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Sample records for class switch recombination

  1. Epigenetic Codes Programing Class Switch Recombination

    PubMed Central

    Vaidyanathan, Bharat; Chaudhuri, Jayanta

    2015-01-01

    Class switch recombination imparts B cells with a fitness-associated adaptive ­advantage during a humoral immune response by using a precision-tailored DNA excision and ligation process to swap the default constant region gene of the antibody with a new one that has unique effector functions. This secondary diversification of the antibody repertoire is a hallmark of the adaptability of B cells when confronted with environmental and pathogenic challenges. Given that the nucleotide sequence of genes during class switching remains unchanged (genetic constraints), it is logical and necessary therefore, to integrate the adaptability of B cells to an epigenetic state, which is dynamic and can be heritably modulated before, after, or even during an antibody-dependent immune response. Epigenetic regulation encompasses heritable changes that affect function (phenotype) without altering the sequence information embedded in a gene, and include histone, DNA and RNA modifications. Here, we review current literature on how B cells use an epigenetic code language as a means to ensure antibody plasticity in light of pathogenic insults. PMID:26441954

  2. Class-switch recombination: after the dawn of AID

    PubMed Central

    Kenter, Amy L

    2015-01-01

    Isotype class switching is central to the humoral immune response. The discovery that mutations in the activation-induced deaminase (AID) gene inhibit class-switch recombination, somatic hypermutation and gene conversion is a major step forward in defining the underlying mechanisms of these gene modification events. The propensity of mutations to occur at dC/dG nucleotides during somatic hypermutation and the homology between AID and cytidine deaminase has resulted in studies demonstrating that AID has the properties of a cytidine-specific mutator and also that elements of the base-excision repair pathway play a central role in class switching and hypermutation. AID is not a promiscuous mutator in the B cell, suggesting that there are specific molecular targeting mechanisms that regulate the accessibility of DNA to AID and differentially regulate class-switch recombination and somatic hypermutation. During class switching, isotype-specific targeting occurs independently of AID and provides another level of specificity to this recombination event. PMID:12633669

  3. IgH chain class switch recombination: mechanism and regulation.

    PubMed

    Stavnezer, Janet; Schrader, Carol E

    2014-12-01

    IgH class switching occurs rapidly after activation of mature naive B cells, resulting in a switch from expression of IgM and IgD to expression of IgG, IgE, or IgA; this switch improves the ability of Abs to remove the pathogen that induces the humoral immune response. Class switching occurs by a deletional recombination between two switch regions, each of which is associated with a H chain constant region gene. Class switch recombination (CSR) is instigated by activation-induced cytidine deaminase, which converts cytosines in switch regions to uracils. The uracils are subsequently removed by two DNA-repair pathways, resulting in mutations, single-strand DNA breaks, and the double-strand breaks required for CSR. We discuss several aspects of CSR, including how CSR is induced, CSR in B cell progenitors, the roles of transcription and chromosomal looping in CSR, and the roles of certain DNA-repair enzymes in CSR. PMID:25411432

  4. Directed Ig class switch recombination in activated murine B cells.

    PubMed Central

    Winter, E; Krawinkel, U; Radbruch, A

    1987-01-01

    Immunoglobulin class switch recombination occurs at frequencies of up to 10%/cell/generation in activated murine B-lymphocytes. We analysed cH gene rearrangements and switch recombinations from active and inactive IgH loci of B-cells activated in various ways and immortalized by cell fusion. Although about half of the IgM+ cells show rearrangement of c mu genes, the deletion of c mu is a rare event. Half of the IgG3+ and IgG1+ cells show rearrangement of c mu genes on the inactive IgH locus and the other half of the IgG+ cells have deleted c mu from both IgH loci by switch recombination. This recombination is directed to the same switch regions on both IgH loci in 60-80% of all cases. Interleukin 4 may play a critical role in programming murine B-lymphocytes for specific switch recombination. Images Fig. 1. Fig. 2. Fig. 6. PMID:3038529

  5. Parp3 negatively regulates immunoglobulin class switch recombination.

    PubMed

    Robert, Isabelle; Gaudot, Léa; Rogier, Mélanie; Heyer, Vincent; Noll, Aurélia; Dantzer, Françoise; Reina-San-Martin, Bernardo

    2015-05-01

    To generate highly specific and adapted immune responses, B cells diversify their antibody repertoire through mechanisms involving the generation of programmed DNA damage. Somatic hypermutation (SHM) and class switch recombination (CSR) are initiated by the recruitment of activation-induced cytidine deaminase (AID) to immunoglobulin loci and by the subsequent generation of DNA lesions, which are differentially processed to mutations during SHM or to double-stranded DNA break intermediates during CSR. The latter activate the DNA damage response and mobilize multiple DNA repair factors, including Parp1 and Parp2, to promote DNA repair and long-range recombination. We examined the contribution of Parp3 in CSR and SHM. We find that deficiency in Parp3 results in enhanced CSR, while SHM remains unaffected. Mechanistically, this is due to increased occupancy of AID at the donor (Sμ) switch region. We also find evidence of increased levels of DNA damage at switch region junctions and a bias towards alternative end joining in the absence of Parp3. We propose that Parp3 plays a CSR-specific role by controlling AID levels at switch regions during CSR. PMID:26000965

  6. The histone methyltransferase MMSET regulates class switch recombination.

    PubMed

    Pei, Huadong; Wu, Xiaosheng; Liu, Tongzheng; Yu, Kefei; Jelinek, Diane F; Lou, Zhenkun

    2013-01-15

    Wolf-Hirschhorn syndrome (WHS) is a genetic disease with characteristic facial features and developmental disorders. Of interest, loss of the MMSET gene (also known as WHSC1) is considered to be responsible for the core phenotypes of this disease. Patients with WHS also display Ab deficiency, although the underlying cause of this deficiency is unclear. Recent studies suggest that the histone methyltransferase activity of MMSET plays an important role in the DNA damage response by facilitating the recruitment of 53BP1 to sites of DNA damage. We hypothesize that MMSET also regulates class switch recombination (CSR) through its effect on 53BP1. In this study, we show that MMSET indeed plays an important role in CSR through its histone methyltransferase activity. Knocking down MMSET expression impaired 53BP1 recruitment as well as the germline transcription of the Igh switch regions, resulting in defective CSR but no effect on cell growth and viability. These results suggest that defective CSR caused by MMSET deficiency could be a cause of Ab deficiency in WHS patients. PMID:23241889

  7. Non-redundancy of cytidine deaminases in class switch recombination.

    PubMed

    Fugmann, Sebastian D; Rush, James S; Schatz, David G

    2004-03-01

    Class switch recombination (CSR), somatic hypermutation, and gene conversion are immunoglobulin diversification mechanisms that are strictly dependent on the activity of the activation-induced cytidine deaminase (AID). The precise role and substrate(s) of AID in these processes remain to be well defined. The closest homologue of AID is APOBEC-1, a bona fide mRNA-editing enzyme, which shares with AID the ability to deaminate cytidines within single-stranded DNA in vitro and in prokaryotic cells. To determine whether APOBEC-1 can therefore substitute for AID in activated B cells, we expressed human AID, a catalytic mutant thereof, and rat APOBEC-1 in AID-deficient murine B cells. Whereas AID rescued CSR, neither the inactive mutant nor APOBEC-1 could complement AID deficiency. This indicates that cytidine deaminase activity is necessary but not sufficient to initiate CSR, and suggests that AID is specifically targeted to its cognate substrate, the immunoglobulin genes or a distinct mRNA, by an as-yet-unknown mechanism. PMID:14991614

  8. Related Mechanisms of Antibody Somatic Hypermutation and Class Switch Recombination

    PubMed Central

    HWANG, JOYCE K.; ALT, FREDERICK W.; YEAP, LENG-SIEW

    2015-01-01

    The primary antibody repertoire is generated by mechanisms involving the assembly of the exons that encode the antigen-binding variable regions of immunoglobulin heavy (IgH) and light (IgL) chains during the early development of B lymphocytes. After antigen-dependent activation, mature B lymphocytes can further alter their IgH and IgL variable region exons by the process of somatic hypermutation (SHM), which allows the selection of B cells in which SHMs resulted in the production of antibodies with increased antigen affinity. In addition, during antigen-dependent activation, B cells can also change the constant region of their IgH chain through a DNA double-strand-break (DSB) dependent process referred to as IgH class switch recombination (CSR), which generates B cell progeny that produce antibodies with different IgH constant region effector functions that are best suited for a elimination of a particular pathogen or in a particular setting. Both the mutations that underlie SHM and the DSBs that underlie CSR are initiated in target genes by activation-induced cytidine deaminase (AID). This review describes in depth the processes of SHM and CSR with a focus on mechanisms that direct AID cytidine deamination in activated B cells and mechanisms that promote the differential outcomes of such cytidine deamination. PMID:26104555

  9. The role of G-density in switch region repeats for immunoglobulin class switch recombination

    PubMed Central

    Zhang, Zheng Z.; Pannunzio, Nicholas R.; Hsieh, Chih-Lin; Yu, Kefei; Lieber, Michael R.

    2014-01-01

    The boundaries of R-loops are well-documented at immunoglobulin heavy chain loci in mammalian B cells. Within primary B cells or B cell lines, the upstream boundaries of R-loops typically begin early in the repetitive portion of the switch regions. Most R-loops terminate within the switch repetitive zone, but the remainder can extend a few hundred base pairs further, where G-density on the non-template DNA strand gradually drops to the genome average. Whether the G-density determines how far the R-loops extend is an important question. We previously studied the role of G-clusters in initiating R-loop formation, but we did not examine the role of G-density in permitting the elongation of the R-loop, after it had initiated. Here, we vary the G-density of different portions of the switch region in a murine B cell line. We find that both class switch recombination (CSR) and R-loop formation decrease significantly when the overall G-density is reduced from 46% to 29%. Short 50 bp insertions with low G-density within switch regions do not appear to affect either CSR or R-loop elongation, whereas a longer (150 bp) insertion impairs both. These results demonstrate that G-density is an important determinant of the length over which mammalian genomic R-loops extend. PMID:25378327

  10. Switch Region Identity Plays an Important Role in Ig Class Switch Recombination

    PubMed Central

    Bhattacharya, Palash; Wuerffel, Robert; Kenter, Amy L.

    2016-01-01

    Ig class switch recombination (CSR) is regulated through long-range intrachromosomal interactions between germline transcript promoters and enhancers to initiate transcription and create chromatin accessible to activation-induced deaminase attack. CSR occurs between switch (S) regions that flank Cμ and downstream CH regions and functions via an intrachromosomal deletional event between the donor Sμ region and a downstream S region. It is unclear to what extent S region primary sequence influences differential targeting of CSR to specific isotypes. We address this issue in this study by generating mutant mice in which the endogenous Sγ3 region was replaced with size-matched Sγ1 sequence. B cell activation conditions are established that support robust γ3 and γ1 germline transcript expression and stimulate IgG1 switching but suppress IgG3 CSR. We found that the Sγ1 replacement allele engages in μ→γ3 CSR, whereas the intact allele is repressed. We conclude that S region identity makes a significant contribution to CSR. We propose that the Sγ1 region is selectively targeted for CSR following the induction of an isotype-specific factor that targets the S region and recruits CSR machinery. PMID:20427773

  11. Efficient AID targeting of switch regions is not sufficient for optimal class switch recombination.

    PubMed

    Bonaud, Amélie; Lechouane, Fabien; Le Noir, Sandrine; Monestier, Olivier; Cogné, Michel; Sirac, Christophe

    2015-01-01

    Antibody affinity maturation relies on activation-induced cytidine deaminase (AID)-dependent somatic hypermutation (SHM) of immunoglobulin (Ig) loci. Class switch recombination (CSR) can in parallel occur between AID-targeted, transcribed, spliced and repetitive switch (S) regions. AID thus initiates not only mutations but also double-strand breaks (DSBs). What governs the choice between those two outcomes remains uncertain. Here we explore whether insertion of transcribed intronic S regions in a locus (Igκ) strongly recruiting AID is sufficient for efficient CSR. Although strongly targeted by AID and carrying internal deletions, the knocked-in S regions only undergo rare CSR-like events. This model confirms S regions as exquisite SHM targets, extending AID activity far from transcription initiation sites, and shows that such spliced and repetitive AID targets are not sufficient by themselves for CSR. Beyond transcription and AID recruitment, additional IgH elements are thus needed for CSR, restricting this hazardous gene remodelling to IgH loci. PMID:26146363

  12. Regulating infidelity: RNA-mediated recruitment of AID to DNA during class switch recombination.

    PubMed

    DiMenna, Lauren J; Chaudhuri, Jayanta

    2016-03-01

    The mechanism by which the DNA deaminase activation-induced cytidine deaminase (AID) is specifically recruited to repetitive switch region DNA during class switch recombination is still poorly understood. Work over the past decade has revealed a strong link between transcription and RNA polymerase-associated factors in AID recruitment, yet none of these processes satisfactorily explain how AID specificity is affected. Here, we review a recent finding wherein AID is guided to switch regions not by a protein factor but by an RNA moiety, and especially one associated with a noncoding RNA that has been long thought of as being inert. This work explains the long-standing requirement of splicing of noncoding transcripts during class switching, and has implications in both B cell-mediated immunity as well as the underlying pathological syndromes associated with the recombination reaction. PMID:26799454

  13. Constraints contributed by chromatin looping limit recombination targeting during immunoglobulin class switch recombination

    PubMed Central

    Feldman, Scott; Achour, Ikbel; Wuerffel, Robert; Kumar, Satyendra; Gerasimova, Tatiana; Sen, Ranjan; Kenter, Amy L.

    2015-01-01

    Engagement of promoters with distal elements in long range looping interactions has been implicated in regulation of Ig class switch recombination (CSR). The principles determining the spatial and regulatory relationships among Igh transcriptional elements remain poorly defined. We examined the chromosome conformation of constant region (CH) loci that are targeted for CSR in a cytokine dependent fashion in mature B lymphocytes. Germline transcription (GLT) of the γ1 and ε CH loci is controlled by two transcription factors, IL4 inducible STAT6 and LPS activated NFκB. We showed that although STAT6 deficiency triggered loss of GLT, deletion of NFκB p50 abolished both GLT and γ1 locus:enhancer looping. Thus, chromatin looping between CH loci and Igh enhancers is independent of GLT production and STAT6, whereas the establishment and maintenance of these chromatin contacts requires NFκB p50. Comparative analysis of the endogenous γ1 locus and a knock-in heterologous promoter in mice identified the promoter per se as the interactive looping element and showed that transcription elongation is dispensable for promoter/enhancer interactions. Interposition of the LPS responsive heterologous promoter between the LPS inducible γ3 and γ2b loci altered GLT expression and essentially abolished direct IgG2b switching while maintaining a sequential μ-> γ3-> γ2b format. Our study provides evidence that promoter/enhancer looping interactions can introduce negative constraints on distal promoters and affect their ability to engage in germline transcription and determine CSR targeting. PMID:25624452

  14. Constraints contributed by chromatin looping limit recombination targeting during Ig class switch recombination.

    PubMed

    Feldman, Scott; Achour, Ikbel; Wuerffel, Robert; Kumar, Satyendra; Gerasimova, Tatiana; Sen, Ranjan; Kenter, Amy L

    2015-03-01

    Engagement of promoters with distal elements in long-range looping interactions has been implicated in regulation of Ig class switch recombination (CSR). The principles determining the spatial and regulatory relationships among Igh transcriptional elements remain poorly defined. We examined the chromosome conformation of C region (CH) loci that are targeted for CSR in a cytokine-dependent fashion in mature B lymphocytes. Germline transcription (GLT) of the γ1 and ε CH loci is controlled by two transcription factors, IL-4-inducible STAT6 and LPS-activated NF-κB. We showed that although STAT6 deficiency triggered loss of GLT, deletion of NF-κB p50 abolished both GLT and γ1 locus:enhancer looping. Thus, chromatin looping between CH loci and Igh enhancers is independent of GLT production and STAT6, whereas the establishment and maintenance of these chromatin contacts requires NF-κB p50. Comparative analysis of the endogenous γ1 locus and a knock-in heterologous promoter in mice identified the promoter per se as the interactive looping element and showed that transcription elongation is dispensable for promoter/enhancer interactions. Interposition of the LPS-responsive heterologous promoter between the LPS-inducible γ3 and γ2b loci altered GLT expression and essentially abolished direct IgG2b switching while maintaining a sequential μ→γ3→γ2b format. Our study provides evidence that promoter/enhancer looping interactions can introduce negative constraints on distal promoters and affect their ability to engage in germline transcription and determine CSR targeting. PMID:25624452

  15. Role for Msh5 in the Regulation of Ig Class Switch Recombination

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Immunoglobulin (Ig) class switch recombination (CSR) and somatic hypermutation serve to diversify antibody responses, and are orchestrated by the activity of activation-induced cytidine deaminase (AID) and a large number of proteins involved in DNA repair and genome surveillance. Here we show that ...

  16. ALKBH1 is dispensable for abasic site cleavage during base excision repair and class switch recombination.

    PubMed

    Müller, Tina A; Yu, Kefei; Hausinger, Robert P; Meek, Katheryn

    2013-01-01

    Potential roles of the abasic site lyase activity associated with AlkB homolog 1 (ALKBH1) were assessed by studies focusing on the two cellular processes that create abasic sites as intermediates: base excision repair and class switch recombination. Alkbh1(-/-) pups (lacking exon 3) were born at a lower than expected frequency from heterozygous parents, suggesting a reduced survival rate and non-Mendelian inheritance, and they exhibited a gender bias in favor of males (70% males and 30% females). To study ALKBH1's potential involvement in DNA repair, fibroblasts were isolated from Alkbh1(-/-) mice, spontaneously immortalized and tested for resistance to DNA damaging agents. Alkbh1(-/-) and isogenic cells expressing hALKBH1 showed no difference in survival to the DNA damaging agents methyl-methionine sulfate or H2O2. This result indicates that ALKBH1 does not play a major role in the base excision repair pathway. To assess ALKBH1's role in class switch recombination, splenic B cells were isolated from Alkbh1(-/-) and Alkbh1(+/+) mice and subjected to switching from IgM to IgG1. No differences were found in IgG1 switching, suggesting that Alkbh1 is not involved in class switch recombination of the immunoglobulin heavy chain during B lymphocyte activation. PMID:23825659

  17. Aberrant recombination and repair during immunoglobulin class switching in BRCA1-deficient human B cells

    PubMed Central

    Björkman, Andrea; Qvist, Per; Du, Likun; Bartish, Margarita; Zaravinos, Apostolos; Georgiou, Konstantinos; Børglum, Anders D.; Gatti, Richard A.; Törngren, Therese; Pan-Hammarström, Qiang

    2015-01-01

    Breast cancer type 1 susceptibility protein (BRCA1) has a multitude of functions that contribute to genome integrity and tumor suppression. Its participation in the repair of DNA double-strand breaks (DSBs) during homologous recombination (HR) is well recognized, whereas its involvement in the second major DSB repair pathway, nonhomologous end-joining (NHEJ), remains controversial. Here we have studied the role of BRCA1 in the repair of DSBs in switch (S) regions during immunoglobulin class switch recombination, a physiological, deletion/recombination process that relies on the classical NHEJ machinery. A shift to the use of microhomology-based, alternative end-joining (A-EJ) and increased frequencies of intra-S region deletions as well as insertions of inverted S sequences were observed at the recombination junctions amplified from BRCA1-deficient human B cells. Furthermore, increased use of long microhomologies was found at recombination junctions derived from E3 ubiquitin-protein ligase RNF168-deficient, Fanconi anemia group J protein (FACJ, BRIP1)-deficient, or DNA endonuclease RBBP8 (CtIP)-compromised cells, whereas an increased frequency of S-region inversions was observed in breast cancer type 2 susceptibility protein (BRCA2)-deficient cells. Thus, BRCA1, together with its interaction partners, seems to play an important role in repairing DSBs generated during class switch recombination by promoting the classical NHEJ pathway. This may not only provide a general mechanism underlying BRCA1’s function in maintaining genome stability and tumor suppression but may also point to a previously unrecognized role of BRCA1 in B-cell lymphomagenesis. PMID:25646469

  18. Effect of CpG dinucleotides within IgH switch region repeats on immunoglobulin class switch recombination.

    PubMed

    Zhang, Zheng Z; Hsieh, Chih-Lin; Okitsu, Cindy Yen; Han, Li; Yu, Kefei; Lieber, Michael R

    2015-08-01

    Immunoglobulin (Ig) heavy chains undergo class switch recombination (CSR) to change the heavy chain isotype from IgM to IgG, A or E. The switch regions are several kilobases long, repetitive, and G-rich on the nontemplate strand. They are also relatively depleted of CpG (also called CG) sites for unknown reasons. Here we use synthetic switch regions at the IgH switch alpha (Sα) locus to test the effect of CpG sites and to try to understand why the IgH switch sequences evolved to be relatively depleted of CpG. We find that even just two CpG sites within an 80 bp synthetic switch repeat iterated 15 times (total switch region length of 1200 bp containing 30 CpG sites) are sufficient to dramatically reduce both Ig CSR and transcription through the switch region from the upstream Iα sterile transcript promoter, which is the promoter that directs transcripts through the Sα region. De novo DNA methylation occurs at the four CpG sites in and around the Iα promoter when each 80 bp Iα switch repeat contains the two CpG sites. Thus, a relatively low density of CpG sites within the switch repeats can induce upstream CpG methylation at the IgH alpha locus, and cause a substantial decrease in transcription from the sterile transcript promoter. This effect is likely the reason that switch regions evolved to contain very few CpG sites. We discuss these findings as they relate to DNA methylation and to Ig CSR. PMID:25899867

  19. Roles of nonhomologous DNA end joining, V(D)J recombination, and class switch recombination in chromosomal translocations.

