evidence showing an inferior endurance of severe combined immunodeficiency (SCID)-derived pancreatic islets when engrafted into syngeneic immune-intact wild-type (WT) mice, relative to islets from WT. Here we extend the experimental exploration of ecoimmunity by searching for the presence of the phenotypic changes suggested by the theory. Immune-related phenotypes of islets, spleen, and bone marrow immune cells were determined, as well as SCID and WT nonlymphocytic cells. Islet submass grafting was performed to depict syngeneic graft functionality. Islet cultures were examined under both resting and inflamed conditions for expression of CD40 and major histocompatibility complex (MHC) class I/II and release of interleukin-1α (IL-1α), IL-1β, IL-6, tumor necrosis factor-α (TNF-α), IL-10, and insulin. Results depict multiple pathways that appear to be related to the sculpting of nonimmune cells by immune cells; 59 SCID islet genes displayed relative expression changes compared with WT islets. SCID cells expressed lower tolerability to inflammation and higher levels of immune-related molecules, including MHC class I. Accordingly, islets exhibited a marked increase in insulin release upon immunocyte depletion, in effect resuming endocrine function that was otherwise suppressed by resident immunocytes. This work provides further support of the ecoimmunity theory and encourages subsequent studies to identify its role in the emergence and treatment of autoimmune pathologies, transplant rejection, and cancer.

Liver Cancer Cell of Origin, Molecular Class, and Effects on Patient Prognosis.

PubMed

Sia, Daniela; Villanueva, Augusto; Friedman, Scott L; Llovet, Josep M

2017-03-01

Primary liver cancer is the second leading cause of cancer-related death worldwide and therefore a major public health challenge. We review hypotheses of the cell of origin of liver tumorigenesis and clarify the classes of liver cancer based on molecular features and how they affect patient prognosis. Primary liver cancer comprises hepatocellular carcinoma (HCC), intrahepatic cholangiocarcinoma (iCCA), and other rare tumors, notably fibrolamellar carcinoma and hepatoblastoma. The molecular and clinical features of HCC versus iCCA are distinct, but these conditions have overlapping risk factors and pathways of oncogenesis. A better understanding of the cell types originating liver cancer can aid in exploring molecular mechanisms of carcinogenesis and therapeutic options. Molecular studies have identified adult hepatocytes as the cell of origin. These cells have been proposed to transform directly into HCC cells (via a sequence of genetic alterations), to dedifferentiate into hepatocyte precursor cells (which then become HCC cells that express progenitor cell markers), or to transdifferentiate into biliary-like cells (which give rise to iCCA). Alternatively, progenitor cells also give rise to HCCs and iCCAs with markers of progenitor cells. Advances in genome profiling and next-generation sequencing have led to the classification of HCCs based on molecular features and assigned them to categories such as proliferation-progenitor, proliferation-transforming growth factor β, and Wnt-catenin β1. iCCAs have been assigned to categories of proliferation and inflammation. Overall, proliferation subclasses are associated with a more aggressive phenotype and poor outcome of patients, although more specific signatures have refined our prognostic abilities. Analyses of genetic alterations have identified those that might be targeted therapeutically, such as fusions in the FGFR2 gene and mutations in genes encoding isocitrate dehydrogenases (in approximately 60% of iCCAs) or amplifications at 11q13 and 6p21 (in approximately 15% of HCCs). Further studies of these alterations are needed before they can be used as biomarkers in clinical decision making. Copyright © 2017 AGA Institute. Published by Elsevier Inc. All rights reserved.

Investigations of Potential Phenotypes of Foot Osteoarthritis: Cross‐Sectional Analysis From the Clinical Assessment Study of the Foot

PubMed Central

Marshall, Michelle; Thomas, Martin J.; Menz, Hylton B.; Myers, Helen L.; Thomas, Elaine; Downes, Thomas; Peat, George; Roddy, Edward

2016-01-01

Objective To investigate the existence of distinct foot osteoarthritis (OA) phenotypes based on pattern of joint involvement and comparative symptom and risk profiles. Methods Participants ages ≥50 years reporting foot pain in the previous year were drawn from a population‐based cohort. Radiographs were scored for OA in the first metatarsophalangeal (MTP) joint, first and second cuneometatarsal, navicular first cuneiform, and talonavicular joints according to a published atlas. Chi‐square tests established clustering, and odds ratios (ORs) examined symmetry and pairwise associations of radiographic OA in the feet. Distinct underlying classes of foot OA were investigated by latent class analysis (LCA) and their association with symptoms and risk factors was assessed. Results In 533 participants (mean age 64.9 years, 55.9% female) radiographic OA clustered across both feet (P < 0.001) and was highly symmetrical (adjusted OR 3.0, 95% confidence interval 2.1, 4.2). LCA identified 3 distinct classes of foot OA: no or minimal foot OA (64%), isolated first MTP joint OA (22%), and polyarticular foot OA (15%). After adjustment for age and sex, polyarticular foot OA was associated with nodal OA, increased body mass index, and more pain and functional limitation compared to the other classes. Conclusion Patterning of radiographic foot OA has provided insight into the existence of 2 forms of foot OA: isolated first MTP joint OA and polyarticular foot OA. The symptom and risk factor profiles in individuals with polyarticular foot OA indicate a possible distinctive phenotype of foot OA, but further research is needed to explore the characteristics of isolated first MTP joint and polyarticular foot OA. PMID:26238801

Virtual libraries of tetrapyrrole macrocycles. Combinatorics, isomers, product distributions, and data mining.

PubMed

Taniguchi, Masahiko; Du, Hai; Lindsey, Jonathan S

2011-09-26

A software program (PorphyrinViLiGe) has been developed to enumerate the type and relative amounts of substituted tetrapyrrole macrocycles in a virtual library formed by one of four different classes of reactions. The classes include (1) 4-fold reaction of n disubstituted heterocycles (e.g., pyrroles or diiminoisoindolines) to form β-substituted porphyrins, β-substituted tetraazaporphyrins, or α- or β-substituted phthalocyanines; (2) combination of m aminoketones and n diones to form m × n pyrroles, which upon 4-fold reaction give β-substituted porphyrins; (3) derivatization of an 8-point tetrapyrrole scaffold with n reagents, and (4) 4-fold reaction of n aldehydes and pyrrole to form meso-substituted porphyrins. The program accommodates variable ratios of reactants, reversible or irreversible reaction (reaction classes 1 and 2), and degenerate modes of formation. Pólya's theorem (for enumeration of cyclic entities) has also been implemented and provides validation for reaction classes 3 and 4. The output includes the number and identity of distinct reaction-accessible substituent combinations, the number and identity of isomers thereof, and the theoretical mass spectrum. Provisions for data mining enable assessment of the number of products having a chosen pattern of substituents. Examples include derivatization of an octa-substituted phthalocyanine with eight reagents to afford a library of 2,099,728 members (yet only 6435 distinct substituent combinations) and reversible reaction of six distinct disubstituted pyrroles to afford 2649 members (yet only 126 distinct substituent combinations). In general, libraries of substituted tetrapyrrole macrocycles occupy a synthetically accessible region of chemical space that is rich in isomers (>99% or 95% for the two examples, respectively).