    PubMed

    Lieber, Michael R; Yu, Kefei; Raghavan, Sathees C

    2006-09-01

    When a single double-strand break arises in the genome, nonhomologous DNA end joining (NHEJ) is a major pathway for its repair. When double-strand breaks arise at two nonhomologous sites in the genome, NHEJ also appears to be a major pathway by which the translocated ends are joined. The mechanism of NHEJ is briefly summarized, and alternative enzymes are also discussed. V(D)J recombination and class switch recombination are specialized processes designed to create double-strand DNA breaks at specific locations in the genomes of lymphoid cells. Sporadic Burkitt's lymphoma and myelomas can arise due to translocation of the c-myc gene into the Ig heavy chain locus during class switch recombination. In other lymphoid neoplasms, the RAG complex can create double-strand breaks that result in a translocation. Such RAG-generated breaks occur at very specific nucleotides that are directly adjacent to sequences that resemble canonical heptamer/nonamer sequences characteristic of normal V(D)J recombination. This occurs in some T cell leukemias and lymphomas. The RAG complex also appears capable of recognizing regions for their altered DNA structure rather than their primary sequence, and this may account for the action by RAGs at some chromosomal translocation sites, such as at the bcl-2 major breakpoint region in the follicular lymphomas that arise in B lymphocytes. PMID:16793349

  20. DNA-PKcs Is Involved in Ig Class Switch Recombination in Human B Cells.

    PubMed

    Björkman, Andrea; Du, Likun; Felgentreff, Kerstin; Rosner, Cornelia; Pankaj Kamdar, Radhika; Kokaraki, Georgia; Matsumoto, Yoshihisa; Davies, E Graham; van der Burg, Mirjam; Notarangelo, Luigi D; Hammarström, Lennart; Pan-Hammarström, Qiang

    2015-12-15

    Nonhomologous end-joining (NHEJ) is one of the major DNA double-strand break repair pathways in mammalian cells and is required for both V(D)J recombination and class switch recombination (CSR), two Ig gene-diversification processes occurring during B cell development. DNA-dependent protein kinase, catalytic subunit (DNA-PKcs) is a component of the classical NHEJ machinery and has a critical function during V(D)J recombination. However, its role in CSR has been controversial. In this study, we examined the pattern of recombination junctions from in vivo-switched B cells from two DNA-PKcs-deficient patients. One of them harbored mutations that did not affect DNA-PKcs kinase activity but caused impaired Artemis activation; the second patient had mutations resulting in diminished DNA-PKcs protein expression and kinase activity. These results were compared with those from DNA-PKcs-deficient mouse B cells. A shift toward the microhomology-based alternative end-joining at the recombination junctions was observed in both human and mouse B cells, suggesting that the classical NHEJ pathway is impaired during CSR when DNA-PKcs is defective. Furthermore, cells from the second patient showed additional or more severe alterations in CSR and/or NHEJ, which may suggest that DNA-PKcs and/or its kinase activity have additional, Artemis-independent functions during these processes. PMID:26546606

  1. A DNA break- and phosphorylation-dependent positive feedback loop promotes immunoglobulin class-switch recombination.

    PubMed

    Vuong, Bao Q; Herrick-Reynolds, Kayleigh; Vaidyanathan, Bharat; Pucella, Joseph N; Ucher, Anna J; Donghia, Nina M; Gu, Xiwen; Nicolas, Laura; Nowak, Urszula; Rahman, Numa; Strout, Matthew P; Mills, Kevin D; Stavnezer, Janet; Chaudhuri, Jayanta

    2013-11-01

    The ability of activation-induced cytidine deaminase (AID) to efficiently mediate class-switch recombination (CSR) is dependent on its phosphorylation at Ser38; however, the trigger that induces AID phosphorylation and the mechanism by which phosphorylated AID drives CSR have not been elucidated. Here we found that phosphorylation of AID at Ser38 was induced by DNA breaks. Conversely, in the absence of AID phosphorylation, DNA breaks were not efficiently generated at switch (S) regions in the immunoglobulin heavy-chain locus (Igh), consistent with a failure of AID to interact with the endonuclease APE1. Additionally, deficiency in the DNA-damage sensor ATM impaired the phosphorylation of AID at Ser38 and the interaction of AID with APE1. Our results identify a positive feedback loop for the amplification of DNA breaks at S regions through the phosphorylation- and ATM-dependent interaction of AID with APE1. PMID:24097111

  2. Interaction between HIV-1 Tat and DNA-PKcs modulates HIV transcription and class switch recombination.

    PubMed

    Zhang, Shi-Meng; Zhang, He; Yang, Tian-Yi; Ying, Tian-Yi; Yang, Pei-Xiang; Liu, Xiao-Dan; Tang, Sheng-Jian; Zhou, Ping-Kun

    2014-01-01

    HIV-1 tat targets a variety of host cell proteins to facilitate viral transcription and disrupts host cellular immunity by inducing lymphocyte apoptosis, but whether it influences humoral immunity remains unclear. Previously, our group demonstrated that tat depresses expression of DNA-PKcs, a critical component of the non-homologous end joining pathway (NHEJ) of DNA double-strand breaks repair, immunoglobulin class switch recombination (CSR) and V(D)J recombination, and sensitizes cells to ionizing radiation. In this study, we demonstrated that HIV-1 Tat down-regulates DNA-PKcs expression by directly binding to the core promoter sequence. In addition, Tat interacts with and activates the kinase activity of DNA-PKcs in a dose-dependent and DNA independent manner. Furthermore, Tat inhibits class switch recombination (CSR) at low concentrations (≤ 4 µg/ml) and stimulates CSR at high concentrations (≥ 8 µg/ml). On the other hand, low protein level and high kinase activity of DNA-PKcs promotes HIV-1 transcription, while high protein level and low kinase activity inhibit HIV-1 transcription. Co-immunoprecipitation results revealed that DNA-PKcs forms a large complex comprised of Cyclin T1, CDK9 and Tat via direct interacting with CDK9 and Tat but not Cyclin T1. Taken together, our results provide new clues that Tat regulates host humoral immunity via both transcriptional depression and kinase activation of DNA-PKcs. We also raise the possibility that inhibitors and interventions directed towards DNA-PKcs may inhibit HIV-1 transcription in AIDS patients. PMID:25332688

  3. Interaction between HIV-1 Tat and DNA-PKcs modulates HIV transcription and class switch recombination

    PubMed Central

    Zhang, Shi-Meng; Zhang, He; Yang, Tian-Yi; Ying, Tian-Yi; Yang, Pei-Xiang; Liu, Xiao-Dan; Tang, Sheng-Jian; Zhou, Ping-Kun

    2014-01-01

    HIV-1 tat targets a variety of host cell proteins to facilitate viral transcription and disrupts host cellular immunity by inducing lymphocyte apoptosis, but whether it influences humoral immunity remains unclear. Previously, our group demonstrated that tat depresses expression of DNA-PKcs, a critical component of the non-homologous end joining pathway (NHEJ) of DNA double-strand breaks repair, immunoglobulin class switch recombination (CSR) and V(D)J recombination, and sensitizes cells to ionizing radiation. In this study, we demonstrated that HIV-1 Tat down-regulates DNA-PKcs expression by directly binding to the core promoter sequence. In addition, Tat interacts with and activates the kinase activity of DNA-PKcs in a dose-dependent and DNA independent manner. Furthermore, Tat inhibits class switch recombination (CSR) at low concentrations (≤4 µg/ml) and stimulates CSR at high concentrations (≥8 µg/ml). On the other hand, low protein level and high kinase activity of DNA-PKcs promotes HIV-1 transcription, while high protein level and low kinase activity inhibit HIV-1 transcription. Co-immunoprecipitation results revealed that DNA-PKcs forms a large complex comprised of Cyclin T1, CDK9 and Tat via direct interacting with CDK9 and Tat but not Cyclin T1. Taken together, our results provide new clues that Tat regulates host humoral immunity via both transcriptional depression and kinase activation of DNA-PKcs. We also raise the possibility that inhibitors and interventions directed towards DNA-PKcs may inhibit HIV-1 transcription in AIDS patients. PMID:25332688

  4. Evaluation of the adverse effect of low concentration of cadmium on interleukin-4 induced class switch recombination in Burkett's lymphoma Raji cell line.

    PubMed

    Poltoratsky, Vladimir

    2014-01-01

    Affinity maturation of B lymphocytes, a process that includes somatic hypermutation and class switch recombination, initiates global DNA rearrangements. The interruption of this process has an adverse effect on human health and results in immunodeficiency and autoimmune disease. Class switch recombination is a fundamental factor of the human adaptive immunity. Evaluation of the class switch recombination efficiency is an important component of laboratory diagnostic of immunotoxic components. Here, we describe a method for testing the efficiency of the class switch recombination. Cultivation of Raji Burkett's lymphoma cell line with anti-CD40 antibodies and recombinant interleukin-4 (IL-4) triggers a cascade of signal transduction network events that lead to switching the immunoglobulin isotopes from IgM to IgE. This chapter describes the methodology of class switch recombination assay for assessment of the effect of the environmental pollutants in toxicological laboratory diagnostics. PMID:24908303

  5. 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.

  6. Cernunnos influences human immunoglobulin class switch recombination and may be associated with B cell lymphomagenesis.

    PubMed

    Du, Likun; Peng, Roujun; Björkman, Andrea; Filipe de Miranda, Noel; Rosner, Cornelia; Kotnis, Ashwin; Berglund, Mattias; Liu, Chonghai; Rosenquist, Richard; Enblad, Gunilla; Sundström, Christer; Hojjat-Farsangi, Mohammad; Rabbani, Hodjattallah; Teixeira, Manuel R; Revy, Patrick; Durandy, Anne; Zeng, Yixin; Gennery, Andrew R; de Villartay, Jean-Pierre; Pan-Hammarström, Qiang

    2012-02-13

    Cernunnos is involved in the nonhomologous end-joining (NHEJ) process during DNA double-strand break (DSB) repair. Here, we studied immunoglobulin (Ig) class switch recombination (CSR), a physiological process which relies on proper repair of the DSBs, in B cells from Cernunnos-deficient patients. The pattern of in vivo generated CSR junctions is altered in these cells, with unusually long microhomologies and a lack of direct end-joining. The CSR junctions from Cernunnos-deficient patients largely resemble those from patients lacking DNA ligase IV, Artemis, or ATM, suggesting that these factors are involved in the same end-joining pathway during CSR. By screening 269 mature B cell lymphoma biopsies, we also identified a somatic missense Cernunnos mutation in a diffuse large B cell lymphoma sample. This mutation has a dominant-negative effect on joining of a subset of DNA ends in an in vitro NHEJ assay. Translocations involving both Ig heavy chain loci and clonal-like, dynamic IgA switching activities were observed in this tumor. Collectively, our results suggest a link between defects in the Cernunnos-dependent NHEJ pathway and aberrant CSR or switch translocations during the development of B cell malignancies. PMID:22312109

  7. AIDing Chromatin and Transcription-Coupled Orchestration of Immunoglobulin Class-Switch Recombination

    PubMed Central

    Vaidyanathan, Bharat; Yen, Wei-Feng; Pucella, Joseph N.; Chaudhuri, Jayanta

    2014-01-01

    Secondary diversification of the antibody repertoire upon antigenic challenge, in the form of immunoglobulin heavy chain (IgH) class-switch recombination (CSR) endows mature, naïve B cells in peripheral lymphoid organs with a limitless ability to mount an optimal humoral immune response, thus expediting pathogen elimination. CSR replaces the default constant (CH) region exons (Cμ) of IgH with any of the downstream CH exons (Cγ, Cε, or Cα), thereby altering effector functions of the antibody molecule. This process depends on, and is orchestrated by, activation-induced deaminase (AID), a DNA cytidine deaminase that acts on single-stranded DNA exposed during transcription of switch (S) region sequences at the IgH locus. DNA lesions thus generated are processed by components of several general DNA repair pathways to drive CSR. Given that AID can instigate DNA lesions and genomic instability, stringent checks are imposed that constrain and restrict its mutagenic potential. In this review, we will discuss how AID expression and substrate specificity and activity is rigorously enforced at the transcriptional, post-transcriptional, post-translational, and epigenetic levels, and how the DNA-damage response is choreographed with precision to permit targeted activity while limiting bystander catastrophe. PMID:24734031

  8. Individual Substitution Mutations in the AID C Terminus That Ablate IgH Class Switch Recombination

    PubMed Central

    Kadungure, Tatenda; Ucher, Anna J.; Linehan, Erin K.; Schrader, Carol E.; Stavnezer, Janet

    2015-01-01

    Activation-induced cytidine deaminase (AID) is essential for class switch recombination (CSR) and somatic hypermutation (SHM) of Ig genes. The C terminus of AID is required for CSR but not for SHM, but the reason for this is not entirely clear. By retroviral transduction of mutant AID proteins into aid-/- mouse splenic B cells, we show that 4 amino acids within the C terminus of mouse AID, when individually mutated to specific amino acids (R190K, A192K, L196S, F198S), reduce CSR about as much or more than deletion of the entire C terminal 10 amino acids. Similar to ΔAID, the substitutions reduce binding of UNG to Ig Sμ regions and some reduce binding of Msh2, both of which are important for introducing S region DNA breaks. Junctions between the IgH donor switch (S)μ and acceptor Sα regions from cells expressing ΔAID or the L196S mutant show increased microhomology compared to junctions in cells expressing wild-type AID, consistent with problems during CSR and the use of alternative end-joining, rather than non-homologous end-joining (NHEJ). Unlike deletion of the AID C terminus, 3 of the substitution mutants reduce DNA double-strand breaks (DSBs) detected within the Sμ region in splenic B cells undergoing CSR. Cells expressing these 3 substitution mutants also have greatly reduced mutations within unrearranged Sμ regions, and they decrease with time after activation. These results might be explained by increased error-free repair, but as the C terminus has been shown to be important for recruitment of NHEJ proteins, this appears unlikely. We hypothesize that Sμ DNA breaks in cells expressing these C terminus substitution mutants are poorly repaired, resulting in destruction of Sμ segments that are deaminated by these mutants. This could explain why these mutants cannot undergo CSR. PMID:26267846

  9. A Damage-Independent Role for 53BP1 that Impacts Break Order and Igh Architecture during Class Switch Recombination.

    PubMed

    Rocha, Pedro P; Raviram, Ramya; Fu, Yi; Kim, JungHyun; Luo, Vincent M; Aljoufi, Arafat; Swanzey, Emily; Pasquarella, Alessandra; Balestrini, Alessia; Miraldi, Emily R; Bonneau, Richard; Petrini, John; Schotta, Gunnar; Skok, Jane A

    2016-06-28

    During class switch recombination (CSR), B cells replace the Igh Cμ or δ exons with another downstream constant region exon (CH), altering the antibody isotype. CSR occurs through the introduction of AID-mediated double-strand breaks (DSBs) in switch regions and subsequent ligation of broken ends. Here, we developed an assay to investigate the dynamics of DSB formation in individual cells. We demonstrate that the upstream switch region Sμ is first targeted during recombination and that the mechanism underlying this control relies on 53BP1. Surprisingly, regulation of break order occurs through residual binding of 53BP1 to chromatin before the introduction of damage and independent of its established role in DNA repair. Using chromosome conformation capture, we show that 53BP1 mediates changes in chromatin architecture that affect break order. Finally, our results explain how changes in Igh architecture in the absence of 53BP1 could promote inversional rearrangements that compromise CSR. PMID:27320916

  10. Polyclonal hyper-IgE mouse model reveals mechanistic insights into antibody class switch recombination

    PubMed Central

    Misaghi, Shahram; Senger, Kate; Sai, Tao; Qu, Yan; Sun, Yonglian; Hamidzadeh, Kajal; Nguyen, Allen; Jin, Zhaoyu; Zhou, Meijuan; Yan, Donghong; Lin, Wei Yu; Lin, Zhonghua; Lorenzo, Maria N.; Sebrell, Andrew; Ding, Jiabing; Xu, Min; Caplazi, Patrick; Austin, Cary D.; Balazs, Mercedesz; Roose-Girma, Merone; DeForge, Laura; Warming, Søren; Lee, Wyne P.; Dixit, Vishva M.; Zarrin, Ali A.

    2013-01-01

    Preceding antibody constant regions are switch (S) regions varying in length and repeat density that are targets of activation-induced cytidine deaminase. We asked how participating S regions influence each other to orchestrate rearrangements at the IgH locus by engineering mice in which the weakest S region, Sε, is replaced with prominent recombination hotspot Sμ. These mice produce copious polyclonal IgE upon challenge, providing a platform to study IgE biology and therapeutic interventions. The insertion enhances ε germ-line transcript levels, shows a preference for direct vs. sequential switching, and reduces intraswitch recombination events at native Sμ. These results suggest that the sufficiency of Sμ to mediate IgH rearrangements may be influenced by context-dependent cues. PMID:24019479

  11. Polyclonal hyper-IgE mouse model reveals mechanistic insights into antibody class switch recombination.

    PubMed

    Misaghi, Shahram; Senger, Kate; Sai, Tao; Qu, Yan; Sun, Yonglian; Hamidzadeh, Kajal; Nguyen, Allen; Jin, Zhaoyu; Zhou, Meijuan; Yan, Donghong; Lin, Wei Yu; Lin, Zhonghua; Lorenzo, Maria N; Sebrell, Andrew; Ding, Jiabing; Xu, Min; Caplazi, Patrick; Austin, Cary D; Balazs, Mercedesz; Roose-Girma, Merone; DeForge, Laura; Warming, Søren; Lee, Wyne P; Dixit, Vishva M; Zarrin, Ali A

    2013-09-24

    Preceding antibody constant regions are switch (S) regions varying in length and repeat density that are targets of activation-induced cytidine deaminase. We asked how participating S regions influence each other to orchestrate rearrangements at the IgH locus by engineering mice in which the weakest S region, Sε, is replaced with prominent recombination hotspot Sμ. These mice produce copious polyclonal IgE upon challenge, providing a platform to study IgE biology and therapeutic interventions. The insertion enhances ε germ-line transcript levels, shows a preference for direct vs. sequential switching, and reduces intraswitch recombination events at native Sμ. These results suggest that the sufficiency of Sμ to mediate IgH rearrangements may be influenced by context-dependent cues. PMID:24019479

  12. Regulation of aicda expression and AID activity: Relevance to somatic hypermutation and class switch DNA recombination

    PubMed Central

    Xu, Zhenming; Pone, Egest J.; Al-Qahtani, Ahmed; Park, Seok-Rae; Zan, Hong; Casali, Paolo

    2010-01-01

    Expression and activity of activation-induced cytidine deaminase (AID) encoded by the aicda gene are essential for immunoglobulin (Ig) gene somatic hypermutation (SHM) and class switch DNA recombination (CSR). SHM and CSR unfold in general in germinal centers and are central to the maturation of effective antibody responses. AID expression is induced by activated B cell CD40 signaling, which is critical for the germinal center reaction, and is further enhanced by other stimuli, including interleukin-4 (IL-4) secreted from CD4+ T cells or Toll-like receptor (TLR)-activating bacterial and/or viral molecules. Integration of different intracellular signal transduction pathways, as activated by these stimuli, leads to a dynamic aicda-regulating program, which involves both positively acting trans-factors, such as Pax5, HoxC4, E47 and Irf8, and negative modulators, such as Blimp1 and Id2, to restrict aicda expression primarily to germinal center B cells. The phosphatidylinositol 3-kinase (PI 3-K), which functions downstream of activated B cell receptor (BCR) signaling, likely plays an important role in triggering the downregulation of aicda expression in post-germinal center B cells and throughout plasmacytoid differentiation. In B cells undergoing SHM and CSR, AID activity and, possibly, AID targeting to the Ig locus are regulated at a post-translational level, including AID dimerization/oligomerization, nuclear/cytoplasmic AID translocation and phosphorylation of the AID Ser38 residue by protein kinase A (PKA). Here, we will discuss the role of B cell activation signals, transcription regulation programs and post-translational modifications in controlling aicda expression and AID activity, thereby delineating an integrated model of modulation of SHM and CSR in the germinal center reaction. PMID:18197815

  13. The Interaction between AID and CIB1 Is Nonessential for Antibody Gene Diversification by Gene Conversion or Class Switch Recombination

    PubMed Central

    Demorest, Zachary L.; MacDuff, Donna A.; Brown, William L.; Morham, Scott G.; Parise, Leslie V.; Harris, Reuben S.

    2010-01-01

    Activation-induced deaminase (AID) initiates somatic hypermutation, gene conversion and class switch recombination by deaminating variable and switch region DNA cytidines to uridines. AID is predominantly cytoplasmic and must enter the nuclear compartment to initiate these distinct antibody gene diversification reactions. Nuclear AID is relatively short-lived, as it is efficiently exported by a CRM1-dependent mechanism and it is susceptible to proteasome-dependent degradation. To help shed light on mechanisms of post-translational regulation, a yeast-based screen was performed to identify AID-interacting proteins. The calcium and integrin binding protein CIB1 was identified by sequencing and the interaction was confirmed by immunoprecipitation experiments. The AID/CIB1 resisted DNase and RNase treatment, and it is therefore unlikely to be mediated by nucleic acid. The requirement for CIB1 in AID-mediated antibody gene diversification reactions was assessed in CIB1-deficient DT40 cells and in knockout mice, but immunoglobulin gene conversion and class switch recombination appeared normal. The DT40 system was also used to show that CIB1 over-expression has no effect on gene conversion and that AID-EGFP subcellular localization is normal. These combined data demonstrate that CIB1 is not required for AID to mediate antibody gene diversification processes. It remains possible that CIB1 has an alternative, a redundant or a subtle non-limiting regulatory role in AID biology. PMID:20652029

  14. AID expression increased by TNF-α is associated with class switch recombination of Igα gene in cancers

    PubMed Central

    Duan, Zhi; Zheng, Hui; Liu, Haidan; Li, Ming; Tang, Min; Weng, Xinxian; Yi, Wei; Bode, Ann M.; Cao, Ya

    2016-01-01

    Recently, immunoglobulins (Igs) were unexpectedly found to be expressed in epithelial cancers. Immunoglobulin class switching or class switch recombination (CSR) is a natural biological process that alters a B cell's production of antibodies (immunoglobulins) from one class to another. However, the mechanism of CSR of Ig genes in cancer is still unknown. Here, we confirmed by detecting the hallmark of CSR that the Igα gene in cancer underwent CSR. Then we focused on activation-induced cytidine deaminase (AID), a crucial factor for initiating CSR. Further studies using tumor necrosis factor (TNF)-α stimulation and specific inhibitor of NF-κB revealed that TNF-α could increase AID expression through NF-κB signaling. Finally, we demonstrated that AID could co-localize with protein kinase A and bind to the switching (Sα) region of the Igα gene. Overexpression of AID obviously enhanced Igα heavy chain expression and its binding ability to the Sα region. These findings indicated that TNF-α-induced AID expression is involved with CSR in cancer. PMID:25849121

  15. AID expression increased by TNF-α is associated with class switch recombination of Igα gene in cancers.

    PubMed

    Duan, Zhi; Zheng, Hui; Liu, Haidan; Li, Ming; Tang, Min; Weng, Xinxian; Yi, Wei; Bode, Ann M; Cao, Ya

    2016-07-01

    Recently, immunoglobulins (Igs) were unexpectedly found to be expressed in epithelial cancers. Immunoglobulin class switching or class switch recombination (CSR) is a natural biological process that alters a B cell's production of antibodies (immunoglobulins) from one class to another. However, the mechanism of CSR of Ig genes in cancer is still unknown. Here, we confirmed by detecting the hallmark of CSR that the Igα gene in cancer underwent CSR. Then we focused on activation-induced cytidine deaminase (AID), a crucial factor for initiating CSR. Further studies using tumor necrosis factor (TNF)-α stimulation and specific inhibitor of NF-κB revealed that TNF-α could increase AID expression through NF-κB signaling. Finally, we demonstrated that AID could co-localize with protein kinase A and bind to the switching (Sα) region of the Igα gene. Overexpression of AID obviously enhanced Igα heavy chain expression and its binding ability to the Sα region. These findings indicated that TNF-α-induced AID expression is involved with CSR in cancer. PMID:25849121

  16. Redundant function of DNA ligase 1 and 3 in alternative end-joining during immunoglobulin class switch recombination.

    PubMed

    Masani, Shahnaz; Han, Li; Meek, Katheryn; Yu, Kefei

    2016-02-01

    Nonhomologous end-joining (NHEJ) is the major DNA double-strand break (DSB) repair pathway in mammals and resolves the DSBs generated during both V(D)J recombination in developing lymphocytes and class switch recombination (CSR) in antigen-stimulated B cells. In contrast to the absolute requirement for NHEJ to resolve DSBs associated with V(D)J recombination, DSBs associated with CSR can be resolved in NHEJ-deficient cells (albeit at a reduced level) by a poorly defined alternative end-joining (A-EJ) pathway. Deletion of DNA ligase IV (Lig4), a core component of the NHEJ pathway, reduces CSR efficiency in a mouse B-cell line capable of robust cytokine-stimulated CSR in cell culture. Here, we report that CSR levels are not further reduced by deletion of either of the two remaining DNA ligases (Lig1 and nuclear Lig3) in Lig4(-/-) cells. We conclude that in the absence of Lig4, Lig1, and Lig3 function in a redundant manner in resolving switch region DSBs during CSR. PMID:26787901

  17. ATM increases activation-induced cytidine deaminase activity at downstream S regions during class-switch recombination.

    PubMed

    Khair, Lyne; Guikema, Jeroen E J; Linehan, Erin K; Ucher, Anna J; Leus, Niek G J; Ogilvie, Colin; Lou, Zhenkun; Schrader, Carol E; Stavnezer, Janet

    2014-05-15

    Activation-induced cytidine deaminase (AID) initiates Ab class-switch recombination (CSR) in activated B cells resulting in exchanging the IgH C region and improved Ab effector function. During CSR, AID instigates DNA double-strand break (DSB) formation in switch (S) regions located upstream of C region genes. DSBs are necessary for CSR, but improper regulation of DSBs can lead to chromosomal translocations that can result in B cell lymphoma. The protein kinase ataxia telangiectasia mutated (ATM) is an important proximal regulator of the DNA damage response (DDR), and translocations involving S regions are increased in its absence. ATM phosphorylates H2AX, which recruits other DNA damage response (DDR) proteins, including mediator of DNA damage checkpoint 1 (Mdc1) and p53 binding protein 1 (53BP1), to sites of DNA damage. As these DDR proteins all function to promote repair and recombination of DSBs during CSR, we examined whether mouse splenic B cells deficient in these proteins would show alterations in S region DSBs when undergoing CSR. We find that in atm(-/-) cells Sμ DSBs are increased, whereas DSBs in downstream Sγ regions are decreased. We also find that mutations in the unrearranged Sγ3 segment are reduced in atm(-/-) cells. Our data suggest that ATM increases AID targeting and activity at downstream acceptor S regions during CSR and that in atm(-/-) cells Sμ DSBs accumulate as they lack a recombination partner. PMID:24729610

  18. Opinion: uracil DNA glycosylase (UNG) plays distinct and non-canonical roles in somatic hypermutation and class switch recombination

    PubMed Central

    Yousif, Ashraf S.; Stanlie, Andre; Begum, Nasim A.

    2014-01-01

    Activation-induced cytidine deaminase (AID) is essential to class switch recombination (CSR) and somatic hypermutation (SHM). Uracil DNA glycosylase (UNG), a member of the base excision repair complex, is required for CSR. The role of UNG in CSR and SHM is extremely controversial. AID deficiency in mice abolishes both CSR and SHM, while UNG-deficient mice have drastically reduced CSR but augmented SHM raising a possibility of differential functions of UNG in CSR and SHM. Interestingly, UNG has been associated with a CSR-specific repair adapter protein Brd4, which interacts with acetyl histone 4, γH2AX and 53BP1 to promote non-homologous end joining during CSR. A non-canonical scaffold function of UNG, but not the catalytic activity, can be attributed to the recruitment of essential repair proteins associated with the error-free repair during SHM, and the end joining during CSR. PMID:24994819

  19. Opinion: uracil DNA glycosylase (UNG) plays distinct and non-canonical roles in somatic hypermutation and class switch recombination.

    PubMed

    Yousif, Ashraf S; Stanlie, Andre; Begum, Nasim A; Honjo, Tasuku

    2014-10-01

    Activation-induced cytidine deaminase (AID) is essential to class switch recombination (CSR) and somatic hypermutation (SHM). Uracil DNA glycosylase (UNG), a member of the base excision repair complex, is required for CSR. The role of UNG in CSR and SHM is extremely controversial. AID deficiency in mice abolishes both CSR and SHM, while UNG-deficient mice have drastically reduced CSR but augmented SHM raising a possibility of differential functions of UNG in CSR and SHM. Interestingly, UNG has been associated with a CSR-specific repair adapter protein Brd4, which interacts with acetyl histone 4, γH2AX and 53BP1 to promote non-homologous end joining during CSR. A non-canonical scaffold function of UNG, but not the catalytic activity, can be attributed to the recruitment of essential repair proteins associated with the error-free repair during SHM, and the end joining during CSR. PMID:24994819

  20. Mediator facilitates transcriptional activation and dynamic long-range contacts at the IgH locus during class switch recombination.

    PubMed

    Thomas-Claudepierre, Anne-Sophie; Robert, Isabelle; Rocha, Pedro P; Raviram, Ramya; Schiavo, Ebe; Heyer, Vincent; Bonneau, Richard; Luo, Vincent M; Reddy, Janardan K; Borggrefe, Tilman; Skok, Jane A; Reina-San-Martin, Bernardo

    2016-03-01

    Immunoglobulin (Ig) class switch recombination (CSR) is initiated by the transcription-coupled recruitment of activation-induced cytidine deaminase (AID) to Ig switch regions (S regions). During CSR, the IgH locus undergoes dynamic three-dimensional structural changes in which promoters, enhancers, and S regions are brought to close proximity. Nevertheless, little is known about the underlying mechanisms. In this study, we show that Med1 and Med12, two subunits of the mediator complex implicated in transcription initiation and long-range enhancer/promoter loop formation, are dynamically recruited to the IgH locus enhancers and the acceptor regions during CSR and that their knockdown in CH12 cells results in impaired CSR. Furthermore, we show that conditional inactivation of Med1 in B cells results in defective CSR and reduced acceptor S region transcription. Finally, we show that in B cells undergoing CSR, the dynamic long-range contacts between the IgH enhancers and the acceptor regions correlate with Med1 and Med12 binding and that they happen at a reduced frequency in Med1-deficient B cells. Our results implicate the mediator complex in the mechanism of CSR and are consistent with a model in which mediator facilitates the long-range contacts between S regions and the IgH locus enhancers during CSR and their transcriptional activation. PMID:26903242

  1. A regulatory role for the cohesin loader NIPBL in nonhomologous end joining during immunoglobulin class switch recombination

    PubMed Central

    Enervald, Elin; Du, Likun; Visnes, Torkild; Björkman, Andrea; Lindgren, Emma; Wincent, Josephine; Borck, Guntram; Colleaux, Laurence; Cormier-Daire, Valerie; van Gent, Dik C.; Pie, Juan; Puisac, Beatriz; de Miranda, Noel FCC; Kracker, Sven; de Villartay, Jean-Pierre; Durandy, Anne; Schoumans, Jacqueline

    2013-01-01

    DNA double strand breaks (DSBs) are mainly repaired via homologous recombination (HR) or nonhomologous end joining (NHEJ). These breaks pose severe threats to genome integrity but can also be necessary intermediates of normal cellular processes such as immunoglobulin class switch recombination (CSR). During CSR, DSBs are produced in the G1 phase of the cell cycle and are repaired by the classical NHEJ machinery. By studying B lymphocytes derived from patients with Cornelia de Lange Syndrome, we observed a strong correlation between heterozygous loss-of-function mutations in the gene encoding the cohesin loading protein NIPBL and a shift toward the use of an alternative, microhomology-based end joining during CSR. Furthermore, the early recruitment of 53BP1 to DSBs was reduced in the NIPBL-deficient patient cells. Association of NIPBL deficiency and impaired NHEJ was also observed in a plasmid-based end-joining assay and a yeast model system. Our results suggest that NIPBL plays an important and evolutionarily conserved role in NHEJ, in addition to its canonical function in sister chromatid cohesion and its recently suggested function in HR. PMID:24145515

  2. APE1 is dispensable for S-region cleavage but required for its repair in class switch recombination

    PubMed Central

    Xu, Jianliang; Husain, Afzal; Hu, Wenjun; Honjo, Tasuku; Kobayashi, Maki

    2014-01-01

    Activation-induced cytidine deaminase (AID) is essential for antibody diversification, namely somatic hypermutation (SHM) and class switch recombination (CSR). The deficiency of apurinic/apyrimidinic endonuclease 1 (Ape1) in CH12F3-2A B cells reduces CSR to ∼20% of wild-type cells, whereas the effect of APE1 loss on SHM has not been examined. Here we show that, although APE1’s endonuclease activity is important for CSR, it is dispensable for SHM as well as IgH/c-myc translocation. Importantly, APE1 deficiency did not show any defect in AID-induced S-region break formation, but blocked both the recruitment of repair protein Ku80 to the S region and the synapse formation between Sμ and Sα. Knockdown of end-processing factors such as meiotic recombination 11 homolog (MRE11) and carboxy-terminal binding protein (CtBP)-interacting protein (CtIP) further reduced the remaining CSR in Ape1-null CH12F3-2A cells. Together, our results show that APE1 is dispensable for SHM and AID-induced DNA breaks and may function as a DNA end-processing enzyme to facilitate the joining of broken ends during CSR. PMID:25404348

  3. Fanca deficiency reduces A/T transitions in somatic hypermutation and alters class switch recombination junctions in mouse B cells.

    PubMed

    Nguyen, Thuy Vy; Riou, Lydia; Aoufouchi, Saïd; Rosselli, Filippo

    2014-06-01

    Fanconi anemia is a rare genetic disorder that can lead to bone marrow failure, congenital abnormalities, and increased risk for leukemia and cancer. Cells with loss-of-function mutations in the FANC pathway are characterized by chromosome fragility, altered mutability, and abnormal regulation of the nonhomologous end-joining (NHEJ) pathway. Somatic hypermutation (SHM) and immunoglobulin (Ig) class switch recombination (CSR) enable B cells to produce high-affinity antibodies of various isotypes. Both processes are initiated after the generation of dG:dU mismatches by activation-induced cytidine deaminase. Whereas SHM involves an error-prone repair process that introduces novel point mutations into the Ig gene, the mismatches generated during CSR are processed to create double-stranded breaks (DSBs) in DNA, which are then repaired by the NHEJ pathway. As several lines of evidence suggest a possible role for the FANC pathway in SHM and CSR, we analyzed both processes in B cells derived from Fanca(-/-) mice. Here we show that Fanca is required for the induction of transition mutations at A/T residues during SHM and that despite globally normal CSR function in splenic B cells, Fanca is required during CSR to stabilize duplexes between pairs of short microhomology regions, thereby impeding short-range recombination downstream of DSB formation. PMID:24799500

  4. Regulation of immunoglobulin class-switch recombination: choreography of noncoding transcription, targeted DNA deamination, and long-range DNA repair.

    PubMed

    Matthews, Allysia J; Zheng, Simin; DiMenna, Lauren J; Chaudhuri, Jayanta

    2014-01-01

    Upon encountering antigens, mature IgM-positive B lymphocytes undergo class-switch recombination (CSR) wherein exons encoding the default Cμ constant coding gene segment of the immunoglobulin (Ig) heavy-chain (Igh) locus are excised and replaced with a new constant gene segment (referred to as "Ch genes", e.g., Cγ, Cɛ, or Cα). The B cell thereby changes from expressing IgM to one producing IgG, IgE, or IgA, with each antibody isotype having a different effector function during an immune reaction. CSR is a DNA deletional-recombination reaction that proceeds through the generation of DNA double-strand breaks (DSBs) in repetitive switch (S) sequences preceding each Ch gene and is completed by end-joining between donor Sμ and acceptor S regions. CSR is a multistep reaction requiring transcription through S regions, the DNA cytidine deaminase AID, and the participation of several general DNA repair pathways including base excision repair, mismatch repair, and classical nonhomologous end-joining. In this review, we discuss our current understanding of how transcription through S regions generates substrates for AID-mediated deamination and how AID participates not only in the initiation of CSR but also in the conversion of deaminated residues into DSBs. Additionally, we review the multiple processes that regulate AID expression and facilitate its recruitment specifically to the Ig loci, and how deregulation of AID specificity leads to oncogenic translocations. Finally, we summarize recent data on the potential role of AID in the maintenance of the pluripotent stem cell state during epigenetic reprogramming. PMID:24507154

  5. NF-κB binds to the immunoglobulin Sγ3 region in vivo during class switch recombination

    PubMed Central

    Wang, Lili; Wuerffel, Robert; Kenter, Amy L.

    2016-01-01

    Ig class switch recombination (CSR) is dependent upon the expression of activation-induced deaminase and targeted to specific isotypes by germ-line transcript expression and isotype-specific factors. NF-κB plays critical roles in multiple aspects of B cell biology and has been implicated in the mechanism of CSR by in vitro binding assays and altered S/S junctions derived from NF-κB p50-deficient mice. However, the pleiotropic contributions of NF-κB to gene expression in B cells has made discerning a direct role for NF-κB in CSR difficult. We now observe that binding of NF-κB components p50 and p65 is detected on Sγ3 in vivo following lipopolysaccharide (LPS) activation and repressed by LPS + IL–4, suggesting a direct role for this factor in CSR. In vivo footprinting confirms occupancy of a previously defined NF-κB recognition site in Sγ3 with the same temporal kinetics as found in the chromatin immunoprecipitation analysis. Binding of NF-κB components p50 and p65 was also detected on Sγ1 following B cell activation. H3 histone hyper acetylation at Sγ1 is strongly correlated with NF-κB binding, suggesting that NF-κB mediates chromatin remodeling in the Sγ3 and Sγ1 region. PMID:17109470

  6. S-S Synapsis during Class Switch Recombination Is Promoted by Distantly Located Transcriptional Elements and Activation-Induced Deaminase

    PubMed Central

    Wuerffel, Robert; Wang, Lili; Grigera, Fernando; Manis, John; Selsing, Erik; Perlot, Thomas; Alt, Frederick W.; Cogne, Michel; Pinaud, Eric; Kenter, Amy L.