Identification and Transcript Analysis of the TCP Transcription Factors in the Diploid Woodland Strawberry Fragaria vesca

PubMed Central

Wei, Wei; Hu, Yang; Cui, Meng-Yuan; Han, Yong-Tao; Gao, Kuan; Feng, Jia-Yue

2016-01-01

Plant-specific TEOSINTE BRANCHED 1, CYCLOIDEA, and PROLIFERATING CELL FACTORS (TCP) transcription factors play versatile functions in multiple processes of plant growth and development. However, no systematic study has been performed in strawberry. In this study, 19 FvTCP genes were identified in the diploid woodland strawberry (Fragaria vesca) accession Heilongjiang-3. Phylogenetic analysis suggested that the FvTCP genes were classified into two main classes, with the second class further divided into two subclasses, which was supported by the exon-intron organizations and the conserved motif structures. Promoter analysis revealed various cis-acting elements related to growth and development, hormone and/or stress responses. We analyzed FvTCP gene transcript accumulation patterns in different tissues and fruit developmental stages. Among them, 12 FvTCP genes exhibited distinct tissue-specific transcript accumulation patterns. Eleven FvTCP genes were down-regulated in different fruit developmental stages, while five FvTCP genes were up-regulated. Transcripts of FvTCP genes also varied with different subcultural propagation periods and were induced by hormone treatments and biotic and abiotic stresses. Subcellular localization analysis showed that six FvTCP-GFP fusion proteins showed distinct localizations in Arabidopsis mesophyll protoplasts. Notably, transient over-expression of FvTCP9 in strawberry fruits dramatically affected the expression of a series of genes implicated in fruit development and ripening. Taken together, the present study may provide the basis for functional studies to reveal the role of this gene family in strawberry growth and development. PMID:28066489

A novel fluorescence-based biosynthetic trafficking method provides pharmacologic evidence that PI4-kinase IIIα is important for protein trafficking from the endoplasmic reticulum to the plasma membrane.

PubMed

Bryant, Kirsten L; Baird, Barbara; Holowka, David

2015-02-27

Biosynthetic trafficking of receptors and other membrane-associated proteins from the endoplasmic reticulum (ER) to the plasma membrane (PM) underlies the capacity of these proteins to participate in crucial cellular roles. Phosphoinositides have been shown to mediate distinct biological functions in cells, and phosphatidylinositol 4-phosphate (PI4P), in particular, has emerged as a key regulator of biosynthetic trafficking. To investigate the source of PI4P that orchestrates trafficking events, we developed a novel flow cytometry based method to monitor biosynthetic trafficking of transiently transfected proteins. We demonstrated that our method can be used to assess the trafficking of both type-1 transmembrane and GPI-linked proteins, and that it can accurately monitor the pharmacological disruption of biosynthetic trafficking with brefeldin A, a well-documented inhibitor of early biosynthetic trafficking. Furthermore, utilizing our newly developed method, we applied pharmacological inhibition of different isoforms of PI 4-kinase to reveal a role for a distinct pool of PI4P, synthesized by PI4KIIIα, in ER-to-PM trafficking. Taken together, these findings provide evidence that a specific pool of PI4P plays a role in biosynthetic trafficking of two different classes of proteins from the ER to the Golgi complex. Furthermore, our simple, flow cytometry-based biosynthetic trafficking assay can be widely applied to the study of multiple classes of proteins and varied pharmacological and genetic perturbations.

Optimizing morphology through blood cell image analysis.

PubMed

Merino, A; Puigví, L; Boldú, L; Alférez, S; Rodellar, J

2018-05-01

Morphological review of the peripheral blood smear is still a crucial diagnostic aid as it provides relevant information related to the diagnosis and is important for selection of additional techniques. Nevertheless, the distinctive cytological characteristics of the blood cells are subjective and influenced by the reviewer's interpretation and, because of that, translating subjective morphological examination into objective parameters is a challenge. The use of digital microscopy systems has been extended in the clinical laboratories. As automatic analyzers have some limitations for abnormal or neoplastic cell detection, it is interesting to identify quantitative features through digital image analysis for morphological characteristics of different cells. Three main classes of features are used as follows: geometric, color, and texture. Geometric parameters (nucleus/cytoplasmic ratio, cellular area, nucleus perimeter, cytoplasmic profile, RBC proximity, and others) are familiar to pathologists, as they are related to the visual cell patterns. Different color spaces can be used to investigate the rich amount of information that color may offer to describe abnormal lymphoid or blast cells. Texture is related to spatial patterns of color or intensities, which can be visually detected and quantitatively represented using statistical tools. This study reviews current and new quantitative features, which can contribute to optimize morphology through blood cell digital image processing techniques. © 2018 John Wiley & Sons Ltd.

A biophysical signature of network affiliation and sensory processing in mitral cells

PubMed Central

Angelo, Kamilla; Rancz, Ede A.; Pimentel, Diogo; Hundahl, Christian; Hannibal, Jens; Fleischmann, Alexander; Pichler, Bruno; Margrie, Troy W.

2012-01-01

One defining characteristic of the mammalian brain is its neuronal diversity1. For a given region, substructure or layer and even cell type2, variability in neuronal morphology and connectivity2-5 persists. While it is well established that such cellular properties vary considerably according to neuronal type, the significant biophysical diversity of neurons of the same morphological class is typically averaged out and ignored. Here we show that the amplitude of hyperpolarization-evoked membrane potential sag recorded in olfactory bulb mitral cells is an emergent, homotypic property of local networks and sensory information processing. Simultaneous whole-cell recordings from pairs of cells reveal that the amount of hyperpolarization-evoked sag potential and current6 is stereotypic for mitral cells belonging to the same glomerular circuit. This is corroborated by a mosaic, glomerulus-based pattern of expression of the HCN2 subunit of the hyperpolarization-activated current (Ih) channel. Furthermore, inter-glomerular differences in both membrane potential sag and HCN2 protein are diminished when sensory input to glomeruli is genetically and globally altered so only one type of odorant receptor is universally expressed7. We therefore suggest that population diversity in the intrinsic profile of mitral cells reflect functional adaptations of distinct local circuits dedicated to processing subtly different odor-related information. PMID:22820253

Atrial natriuretic peptide receptor heterogeneity and effects on cyclic GMP accumulation

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

Leitman, D.C.

1988-01-01

The effects of atrial natriuretic peptide (ANP), oxytocin (OT) and vasopressin (AVP) on guanylate cyclase activity and cyclic GMP accumulation were examined, since these hormones appear to be intimately associated with blood pressure and intravascular volume homeostasis. ANP was found to increase cyclic GMP accumulation in ten cell culture systems, which were derived from blood vessels, adrenal cortex, kidney, lung, testes and mammary gland. ANP receptors were characterized in intact cultured cells using {sup 125}I-ANP{sub 8-33}. Specific {sup 125}I-ANP binding was saturable and of high affinity. Scratchard analysis of the binding data for all cell types exhibited a straight line,more » indicating that these cells possessed a single class of binding sites. Despite the presence of linear Scatchard plots, these studies demonstrated that cultured cells possess two functionally and physically distinct ANP-binding sites. Most of the ANP-binding sites in cultured cells have a molecular size of 66,000 daltons under reducing conditions. The identification of cultured cell types in which hormones (ANP and oxytocin) regulate guanylate cyclase activity and increase cyclic GMP synthesis will provide valuable systems to determine the mechanisms of hormone-receptor coupling to guanylate cyclase and the cellular processes regulated by cyclic GMP.« less

High definition infrared spectroscopic imaging for lymph node histopathology.

PubMed

Leslie, L Suzanne; Wrobel, Tomasz P; Mayerich, David; Bindra, Snehal; Emmadi, Rajyasree; Bhargava, Rohit

2015-01-01

Chemical imaging is a rapidly emerging field in which molecular information within samples can be used to predict biological function and recognize disease without the use of stains or manual identification. In Fourier transform infrared (FT-IR) spectroscopic imaging, molecular absorption contrast provides a large signal relative to noise. Due to the long mid-IR wavelengths and sub-optimal instrument design, however, pixel sizes have historically been much larger than cells. This limits both the accuracy of the technique in identifying small regions, as well as the ability to visualize single cells. Here we obtain data with micron-sized sampling using a tabletop FT-IR instrument, and demonstrate that the high-definition (HD) data lead to accurate identification of multiple cells in lymph nodes that was not previously possible. Highly accurate recognition of eight distinct classes - naïve and memory B cells, T cells, erythrocytes, connective tissue, fibrovascular network, smooth muscle, and light and dark zone activated B cells was achieved in healthy, reactive, and malignant lymph node biopsies using a random forest classifier. The results demonstrate that cells currently identifiable only through immunohistochemical stains and cumbersome manual recognition of optical microscopy images can now be distinguished to a similar level through a single IR spectroscopic image from a lymph node biopsy.