    2016-01-01

    SUMMARY Molecular mechanisms underlying synapsis of activation-induced deaminase (AID)-targeted S regions during class switch recombination (CSR) are poorly understood. By using chromosome conformation capture techniques, we found that in B cells, the Eμ and 3′Eα enhancers were in close spatial proximity, forming a unique chromosomal loop configuration. B cell activation led to recruitment of the germline transcript (GLT) promoters to the Eμ:3′Eα complex in a cytokine-dependent fashion. This structure facilitated S-S synapsis because Sμ was proximal to Eμ and a downstream S region was corecruited with the targeted GLT promoter to Eμ:3′Eα. We propose that GLT promoter association with the Eμ:3′Eα complex creates an architectural scaffolding that promotes S-S synapsis during CSR and that these interactions are stabilized by AID. Thus, the S-S synaptosome is formed as a result of the self-organizing transcription system that regulates GLT expression and may serve to guard against spurious chromosomal translocations. PMID:17980632

  7. Differential regulation of S-region hypermutation and class-switch recombination by noncanonical functions of uracil DNA glycosylase

    PubMed Central

    Yousif, Ashraf S.; Stanlie, Andre; Mondal, Samiran; Honjo, Tasuku; Begum, Nasim A.

    2014-01-01

    Activation-induced cytidine deaminase (AID) is essential to class-switch recombination (CSR) and somatic hypermutation (SHM) in both V region SHM and S region SHM (s-SHM). Uracil DNA glycosylase (UNG), a member of the base excision repair (BER) complex, is required for CSR. Strikingly, however, UNG deficiency causes augmentation of SHM, suggesting involvement of distinct functions of UNG in SHM and CSR. Here, we show that noncanonical scaffold functions of UNG regulate s-SHM negatively and CSR positively. The s-SHM suppressive function of UNG is attributed to the recruitment of faithful BER components at the cleaved DNA locus, with competition against error-prone polymerases. By contrast, the CSR-promoting function of UNG enhances AID-dependent S-S synapse formation by recruiting p53-binding protein 1 and DNA-dependent protein kinase, catalytic subunit. Several loss-of-catalysis mutants of UNG discriminated CSR-promoting activity from s-SHM suppressive activity. Taken together, the noncanonical function of UNG regulates the steps after AID-induced DNA cleavage: error-prone repair suppression in s-SHM and end-joining promotion in CSR. PMID:24591630

  8. Differential regulation of S-region hypermutation and class-switch recombination by noncanonical functions of uracil DNA glycosylase.

    PubMed

    Yousif, Ashraf S; Stanlie, Andre; Mondal, Samiran; Honjo, Tasuku; Begum, Nasim A

    2014-03-18

    Activation-induced cytidine deaminase (AID) is essential to class-switch recombination (CSR) and somatic hypermutation (SHM) in both V region SHM and S region SHM (s-SHM). Uracil DNA glycosylase (UNG), a member of the base excision repair (BER) complex, is required for CSR. Strikingly, however, UNG deficiency causes augmentation of SHM, suggesting involvement of distinct functions of UNG in SHM and CSR. Here, we show that noncanonical scaffold functions of UNG regulate s-SHM negatively and CSR positively. The s-SHM suppressive function of UNG is attributed to the recruitment of faithful BER components at the cleaved DNA locus, with competition against error-prone polymerases. By contrast, the CSR-promoting function of UNG enhances AID-dependent S-S synapse formation by recruiting p53-binding protein 1 and DNA-dependent protein kinase, catalytic subunit. Several loss-of-catalysis mutants of UNG discriminated CSR-promoting activity from s-SHM suppressive activity. Taken together, the noncanonical function of UNG regulates the steps after AID-induced DNA cleavage: error-prone repair suppression in s-SHM and end-joining promotion in CSR. PMID:24591630

  9. Exosomes Derived from Burkitt’s Lymphoma Cell Lines Induce Proliferation, Differentiation, and Class-Switch Recombination in B Cells

    PubMed Central

    Gutzeit, Cindy; Nagy, Noemi; Gentile, Maurizio; Lyberg, Katarina; Gumz, Janine; Vallhov, Helen; Puga, Irene; Klein, Eva; Gabrielsson, Susanne; Cerutti, Andrea; Scheynius, Annika

    2014-01-01

    Exosomes, nano-sized membrane vesicles, are released by various cells and are found in many human body fluids. They are active players in intercellular communication and have immune-suppressive, immune-regulatory, and immune-stimulatory functions. EBV is a ubiquitous human herpesvirus that is associated with various lymphoid and epithelial malignancies. EBV infection of B cells in vitro induces the release of exosomes that harbor the viral latent membrane protein 1 (LMP1). LMP1 per se mimics CD40 signaling and induces proliferation of B lymphocytes and T cell–independent class-switch recombination. Constitutive LMP1 signaling within B cells is blunted through the shedding of LMP1 via exosomes. In this study, we investigated the functional effect of exosomes derived from the DG75 Burkitt’s lymphoma cell line and its sublines (LMP1 transfected and EBV infected), with the hypothesis that they might mimic exosomes released during EBV-associated diseases. We show that exosomes released during primary EBV infection of B cells harbored LMP1, and similar levels were detected in exosomes from LMP1-transfected DG75 cells. DG75 exosomes efficiently bound to human B cells within PBMCs and were internalized by isolated B cells. In turn, this led to proliferation, induction of activation-induced cytidine deaminase, and the production of circle and germline transcripts for IgG1 in B cells. Finally, exosomes harboring LMP1 enhanced proliferation and drove B cell differentiation toward a plasmablast-like phenotype. In conclusion, our results suggest that exosomes released from EBV-infected B cells have a stimulatory capacity and interfere with the fate of human B cells. PMID:24829410

  10. Non-canonical uracil processing in DNA gives rise to double-strand breaks and deletions: relevance to class switch recombination.

    PubMed

    Bregenhorn, Stephanie; Kallenberger, Lia; Artola-Borán, Mariela; Peña-Diaz, Javier; Jiricny, Josef

    2016-04-01

    During class switch recombination (CSR), antigen-stimulated B-cells rearrange their immunoglobulin constant heavy chain (CH) loci to generate antibodies with different effector functions. CSR is initiated by activation-induced deaminase (AID), which converts cytosines in switch (S) regions, repetitive sequences flanking the CH loci, to uracils. Although U/G mispairs arising in this way are generally efficiently repaired to C/Gs by uracil DNA glycosylase (UNG)-initiated base excision repair (BER), uracil processing in S-regions of activated B-cells occasionally gives rise to double strand breaks (DSBs), which trigger CSR. Surprisingly, genetic experiments revealed that CSR is dependent not only on AID and UNG, but also on mismatch repair (MMR). To elucidate the role of MMR in CSR, we studied the processing of uracil-containing DNA substrates in extracts of MMR-proficient and -deficient human cells, as well as in a system reconstituted from recombinant BER and MMR proteins. Here, we show that the interplay of these repair systems gives rise to DSBs in vitro and to genomic deletions and mutations in vivo, particularly in an S-region sequence. Our findings further suggest that MMR affects pathway choice in DSB repair. Given its amenability to manipulation, our system represents a powerful tool for the molecular dissection of CSR. PMID:26743004

  11. Non-canonical uracil processing in DNA gives rise to double-strand breaks and deletions: relevance to class switch recombination

    PubMed Central

    Bregenhorn, Stephanie; Kallenberger, Lia; Artola-Borán, Mariela; Peña-Diaz, Javier; Jiricny, Josef

    2016-01-01

    During class switch recombination (CSR), antigen-stimulated B-cells rearrange their immunoglobulin constant heavy chain (CH) loci to generate antibodies with different effector functions. CSR is initiated by activation-induced deaminase (AID), which converts cytosines in switch (S) regions, repetitive sequences flanking the CH loci, to uracils. Although U/G mispairs arising in this way are generally efficiently repaired to C/Gs by uracil DNA glycosylase (UNG)-initiated base excision repair (BER), uracil processing in S-regions of activated B-cells occasionally gives rise to double strand breaks (DSBs), which trigger CSR. Surprisingly, genetic experiments revealed that CSR is dependent not only on AID and UNG, but also on mismatch repair (MMR). To elucidate the role of MMR in CSR, we studied the processing of uracil-containing DNA substrates in extracts of MMR-proficient and –deficient human cells, as well as in a system reconstituted from recombinant BER and MMR proteins. Here, we show that the interplay of these repair systems gives rise to DSBs in vitro and to genomic deletions and mutations in vivo, particularly in an S-region sequence. Our findings further suggest that MMR affects pathway choice in DSB repair. Given its amenability to manipulation, our system represents a powerful tool for the molecular dissection of CSR. PMID:26743004

  12. Microbiota regulate the ability of lung dendritic cells to induce IgA class-switch recombination and generate protective gastrointestinal immune responses

    PubMed Central

    Ruane, Darren; Chorny, Alejo; Lee, Haekyung; Faith, Jeremiah; Pandey, Gaurav; Shan, Meimei; Simchoni, Noa; Rahman, Adeeb; Garg, Aakash; Weinstein, Erica G.; Oropallo, Michael; Gaylord, Michelle; Ungaro, Ryan; Cunningham-Rundles, Charlotte; Alexandropoulos, Konstantina; Mucida, Daniel; Merad, Miriam; Cerutti, Andrea

    2016-01-01

    Protective immunoglobulin A (IgA) responses to oral antigens are usually orchestrated by gut dendritic cells (DCs). Here, we show that lung CD103+ and CD24+CD11b+ DCs induced IgA class-switch recombination (CSR) by activating B cells through T cell–dependent or –independent pathways. Compared with lung DCs (LDC), lung CD64+ macrophages had decreased expression of B cell activation genes and induced significantly less IgA production. Microbial stimuli, acting through Toll-like receptors, induced transforming growth factor-β (TGF-β) production by LDCs and exerted a profound influence on LDC-mediated IgA CSR. After intranasal immunization with inactive cholera toxin (CT), LDCs stimulated retinoic acid–dependent up-regulation of α4β7 and CCR9 gut-homing receptors on local IgA-expressing B cells. Migration of these B cells to the gut resulted in IgA-mediated protection against an oral challenge with active CT. However, in germ-free mice, the levels of LDC-induced, CT–specific IgA in the gut are significantly reduced. Herein, we demonstrate an unexpected role of the microbiota in modulating the protective efficacy of intranasal vaccination through their effect on the IgA class-switching function of LDCs. PMID:26712806

  13. Microbiota regulate the ability of lung dendritic cells to induce IgA class-switch recombination and generate protective gastrointestinal immune responses.

    PubMed

    Ruane, Darren; Chorny, Alejo; Lee, Haekyung; Faith, Jeremiah; Pandey, Gaurav; Shan, Meimei; Simchoni, Noa; Rahman, Adeeb; Garg, Aakash; Weinstein, Erica G; Oropallo, Michael; Gaylord, Michelle; Ungaro, Ryan; Cunningham-Rundles, Charlotte; Alexandropoulos, Konstantina; Mucida, Daniel; Merad, Miriam; Cerutti, Andrea; Mehandru, Saurabh

    2016-01-11

    Protective immunoglobulin A (IgA) responses to oral antigens are usually orchestrated by gut dendritic cells (DCs). Here, we show that lung CD103(+) and CD24(+)CD11b(+) DCs induced IgA class-switch recombination (CSR) by activating B cells through T cell-dependent or -independent pathways. Compared with lung DCs (LDC), lung CD64(+) macrophages had decreased expression of B cell activation genes and induced significantly less IgA production. Microbial stimuli, acting through Toll-like receptors, induced transforming growth factor-β (TGF-β) production by LDCs and exerted a profound influence on LDC-mediated IgA CSR. After intranasal immunization with inactive cholera toxin (CT), LDCs stimulated retinoic acid-dependent up-regulation of α4β7 and CCR9 gut-homing receptors on local IgA-expressing B cells. Migration of these B cells to the gut resulted in IgA-mediated protection against an oral challenge with active CT. However, in germ-free mice, the levels of LDC-induced, CT-specific IgA in the gut are significantly reduced. Herein, we demonstrate an unexpected role of the microbiota in modulating the protective efficacy of intranasal vaccination through their effect on the IgA class-switching function of LDCs. PMID:26712806

  14. Murine germinal center B cells require functional Fms-like tyrosine kinase 3 signaling for IgG1 class-switch recombination.

    PubMed

    Svensson, Mattias N D; Andersson, Karin M E; Wasén, Caroline; Erlandsson, Malin C; Nurkkala-Karlsson, Merja; Jonsson, Ing-Marie; Brisslert, Mikael; Bemark, Mats; Bokarewa, Maria I

    2015-12-01

    Switched antibody classes are important for efficient immune responses. Aberrant antibody production to otherwise harmless antigens may result in autoimmunity. The protein kinase fms-like tyrosine kinase 3 receptor (Flt3) has an important role during early B-cell development, but the role of Flt3 in peripheral B cells has not been assessed before. Herein we describe a previously unappreciated role for Flt3 in IgG1 class-switch recombination (CSR) and production. We show that Flt3 is reexpressed on B-cell lymphoma 6(+) germinal center B cells in vivo and following LPS activation of peripheral B cells in vitro. Absence of Flt3 signaling in Flt3 ligand-deficient mice results in impaired IgG1 CSR and accumulation of IgM-secreting plasma cells. On activated B cells, Flt3 is coexpressed and functions in synergy with the common-gamma chain receptor family. B cells from Flt3 ligand-deficient mice have impaired IL-4R signaling, with reduced phosphorylation of signal transducer and activator of transcription (Stat) 6, and demonstrate a failure to initiate CSR to IgG1 with low expression of γ1 germ-line transcripts, resulting in impaired IgG1 production. Thus, functional synergy between Flt3 and IL-4R signaling is critical for Stat-mediated regulation of sterile γ1 germ-line transcripts and CSR to IgG1. PMID:26627255

  15. Staggered AID-dependent DNA double strand breaks are the predominant DNA lesions targeted to S mu in Ig class switch recombination.

    PubMed

    Rush, James S; Fugmann, Sebastian D; Schatz, David G

    2004-04-01

    Class switch recombination (CSR) is the process whereby B cells alter the effector properties of their Ig molecules. Whilst much is known about the cellular regulation of this process, many of the molecular details remain elusive. Recent evidence suggests that CSR involves blunt DNA double strand breaks (dsbs), and that formation of these dsbs requires the function of the activation-induced cytidine deaminase (AID). We sought to characterize the structural properties and kinetics of induction of the DNA lesions associated with CSR. Using ligation-mediated PCR, we found that AID-dependent DNA dsbs were specifically induced in the S mu region of murine B cells stimulated to undergo CSR. While blunt dsbs were detected, they were only a minor species, with staggered breaks being more than an order of magnitude more abundant. In addition, these breaks could be detected at equal frequency at upstream and downstream portions of S mu, and were induced prior to expression of newly switched isotypes. Collectively, these results provide direct evidence that staggered, S mu-targeted AID-dependent dsbs are the predominant DNA lesion associated with CSR, with important implications for the mechanisms by which CSR DNA lesions are made and processed. PMID:15039385

  16. Elucidation of the enigmatic IgD class-switch recombination via germline deletion of the IgH 3′ regulatory region

    PubMed Central

    Rouaud, Pauline; Saintamand, Alexis; Saad, Faten; Carrion, Claire; Lecardeur, Sandrine; Cogné, Michel

    2014-01-01

    Classical class-switch recombination (cCSR) substitutes the Cμ gene with Cγ, Cε, or Cα, thereby generating IgG, IgE, or IgA classes, respectively. This activation-induced deaminase (AID)–driven process is controlled by the IgH 3′ regulatory region (3′RR). Regulation of rare IgD CSR events has been enigmatic. We show that μδCSR occurs in mouse mesenteric lymph node (MLN) B cells and is AID-dependent. AID attacks differ from those in cCSR because they are not accompanied by extensive somatic hypermutation (SHM) of targeted regions and because repaired junctions exhibit features of the alternative end-joining (A-EJ) pathway. In contrast to cCSR and SHM, μδCSR is 3′RR-independent, as its absence affects neither breakpoint locations in Sμ- and Sδ-like (σδ) nor mutation patterns at Sμ-σδ junctions. Although mutations occur in the immediate proximity of the μδ junctions, SHM is absent distal to the junctions within both Sμ and rearranged VDJ regions. In conclusion, μδCSR is active in MLNs, occurs independently of 3′RR-driven assembly, and is even dramatically increased in 3′RR-deficient mice, further showing that its regulation differs from cCSR. PMID:24752300

  17. Elucidation of the enigmatic IgD class-switch recombination via germline deletion of the IgH 3' regulatory region.

    PubMed

    Rouaud, Pauline; Saintamand, Alexis; Saad, Faten; Carrion, Claire; Lecardeur, Sandrine; Cogné, Michel; Denizot, Yves

    2014-05-01

    Classical class-switch recombination (cCSR) substitutes the Cμ gene with Cγ, Cε, or Cα, thereby generating IgG, IgE, or IgA classes, respectively. This activation-induced deaminase (AID)-driven process is controlled by the IgH 3' regulatory region (3'RR). Regulation of rare IgD CSR events has been enigmatic. We show that μδCSR occurs in mouse mesenteric lymph node (MLN) B cells and is AID-dependent. AID attacks differ from those in cCSR because they are not accompanied by extensive somatic hypermutation (SHM) of targeted regions and because repaired junctions exhibit features of the alternative end-joining (A-EJ) pathway. In contrast to cCSR and SHM, μδCSR is 3'RR-independent, as its absence affects neither breakpoint locations in Sμ- and Sδ-like (σ(δ)) nor mutation patterns at Sμ-σ(δ) junctions. Although mutations occur in the immediate proximity of the μδ junctions, SHM is absent distal to the junctions within both Sμ and rearranged VDJ regions. In conclusion, μδCSR is active in MLNs, occurs independently of 3'RR-driven assembly, and is even dramatically increased in 3'RR-deficient mice, further showing that its regulation differs from cCSR. PMID:24752300

  18. Hotspots for Vitamin-Steroid-Thyroid Hormone Response Elements Within Switch Regions of Immunoglobulin Heavy Chain Loci Predict a Direct Influence of Vitamins and Hormones on B Cell Class Switch Recombination.

    PubMed

    Hurwitz, Julia L; Penkert, Rhiannon R; Xu, Beisi; Fan, Yiping; Partridge, Janet F; Maul, Robert W; Gearhart, Patricia J

    2016-03-01

    Vitamin A deficiencies are common throughout the world and have a significant negative influence on immune protection against viral infections. Mouse models demonstrate that the production of IgA, a first line of defense against viruses at mucosal sites, is inhibited in the context of vitamin A deficiency. In vitro, the addition of vitamin A to activated B cells can enhance IgA expression, but downregulate IgE. Previous reports have demonstrated that vitamin A modifies cytokine patterns, and in so doing may influence antibody isotype expression by an indirect mechanism. However, we have now discovered hundreds of potential response elements among Sμ, Sɛ, and Sα switch sites within immunoglobulin heavy chain loci. These hotspots appear in both mouse and human loci and include targets for vitamin receptors and related proteins (e.g., estrogen receptors) in the nuclear receptor superfamily. Full response elements with direct repeats are relatively infrequent or absent in Sγ regions although half-sites are present. Based on these results, we pose a hypothesis that nuclear receptors have a direct effect on the immunoglobulin heavy chain class switch recombination event. We propose that vitamin A may alter S site accessibility to activation-induced deaminase and nonhomologous end-joining machinery, thereby influencing the isotype switch, antibody production, and protection against viral infections at mucosal sites. PMID:26741514

  19. The SAGA Deubiquitination Module Promotes DNA Repair and Class Switch Recombination through ATM and DNAPK-Mediated γH2AX Formation.