Classes and continua of hippocampal CA1 inhibitory neurons revealed by single-cell transcriptomics.

PubMed

Harris, Kenneth D; Hochgerner, Hannah; Skene, Nathan G; Magno, Lorenza; Katona, Linda; Bengtsson Gonzales, Carolina; Somogyi, Peter; Kessaris, Nicoletta; Linnarsson, Sten; Hjerling-Leffler, Jens

2018-06-18

Understanding any brain circuit will require a categorization of its constituent neurons. In hippocampal area CA1, at least 23 classes of GABAergic neuron have been proposed to date. However, this list may be incomplete; additionally, it is unclear whether discrete classes are sufficient to describe the diversity of cortical inhibitory neurons or whether continuous modes of variability are also required. We studied the transcriptomes of 3,663 CA1 inhibitory cells, revealing 10 major GABAergic groups that divided into 49 fine-scale clusters. All previously described and several novel cell classes were identified, with three previously described classes unexpectedly found to be identical. A division into discrete classes, however, was not sufficient to describe the diversity of these cells, as continuous variation also occurred between and within classes. Latent factor analysis revealed that a single continuous variable could predict the expression levels of several genes, which correlated similarly with it across multiple cell types. Analysis of the genes correlating with this variable suggested it reflects a range from metabolically highly active faster-spiking cells that proximally target pyramidal cells to slower-spiking cells targeting distal dendrites or interneurons. These results elucidate the complexity of inhibitory neurons in one of the simplest cortical structures and show that characterizing these cells requires continuous modes of variation as well as discrete cell classes.

Normal adult ramified microglia separated from other central nervous system macrophages by flow cytometric sorting: Phenotypic differences defined and direct ex vivo antigen presentation to myelin basic protein-reactive CD4{sup +} T cells compared

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

Ford, A.L.; Goodsall, A.L.; Sedgwick, J.D.

1995-05-01

Ramified microglia in the adult central nervous system (CNS) are the principal glial element up-regulating MHC class I and II expression in response to inflammatory events or neuronal damage. A proportion of these cells also express MHC class II constitutively in the normal CNS. The role of microglia as APCs for CD4{sup +} cells extravasating into the CNS remains undefined. In this study, using irradiation bone marrow chimeras in CD45-congenic rats, the phenotype CD45{sup low}CD11b/c{sup +} is shown to identify microglial cells specifically within the CNS. Highly purified populations of microglia and nonmicroglial but CNS-associated macrophages (CD45{sup high}CD11b/c{sup +}) havemore » been obtained directly from the adult CNS, by using flow cytometric sorting. Morphologically, freshly isolated microglia vs other CNS macrophages are quite distinct. Of the two populations recovered from the normal CNS, it is the minority CD45{sup high}CD11 b/c{sup +} transitional macrophage population, and not microglia, that is the effective APC for experimental autoimmune encephalomyelitis-inducing CD4{sup +} myelin basic protein (MBP)-reactive T cells. CD45{sup high}CD11b/c{sup +} CNS macrophages also stimulate MBP-reactive T cells without addition of MBP to culture suggesting presentation of endogenous Ag. This is the first study in which microglia vs other CNS macrophages have been analyzed for APC ability directly from the CNS, with substantial cross-contamination between the two populations eliminated. The heterogeneity of these populations in terms of APC function is clearly demonstrated. Evidence is still lacking that adult CNS microglia have the capacity to interact with and stimulate CD4{sup +} T cells to proliferate or secrete IL-2. 60 refs., 6 figs., 1 tab.« less

Structure and function of three novel MHC class I antigens derived from a C3H ultraviolet-induced fibrosarcoma

PubMed Central

1986-01-01

The UV-induced, C3H fibrosarcoma, 1591, expresses at least three unique MHC class I antigens not found on normal C3H tissue. Here we report the complete DNA sequence of the three novel class I genes encoding these molecules, and describe in detail the recognition of the individual products by tumor-reactive and allospecific CTL. Remarkably, although C3H does not appear to express H-2L locus information, this C3H tumor expresses two distinct antigens, termed A149 and A166, which are extremely homologous to each other and to the H-2Ld antigen from BALB/c. The gene encoding the third novel class I antigen from 1591, A216, is quite homologous to H-2Kk) throughout its 3' end. Since all three of these genes account for polymorphic restriction fragments not found in C3H, it is likely that they were derived by recombination from the endogenous class I genes of C3H. The DNA sequence homology of A149, A166, and H-2Ld is especially significant given the functional conservation observed between the products of these genes. Limited sequence substitutions appear to correlate with some of the discrete serological differences observed between these molecules. In addition, both A149 and A166 crossreact, but to differing extents, with H-2Ld at the level of T cell recognition. Our results are consistent with the view that CTL recognize complex conformational determinants on class I molecules, but extend previous observations by comparing a set of antigens with discrete and overlapping structural and functional differences. PMID:3489061

Can the dissociative PTSD subtype be identified across two distinct trauma samples meeting caseness for PTSD?

PubMed

Hansen, Maj; Műllerová, Jana; Elklit, Ask; Armour, Cherie

2016-08-01

For over a century, the occurrence of dissociative symptoms in connection to traumatic exposure has been acknowledged in the scientific literature. Recently, the importance of dissociation has also been recognized in the long-term traumatic response within the DSM-5 nomenclature. Several studies have confirmed the existence of the dissociative posttraumatic stress disorder (PTSD) subtype. However, there is a lack of studies investigating latent profiles of PTSD solely in victims with PTSD. This study investigates the possible presence of PTSD subtypes using latent class analysis (LCA) across two distinct trauma samples meeting caseness for DSM-5 PTSD based on self-reports (N = 787). Moreover, we assessed if a number of risk factors resulted in an increased probability of membership in a dissociative compared with a non-dissociative PTSD class. The results of LCA revealed a two-class solution with two highly symptomatic classes: a dissociative class and a non-dissociative class across both samples. Increased emotion-focused coping increased the probability of individuals being grouped into the dissociative class across both samples. Social support reduced the probability of individuals being grouped into the dissociative class but only in the victims of motor vehicle accidents (MVAs) suffering from whiplash. The results are discussed in light of their clinical implications and suggest that the dissociative subtype can be identified in victims of incest and victims of MVA suffering from whiplash meeting caseness for DSM-5 PTSD.

Nottingham Prognostic Index Plus (NPI+): a modern clinical decision making tool in breast cancer.