    PubMed

    Ramachandran, Shaliny; Haddad, Dania; Li, Conglei; Le, Michael X; Ling, Alexanda K; So, Clare C; Nepal, Rajeev M; Gommerman, Jennifer L; Yu, Kefei; Ketela, Troy; Moffat, Jason; Martin, Alberto

    2016-05-17

    Class switch recombination (CSR) requires activation-induced deaminase (AID) to instigate double-stranded DNA breaks at the immunoglobulin locus. DNA breaks activate the DNA damage response (DDR) by inducing phosphorylation of histone H2AX followed by non-homologous end joining (NHEJ) repair. We carried out a genome-wide screen to identify CSR factors. We found that Usp22, Eny2, and Atxn7, members of the Spt-Ada-Gcn5-acetyltransferase (SAGA) deubiquitination module, are required for deubiquitination of H2BK120ub following DNA damage, are critical for CSR, and function downstream of AID. The SAGA deubiquitinase activity was required for optimal irradiation-induced γH2AX formation, and failure to remove H2BK120ub inhibits ATM- and DNAPK-induced γH2AX formation. Consistent with this effect, these proteins were found to function upstream of various double-stranded DNA repair pathways. This report demonstrates that deubiquitination of histone H2B impacts the early stages of the DDR and is required for the DNA repair phase of CSR. PMID:27160905

  20. Toll-like Receptors and B-cell Receptors Synergize to Induce Immunoglobulin Class Switch DNA Recombination: Relevance to Microbial Antibody Responses

    PubMed Central

    Pone, Egest J.; Zan, Hong; Zhang, Jinsong; Al-Qahtani, Ahmed; Xu, Zhenming; Casali, Paolo

    2011-01-01

    Differentiation of naïve B cells, including immunoglobulin (Ig) class switch DNA recombination (CSR), is critical for the immune response and depends on the extensive integration of signals from the B cell receptor (BCR), tumor necrosis factor (TNF) receptor family members, Toll-like receptors (TLRs) and cytokine receptors. TLRs and BCR synergize to induce CSR in T cell-dependent and T cell-independent antibody responses to microbial pathogens. BCR triggering together with simultaneous endosomal TLR engagement leads to enhanced B cell differentiation and antibody responses. The requirement of both BCR and TLR engagement would ensure appropriate antigen-specific activation in an infection. Co-stimulation of TLRs and BCR likely plays a significant role in anti-microbial antibody responses to contain pathogen loads until the T cell-dependent antibody responses peak. Furthermore, the temporal sequence of different signals is also critical for optimal B cell responses, as exemplified by the activation of B cells by initial TLR engagement, leading to the upregulation of co-stimulatory CD80 and MHC-II receptors, which, in turn, result in more efficient interactions with T cells, thereby enhancing the germinal center (GC) reaction and antibody affinity maturation. Overall, BCR and TLR stimulation and the integration with signals from the pathogen or immune cells and their products, determine the ensuing B cell antibody response. PMID:20370617

  1. Class E/F switching power amplifiers

    NASA Technical Reports Server (NTRS)

    Hajimiri, Seyed-Ali (Inventor); Aoki, Ichiro (Inventor); Rutledge, David B. (Inventor); Kee, Scott David (Inventor)

    2004-01-01

    The present invention discloses a new family of switching amplifier classes called class E/F amplifiers. These amplifiers are generally characterized by their use of the zero-voltage-switching (ZVS) phase correction technique to eliminate of the loss normally associated with the inherent capacitance of the switching device as utilized in class-E amplifiers, together with a load network for improved voltage and current wave-shaping by presenting class-F.sup.-1 impedances at selected overtones and class-E impedances at the remaining overtones. The present invention discloses a several topologies and specific circuit implementations for achieving such performance.

  2. Lineage tracing of human B cells reveals the in vivo landscape of human antibody class switching.

    PubMed

    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. PMID:27481325

  3. B cell TLR1/2, TLR4, TLR7 and TLR9 interact in induction of class switch DNA recombination: modulation by BCR and CD40, and relevance to T-independent antibody responses.

    PubMed

    Pone, Egest J; Lou, Zheng; Lam, Tonika; Greenberg, Milton L; Wang, Rui; Xu, Zhenming; Casali, Paolo

    2015-02-01

    Ig class switch DNA recombination (CSR) in B cells is crucial to the maturation of antibody responses. It requires IgH germline IH-CH transcription and expression of AID, both of which are induced by engagement of CD40 or dual engagement of a Toll-like receptor (TLR) and B cell receptor (BCR). Here, we have addressed cross-regulation between two different TLRs or between a TLR and CD40 in CSR induction by using a B cell stimulation system involving lipopolysaccharides (LPS). LPS-mediated long-term primary class-switched antibody responses and memory-like antibody responses in vivo and induced generation of class-switched B cells and plasma cells in vitro. Consistent with the requirement for dual TLR and BCR engagement in CSR induction, LPS, which engages TLR4 through its lipid A moiety, triggered cytosolic Ca2+ flux in B cells through its BCR-engaging polysaccharidic moiety. In the presence of BCR crosslinking, LPS synergized with a TLR1/2 ligand (Pam3CSK4) in CSR induction, but much less efficiently with a TLR7 (R-848) or TLR9 (CpG) ligand. In the absence of BCR crosslinking, R-848 and CpG, which per se induced marginal CSR, virtually abrogated CSR to IgG1, IgG2a, IgG2b, IgG3 and/or IgA, as induced by LPS or CD154 (CD40 ligand) plus IL-4, IFN-γ or TGF-β, and reduced secretion of class-switched Igs, without affecting B cell proliferation or IgM expression. The CSR inhibition by TLR9 was associated with the reduction in AID expression and/or IgH germline IH-S-CH transcription, and required co-stimulation of B cells by CpG with LPS or CD154. Unexpectedly, B cells also failed to undergo CSR or plasma cell differentiation when co-stimulated by LPS and CD154. Overall, by addressing the interaction of TLR1/2, TLR4, TLR7 and TLR9 in the induction of CSR and modulation of TLR-dependent CSR by BCR and CD40, our study suggests the complexity of how different stimuli cross-regulate an important B cell differentiation process and an important role of TLRs in inducing

  4. Homologous Elements hs3a and hs3b in the 3′ Regulatory Region of the Murine Immunoglobulin Heavy Chain (Igh) Locus Are Both Dispensable for Class-switch Recombination*

    PubMed Central

    Yan, Yi; Pieretti, Joyce; Ju, Zhongliang; Wei, Shiniu; Christin, John R.; Bah, Fatmata; Birshtein, Barbara K.; Eckhardt, Laurel A.

    2011-01-01

    Immunoglobulin heavy chain (IgH) genes are formed, tested, and modified to yield diverse, specific, and high affinity antibody responses to antigen. The processes involved must be regulated, however, to avoid unintended damage to chromosomes. The 3′ regulatory region of the Igh locus plays a major role in regulating class-switch recombination (CSR), the process by which antibody effector functions are modified during an immune response. Loss of all known enhancer-like elements in this region dramatically impairs CSR, but individual element deletions have no effect on this process. In the present study, we explored the hypothesis that an underlying functional redundancy in the homologous elements hs3a and hs3b was masking the importance of either element to CSR. Several transgenic mouse lines were generated, each carrying a bacterial artificial chromosome transgene that mimicked Igh locus structure but in which hs3a was missing and hs3b was flanked by loxP sites. Matings to Cyclization Recombination Enzyme-expressing mice established “pairs” of lines that differed only in the presence or absence of hs3b. Remarkably, CSR remained robust in the absence of both hs3a and hs3b, suggesting that the remaining two elements of the 3′ regulatory region, hs1.2 and hs4, although individually dispensable for CSR, are, together, sufficient to support CSR. PMID:21673112

  5. Combinatorial H3K9acS10ph histone modification in IgH locus S regions targets 14-3-3 adaptors and AID to specify antibody class-switch DNA recombination.

    PubMed

    Li, Guideng; White, Clayton A; Lam, Tonika; Pone, Egest J; Tran, Daniel C; Hayama, Ken L; Zan, Hong; Xu, Zhenming; Casali, Paolo

    2013-11-14

    Class-switch DNA recombination (CSR) is central to the antibody response, in that it changes the immunoglobulin heavy chain (IgH) constant region, thereby diversifying biological effector functions of antibodies. The activation-induced cytidine deaminase (AID)-centered CSR machinery excises and rejoins DNA between an upstream (donor) and a downstream (acceptor) S region, which precede the respective constant region DNA. AID is stabilized on S regions by 14-3-3 adaptors. These adaptors display a high affinity for 5'-AGCT-3' repeats, which recur in all S regions. However, how 14-3-3, AID, and the CSR machinery target exclusively the donor and acceptor S regions is poorly understood. Here, we show that histone methyltransferases and acetyltransferases are induced by CD40 or Toll-like receptor signaling and catalyze H3K4me3 and H3K9ac/K14ac histone modifications, which are enriched in S regions but do not specify the S region targets of CSR. By contrast, the combinatorial H3K9acS10ph modification specifically marks the S regions set to recombine and directly recruits 14-3-3 adaptors for AID stabilization there. Inhibition of the enzymatic activity of GCN5 and PCAF histone acetyltransferases reduces H3K9acS10ph in S regions, 14-3-3 and AID stabilization, and CSR. Thus, H3K9acS10ph is a histone code that is "written" specifically in S regions and is "read" by 14-3-3 adaptors to target AID for CSR as an important biological outcome. PMID:24209747

  6. Estrogen receptors bind to and activate the HOXC4/HoxC4 promoter to potentiate HoxC4-mediated activation-induced cytosine deaminase induction, immunoglobulin class switch DNA recombination, and somatic hypermutation.

    PubMed

    Mai, Thach; Zan, Hong; Zhang, Jinsong; Hawkins, J Seth; Xu, Zhenming; Casali, Paolo

    2010-11-26

    Estrogen enhances antibody and autoantibody responses through yet to be defined mechanisms. It has been suggested that estrogen up-regulates the expression of activation-induced cytosine deaminase (AID), which is critical for antibody class switch DNA recombination (CSR) and somatic hypermutation (SHM), through direct activation of this gene. AID, as we have shown, is induced by the HoxC4 homeodomain transcription factor, which binds to a conserved HoxC4/Oct site in the AICDA/Aicda promoter. Here we show that estrogen-estrogen receptor (ER) complexes do not directly activate the AID gene promoter in B cells undergoing CSR. Rather, they bind to three evolutionarily conserved and cooperative estrogen response elements (EREs) we identified in the HOXC4/HoxC4 promoter. By binding to these EREs, ERs synergized with CD154 or LPS and IL-4 signaling to up-regulate HoxC4 expression, thereby inducing AID and CSR without affecting B cell proliferation or plasmacytoid differentiation. Estrogen administration in vivo significantly potentiated CSR and SHM in the specific antibody response to the 4-hydroxy-3-nitrophenylacetyl hapten conjugated with chicken γ-globulin. Ablation of HoxC4 (HoxC4(-/-)) abrogated the estrogen-mediated enhancement of AID gene expression and decreased CSR and SHM. Thus, estrogen enhances AID expression by activating the HOXC4/HoxC4 promoter and inducing the critical AID gene activator, HoxC4. PMID:20855884

  7. 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

  8. Genome-Wide Analysis Reveals Selective Modulation of microRNAs and mRNAs by Histone Deacetylase Inhibitor in B Cells Induced to Undergo Class-Switch DNA Recombination and Plasma Cell Differentiation

    PubMed Central

    Shen, Tian; Sanchez, Helia N.; Zan, Hong; Casali, Paolo

    2015-01-01

    As we have suggested, epigenetic factors, such as microRNAs (miRNAs), can interact with genetic programs to regulate B cell functions, thereby informing antibody and autoantibody responses. We have shown that histone deacetylase (HDAC) inhibitors (HDI) inhibit the differentiation events critical to the maturation of the antibody response: class-switch DNA recombination (CSR), somatic hypermutation (SHM), and plasma cell differentiation, by modulating intrinsic B cell mechanisms. HDI repress the expression of AID and Blimp-1, which are critical for CSR/SHM and plasma cell differentiation, respectively, in mouse and human B cells by upregulating selected miRNAs that silenced AICDA/Aicda and PRDM1/Prdm1 mRNAs, as demonstrated by multiple qRT-PCRs (J Immunol 193:5933–5950, 2014). To further define the selectivity of HDI-mediated modulation of miRNA and gene expression, we performed genome-wide miRNA-Seq and mRNA-Seq analysis in B cells stimulated by LPS plus IL-4 and treated with HDI or nil. Consistent with what we have shown using qRT-PCR, these HDI-treated B cells displayed reduced expression of Aicda and Prdm1, and increased expression of miR-155, miR-181b, and miR-361, which target Aicda, and miR-23b, miR-30a, and miR-125b, which target Prdm1. In B cells induced to undergo CSR and plasma cell differentiation, about 23% of over 22,000 mRNAs analyzed were expressed at a significantly high copy number (more than 20 copies/cell). Only 18 (0.36%) of these highly expressed mRNAs, including Aicda, Prdm1, and Xbp1, were downregulated by HDI by 50% or more. Further, only 16 (0.30%) of the highly expressed mRNAs were upregulated (more than twofold) by HDI. The selectivity of HDI-mediated modulation of gene expression was emphasized by unchanged expression of the genes that are involved in regulation, targeting, or DNA repair processes of CSR, as well as unchanged expression of the genes encoding epigenetic regulators and factors that are important for cell signaling or

  9. Genome-Wide Analysis Reveals Selective Modulation of microRNAs and mRNAs by Histone Deacetylase Inhibitor in B Cells Induced to Undergo Class-Switch DNA Recombination and Plasma Cell Differentiation.

    PubMed

    Shen, Tian; Sanchez, Helia N; Zan, Hong; Casali, Paolo

    2015-01-01

    As we have suggested, epigenetic factors, such as microRNAs (miRNAs), can interact with genetic programs to regulate B cell functions, thereby informing antibody and autoantibody responses. We have shown that histone deacetylase (HDAC) inhibitors (HDI) inhibit the differentiation events critical to the maturation of the antibody response: class-switch DNA recombination (CSR), somatic hypermutation (SHM), and plasma cell differentiation, by modulating intrinsic B cell mechanisms. HDI repress the expression of AID and Blimp-1, which are critical for CSR/SHM and plasma cell differentiation, respectively, in mouse and human B cells by upregulating selected miRNAs that silenced AICDA/Aicda and PRDM1/Prdm1 mRNAs, as demonstrated by multiple qRT-PCRs (J Immunol 193:5933-5950, 2014). To further define the selectivity of HDI-mediated modulation of miRNA and gene expression, we performed genome-wide miRNA-Seq and mRNA-Seq analysis in B cells stimulated by LPS plus IL-4 and treated with HDI or nil. Consistent with what we have shown using qRT-PCR, these HDI-treated B cells displayed reduced expression of Aicda and Prdm1, and increased expression of miR-155, miR-181b, and miR-361, which target Aicda, and miR-23b, miR-30a, and miR-125b, which target Prdm1. In B cells induced to undergo CSR and plasma cell differentiation, about 23% of over 22,000 mRNAs analyzed were expressed at a significantly high copy number (more than 20 copies/cell). Only 18 (0.36%) of these highly expressed mRNAs, including Aicda, Prdm1, and Xbp1, were downregulated by HDI by 50% or more. Further, only 16 (0.30%) of the highly expressed mRNAs were upregulated (more than twofold) by HDI. The selectivity of HDI-mediated modulation of gene expression was emphasized by unchanged expression of the genes that are involved in regulation, targeting, or DNA repair processes of CSR, as well as unchanged expression of the genes encoding epigenetic regulators and factors that are important for cell signaling or

  10. Molecular requirements for immunoglobulin heavy chain constant region gene switch-recombination revealed with switch-substrate retroviruses.

    PubMed

    Ott, D E; Marcu, K B

    1989-01-01

    We have employed a retroviral vector, ZN(Smu/S gamma 2b)tk1, as a means of introducing immunoglobulin heavy chain (IgH) switch (S) region sequences into B cell lines to directly measure their switch-recombinase activities. In an earlier study, we demonstrated that retrovector Smu-S gamma 2b recombination events occurred in two thymidine kinase (tk)-negative murine pre-B cell lines (18-8 and 38B9) upon selection in bromodeoxyuridine (BUdR) media for the loss of an Htk gene inserted in between the vector's Smu and S gamma 2b sequences. Here we have used this assay system to show that the 300-18 murine pre-B cell line possesses a very high level of switch-recombinase activity (greater than 1 event in 2500 cells/generation) while a terminally differentiated, antibody-secreting hybridoma line (A39R 1.1) has no detectable recombinase activity. Both S mu and S gamma 2b segments are required for switch region-mediated deletions. Retrovectors harboring only an Smu segment or an Smu segment and a portion of the murine c-myc gene in place of S gamma 2b sequences were both non-recombinagenic in this assay system. Nucleotide sequence analysis of six retrovector S segment recombinants, recovered from ZN(Smu/S gamma 2b) tk1-infected 18-8 and 39B9 pre-B lines, did not reveal homology at their sites of recombination. We conclude that: (1) S segment repetitive sequences play an essential but indirect role in IgCH gene switch-recombination, which occurs by an illegitimate, non-homologous mechanism; (2) the c-myc gene is not a significant target for switch-recombination; and (3) since endogenous Smu and S gamma 2b rearrangements were not observed in populations and clones of pre-B cells expressing a high level of switch-recombinase activity, multiple factors (presumably contributed in part by the degree of S segment accessibility) in addition to S recombinase activity are required for CH class switching. PMID:2489045

  11. The Saccharomyces cerevisiae recombination enhancer biases recombination during interchromosomal mating-type switching but not in interchromosomal homologous recombination.

    PubMed Central

    Houston, Peter; Simon, Peter J; Broach, James R

    2004-01-01

    Haploid Saccharomyces can change mating type through HO-endonuclease cleavage of an expressor locus, MAT, followed by gene conversion using one of two repository loci, HML or HMR, as donor. The mating type of a cell dictates which repository locus is used as donor, with a cells using HML and alpha cells using HMR. This preference is established in part by RE, a locus on the left arm of chromosome III that activates the surrounding region, including HML, for recombination in a cells, an activity suppressed by alpha 2 protein in alpha cells. We have examined the ability of RE to stimulate different forms of interchromosomal recombination. We found that RE exerted an effect on interchromosomal mating-type switching and on intrachromosomal homologous recombination but not on interchromosomal homologous recombination. Also, even in the absence of RE, MAT alpha still influenced donor preference in interchromosomal mating-type switching, supporting a role of alpha 2 in donor preference independent of RE. These results suggest a model in which RE affects competition between productive and nonproductive recombination outcomes. In interchromosome gene conversion, RE enhances both productive and nonproductive pathways, whereas in intrachromosomal gene conversion and mating-type switching, RE enhances only the productive pathway. PMID:15082540

  12. Regulation and targeting of recombination in extrachromosomal substrates carrying immunoglobulin switch region sequences.