PubMed

Rakha, E A; Soria, D; Green, A R; Lemetre, C; Powe, D G; Nolan, C C; Garibaldi, J M; Ball, G; Ellis, I O

2014-04-02

Current management of breast cancer (BC) relies on risk stratification based on well-defined clinicopathologic factors. Global gene expression profiling studies have demonstrated that BC comprises distinct molecular classes with clinical relevance. In this study, we hypothesised that molecular features of BC are a key driver of tumour behaviour and when coupled with a novel and bespoke application of established clinicopathologic prognostic variables can predict both clinical outcome and relevant therapeutic options more accurately than existing methods. In the current study, a comprehensive panel of biomarkers with relevance to BC was applied to a large and well-characterised series of BC, using immunohistochemistry and different multivariate clustering techniques, to identify the key molecular classes. Subsequently, each class was further stratified using a set of well-defined prognostic clinicopathologic variables. These variables were combined in formulae to prognostically stratify different molecular classes, collectively known as the Nottingham Prognostic Index Plus (NPI+). The NPI+ was then used to predict outcome in the different molecular classes. Seven core molecular classes were identified using a selective panel of 10 biomarkers. Incorporation of clinicopathologic variables in a second-stage analysis resulted in identification of distinct prognostic groups within each molecular class (NPI+). Outcome analysis showed that using the bespoke NPI formulae for each biological BC class provides improved patient outcome stratification superior to the traditional NPI. This study provides proof-of-principle evidence for the use of NPI+ in supporting improved individualised clinical decision making.

Evolution of vertebrate visual pigments.

PubMed

Bowmaker, James K

2008-09-01

The visual pigments of vertebrates evolved about 500 million years ago, before the major evolutionary step of the development of jaws. Four spectrally distinct classes of cone opsin evolved through gene duplication, followed by the rod opsin class that arose from the duplication of the middle-wave-sensitive cone opsin. All four cone classes are present in many extant teleost fish, reptiles and birds, but one or more classes have been lost in primitive fish, amphibians and mammals. Gene duplication within the cone classes, especially in teleosts, has resulted in multiple opsins being available, both temporally and spatially, during development.

LIPID CLASS DISTRIBUTION OF HIGHLY UNSATURATED LONG CHAIN FATTY ACIDS IN MARINE DINOFLAGELLATES.

EPA Science Inventory

The very long chain highly unsaturated C28 fatty acids, octacosaheptaenoic [28:7(n-6)] and octacosaoctaenoic acid [28:8(n-3)], were found to be associated with phospholipids, obtained by fractionation of total lipid extracts into distinct lipid classes, in 4 and 6, respectively, ...

Integrating Single-System Design Research into the Clinical Practice Class

ERIC Educational Resources Information Center

Cooper, Marlene G.

2006-01-01

Clinical practice and research are generally taught separately in Master of Social Work programs by faculty with distinct areas of expertise. This paper discusses the teaching of single-subject design research methodology by clinical faculty, in the clinical practice class. Examples from student papers demonstrate the effectiveness of integrating…

Critical Analysis: A Comparison of Critical Thinking Changes in Psychology and Philosophy Classes

ERIC Educational Resources Information Center

Burke, Brian L.; Sears, Sharon R.; Kraus, Sue; Roberts-Cady, Sarah

2014-01-01

This study compared changes in psychology and philosophy classes in two distinct components of critical thinking (CT): general skills and personal beliefs. Participants were 128 undergraduates enrolled in CT in psychology, other psychology courses, or philosophy courses. CT and philosophy students significantly reduced beliefs in paranormal…

Women and Men in the Class Structure.

ERIC Educational Resources Information Center

Collins, Randall

1988-01-01

Describes the stratification position of women as generally more complex than that of men, because of class distinction of organizational power. Asserts that many women, in presenting the front-stage image of an organization, in performing considerable surplus domestic labor, and in employment and leisure activities, are involved primarily in…

The Influence of Semantic Property and Grammatical Class on Semantic Selection

ERIC Educational Resources Information Center

Yang, Fan-pei Gloria; Khodaparast, Navid; Bradley, Kailyn; Fang, Min-Chieh; Bernstein, Ari; Krawczyk, Daniel C.

2013-01-01

Research to-date has not successfully demonstrated consistent neural distinctions for different types of ambiguity or explored the effect of grammatical class on semantic selection. We conducted a relatedness judgment task using event-related functional magnetic resonance imaging (fMRI) to further explore these topics. Participants judged…

Altering Methods to Fill the English Curriculum Gap in Japan

ERIC Educational Resources Information Center

Zinck, Gerald W.

2017-01-01

In the Japanese English education system, a distinct disconnect exists between the elementary and secondary education curricula. Elementary schools across Japan offer English classes, but adjusting to junior high English classes is often difficult for students. While the Japanese government reformed junior high school tests to aid student…

Downregulation of peptide transporter genes in cell lines transformed with the highly oncogenic adenovirus 12

PubMed Central

1994-01-01

The expression of class I major histocompatibility complex antigens on the surface of cells transformed by adenovirus 12 (Ad12) is generally very low, and correlates with the high oncogenicity of this virus. In primary embryonal fibroblasts from transgenic mice that express both endogenous H-2 genes and a miniature swine class I gene (PD1), Ad12- mediated transformation results in suppression of cell surface expression of all class I antigens. Although class I mRNA levels of PD1 and H-2Db are similar to those in nonvirally transformed cells, recognition of newly synthesized class I molecules by a panel of monoclonal antibodies is impaired, presumably as a result of inefficient assembly and transport of the class I molecules. Class I expression can be partially induced by culturing cells at 26 degrees C, or by coculture of cells with class I binding peptides at 37 degrees C. Analysis of steady state mRNA levels of the TAP1 and TAP2 transporter genes for Ad12-transformed cell lines revealed that they both are significantly reduced, TAP2 by about 100-fold and TAP1 by 5-10-fold. Reconstitution of PD1 and H-2Db, but not H-2Kb, expression is achieved in an Ad12-transformed cell line by stable transfection with a TAP2, but not a TAP1, expression construct. From these data it may be concluded that suppressed expression of peptide transporter genes, especially TAP2, in Ad12-transformed cells inhibits cell surface expression of class I molecules. The failure to fully reconstitute H- 2Db and H-2Kb expression indicates that additional factors are involved in controlling class I gene expression in Ad12-transformed cells. Nevertheless, these results suggest that suppression of peptide transporter genes might be an important mechanism whereby virus- transformed cells escape immune recognition in vivo. PMID:7519239

Immunological Functions of the Membrane Proximal Region of MHC Class II Molecules

PubMed Central

Harton, Jonathan; Jin, Lei; Hahn, Amy; Drake, Jim

2016-01-01

Major histocompatibility complex (MHC) class II molecules present exogenously derived antigen peptides to CD4 T cells, driving activation of naïve T cells and supporting CD4-driven immune functions. However, MHC class II molecules are not inert protein pedestals that simply bind and present peptides. These molecules also serve as multi-functional signaling molecules delivering activation, differentiation, or death signals (or a combination of these) to B cells, macrophages, as well as MHC class II-expressing T cells and tumor cells. Although multiple proteins are known to associate with MHC class II, interaction with STING (stimulator of interferon genes) and CD79 is essential for signaling. In addition, alternative transmembrane domain pairing between class II α and β chains influences association with membrane lipid sub-domains, impacting both signaling and antigen presentation. In contrast to the membrane-distal region of the class II molecule responsible for peptide binding and T-cell receptor engagement, the membrane-proximal region (composed of the connecting peptide, transmembrane domain, and cytoplasmic tail) mediates these “non-traditional” class II functions. Here, we review the literature on the function of the membrane-proximal region of the MHC class II molecule and discuss the impact of this aspect of class II immunobiology on immune regulation and human disease. PMID:27006762

Herpes B Virus, Macacine Herpesvirus 1, Breaks Simplex Virus Tradition via Major Histocompatibility Complex Class I Expression in Cells from Human and Macaque Hosts

PubMed Central

Vasireddi, Mugdha

2012-01-01

B virus of the family Herpesviridae is endemic to rhesus macaques but results in 80% fatality in untreated humans who are zoonotically infected. Downregulation of major histocompatibility complex (MHC) class I in order to evade CD8+ T-cell activation is characteristic of most herpesviruses. Here we examined the cell surface presence and total protein expression of MHC class I molecules in B virus-infected human foreskin fibroblast cells and macaque kidney epithelial cells in culture, which are representative of foreign and natural host initial target cells of B virus. Our results show <20% downregulation of surface MHC class I molecules in either type of host cells infected with B virus, which is statistically insignificantly different from that observed in uninfected cells. We also examined the surface expression of MHC class Ib molecules, HLA-E and HLA-G, involved in NK cell inhibition. Our results showed significant upregulation of HLA-E and HLA-G in host cells infected with B virus relative to the amounts observed in other herpesvirus-infected cells. These results suggest that B virus-infected cell surfaces maintain normal levels of MHC class Ia molecules, a finding unique among simplex viruses. This is a unique divergence in immune evasion for B virus, which, unlike human simplex viruses, does not inhibit the transport of peptides for loading onto MHC class Ia molecules because B virus ICP47 lacks a transporter-associated protein binding domain. The fact that MHC class Ib molecules were significantly upregulated has additional implications for host-pathogen interactions. PMID:22973043

Evolution of the deaminase fold and multiple origins of eukaryotic editing and mutagenic nucleic acid deaminases from bacterial toxin systems

PubMed Central

Iyer, Lakshminarayan M.; Zhang, Dapeng; Rogozin, Igor B.; Aravind, L.

2011-01-01

The deaminase-like fold includes, in addition to nucleic acid/nucleotide deaminases, several catalytic domains such as the JAB domain, and others involved in nucleotide and ADP-ribose metabolism. Using sensitive sequence and structural comparison methods, we develop a comprehensive natural classification of the deaminase-like fold and show that its ancestral version was likely to operate on nucleotides or nucleic acids. Consequently, we present evidence that a specific group of JAB domains are likely to possess a DNA repair function, distinct from the previously known deubiquitinating peptidase activity. We also identified numerous previously unknown clades of nucleic acid deaminases. Using inference based on contextual information, we suggest that most of these clades are toxin domains of two distinct classes of bacterial toxin systems, namely polymorphic toxins implicated in bacterial interstrain competition and those that target distantly related cells. Genome context information suggests that these toxins might be delivered via diverse secretory systems, such as Type V, Type VI, PVC and a novel PrsW-like intramembrane peptidase-dependent mechanism. We propose that certain deaminase toxins might be deployed by diverse extracellular and intracellular pathogens as also endosymbionts as effectors targeting nucleic acids of host cells. Our analysis suggests that these toxin deaminases have been acquired by eukaryotes on several independent occasions and recruited as organellar or nucleo-cytoplasmic RNA modifiers, operating on tRNAs, mRNAs and short non-coding RNAs, and also as mutators of hyper-variable genes, viruses and selfish elements. This scenario potentially explains the origin of mutagenic AID/APOBEC-like deaminases, including novel versions from Caenorhabditis, Nematostella and diverse algae and a large class of fast-evolving fungal deaminases. These observations greatly expand the distribution of possible unidentified mutagenic processes catalyzed by nucleic acid deaminases. PMID:21890906

CD34 Antigen and the MPL Receptor Expression Defines a Novel Class of Human Cord Blood-Derived Primitive Hematopoietic Stem Cells.

PubMed

Matsuoka, Yoshikazu; Takahashi, Masaya; Sumide, Keisuke; Kawamura, Hiroshi; Nakatsuka, Ryusuke; Fujioka, Tatsuya; Sonoda, Yoshiaki

2017-06-09

In the murine hematopoietic stem cell (HSC) compartment, thrombopoietin (THPO)/MPL (THPO receptor) signaling plays an important role in the maintenance of adult quiescent HSCs. However, the role of THPO/MPL signaling in the human primitive HSC compartment has not yet been elucidated. We have identified very primitive human cord blood (CB)-derived CD34- severe combined immunodeficiency (SCID)-repopulating cells (SRCs) using the intra-bone marrow injection method. In this study, we investigated the roles of the MPL expression in the human primitive HSC compartment. The SRC activities of the highly purified CB-derived 18Lin-CD34+/-MPL+/- cells were analyzed using NOG mice. In the primary recipient mice, nearly all mice that received CD34+/-MPL+/- cells were repopulated with human CD45+ cells. Nearly all of these mice that received CD34+MPL+/- and CD34-MPL- cells showed a secondary repopulation. Interestingly, the secondary recipient mice that received CD34+/-MPL- cells showed a distinct tertiary repopulation. These results clearly indicate that the CD34+/- SRCs not expressing MPL sustain a long-term (LT) (>1 year) human cell repopulation in NOG mice. Moreover, CD34- SRCs generate CD34+CD38-CD90+ SRCs in vitro and in vivo. These findings provide a new concept that CD34-MPL- SRCs reside at the apex of the human HSC hierarchy.

Signatures of cytoplasmic proteins in the exoproteome distinguish community- and hospital-associated methicillin-resistant Staphylococcus aureus USA300 lineages.

PubMed

Mekonnen, Solomon A; Palma Medina, Laura M; Glasner, Corinna; Tsompanidou, Eleni; de Jong, Anne; Grasso, Stefano; Schaffer, Marc; Mäder, Ulrike; Larsen, Anders R; Gumpert, Heidi; Westh, Henrik; Völker, Uwe; Otto, Andreas; Becher, Dörte; van Dijl, Jan Maarten

2017-08-18

Methicillin-resistant Staphylococcus aureus (MRSA) is the common name for a heterogeneous group of highly drug-resistant staphylococci. Two major MRSA classes are distinguished based on epidemiology, namely community-associated (CA) and hospital-associated (HA) MRSA. Notably, the distinction of CA- and HA-MRSA based on molecular traits remains difficult due to the high genomic plasticity of S. aureus. Here we sought to pinpoint global distinguishing features of CA- and HA-MRSA through a comparative genome and proteome analysis of the notorious MRSA lineage USA300. We show for the first time that CA- and HA-MRSA isolates can be distinguished by 2 distinct extracellular protein abundance clusters that are predictive not only for epidemiologic behavior, but also for their growth and survival within epithelial cells. This 'exoproteome profiling' also groups more distantly related HA-MRSA isolates into the HA exoproteome cluster. Comparative genome analysis suggests that these distinctive features of CA- and HA-MRSA isolates relate predominantly to the accessory genome. Intriguingly, the identified exoproteome clusters differ in the relative abundance of typical cytoplasmic proteins, suggesting that signatures of cytoplasmic proteins in the exoproteome represent a new distinguishing feature of CA- and HA-MRSA. Our comparative genome and proteome analysis focuses attention on potentially distinctive roles of 'liberated' cytoplasmic proteins in the epidemiology and intracellular survival of CA- and HA-MRSA isolates. Such extracellular cytoplasmic proteins were recently invoked in staphylococcal virulence, but their implication in the epidemiology of MRSA is unprecedented.

Positioning Food Cultures: ‘Alternative’ Food as Distinctive Consumer Practice

PubMed Central

Paddock, Jessica

2015-01-01

Many sociological studies to date have explored the role of food in marking distinctions between groups. Less well understood is how ‘alternative’ means of food consumption become figured in such relations. Drawing on accounts of food practice derived from 20 in-depth interviews and a two-year period of participant observation, this article considers the role of class culture in the practice of alternative food consumption. As participants speak their position, expressions of class arise through discussions of food practice. Having explored how food plays a part in marking boundaries of distinction between foods ‘for us’ and ‘for them’, we are reminded that in reproducing certain ideas about proper eating, we confine our imagining of alternative food futures to a limited politics of the possible. The article highlights implications for future development of equitable alternatives to conventional foodways. PMID:28018005

Major histocompatibility complex class II molecule expression on muscle cells is regulated by differentiation: implications for the immunopathogenesis of muscle autoimmune diseases.