    PubMed Central

    Leung, H; Maizels, N

    1994-01-01

    We have used extrachromosomal substrates carrying immunoglobulin heavy-chain S mu and S gamma 3 switch region sequences to study activation and targeting of recombination by a transcriptional enhancer element. Substrates are transiently introduced into activated primary murine B cells, in which recombination involving S-region sequences deletes a conditionally lethal marker, and recombination is measured by transformation of Escherichia coli in the second step of the assay. Previously we found that as many as 25% of replicated substrates recombined during 40-h transfection of lipopolysaccharide (LPS)-stimulated primary cells and that efficient recombination was dependent on the presence of S-region sequences as well as a transcriptional activator region in the constructs (H. Leung and N. Maizels, Proc. Natl. Acad. Sci. USA 89:4154-4158, 1992). Here we show that recombination of the switch substrates is threefold more efficient in LPS-cultured primary B cells than in the T-cell line EL4; the activities responsible for switch substrate recombination thus appear to be more abundant or more active in cells which can carry out chromosomal switch recombination. We test the role of the transcriptional activator region and show that the immunoglobulin heavy-chain intron enhancer (E mu) alone stimulates recombination as well as E mu combined with a heavy-chain promoter and that mutations that diminish enhancer-dependent transcription 500-fold diminish recombinational activation less than 2-fold. These observations suggest that the enhancer stimulates recombination by a mechanism that does not depend on transcript production or that is insensitive to the level of transcript production over a very broad range. Furthermore, we find that E mu stimulates recombination when located either upstream or downstream of S mu but that the position of the recombinational activator does affect the targeting of recombination junctions, suggesting that the relatively imprecise targeting of

  13. Innate signals in mucosal immunoglobulin class switching.

    PubMed

    Puga, Irene; Cols, Montserrat; Cerutti, Andrea

    2010-11-01

    The intestinal mucosa contains large communities of commensal bacteria that process otherwise indigestible food components, synthesize essential vitamins, stimulate the maturation of the immune system, and form an ecologic niche that prevents the growth of pathogenic species. Conversely, the intestine provides the commensals with a stable habitat rich in energy derived from the ingested food. A delicate homeostatic balance maintains this mutualistic relationship without triggering a destructive inflammatory response. Commensals orchestrate intestinal homeostasis by entertaining an intimate dialogue with epithelial cells and immune cells lodged in the mucosa. Such a dialogue generates finely tuned signaling programs that ensure a state of hyporesponsiveness against noninvasive commensals and a state of active readiness against invasive pathogens. In this dialogue epithelial cells function as "interpreters" that continuously translate microbial messages to "instruct" immune cells as to the antigenic composition of the intestinal lumen. This education process initiates sophisticated defensive strategies that comprise massive production of IgA, a noninflammatory mucosal antibody class that generates immunity while preserving homeostasis. PMID:21050939

  14. Immature B cells preferentially switch to IgE with increased direct Sμ to Sε recombination

    PubMed Central

    Wesemann, Duane R.; Magee, Jennifer M.; Boboila, Cristian; Calado, Dinis Pedro; Gallagher, Michael P.; Portuguese, Andrew J.; Manis, John P.; Zhou, Xiaolong; Recher, Mike; Rajewsky, Klaus; Notarangelo, Luigi D.

    2011-01-01

    Immunoglobulin heavy chain (IgH) class-switch recombination (CSR) replaces initially expressed Cμ (IgM) constant regions (CH) exons with downstream CH exons. Stimulation of B cells with anti-CD40 plus interleukin-4 induces CSR from Cμ to Cγ1 (IgG1) and Cε (IgE), the latter of which contributes to the pathogenesis of atopic diseases. Although Cε CSR can occur directly from Cμ, most mature peripheral B cells undergo CSR to Cε indirectly, namely from Cμ to Cγ1, and subsequently to Cε. Physiological mechanisms that influence CSR to Cγ1 versus Cε are incompletely understood. In this study, we report a role for B cell developmental maturity in IgE CSR. Based in part on a novel flow cytometric IgE CSR assay, we show that immature B cells preferentially switch to IgE versus IgG1 through a mechanism involving increased direct CSR from Cμ to Cε. Our findings suggest that IgE dysregulation in certain immunodeficiencies may be related to impaired B cell maturation. PMID:22143888

  15. Fission yeast switches mating type by a replication-recombination coupled process.

    PubMed

    Arcangioli, B; de Lahondès, R

    2000-03-15

    Fission yeast exhibits a homothallic life cycle, in which the mating type of the cell mitotically alternates in a highly regulated fashion. Pedigree analysis of dividing cells has shown that only one of the two sister cells switches mating type. It was shown recently that a site- and strand-specific DNA modification at the mat1 locus precedes mating-type switching. By tracking the fate of mat1 DNA throughout the cell cycle with a PCR assay, we identified a novel DNA intermediate of mating-type switching in S-phase. The time and rate of appearance and disappearance of this DNA intermediate are consistent with a model in which mating-type switching occurs through a replication-recombination coupled pathway. Such a process provides experimental evidence in support of a copy choice recombination model in Schizosaccharomyces pombe mating-type switching and is reminiscent of the sister chromatid recombination used to complete replication in the presence of certain types of DNA damage. PMID:10716938

  16. Bach2 represses plasma cell gene regulatory network in B cells to promote antibody class switch

    PubMed Central

    Muto, Akihiko; Ochiai, Kyoko; Kimura, Yoshitaka; Itoh-Nakadai, Ari; Calame, Kathryn L; Ikebe, Dai; Tashiro, Satoshi; Igarashi, Kazuhiko

    2010-01-01

    Two transcription factors, Pax5 and Blimp-1, form a gene regulatory network (GRN) with a double-negative loop, which defines either B-cell (Pax5 high) or plasma cell (Blimp-1 high) status as a binary switch. However, it is unclear how this B-cell GRN registers class switch DNA recombination (CSR), an event that takes place before the terminal differentiation to plasma cells. In the absence of Bach2 encoding a transcription factor required for CSR, mouse splenic B cells more frequently and rapidly expressed Blimp-1 and differentiated to IgM plasma cells as compared with wild-type cells. Genetic loss of Blimp-1 in Bach2−/− B cells was sufficient to restore CSR. These data with mathematical modelling of the GRN indicate that Bach2 achieves a time delay in Blimp-1 induction, which inhibits plasma cell differentiation and promotes CSR (Delay-Driven Diversity model for CSR). Reduction in mature B-cell numbers in Bach2−/− mice was not rescued by Blimp-1 ablation, indicating that Bach2 regulates B-cell differentiation and function through Blimp-1-dependent and -independent GRNs. PMID:20953163

  17. Stabilization for a Class of Switched Nonlinear Systems With Novel Average Dwell Time Switching by T-S Fuzzy Modeling.

    PubMed

    Zhao, Xudong; Yin, Yunfei; Niu, Ben; Zheng, Xiaolong

    2016-08-01

    In this paper, the problem of switching stabilization for a class of switched nonlinear systems is studied by using average dwell time (ADT) switching, where the subsystems are possibly all unstable. First, a new concept of ADT is given, which is different from the traditional definition of ADT. Based on the new proposed switching signals, a sufficient condition of stabilization for switched nonlinear systems with unstable subsystems is derived. Then, the T-S fuzzy modeling approach is applied to represent the underlying nonlinear system to make the obtained condition easily verified. A novel multiple quadratic Lyapunov function approach is also proposed, by which some conditions are provided in terms of a set of linear matrix inequalities to guarantee the derived T-S fuzzy system to be asymptotically stable. Finally, a numerical example is given to demonstrate the effectiveness of our developed results. PMID:26316288

  18. Orientation-Specific Joining of AID-initiated DNA Breaks Promotes Antibody Class Switching

    PubMed Central

    Zhang, Yu; Hu, Jiazhi; Volpi, Sabrina A.; Meyers, Robin M.; Ho, Yu-Jui; Du, Zhou; Robbiani, Davide F.; Meng, Feilong; Gostissa, Monica; Nussenzweig, Michel C.; Manis, John P.; Alt, Frederick W.

    2015-01-01

    During B cell development, RAG endonuclease cleaves immunoglobulin heavy chain (IgH) V, D, and J gene segments and orchestrates their fusion as deletional events that assemble a V(D)J exon in the same transcriptional orientation as adjacent Cμ constant region exons1,2. In mice, six additional sets of constant region exons (CHs) lie 100-200 kb downstream in the same transcriptional orientation as V(D)J and Cμ exons2. Long repetitive switch (S) regions precede Cμ and downstream CHs. In mature B cells, class switch recombination (CSR) generates different antibody classes by replacing Cμ with a downstream CH2. Activation-Induced Cytidine Deaminase (AID) initiates CSR by promoting deamination lesions within Sμ and a downstream acceptor S region2,3; these lesions are converted into DNA double-strand breaks (DSBs) by general DNA repair factors3. Productive CSR must occur in a deletional orientation by joining the upstream end of an Sμ DSB to the downstream end of an acceptor S region DSB (Fig. 1a). However, the relative frequency of deletional to inversional CSR junctions had not been measured. Thus, whether orientation-specific joining is a programmed mechanistic feature of CSR as it is for V(D)J recombination and, if so, how this is achieved was unknown. To address this question, we adapted high-throughput genome-wide translocation sequencing (HTGTS)4 into a highly sensitive DSB end-joining assay and applied it to endogenous AID-initiated S region DSBs. We find that CSR indeed is programmed to occur in a productive deletional orientation and does so via an unprecedented mechanism that involves in cis IgH organizational features in combination with frequent S region DSBs initiated by AID. We further implicate ATM-dependent DSB response (DSBR) factors in enforcing this mechanism and provide a solution to the enigma of why CSR is so reliant on the 53BP1 DSBR factor. PMID:26308889

  19. Investigation of a hybrid optical-electronic switch supporting different service classes

    NASA Astrophysics Data System (ADS)

    Samoud, Wiem; Ware, Cédric; Lourdiane, Mounia

    2014-09-01

    Optical fiber is considered the most competitive wired transmission support thanks to its low attenuation, wide optical bandwidth, long reach, and low cost. However, optics do not yet perform higher functionalities such as switching. In fact, all-optical switches face a contention issue, due to the lack of practical optical buffers. Thus, the switching function is still performed electronically, which requires energetically costly optical-to-electronic conversions. The energy consumption is a critical issue within the growing data traffic. Thus, a proposition of hybrid switch architecture supplementing optical switch with an electronic buffer. In this paper, we propose to investigate the performance of hybrid switch that supports different priority classes where the priority is defined in terms of Packet Loss Ratio (PLR). We show that the hybrid switch is a good trade off since it allows significant performance improvements towards a buffer-less all optical switch in terms of PLR and sustainable load, for relatively few electronic ports of the buffer, which would reduce energy consumption compared to an electronic switch.

  20. Ectopic Lymphoid Structures Support Ongoing Production of Class-Switched Autoantibodies in Rheumatoid Synovium

    PubMed Central

    Manzo, Antonio; Kelly, Stephen; Blades, Mark C; Kirkham, Bruce; Spencer, Jo; Pitzalis, Costantino

    2009-01-01

    Background Follicular structures resembling germinal centres (GCs) that are characterized by follicular dendritic cell (FDC) networks have long been recognized in chronically inflamed tissues in autoimmune diseases, including the synovium of rheumatoid arthritis (RA). However, it is debated whether these ectopic structures promote autoimmunity and chronic inflammation driving the production of pathogenic autoantibodies. Anti-citrullinated protein/peptide antibodies (ACPA) are highly specific markers of RA, predict a poor prognosis, and have been suggested to be pathogenic. Therefore, the main study objectives were to determine whether ectopic lymphoid structures in RA synovium: (i) express activation-induced cytidine deaminase (AID), the enzyme required for somatic hypermutation and class-switch recombination (CSR) of Ig genes; (ii) support ongoing CSR and ACPA production; and (iii) remain functional in a RA/severe combined immunodeficiency (SCID) chimera model devoid of new immune cell influx into the synovium. Methods and Findings Using immunohistochemistry (IHC) and quantitative Taqman real-time PCR (QT-PCR) in synovial tissue from 55 patients with RA, we demonstrated that FDC+ structures invariably expressed AID with a distribution resembling secondary lymphoid organs. Further, AID+/CD21+ follicular structures were surrounded by ACPA+/CD138+ plasma cells, as demonstrated by immune reactivity to citrullinated fibrinogen. Moreover, we identified a novel subset of synovial AID+/CD20+ B cells outside GCs resembling interfollicular large B cells. In order to gain direct functional evidence that AID+ structures support CSR and in situ manufacturing of class-switched ACPA, 34 SCID mice were transplanted with RA synovium and humanely killed at 4 wk for harvesting of transplants and sera. Persistent expression of AID and Iγ-Cμ circular transcripts (identifying ongoing IgM-IgG class-switching) was observed in synovial grafts expressing FDCs/CD21L. Furthermore, synovial

  1. Two classes of ODE models with switch-like behavior

    PubMed Central

    Just, Winfried; Korb, Mason; Elbert, Ben; Young, Todd

    2013-01-01

    In cases where the same real-world system can be modeled both by an ODE system ⅅ and a Boolean system 𝔹, it is of interest to identify conditions under which the two systems will be consistent, that is, will make qualitatively equivalent predictions. In this note we introduce two broad classes of relatively simple models that provide a convenient framework for studying such questions. In contrast to the widely known class of Glass networks, the right-hand sides of our ODEs are Lipschitz-continuous. We prove that if 𝔹 has certain structures, consistency between ⅅ and 𝔹 is implied by sufficient separation of time scales in one class of our models. Namely, if the trajectories of 𝔹 are “one-stepping” then we prove a strong form of consistency and if 𝔹 has a certain monotonicity property then there is a weaker consistency between ⅅ and 𝔹. These results appear to point to more general structure properties that favor consistency between ODE and Boolean models. PMID:24244061

  2. Whole-class interactions and code-switching in secondary mathematics teaching in Mauritius

    NASA Astrophysics Data System (ADS)

    Salehmohamed, Asifa; Rowland, Tim

    2014-09-01

    This paper reports a study of whole-class interactions in mathematics classrooms in a girls' secondary school in Mauritius. It focuses on three teachers and their instructional language practices. Analysis of audio-recordings of lessons showed that code-switching was commonly practised by all the teachers in the study. The teachers' comments on their use of language within the classroom show that although they are aware of the languages they use, they are not always conscious of their code-switching. Different functions of the teachers' code-switching practices were identified, indicating it can be an important support for learning mathematics, despite some related tensions that teachers face in using code-switch in their teaching. The paper concludes with some implications for national policy and for teacher education.

  3. Identity and Language Functions: High School Chinese Immigrant Students' Code-Switching Dilemmas in ESL Classes

    ERIC Educational Resources Information Center

    Liang, Xiaoping

    2006-01-01

    This study examines classroom code-switching in relation to individual and group identity and to functional use of two languages. It investigates how high school Chinese immigrant students perceive the use of first language (L1) and second language (L2) in class, and how they use these languages during group activities. The interview data…

  4. Polymorphism, recombination, and linkage disequilibrium within the HLA class II region

    SciTech Connect

    Begovich, A.B.; McClure, G.R.; Suraj, V.C.; Helmuth, R.C.; Fildes, N.; Bugawan, T.L.; Erlich, H.A. ); Klitz, W. )

    1992-01-01

    Thirty-nine CEPH families, comprised of 502 individuals, have been typed for the HLA class II genes DRB1, DQA1, DQB1, and DPB1 using nonradioactive sequence-specific oligonucleotide probes to analyze polymerase chain reaction amplified DNA. This population, which consists of 266 independent chromosomes, contains 27 DRB1, 7 DQA1, 12 DQB1, and 17 DPB1 alleles. Analysis of the distribution of allele frequencies using the homozygosity statistic, which gives an indication of past selection pressures, suggests that balancing selection has acted on the DRB1, DQA1, and DQB1 loci. The distribution of DPB1 alleles, however, suggests a different evolutionary past. Family data permits the estimation of recombination rates and the unambiguous assignment of haplotypes. No recombinants were found between DRB1, DQA1, and DQB1; however, recombinants were detected between DQB1 and DPB1, resulting in an estimated recombination fraction of [ge]0.008 [+-] 0.004. Only 33 distinct DRB1-DQA1-DQB1 haplotypes were found in this population which illustrates the extreme nonrandom haplotypic association of alleles at these loci. A few of these haplotypes are unusual (previously unreported) for a Caucasian population and most likely result from past recombination events between the DR and DQ subregions. Examination of disequilibrium across the HLA region using these data and the available serologic HLA-A and HLA-B types of these samples shows that global disequilibrium between these loci declines with the recombination fraction, approaching statistic nonsignificance at the most distant interval, HLA-A and HLA-DP. DR-DQ haplotypes in linkage disequilibrium with DPB1 and B are noted and, finally, the evolutionary origin of certain class II haplotypes is addressed. 63 refs., 3 figs., 8 tabs.

  5. Random parameter-switching synthesis of a class of hyperbolic attractors.

    PubMed

    Danca, Marius-F

    2008-09-01

    The parameter perturbation methods (the most known being the OGY method) apply small wisely chosen swift kicks to the system once per cycle, to maintain it near the desired unstable periodic orbit. Thus, one can consider that a new attractor is finally generated. Another class of methods which allow the attractors born, imply small perturbations of the state variable [see, e.g., J. Güémez and M. A. Matías, Phys. Lett. A 181, 29 (1993)]. Whatever technique is utilized, generating any targeted attractor starting from a set of two or more of any kind of attractors (stable or not) of a considered dissipative continuous-time system cannot be achieved with these techniques. This kind of attractor synthesis [introduced in M.-F. Danca, W. K. S. Tang, and G. Chen, Appl. Math. Comput. 201, 650 (2008) and proved analytically in Y. Mao, W. K. S. Tang, and M.-F. Danca, Appl. Math. Comput. (submitted)] which starts from a set of given attractors, allows us, via periodic parameter-switching, to generate any of the set of all possible attractors of a class of continuous-time dissipative dynamical systems, depending linearly on the control parameter. In this paper we extend this technique proving empirically that even random manners for switching can be utilized for this purpose. These parameter-switches schemes are very easy to implement and require only the mathematical model of the underlying dynamical system, a convergent numerical method to integrate the system, and the bifurcation diagram to choose specific attractors. Relatively large parameter switches are admitted. As a main result, these switching algorithms (deterministic or random) offer a new perspective on the set of all attractors of a class of dissipative continuous-time dynamical systems. PMID:19045449

  6. Random parameter-switching synthesis of a class of hyperbolic attractors

    NASA Astrophysics Data System (ADS)

    Danca, Marius-F.

    2008-09-01

    The parameter perturbation methods (the most known being the OGY method) apply small wisely chosen swift kicks to the system once per cycle, to maintain it near the desired unstable periodic orbit. Thus, one can consider that a new attractor is finally generated. Another class of methods which allow the attractors born, imply small perturbations of the state variable [see, e.g., J. Güémez and M. A. Matías, Phys. Lett. A 181, 29 (1993)]. Whatever technique is utilized, generating any targeted attractor starting from a set of two or more of any kind of attractors (stable or not) of a considered dissipative continuous-time system cannot be achieved with these techniques. This kind of attractor synthesis [introduced in M.-F. Danca, W. K. S. Tang, and G. Chen, Appl. Math. Comput. 201, 650 (2008) and proved analytically in Y. Mao, W. K. S. Tang, and M.-F. Danca, Appl. Math. Comput. (submitted)] which starts from a set of given attractors, allows us, via periodic parameter-switching, to generate any of the set of all possible attractors of a class of continuous-time dissipative dynamical systems, depending linearly on the control parameter. In this paper we extend this technique proving empirically that even random manners for switching can be utilized for this purpose. These parameter-switches schemes are very easy to implement and require only the mathematical model of the underlying dynamical system, a convergent numerical method to integrate the system, and the bifurcation diagram to choose specific attractors. Relatively large parameter switches are admitted. As a main result, these switching algorithms (deterministic or random) offer a new perspective on the set of all attractors of a class of dissipative continuous-time dynamical systems.

  7. Swi6, a Gene Required for Mating-Type Switching, Prohibits Meiotic Recombination in the Mat2-Mat3 ``cold Spot'' of Fission Yeast

    PubMed Central

    Klar, AJS.; Bonaduce, M. J.

    1991-01-01

    Mitotic interconversion of the mating-type locus (mat1) of the fission yeast Schizosaccharomyces pombe is initiated by a double-strand break at mat1. The mat2 and mat3 loci act as nonrandom donors of genetic information for mat1 switching such that switches occur primarily (or only) to the opposite mat1 allele. Location of the mat1 ``hot spot'' for transposition should be contrasted with the ``cold spot'' of meiotic recombination located within the adjoining mat2-mat3 interval. That is, meiotic interchromosomal recombination in mat2, mat3 and the intervening 15-kilobase region does not occur at all. swi2 and swi6 switching-deficient mutants possess the normal level of double-strand break at mat1, yet they fail to switch efficiently. By testing for meiotic recombination in the cold spot, we found the usual lack of recombination in a swi2 mutant but a significant level of recombination in a swi6 mutant. Therefore, the swi6 gene function is required to keep the donor loci inert for interchromosomal recombination. This finding, combined with the additional result that switching primarily occurs intrachromosomally, suggests that the donor loci are made accessible for switching by folding them onto mat1, thus causing the cold spot of recombination. PMID:1783290

  8. Identifying Effective Enzyme Activity Targets for Recombinant Class I and Class II Collagenase for Successful Human Islet Isolation

    PubMed Central

    Balamurugan, Appakalai N.; Green, Michael L.; Breite, Andrew G.; Loganathan, Gopalakrishnan; Wilhelm, Joshua J.; Tweed, Benjamin; Vargova, Lenka; Lockridge, Amber; Kuriti, Manikya; Hughes, Michael G.; Williams, Stuart K.; Hering, Bernhard J.; Dwulet, Francis E.; McCarthy, Robert C.