PubMed

Mantegazza, R; Gebbia, M; Mora, M; Barresi, R; Bernasconi, P; Baggi, F; Cornelio, F

1996-08-01

Major histocompatibility complex (MHC) class II molecules are expressed on myoblasts after interferon-gamma (IFN-gamma) treatment, suggesting a muscle cell involvement in antigen presentation in inflammatory myopathies. However, they were not observed on normal or pathological myofibers. This discrepancy might be related to different responsiveness of developmentally differentiated muscle cells to IFN-gamma. Myoblasts expressed class II transcripts and proteins after IFN-gamma, while myotubes and innervated contracting muscle cells did not show staining for class II molecules. At all cell stages no loss of IFN-gamma receptor was detected indicating that myofiber maturation blocks their capacity to express MHC class II molecules. This suggests that completely differentiated myofibers cannot participate in class II restricted immunological reactions.

Reducing juvenile recidivism with cognitive training and a cell phone follow-up: an evaluation of the realvictory program.

PubMed

Burraston, Bert O; Cherrington, David J; Bahr, Stephen J

2012-02-01

The purpose of this research was to evaluate the effects of a cognitive training and cell phone intervention on the recidivism of 70 juvenile offenders. Median days to rearrest were 106 for the control group, 191 for the class-only group, and 278 for the class plus cell phone group. Using rearrest as the survival criterion, the survival ratios of the class-only and class plus cell phone groups were 2.64 and 2.94 times longer than the control group, respectively. After controlling for gender, prior arrests, and risk score, the Poisson regression indicated that the class-only and class plus cell phone groups were 51% lower in total arrests than the control group. These results suggest that cognitive training supplemented with a cell phone coach is an effective and cost-efficient intervention for reducing recidivism.

Mutant Kras copy number defines metabolic reprogramming and therapeutic susceptibilities

PubMed Central

Kerr, Emma; Gaude, Edoardo; Turrell, Frances; Frezza, Christian; Martins, Carla P

2016-01-01

Summary The RAS/MAPK-signalling pathway is frequently deregulated in non-small cell lung cancer (NSCLC), often through KRAS activating mutations1-3. A single endogenous mutant Kras allele is sufficient to promote lung tumour formation in mice but malignant progression requires additional genetic alterations4-7. We recently showed that advanced lung tumours from KrasG12D/+;p53-null mice frequently exhibit KrasG12D allelic enrichment (KrasG12D/Kraswild-type>1)7, implying that mutant Kras copy gains are positively selected during progression. Through a comprehensive analysis of mutant Kras homozygous and heterozygous MEFs and lung cancer cells we now show that these genotypes are phenotypically distinct. In particular, KrasG12D/G12D cells exhibit a glycolytic switch coupled to increased channelling of glucose-derived metabolites into the TCA cycle and glutathione biosynthesis, resulting in enhanced glutathione-mediated detoxification. This metabolic rewiring is recapitulated in mutant KRAS homozygous NSCLC cells and in vivo, in spontaneous advanced murine lung tumours (which display a high frequency of KrasG12D copy gain), but not in the corresponding early tumours (KrasG12D heterozygous). Finally, we demonstrate that mutant Kras copy gain creates unique metabolic dependences that can be exploited to selectively target these aggressive mutant Kras tumours. Our data demonstrate that mutant Kras lung tumours are not a single disease but rather a heterogeneous group comprised of two classes of tumours with distinct metabolic profiles, prognosis and therapeutic susceptibility, which can be discriminated based on their relative mutant allelic content. We also provide the first in vivo evidence of metabolic rewiring during lung cancer malignant progression. PMID:26909577

Tissue-resident natural killer (NK) cells are cell lineages distinct from thymic and conventional splenic NK cells

PubMed Central

Sojka, Dorothy K; Plougastel-Douglas, Beatrice; Yang, Liping; Pak-Wittel, Melissa A; Artyomov, Maxim N; Ivanova, Yulia; Zhong, Chao; Chase, Julie M; Rothman, Paul B; Yu, Jenny; Riley, Joan K; Zhu, Jinfang; Tian, Zhigang; Yokoyama, Wayne M

2014-01-01

Natural killer (NK) cells belong to the innate immune system; they can control virus infections and developing tumors by cytotoxicity and producing inflammatory cytokines. Most studies of mouse NK cells, however, have focused on conventional NK (cNK) cells in the spleen. Recently, we described two populations of liver NK cells, tissue-resident NK (trNK) cells and those resembling splenic cNK cells. However, their lineage relationship was unclear; trNK cells could be developing cNK cells, related to thymic NK cells, or a lineage distinct from both cNK and thymic NK cells. Herein we used detailed transcriptomic, flow cytometric, and functional analysis and transcription factor-deficient mice to determine that liver trNK cells form a distinct lineage from cNK and thymic NK cells. Taken together with analysis of trNK cells in other tissues, there are at least four distinct lineages of NK cells: cNK, thymic, liver (and skin) trNK, and uterine trNK cells. DOI: http://dx.doi.org/10.7554/eLife.01659.001 PMID:24714492

Epidermotropic presentation by splenic B-cell lymphoma: The importance of clinical-pathologic correlation.

PubMed

Hedayat, Amin A; Carter, Joi B; Lansigan, Frederick; LeBlanc, Robert E

2018-04-01

There are exceedingly rare reports of patients with epidermotropic B-cell lymphomas. A subset presented with intermittent, variably pruritic papular eruptions and involvement of their spleens, peripheral blood and bone marrow at the time of diagnosis. Furthermore, some experienced an indolent course despite dissemination of their lymphomas. We report a 66-year-old woman with a 12-year history of intermittent eruptions of non-pruritic, salmon-colored papules on her torso and proximal extremities that occurred in winter and resolved with outdoor activity in spring. Skin biopsy revealed an epidermotropic B-cell lymphoma with a non-specific B-cell phenotype and heavy chain class switching with IgG expression. On workup, our patient exhibited mild splenomegaly and low-level involvement of her peripheral blood and bone marrow by a kappa-restricted B-cell population. A splenic B-cell lymphoma was diagnosed. Considering her longstanding history and absences of cytopenias, our patient has been followed without splenectomy or systemic therapy. Furthermore, the papules have responded dramatically to narrowband UVB. Our case and a review of similar rare reports aim to raise awareness among dermatopathologists and dermatologists of a clinically distinct and indolent subset of epidermotropic splenic lymphomas with characteristic clinical and histologic findings. © 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Heterogeneity of cell firing properties and opioid sensitivity in the thalamic reticular nucleus.

PubMed

Brunton, J; Charpak, S

1997-05-01

The thalamic reticular nucleus receives afferents from the dorsal thalamus, cortex and brainstem, and projects back onto most cortically projecting thalamic nuclei thus playing a key role in the synchronization of the thalamocortical network. Although this nucleus was initially thought to consist of a homogeneous population of cells using GABA as a transmitter, and sharing identical intrinsic membrane properties, some heterogeneity was subsequently reported. The morphological diversity is generally acknowledged, but only two studies have shown functional differences between two classes of cells which vary in their ability to discharge in bursts. However, the location of the non-bursting cells was not characterized with anatomical techniques. Our recent work on the action of mu-opioid agonists in the thalamus revealed a widespread K+-mediated inhibition of most, if not all, thalamic relay and diffuse projection neurons. However, in the reticular nucleus, preliminary experiments suggested that the opioid sensitivity was variable. Based on these results and on observations of a discrete localization of mu-opioid receptors in the reticular nucleus, we investigated cellular heterogeneity within the nucleus using opioid agonists as markers. Using the whole cell patch clamp technique in young rat thalamic slices, we tested the responses of 28 neurons to opioids, the intrinsic membrane properties of each cell, and their relative location within the nucleus. Two types of intrinsic membrane properties underlying distinct discharge behaviours were seen in neurobiotin-labelled cells clearly located in the reticular nucleus: type I with the typical bursting behaviour previously reported in reticularis neurons, and type II in which bursting was greatly reduced or absent. Each class of cell could be further divided into subpopulations based on their opioid sensitivity. About half of both bursting (20) and non-bursting or tonic (8) cells were strongly inhibited by the mu-opioid receptor agonist D-Ala2,N-Me-Phe4,glycinol5-enkephalin, an effect mediated by an increase in K+ conductance. At no time was inhibition by delta- or kappa-receptor agonists seen. Our work therefore further demonstrates that the reticular nucleus is functionally heterogeneous, although the role of such cell diversity has still to be determined.