    2016-01-01

    Background Isolation following a good manufacturing practice-compliant, human islet product requires development of a robust islet isolation procedure where effective limits of key reagents are known. The enzymes used for islet isolation are critical but little is known about the doses of class I and class II collagenase required for successful islet isolation. Methods We used a factorial approach to evaluate the effect of high and low target activities of recombinant class I (rC1) and class II (rC2) collagenase on human islet yield. Consequently, 4 different enzyme formulations with divergent C1:C2 collagenase mass ratios were assessed, each supplemented with the same dose of neutral protease. Both split pancreas and whole pancreas models were used to test enzyme targets (n = 20). Islet yield/g pancreas was compared with historical enzymes (n = 42). Results Varying the Wunsch (rC2) and collagen degradation activity (CDA, rC1) target dose, and consequently the C1:C2 mass ratio, had no significant effect on tissue digestion. Digestions using higher doses of Wunsch and CDA resulted in comparable islet yields to those obtained with 60% and 50% of those activities, respectively. Factorial analysis revealed no significant main effect of Wunsch activity or CDA for any parameter measured. Aggregate results from 4 different collagenase formulations gave 44% higher islet yield (>5000 islet equivalents/g) in the body/tail of the pancreas (n = 12) when compared with those from the same segment using a standard natural collagenase/protease mixture (n = 6). Additionally, islet yields greater than 5000 islet equivalents/g pancreas were also obtained in whole human pancreas. Conclusions A broader C1:C2 ratio can be used for human islet isolation than has been used in the past. Recombinant collagenase is an effective replacement for the natural enzyme and we have determined that high islet yield can be obtained even with low doses of rC1:rC2, which is beneficial for the survival

  9. A combined nuclear and nucleolar localization motif in activation-induced cytidine deaminase (AID) controls immunoglobulin class switching.

    PubMed

    Hu, Yi; Ericsson, Ida; Torseth, Kathrin; Methot, Stephen P; Sundheim, Ottar; Liabakk, Nina B; Slupphaug, Geir; Di Noia, Javier M; Krokan, Hans E; Kavli, Bodil

    2013-01-23

    Activation-induced cytidine deaminase (AID) is a DNA mutator enzyme essential for adaptive immunity. AID initiates somatic hypermutation and class switch recombination (CSR) by deaminating cytosine to uracil in specific immunoglobulin (Ig) gene regions. However, other loci, including cancer-related genes, are also targeted. Thus, tight regulation of AID is crucial to balance immunity versus disease such as cancer. AID is regulated by several mechanisms including nucleocytoplasmic shuttling. Here we have studied nuclear import kinetics and subnuclear trafficking of AID in live cells and characterized in detail its nuclear localization signal. Importantly, we find that the nuclear localization signal motif also directs AID to nucleoli where it colocalizes with its interaction partner, catenin-β-like 1 (CTNNBL1), and physically associates with nucleolin and nucleophosmin. Moreover, we demonstrate that release of AID from nucleoli is dependent on its C-terminal motif. Finally, we find that CSR efficiency correlates strongly with the arithmetic product of AID nuclear import rate and DNA deamination activity. Our findings suggest that directional nucleolar transit is important for the physiological function of AID and demonstrate that nuclear/nucleolar import and DNA cytosine deamination together define the biological activity of AID. This is the first study on subnuclear trafficking of AID and demonstrates a new level in its complex regulation. In addition, our results resolve the problem related to dissociation of deamination activity and CSR activity of AID mutants. PMID:23183374

  10. A Butter Aroma Recombinate Activates Human Class-I Odorant Receptors.

    PubMed

    Geithe, Christiane; Andersen, Gaby; Malki, Agne; Krautwurst, Dietmar

    2015-11-01

    With ∼400 olfactory G protein-coupled receptors (GPCR), humans sensitively perceive ∼230 key aroma compounds as best natural agonists of ∼10000 food volatiles. An understanding of odorant coding, thus, critically depends on the knowledge about interactions of key food aroma chemicals and their mixtures with their cognate receptors. Genetically designed test cell systems enable the screening, deorphaning, and characterization of single odorant receptors (OR). This study shows for the food aroma-specific and quantitative butter aroma recombinate, and its single components, specific in vitro class-I OR activity patterns, as well as the activation of selected OR in a concentration-dependent manner. Recently, chemosensory receptors, especially class-I OR, were demonstrated to be expressed on blood leukocytes, which may encounter foodborne aroma compounds postprandially. This study shows that butter aroma recombinate induced chemotaxis of isolated human neutrophils in a defined gradient, and in a concentration-dependent and pertussis toxin-sensitive manner, suggesting at least a GPCR-mediated activation of blood leukocytes by key food odorants. PMID:26451762

  11. Stability and stabilization studies for a class of switched nonlinear systems via vector norms approach.

    PubMed

    Sakly, Anis; Kermani, Marwen

    2015-07-01

    This paper is concerned with the problems of stability analysis and stabilization with a state feedback controller through pole placement for a class of both continuous and discrete-time switched nonlinear systems. These systems are modeled by differential or difference equations. Then, a transformation under the arrow form is employed. Note that, the main contribution in this work is twofold: firstly, based on the construction of an appropriated common Lyapunov function, as well the use of the vector norms notion, the recourse to the Kotelyanski lemma, the M-matrix proprieties, the aggregation techniques and the application of the Borne-Gentina criterion, new sufficient stability conditions under arbitrary switching for the autonomous system are deduced. Secondly, this result is extended for designing a state feedback controller by using pole assignment control, which guarantee that the corresponding closed-loop system is globally asymptotically stable under arbitrary switching. The main novelties features of these obtained results are the explicitness and the simplicity in their application. Moreover, they allow us to avoid the search of a common Lyapunov function which is a difficult matter. Finally, as validation to stabilize a shunt DC motor under variable mechanical loads is performed to demonstrate the effectiveness of the proposed results. PMID:25701192

  12. Immunogenicity of recombinant class 1 protein from Neisseria meningitidis refolded into phospholipid vesicles and detergent.

    PubMed

    Niebla, O; Alvarez, A; Martín, A; Rodríguez, A; Delgado, M; Falcón, V; Guillén, G

    2001-05-14

    The possibility of eliciting bactericidal antibodies against a recombinant class 1 protein (P1) from Neisseria meningitidis, joined to the first 45 amino acids of the neisserial LpdA protein (PM82), was examined. P1 was produced in Escherichia coli as intracellular inclusion bodies, from which it was purified and reconstituted by (a) inclusion into phospholipid vesicles and detergent and (b) refolding in 0.1% SDS. When Balb/c mice were immunised, high titres of subtype-specific bactericidal antibodies against P1 were obtained in both cases. These results suggest that in spite of being a denaturing agent, it is possible to use SDS to reconstitute the P1 protein in a conformation that exposes the immunodominat regions. PMID:11348724

  13. Saccharomyces forkhead protein Fkh1 regulates donor preference during mating-type switching through the recombination enhancer

    PubMed Central

    Sun, Kaiming; Coïc, Eric; Zhou, Zhiqi; Durrens, Pascal; Haber, James E.

    2002-01-01

    Saccharomyces mating-type switching results from replacement by gene conversion of the MAT locus with sequences copied from one of two unexpressed donor loci, HML or HMR. MATa cells recombine with HMLα ∼90% of the time, whereas MATα cells choose HMRa 80%–90% of the time. HML preference in MATa is controlled by the cis-acting recombination enhancer (RE) that regulates recombination along the entire left arm of chromosome III. Comparison of RE sequences between S. cerevisiae, S. carlsbergensis, and S. bayanus defines four highly conserved regions (A, B, C, and D) within a 270-bp minimum RE. An adjacent E region enhances RE activity. Multimers of region A, D, or E are sufficient to promote selective use of HML. Regions A, D, and E each bind in vivo the transcription activator forkhead proteins Fkh1p and Fkh2p and their associated Ndd1p, although there are no adjacent open reading frames (ORFs). Deletion of FKH1 significantly reduces MATa's use of HML, as does mutation of the Fkh1/Fkh2-binding sites in a multimer of region A. We conclude that Fkh1p regulates MATa donor preference through direct interaction with RE. PMID:12183363

  14. Saccharomyces forkhead protein Fkh1 regulates donor preference during mating-type switching through the recombination enhancer.

    PubMed

    Sun, Kaiming; Coïc, Eric; Zhou, Zhiqi; Durrens, Pascal; Haber, James E

    2002-08-15

    Saccharomyces mating-type switching results from replacement by gene conversion of the MAT locus with sequences copied from one of two unexpressed donor loci, HML or HMR. MATa cells recombine with HMLalpha approximately 90% of the time, whereas MATalpha cells choose HMRa 80%-90% of the time. HML preference in MATa is controlled by the cis-acting recombination enhancer (RE) that regulates recombination along the entire left arm of chromosome III. Comparison of RE sequences between S. cerevisiae, S. carlsbergensis, and S. bayanus defines four highly conserved regions (A, B, C, and D) within a 270-bp minimum RE. An adjacent E region enhances RE activity. Multimers of region A, D, or E are sufficient to promote selective use of HML. Regions A, D, and E each bind in vivo the transcription activator forkhead proteins Fkh1p and Fkh2p and their associated Ndd1p, although there are no adjacent open reading frames (ORFs). Deletion of FKH1 significantly reduces MATa's use of HML, as does mutation of the Fkh1/Fkh2-binding sites in a multimer of region A. We conclude that Fkh1p regulates MATa donor preference through direct interaction with RE. PMID:12183363

  15. Genomic Investigation Reveals Highly Conserved, Mosaic, Recombination Events Associated with Capsular Switching among Invasive Neisseria meningitidis Serogroup W Sequence Type (ST)-11 Strains

    PubMed Central

    Mustapha, Mustapha M.; Marsh, Jane W.; Krauland, Mary G.; Fernandez, Jorge O.; de Lemos, Ana Paula S.; Hotopp, Julie C. Dunning; Wang, Xin; Mayer, Leonard W.; Lawrence, Jeffrey G.; Hiller, N. Luisa; Harrison, Lee H.

    2016-01-01

    Neisseria meningitidis is an important cause of meningococcal disease globally. Sequence type (ST)-11 clonal complex (cc11) is a hypervirulent meningococcal lineage historically associated with serogroup C capsule and is believed to have acquired the W capsule through a C to W capsular switching event. We studied the sequence of capsule gene cluster (cps) and adjoining genomic regions of 524 invasive W cc11 strains isolated globally. We identified recombination breakpoints corresponding to two distinct recombination events within W cc11: A 8.4-kb recombinant region likely acquired from W cc22 including the sialic acid/glycosyl-transferase gene, csw resulted in a C→W change in capsular phenotype and a 13.7-kb recombinant segment likely acquired from Y cc23 lineage includes 4.5 kb of cps genes and 8.2 kb downstream of the cps cluster resulting in allelic changes in capsule translocation genes. A vast majority of W cc11 strains (497/524, 94.8%) retain both recombination events as evidenced by sharing identical or very closely related capsular allelic profiles. These data suggest that the W cc11 capsular switch involved two separate recombination events and that current global W cc11 meningococcal disease is caused by strains bearing this mosaic capsular switch. PMID:27289093

  16. Identification properties of a recombinant class I hydrophobin rHGFI.

    PubMed

    Li, Wenfeng; Gong, Yanbo; Xu, Haijin; Qiao, Mingqiang; Niu, Baolong

    2015-01-01

    Hydrophobins fulfill various functions in fungal growth and morphology. These proteins can self-assemble at hydrophilic/hydrophobic interfaces and form amphipathic membranes. Based on their physical properties and hydropathy patterns, hydrophobins are divided into two classes (I and II). In order to identify the recombinant class I hydrophobin rHGFI, the different properties between rHGFI and the typical class II hydrophobin rHFBI were investigated. In contrast to rHGFI, no rodlet structure was observed on rHFBI coated mica surface, and the membranes formed on siliconized glass surfaces by rHFBI were not robust enough to resist treatment with 60% ethanol and 2% hot SDS. In contrast, the membranes formed by rHGFI on siliconized glass surfaces were so strong that could resist hot detergent and alcohol solution washing. Moreover, self-assembly of rHFBI at the water-air interface was not accompanied by a change in secondary structure. Meanwhile, β-sheet structures dramatically increased after rHGFI self-assembled at water-air interface, which could cause the fluorescence intensity of Thioflavin T increased and Congo Red and CD absorption spectra shift. Water-insoluble erythrosin B dispersion prepared with rHGFI and rHFBI were both stable for more than one month, which indicated that the interaction between erythrosin B and rHGFI/rHFBI was strong. This might promote rHGFI and rHFBI to be considered as potential dispersing agents to stabilize water-insoluble erythrosin B. PMID:25241920

  17. Expression, purification and characterization of recombinant Jerdonitin, a P-II class snake venom metalloproteinase comprising metalloproteinase and disintegrin domains.

    PubMed

    Zhu, Lili; Yuan, Cai; Chen, Zhuo; Wang, Wanyu; Huang, Mingdong

    2010-01-01

    Jerdonitin is a P-II class snake venom metalloproteinase comprising metalloproteinase and disintegrin domains. In this study, we established a high-level expression system in Pichia pastoris and developed a purification strategy for the recombinant Jerdonitin. This recombinant Jerdonitin degraded fibrinogen at a level of activity comparable with its wild type. The effects of recombinant Jerdonitin on inhibiting ADP-induced human platelet aggregation were in a dose-dependent manner with an IC(50) of 248nM. In addition, we reported here that Jerdonitin can significantly inhibit the growth of several cell lines, including human liver cancer cells (Bel7402), human leukemia cells (K562) and human gastric carcinoma cells (BGC823). This study offers recombinant Jerdonitin that will be valuable for further functional and structural studies of Jerdonitin. PMID:19732785

  18. IL-4-induced transcription factor NFIL3/E4BP4 controls IgE class switching.

    PubMed

    Kashiwada, Masaki; Levy, Deborah M; McKeag, Lisa; Murray, Keri; Schröder, Andreas J; Canfield, Stephen M; Traver, Geri; Rothman, Paul B

    2010-01-12

    IL-4 signaling promotes IgE class switching through STAT6 activation and the induction of Ig germ-line epsilon (GLepsilon) transcription. Previously, we and others identified a transcription factor, Nfil3, as a gene induced by IL-4 stimulation in B cells. However, the precise roles of nuclear factor, IL-3-regulated (NFIL3) in IL-4 signaling are unknown. Here, we report that NFIL3 is important for IgE class switching. NFIL3-deficient mice show impaired IgE class switching, and this defect is B-cell intrinsic. The induction of GLepsilon transcripts after LPS and IL-4 stimulation is significantly reduced in NFIL3-deficient B cells. Expression of NFIL3 in NFIL3-deficient B cells restores the impairment of IgE production, and overexpression of NFIL3 in the presence of cycloheximide induces GLepsilon transcripts. Moreover, NFIL3 binds to Iepsilon promoter in vivo. Together, these results identify NFIL3 as a key regulator of IL-4-induced GLepsilon transcription in response to IL-4 and subsequent IgE class switching. PMID:20080759

  19. A Recombinationally Repressed Region between Mat2 and Mat3 Loci Shares Homology to Centromeric Repeats and Regulates Directionality of Mating-Type Switching in Fission Yeast

    PubMed Central

    Grewal, SIS.; Klar, AJS.

    1997-01-01

    Cells of the fission yeast Schizosaccharomyces pombe switch mating type by replacing genetic information at the transcriptionally active mat1 locus with sequences copied from one of two closely linked silent loci, mat2-P or mat3-M. By a process referred to as directionality of switching, cells predominantly switch to the opposite mat1 allele; the mat1-P allele preferentially recombines with mat3, while mat1-M selects the mat2. In contrast to efficient recombination at mat1, recombination within the adjoining mat2-mat3 interval is undetectable. We defined the role of sequences between mat2 and mat3, designated the K-region, in directionality as well as recombinational suppression. Cloning and sequencing analysis revealed that a part of the K-region is homologous to repeat sequences present at centromeres, which also display transcriptional and recombinational suppression. Replacement of 7.5 kb of the K-region with the ura4(+) gene affected directionality in a variegated manner. Analysis of the swi6-mod locus, which was previously shown to affect directionality, in KΔ::ura4(+) strains suggested the existence of at least two overlapping directionality mechanisms. Our work furthers the model that directionality is regulated by cell-type-specific organization of the heterochromatin-like structure in the mating-type region and provides evidence that the K-region contributes to silencing of the mat2-mat3 interval. PMID:9258669

  20. Restoration of Retarded Influenza Virus-specific Immunoglobulin Class Switch in Aged Mice

    PubMed Central

    Zhang, Yongxin; Wang, Ying; Zhang, Monica; Liu, Lin; Mbawuike, Innocent N

    2016-01-01

    Objective The declined immune response to infection causes significant higher morbidity and mortality in aging in spite of the coexisted hyperimmunoglobulinemia (HIG). This study is to reveal the cellular basis of HIG and mechanism of weakened HA-specific IgG response in aged mice and to test cell therapy in the treatment of age-related IgG antibody production deficiency with immunocyte adoptive transfer. Methods BALB/c mice was immunized with Influenza A/Taiwan vaccine and challenged with the same strain of virus. ELISA was used to assess the levels of total immunoglobulins and antigen specific antibody response. The flow cytometry and ELISPOT were used to evaluate the frequencies of total immunoglobulin- and specific antibody-producing and secreting B lymphocytes. In vitro expanded mononuclear cells, CD4+ T lymphocytes and CD20+ B lymphocytes from old and young mice were adoptively transferred into influenza virus-challenged aged mice, and HA-specific IgG responses were observed. Results It is found that old mice exhibited higher levels of total serum IgG, IgM and IgA, higher frequencies of IgG+, IgM+ and IgA+ cells, and greater antigen-specific IgM and IgA responses to influenza infection, in comparison to young mice. However, influenza antigen- specific IgG and its subclass responses in old mice were significantly lower. Conclusion The retarded specific IgG response could be attributed to an insufficiency of immunoglobulin class switch in aging. Correlation analysis indicated that HIG and deficient specific IgG production in aged mice could be independent to each other in their pathogenesis. Correction of deficient specific IgG production by adoptive transfer of in vitro expanded and unexpanded CD4+ cells from immunized young mice suggests the CD4+ cell dysfunction contributes to the insufficiency of immunoglobulin class switch in aged mice. The transfusion of in vitro expanded lymphocytes could be a potential effective therapy for the age

  1. Diversity of metalloproteinases in Bothrops neuwiedi snake venom transcripts: evidences for recombination between different classes of SVMPs

    PubMed Central

    2011-01-01

    Background Snake venom metalloproteinases (SVMPs) are widely distributed in snake venoms and are versatile toxins, targeting many important elements involved in hemostasis, such as basement membrane proteins, clotting proteins, platelets, endothelial and inflammatory cells. The functional diversity of SVMPs is in part due to the structural organization of different combinations of catalytic, disintegrin, disintegrin-like and cysteine-rich domains, which categorizes SVMPs in 3 classes of precursor molecules (PI, PII and PIII) further divided in 11 subclasses, 6 of them belonging to PII group. This heterogeneity is currently correlated to genetic accelerated evolution and post-translational modifications. Results Thirty-one SVMP cDNAs were full length cloned from a single specimen of Bothrops neuwiedi snake, sequenced and grouped in eleven distinct sequences and further analyzed by cladistic analysis. Class P-I and class P-III sequences presented the expected tree topology for fibrinolytic and hemorrhagic SVMPs, respectively. In opposition, three distinct segregations were observed for class P-II sequences. P-IIb showed the typical segregation of class P-II SVMPs. However, P-IIa grouped with class P-I cDNAs presenting a 100% identity in the 365 bp at their 5' ends, suggesting post-transcription events for interclass recombination. In addition, catalytic domain of P-IIx sequences segregated with non-hemorrhagic class P-III SVMPs while their disintegrin domain grouped with other class P-II disintegrin domains suggesting independent evolution of catalytic and disintegrin domains. Complementary regions within cDNA sequences were noted and may participate in recombination either at DNA or RNA levels. Proteins predicted by these cDNAs show the main features of the correspondent classes of SVMP, but P-IIb and P-IIx included two additional cysteines cysteines at the C-termini of the disintegrin domains in positions not yet described. Conclusions In B. neuwiedi venom gland

  2. A DFT study of a new class of gold nanocluster-photochrome multi-functional switches.

    PubMed

    Fihey, Arnaud; Maurel, François; Perrier, Aurélie

    2014-12-21

    With the help of a computational scheme combining molecular dynamics, DFT and TD-DFT methods, the conformational, electronic and optical properties of a new class of hybrid compounds where a photochromic molecule belonging to the dithienylethene family (DTE) is covalently linked to a Au25 nanocluster (gold nanocluster or GNC) are investigated. We compare two types of hybrid GNC-DTE systems where the aromatic linker between the metallic and the DTE moieties is either a phenyl or a thiophene ring. By examining the perturbation of the DTE electronic structure after grafting upon the GNC, we show that the hybrid system with a phenyl linker should preserve its photochromic activity. For the latter system, we have then studied the possible energy and electron transfer between the GNC and the DTE units. The energy transfer between the two moieties can be a priori discarded while a uni-directional electron transfer should take place from the GNC to the excited DTE. We show that this transfer can be controlled by switching the state of the molecule. PMID:25363237

  3. An approach to mapping haplotype-specific recombination sites in human MHC class III

    SciTech Connect

    Levo, A.; Westman, P.; Partanen, J.