Organization of the serotonergic system in the central nervous system of two basal actinopterygian fishes: the Cladistians Polypterus senegalus and Erpetoichthys calabaricus.

PubMed

López, Jesús M; González, Agustín

2014-01-01

Cladistians (Polypteriformes) are currently considered basal to other living ray-finned fishes (actinopterygians), and their brain organization is therefore critical to providing information about the primitive neural characters that existed in the earliest ray-finned fishes. The organization of the serotonergic system in the brain has been carefully analyzed in most vertebrate groups, and in the present study we provide the first detailed information on the distribution of serotonergic cell bodies and fibers in the central nervous system of representative species of the two extant genera of cladistians, i.e. Polypterus senegalus and Erpetoichthys calabaricus, by means of immunohistochemistry against serotonin (5-HT). Distinct groups of immunoreactive cells were detected in the preoptic area, the hypothalamic paraventricular organ, the pineal organ, the pretectal region, the long column of the raphe in the rhombencephalic midline, the spinal cord, and amacrine cells in the inner nuclear layer of the retina. Fiber labeling was widely distributed in all main brain subdivisions but was more abundant in distinct pallial and subpallial areas, the preoptic area, the thalamus, the optic tectum, the tori semicircularis and lateralis, the rhombencephalic reticular formation, the nucleus of the solitary tract, and the dorsal aspect of the spinal cord. Our analysis makes it possible to establish which serotonergic structures characterized the earliest ray-finned fishes, and a comparison of these results with those from other classes of vertebrates, including a segmental analysis to correlate cell populations, reveals that most characteristics, such as the presence of serotonergic cells in the preoptic area and the basal hypothalamus, are preserved in all anamniotes. However, this system seems to be reduced in amniotes, mainly mammals, although important features are shared, such as the presence of serotonergic cells in the pineal organ, the retina, and the raphe nuclei.

Dual MAPK inhibition is an effective therapeutic strategy for a subset of class II BRAF mutant melanoma.

PubMed

Dankner, Matthew; Lajoie, Mathieu; Moldoveanu, Dan; Nguyen, Tan-Trieu; Savage, Paul; Rajkumar, Shivshankari; Huang, Xiu; Lvova, Maria; Protopopov, Alexei; Vuzman, Dana; Hogg, David; Park, Morag; Guiot, Marie-Christine; Petrecca, Kevin; Mihalcioiu, Catalin; Watson, Ian R; Siegel, Peter M; Rose, April A N

2018-06-14

Dual MAPK pathway inhibition (dMAPKi) with BRAF and MEK inhibitors improves survival in BRAF V600E/K mutant melanoma, but the efficacy of dMAPKi in non-V600 BRAF mutant tumors is poorly understood. We sought to characterize the responsiveness of class II (enhanced kinase activity, dimerization dependent) BRAF mutant melanoma to dMAPKi. Tumors from patients with BRAF WT, V600E (class I) and L597S (class II) metastatic melanoma were used to generate patient-derived-xenografts (PDX). We assembled a panel of melanoma cell lines with class IIa (activation segment) or IIb (p-loop) mutations and compared these to wild-type or V600E/K BRAF mutant cells. Cell lines and PDXs were treated with BRAFi (vemurafenib, dabrafenib, encorafenib, LY3009120), MEKi (cobimetinib, trametinib, binimetinib) or the combination. We identified two patients with BRAF L597S metastatic melanoma who were treated with dMAPKi. BRAFi impaired MAPK signalling and cell growth in class I and II BRAF mutant cells. dMAPKi was more effective than either single MAPKi at inhibiting cell growth in all class II BRAF mutant cells tested. dMAPKi caused tumor regression in two melanoma PDXs with class II BRAF mutations, and prolonged survival of mice with class II BRAF mutant melanoma brain metastases. Two patients with BRAF L597S mutant melanoma clinically responded to dMAPKi. Class II BRAF mutant melanoma are growth inhibited by dMAPKi. Responses to dMAPKi have been observed in two patients with class II BRAF mutant melanoma. This data provides rationale for clinical investigation of dMAPKi in patients with class II BRAF mutant metastatic melanoma. Copyright ©2018, American Association for Cancer Research.

Marital Happiness and Psychological Well-Being across the Life Course

ERIC Educational Resources Information Center

Kamp Dush, Claire M.; Taylor, Miles G.; Kroeger, Rhiannon A.

2008-01-01

Using data from six waves of the Study of Marital Instability over the Life Course (N = 1,998), we conducted a latent class analysis to test for distinct marital happiness trajectories. We found three distinct marital happiness trajectories: low, middle, and high happiness. Initial levels of life happiness were strongly associated with membership…

A novel class of pyranocoumarin anti-androgen receptor signaling compounds.

PubMed

Guo, Junming; Jiang, Cheng; Wang, Zhe; Lee, Hyo-Jeong; Hu, Hongbo; Malewicz, Barbara; Lee, Hyo-Jung; Lee, Jae-Ho; Baek, Nam-In; Jeong, Jin-Hyun; Kim, Dae-Keun; Kang, Kyung-Sun; Kim, Sung-Hoon; Lu, Junxuan

2007-03-01

Androgen and the androgen receptor (AR)-mediated signaling are crucial for prostate cancer development. Novel agents that can inhibit AR signaling in ligand-dependent and ligand-independent manners are desirable for the chemoprevention of prostate carcinogenesis and for the treatment of advanced prostate cancer. We have shown recently that the pyranocoumarin compound decursin from the herb Angelica gigas possesses potent anti-AR activities distinct from the anti-androgen bicalutamide. Here, we compared the anti-AR activities and the cell cycle arrest and apoptotic effects of decursin and two natural analogues in the androgen-dependent LNCaP human prostate cancer cell culture model to identify structure-activity relationships and mechanisms. Decursin and its isomer decursinol angelate decreased prostate-specific antigen expression with IC(50) of approximately 1 mumol/L. Both inhibited the androgen-stimulated AR nuclear translocation and transactivation, decreased AR protein abundance through proteasomal degradation, and induced G(0/1) arrest and morphologic differentiation. They also induced caspase-mediated apoptosis and reactive oxygen species at higher concentrations. Furthermore, they lacked the agonist activity of bicalutamide in the absence of androgen and were more potent than bicalutamide for suppressing androgen-stimulated cell growth. Decursinol, which does not contain a side chain, lacked the reactive oxygen species induction and apoptotic activities and exerted paradoxically an inhibitory and a stimulatory effect on AR signaling and cell growth. In conclusion, decursin and decursinol angelate are members of a novel class of nonsteroidal compounds that exert a long-lasting inhibition of both ligand-dependent and ligand-independent AR signaling. The side chain is critical for sustaining the anti-AR activities and the growth arrest and apoptotic effects.

Patient factors and quality of life outcomes differ among four subgroups of oncology patients based on symptom occurrence.

PubMed

Astrup, Guro Lindviksmoen; Hofsø, Kristin; Bjordal, Kristin; Guren, Marianne Grønlie; Vistad, Ingvild; Cooper, Bruce; Miaskowski, Christine; Rustøen, Tone

2017-03-01

Reviews of the literature on symptoms in oncology patients undergoing curative treatment, as well as patients receiving palliative care, suggest that they experience multiple, co-occurring symptoms and side effects. The purposes of this study were to determine if subgroups of oncology patients could be identified based on symptom occurrence rates and if these subgroups differed on a number of demographic and clinical characteristics, as well as on quality of life (QoL) outcomes. Latent class analysis (LCA) was used to identify subgroups (i.e. latent classes) of patients with distinct symptom experiences based on the occurrence rates for the 13 most common symptoms from the Memorial Symptom Assessment Scale. In total, 534 patients with breast, head and neck, colorectal, or ovarian cancer participated. Four latent classes of patients were identified based on probability of symptom occurrence: all low class [i.e. low probability for all symptoms (n = 152)], all high class (n = 149), high psychological class (n = 121), and low psychological class (n = 112). Patients in the all high class were significantly younger compared with patients in the all low class. Furthermore, compared to the other three classes, patients in the all high class had lower functional status and higher comorbidity scores, and reported poorer QoL scores. Patients in the high and low psychological classes had a moderate probability of reporting physical symptoms. Patients in the low psychological class reported a higher number of symptoms, a lower functional status, and poorer physical and total QoL scores. Distinct subgroups of oncology patients can be identified based on symptom occurrence rates. Patient characteristics that are associated with these subgroups can be used to identify patients who are at greater risk for multiple co-occurring symptoms and diminished QoL, so that these patients can be offered appropriate symptom management interventions.

The Latent Classes of Subclinical ADHD Symptoms: Convergences of Multiple Informant Reports

ERIC Educational Resources Information Center

Kobor, Andrea; Takacs, Adam; Urban, Robert; Csepe, Valeria

2012-01-01

The purpose of the present study was to conduct latent class analysis on the Hyperactivity scale of the Strengths and Difficulties Questionnaire in order to identify distinct subgroups of subclinical ADHD in a multi-informant framework. We hypothesized a similar structure between teachers and parents, and differences in symptom severity across…

The Role of Internationalisation in the Schooling of Brazilian Elites: Distinctions between Two Class Fractions

ERIC Educational Resources Information Center

Windle, Joel; Nogueira, Maria Alice

2015-01-01

This paper analyses tendencies that distinguish the internationalisation of education for two class fractions--owners of medium to large businesses and highly qualified university professors and researchers. We identify the importance of cosmopolitan cultural capital, particularly fluency in English, in strengthening the position of both groups…

Contextual Control by Function and Form of Transfer of Functions

ERIC Educational Resources Information Center

Perkins, David R.; Dougher, Michael J.; Greenway, David E.

2007-01-01

This study investigated conditions leading to contextual control by stimulus topography over transfer of functions. Three 4-member stimulus equivalence classes, each consisting of four (A, B, C, D) topographically distinct visual stimuli, were established for 5 college students. Across classes, designated A stimuli were open-ended linear figures,…

"I Can't Do Any More Education": Class, Individualisation and Educational Decision-Making

ERIC Educational Resources Information Center

Morrison, Andrew

2008-01-01

This paper, drawing upon a recent study of youth post-compulsory educational and occupational decision-making, argues for a culturalist perspective to understand the persistence of class-based inequalities within VET. The paper begins by outlining two broadly distinct perspectives within current research into youth: an "individualist" approach…

Informing the Discussion on Evaluator Training: A Look at Evaluators' Course Taking and Professional Practice

ERIC Educational Resources Information Center

Christie, Christina A.; Quiñones, Patricia; Fierro, Leslie

2014-01-01

This classification study examines evaluators' coursework training as a way of understanding evaluation practice. Data regarding courses that span methods and evaluation topics were collected from evaluation practitioners. Using latent class analysis, we establish four distinct classes of evaluator course-taking patterns: quantitative,…

A Comparison of Classification Approaches for Cyberbullying and Traditional Bullying Using Data from Six European Countries

ERIC Educational Resources Information Center

Schultze-Krumbholz, Anja; Göbel, Kristin; Scheithauer, Herbert; Brighi, Antonella; Guarini, Annalisa; Tsorbatzoudis, Haralambos; Barkoukis, Vassilis; Pyzalski, Jacek; Plichta, Piotr; Del Rey, Rosario; Casas, José A.; Thompson, Fran; Smith, Peter K.

2015-01-01

In recently published studies on cyberbullying, students are frequently categorized into distinct (cyber)bully and (cyber)victim clusters based on theoretical assumptions and arbitrary cut-off scores adapted from traditional bullying research. The present study identified involvement classes empirically using latent class analysis (LCA), to…

Ribosomally synthesized and post-translationally modified peptide natural products: overview and recommendations for a universal nomenclature

PubMed Central

Arnison, Paul G.; Bibb, Mervyn J.; Bierbaum, Gabriele; Bowers, Albert A.; Bugni, Tim S.; Bulaj, Grzegorz; Camarero, Julio A.; Campopiano, Dominic J.; Challis, Gregory L.; Clardy, Jon; Cotter, Paul D.; Craik, David J.; Dawson, Michael; Dittmann, Elke; Donadio, Stefano; Dorrestein, Pieter C.; Entian, Karl-Dieter; Fischbach, Michael A.; Garavelli, John S.; Göransson, Ulf; Gruber, Christian W.; Haft, Daniel H.; Hemscheidt, Thomas K.; Hertweck, Christian; Hill, Colin; Horswill, Alexander R.; Jaspars, Marcel; Kelly, Wendy L.; Klinman, Judith P.; Kuipers, Oscar P.; Link, A. James; Liu, Wen; Marahiel, Mohamed A.; Mitchell, Douglas A.; Moll, Gert N.; Moore, Bradley S.; Müller, Rolf; Nair, Satish K.; Nes, Ingolf F.; Norris, Gillian E.; Olivera, Baldomero M.; Onaka, Hiroyasu; Patchett, Mark L.; Piel, Joern; Reaney, Martin J. T.; Rebuffat, Sylvie; Ross, R. Paul; Sahl, Hans-Georg; Schmidt, Eric W.; Selsted, Michael E.; Severinov, Konstantin; Shen, Ben; Sivonen, Kaarina; Smith, Leif; Stein, Torsten; Süssmuth, Roderich D.; Tagg, John R.; Tang, Gong-Li; Truman, Andrew W.; Vederas, John C.; Walsh, Christopher T.; Walton, Jonathan D.; Wenzel, Silke C.; Willey, Joanne M.; van der Donk, Wilfred A.

2014-01-01

This review presents recommended nomenclature for the biosynthesis of ribosomally synthesized and post-translationally modified peptides (RiPPs), a rapidly growing class of natural products. The current knowledge regarding the biosynthesis of the >20 distinct compound classes is also reviewed, and commonalities are discussed. PMID:23165928

Value Preferences Associated With Social Class, Sex, and Race

ERIC Educational Resources Information Center

Malpass, Roy S.; Symonds, John D.

1974-01-01

Preferences for 92 values were measured for 10 groups of black and white males and females of lower and middle class status in two geographically distinct settings in the United States. Factor analysis showed five shared value composites: (1) the good life, (2) pleasant working companions, (3) balance and adjustment; (4) artistic creativity, and…

Co-Ethnic Network, Social Class, and Heritage Language Maintenance among Chinese Immigrant Families

ERIC Educational Resources Information Center

Zhang, Donghui

2012-01-01

This ethnographic study investigated heritage language maintenance among two distinct groups of Chinese immigrant families (Mandarin and Fujianese) from the social network perspective. The results indicated that a co-ethnic network could be a double-edged sword, which works differently on children from different social classes. While the Mandarin…