    1996-12-31

    Studies of the major histocompatibility complex (MHC) in mouse indicate that the recombination sites are not randomly distributed and their occurrence is haplotype-dependent. No data concerning haplotype-specific recombination sites in human are available due to the low number of informative families. To investigate haplotype-specific recombination sites in human MHC, we describe an approach based on identification of recombinant haplotypes derived from one conserved haplotype at the population level. The recombination sites were mapped by comparing polymorphic markers between the recombinant and assumed original haplotypes. We tested this approach on the extended haplotype HLA A3; B47; Bf{sup *}F; C4A{sup *}1; C4B{sup *}Q0; DR7, which is most suitable for this analysis. First, it carries a number of rare markers, and second, the haplotype, albeit rare in the general population, is frequent in patients with 21-hydroxylase (21OH) defect. We observed recombinants derived from this haplotype in patients with 21OH defect. All these haplotypes had the centromeric part (from Bf to DR) identical to the original haplotype, but they differed in HLA A and B. We therefore assumed that they underwent recombinations in the segment that separates the Bf and HLA B genes. Polymorphic markers indicated that all break points mapped to two segments near the TNF locus. This approach makes possible the mapping of preferential recombination sites in different haplotypes. 20 refs., 1 fig., 1 tab.

  4. Substrate stiffness regulates B-cell activation, proliferation, class switch, and T-cell-independent antibody responses in vivo.

    PubMed

    Zeng, Yingyue; Yi, Junyang; Wan, Zhengpeng; Liu, Kai; Song, Ping; Chau, Alicia; Wang, Fei; Chang, Zai; Han, Weidong; Zheng, Wenjie; Chen, Ying-Hua; Xiong, Chunyang; Liu, Wanli

    2015-06-01

    B cells use B-cell receptors (BCRs) to sense antigens that are usually presented on substrates with different stiffness. However, it is not known how substrate stiffness affects B-cell proliferation, class switch, and in vivo antibody responses. We addressed these questions using polydimethylsiloxane (PDMS) substrates with different stiffness (20 or 1100 kPa). Live cell imaging experiments suggested that antigens on stiffer substrates more efficiently trigger the synaptic accumulation of BCR and phospho-Syk molecules compared with antigens on softer substrates. In vitro expansion of mouse primary B cells shows different preferences for substrate stiffness when stimulated by different expansion stimuli. LPS equally drives B-cell proliferation on stiffer or softer substrates. Anti-CD40 antibodies enhance B-cell proliferation on stiffer substrates, while antigens enhance B-cell proliferation on softer substrates through a mechanism involving the enhanced phosphorylation of PI3K, Akt, and FoxO1. In vitro class switch differentiation of B cells prefers softer substrates. Lastly, NP67-Ficoll on softer substrates accounted for an enhanced antibody response in vivo. Thus, substrate stiffness regulates B-cell activation, proliferation, class switch, and T cell independent antibody responses in vivo, suggesting its broad application in manipulating the fate of B cells in vitro and in vivo. PMID:25756957

  5. Classes in Translating and Interpreting Produce Differential Gains in Switching and Updating

    PubMed Central

    Dong, Yanping; Liu, Yuhua

    2016-01-01

    The present longitudinal study was intended to investigate whether the two bilingual experiences of written translation and consecutive interpreting (featured with similar language switching experience but different processing demands) would produce different cognitive control effects in young adults. Three groups of Chinese–English young adult bilinguals, who differed mainly in their half-year long bilingual experience: one for general L2 training, one for written translation and one for oral consecutive interpreting, were tested twice on the number Stroop, switching color-shape and N-back tasks. The results show that the interpreting experience produced significant cognitive advantages in switching (switch cost) and updating, while the translating experience produced marginally significant improvements in updating. The findings indicate that the experience of language switching under higher processing demands brings more domain-general advantages, suggesting that processing demand may be a decisive factor for the presence or absence of the hot-debated bilingual advantages. PMID:27625620

  6. Classes in Translating and Interpreting Produce Differential Gains in Switching and Updating.

    PubMed

    Dong, Yanping; Liu, Yuhua

    2016-01-01

    The present longitudinal study was intended to investigate whether the two bilingual experiences of written translation and consecutive interpreting (featured with similar language switching experience but different processing demands) would produce different cognitive control effects in young adults. Three groups of Chinese-English young adult bilinguals, who differed mainly in their half-year long bilingual experience: one for general L2 training, one for written translation and one for oral consecutive interpreting, were tested twice on the number Stroop, switching color-shape and N-back tasks. The results show that the interpreting experience produced significant cognitive advantages in switching (switch cost) and updating, while the translating experience produced marginally significant improvements in updating. The findings indicate that the experience of language switching under higher processing demands brings more domain-general advantages, suggesting that processing demand may be a decisive factor for the presence or absence of the hot-debated bilingual advantages. PMID:27625620

  7. Laser triggering of water switches in terrawatt-class pulse power accelerators.

    SciTech Connect

    Woodworth, Joseph Ray; Johnson, David Lee (Titan Pulse Sciences, San Leandro, CA); Wilkins, Frank (Bechtel Nevada, Las Vegas, NV); Van De Valde, David (EG&G Technical Services, Albuquerque, NM); Sarkisov, Gennady Sergeevich; Zameroski, Nathan D.; Starbird, Robert L.

    2005-12-01

    Focused Beams from high-power lasers have been used to command trigger gas switches in pulse power accelerators for more than two decades. This Laboratory-Directed Research and Development project was aimed at determining whether high power lasers could also command trigger water switches on high-power accelerators. In initial work, we determined that focused light from three harmonics of a small pulsed Nd:YAG laser at 1064 nm, 532 nm, and 355 nm could be used to form breakdown arcs in water, with the lowest breakdown thresholds of 110 J/cm{sup 2} or 14 GW/cm{sup 2} at 532 nm in the green. In laboratory-scale laser triggering experiments with a 170-kV pulse-charged water switch with a 3-mm anode-cathode gap, we demonstrated that {approx}90 mJ of green laser energy could trigger the gap with a 1-{sigma} jitter of less than 2ns, a factor of 10 improvement over the jitter of the switch in its self breaking mode. In the laboratory-scale experiments we developed optical techniques utilizing polarization rotation of a probe laser beam to measure current in switch channels and electric field enhancements near streamer heads. In the final year of the project, we constructed a pulse-power facility to allow us to test laser triggering of water switches from 0.6- MV to 2.0 MV. Triggering experiments on this facility using an axicon lens for focusing the laser and a switch with a 740 kV self-break voltage produced consistent laser triggering with a {+-} 16-ns 1-{sigma} jitter, a significant improvement over the {+-} 24-ns jitter in the self-breaking mode.

  8. Global tracking for a class of uncertain nonlinear systems with unknown sign-switching control direction by output feedback

    NASA Astrophysics Data System (ADS)

    Roux Oliveira, Tiago; Jacoud Peixoto, Alessandro; Hsu, Liu

    2015-09-01

    This paper addresses the design of a sliding mode controller for a class of high-order uncertain nonlinear plants with unmatched state-dependent nonlinearities and unknown sign of the high frequency gain, i.e., the control direction is assumed unknown. Differently from most previous studies, the control direction is allowed to switch its sign. We show that it is possible to obtain global exact tracking using only output-feedback by coupling a relay periodic switching function with a norm state observer. One significant advantage of the new scheme is its robustness and improved transient response under arbitrary changes of the control direction which have been theoretically demonstrated for jump variations and successfully tested by simulations. The proposed controller is also evaluated with a DC motor control experiment.

  9. NF-κB p50-Dependent In Vivo Footprints at Ig Sγ3 DNA Are Correlated with μ→γ3 Switch Recombination1

    PubMed Central

    Wuerffel, Robert A.; Ma, Limei; Kenter, Amy L.

    2015-01-01

    NF-g=kB has been demonstrated to play critical roles in multiple aspects of immune responses including Ig H chain isotype switching. To better define the specific roles the p50 subunit of NF-κB plays in μ→γ3 switch recombination (SR), we systematically evaluated p50-deficient B cells for activities that are strongly correlated with SR. B cell activation with LPS plus anti-IgD-dextran plus IL-5 plus IL-4 plus TGF-β produced normal levels of proliferation and γ3 germline transcripts in p50-deficient B cells, but μ→γ3 SR was impaired. In vitro binding studies previously showed that NF-kB p50 homodimer binds the switch nuclear B-site protein (SNIP) of the Sγ3 tandem repeat. Ligation-mediated PCR in vivo footprint analysis demonstrates that the region spanning the SNIP and switch nuclear A-site protein (SNAP) binding sites of the Sγ3 region are contacted by protein in normal resting splenic B cells. B cells that are homozygous for the targeted disruption of the gene encoding p50 (–/–) show strong aberrant footprints, whereas heterozygous cells (+/–) reveal a partial effect in Sγ3 DNA. These studies provide evidence of nucleoprotein interactions at switch DNA in vivo and suggest a direct interaction of p50 with Sγ3 DNA that is strongly correlated with SR competence. PMID:11254712

  10. The Ets-1 transcription factor is required for Stat1-mediated T-bet expression and IgG2a class switching in mouse B cells.

    PubMed

    Nguyen, Hai Vu; Mouly, Enguerran; Chemin, Karine; Luinaud, Romain; Despres, Raymonde; Fermand, Jean-Paul; Arnulf, Bertrand; Bories, Jean-Christophe

    2012-05-01

    In response to antigens and cytokines, mouse B cells undergo class-switch recombination (CSR) and differentiate into Ig-secreting cells. T-bet, a T-box transcription factor that is up-regulated in lymphocytes by IFN-γ or IL-27, was shown to regulate CSR to IgG2a after T cell-independent B-cell stimulations. However, the molecular mechanisms controlling this process remain unclear. In the present study, we show that inactivation of the Ets-1 transcription factor results in a severe decrease in IgG2a secretion in vivo and in vitro. No T-bet expression was observed in Ets-1-deficient (Ets-1(-/-)) B cells stimulated with IFN-γ and lipopolysaccharide, and forced expression of T-bet in these cells rescued IgG2a secretion. Furthermore, we identified a transcriptional enhancer in the T-bet locus with an activity in B cells that relies on ETS-binding sites. After IFN-γ stimulation of Ets-1(-/-) B cells, activated Stat1, which forms a complex with Ets-1 in wild-type cells, no longer binds to the T-bet enhancer or promotes histone modifications at this site. These results demonstrate that Ets-1 is critical for IgG2a CSR and acts as an essential cofactor for Stat1 in the regulation of T-bet expression in B cells. PMID:22438254

  11. A PTIP-PA1 subcomplex promotes transcription for IgH class switching independently from the associated MLL3/MLL4 methyltransferase complex.

    PubMed

    Starnes, Linda M; Su, Dan; Pikkupeura, Laura M; Weinert, Brian T; Santos, Margarida A; Mund, Andreas; Soria, Rebeca; Cho, Young-Wook; Pozdnyakova, Irina; Kubec Højfeldt, Martina; Vala, Andrea; Yang, Wenjing; López-Méndez, Blanca; Lee, Ji-Eun; Peng, Weiqun; Yuan, Joan; Ge, Kai; Montoya, Guillermo; Nussenzweig, André; Choudhary, Chunaram; Daniel, Jeremy A

    2016-01-15

    Class switch recombination (CSR) diversifies antibodies for productive immune responses while maintaining stability of the B-cell genome. Transcription at the immunoglobulin heavy chain (Igh) locus targets CSR-associated DNA damage and is promoted by the BRCT domain-containing PTIP (Pax transactivation domain-interacting protein). Although PTIP is a unique component of the mixed-lineage leukemia 3 (MLL3)/MLL4 chromatin-modifying complex, the mechanisms for how PTIP promotes transcription remain unclear. Here we dissected the minimal structural requirements of PTIP and its different protein complexes using quantitative proteomics in primary lymphocytes. We found that PTIP functions in transcription and CSR separately from its association with the MLL3/MLL4 complex and from its localization to sites of DNA damage. We identified a tandem BRCT domain of PTIP that is sufficient for CSR and identified PA1 as its main functional protein partner. Collectively, we provide genetic and biochemical evidence that a PTIP-PA1 subcomplex functions independently from the MLL3/MLL4 complex to mediate transcription during CSR. These results further our understanding of how multifunctional chromatin-modifying complexes are organized by subcomplexes that harbor unique and distinct activities. PMID:26744420

  12. A PTIP–PA1 subcomplex promotes transcription for IgH class switching independently from the associated MLL3/MLL4 methyltransferase complex

    PubMed Central

    Starnes, Linda M.; Su, Dan; Pikkupeura, Laura M.; Weinert, Brian T.; Santos, Margarida A.; Mund, Andreas; Soria, Rebeca; Cho, Young-Wook; Pozdnyakova, Irina; Kubec Højfeldt, Martina; Vala, Andrea; Yang, Wenjing; López-Méndez, Blanca; Lee, Ji-Eun; Peng, Weiqun; Yuan, Joan; Ge, Kai; Montoya, Guillermo; Nussenzweig, André; Choudhary, Chunaram; Daniel, Jeremy A.

    2016-01-01

    Class switch recombination (CSR) diversifies antibodies for productive immune responses while maintaining stability of the B-cell genome. Transcription at the immunoglobulin heavy chain (Igh) locus targets CSR-associated DNA damage and is promoted by the BRCT domain-containing PTIP (Pax transactivation domain-interacting protein). Although PTIP is a unique component of the mixed-lineage leukemia 3 (MLL3)/MLL4 chromatin-modifying complex, the mechanisms for how PTIP promotes transcription remain unclear. Here we dissected the minimal structural requirements of PTIP and its different protein complexes using quantitative proteomics in primary lymphocytes. We found that PTIP functions in transcription and CSR separately from its association with the MLL3/MLL4 complex and from its localization to sites of DNA damage. We identified a tandem BRCT domain of PTIP that is sufficient for CSR and identified PA1 as its main functional protein partner. Collectively, we provide genetic and biochemical evidence that a PTIP–PA1 subcomplex functions independently from the MLL3/MLL4 complex to mediate transcription during CSR. These results further our understanding of how multifunctional chromatin-modifying complexes are organized by subcomplexes that harbor unique and distinct activities. PMID:26744420

  13. Addressing the Problem of Switched Class Labels in Latent Variable Mixture Model Simulation Studies

    ERIC Educational Resources Information Center

    Tueller, Stephen J.; Drotar, Scott; Lubke, Gitta H.

    2011-01-01

    The discrimination between alternative models and the detection of latent classes in the context of latent variable mixture modeling depends on sample size, class separation, and other aspects that are related to power. Prior to a mixture analysis it is useful to investigate model performance in a simulation study that reflects the research…

  14. B cell Rab7 mediates induction of activation-induced cytidine deaminase expression and class-switching in T-dependent and T-independent antibody responses.

    PubMed

    Pone, Egest J; Lam, Tonika; Lou, Zheng; Wang, Rui; Chen, Yuhui; Liu, Dongfang; Edinger, Aimee L; Xu, Zhenming; Casali, Paolo

    2015-04-01

    Class switch DNA recombination (CSR) is central to the maturation of the Ab response because it diversifies Ab effector functions. Like somatic hypermutation, CSR requires activation-induced cytidine deaminase (AID), whose expression is restricted to B cells, as induced by CD40 engagement or dual TLR-BCR engagement (primary CSR-inducing stimuli). By constructing conditional knockout Igh(+/C)γ(1-cre)Rab7(fl/fl) mice, we identified a B cell-intrinsic role for Rab7, a small GTPase involved in intracellular membrane functions, in mediating AID induction and CSR. Igh(+/C)γ(1-cre)Rab7(fl/fl) mice displayed normal B and T cell development and were deficient in Rab7 only in B cells undergoing Igh(C)γ(1-cre) Iγ1-Sγ1-Cγ1-cre transcription, as induced--like Igh germline Iγ1-Sγ1-Cγ1 and Iε-Sε-Cε transcription--by IL-4 in conjunction with a primary CSR-inducing stimulus. These mice could not mount T-independent or T-dependent class-switched IgG1 or IgE responses while maintaining normal IgM levels. Igh(+/C)γ(1-cre)Rab7(fl/fl) B cells showed, in vivo and in vitro, normal proliferation and survival, normal Blimp-1 expression and plasma cell differentiation, as well as intact activation of the noncanonical NF-κB, p38 kinase, and ERK1/2 kinase pathways. They, however, were defective in AID expression and CSR in vivo and in vitro, as induced by CD40 engagement or dual TLR1/2-, TLR4-, TLR7-, or TLR9-BCR engagement. In Igh(+/C)γ(1-cre)Rab7(fl/fl) B cells, CSR was rescued by enforced AID expression. These findings, together with our demonstration that Rab7-mediated canonical NF-κB activation, as critical to AID induction, outline a novel role of Rab7 in signaling pathways that lead to AID expression and CSR, likely by promoting assembly of signaling complexes along intracellular membranes. PMID:25740947

  15. MHC Class IIB Exon 2 Polymorphism in the Grey Partridge (Perdix perdix) Is Shaped by Selection, Recombination and Gene Conversion

    PubMed Central

    Bryjová, Anna; Albrecht, Tomáš; Bryja, Josef

    2013-01-01

    Among bird species, the most studied major histocompatibility complex (MHC) is the chicken MHC. Although the number of studies on MHC in free-ranging species is increasing, the knowledge on MHC variation in species closely related to chicken is required to understand the peculiarities of bird MHC evolution. Here we describe the variation of MHC class IIB (MHCIIB) exon 2 in a population of the Grey partridge (Perdix perdix), a species of high conservation concern throughout Europe and an emerging galliform model in studies of sexual selection. We found 12 alleles in 108 individuals, but in comparison to other birds surprisingly many sites show signatures of historical positive selection. Individuals displayed between two to four alleles both on genomic and complementary DNA, suggesting the presence of two functional MHCIIB loci. Recombination and gene conversion appear to be involved in generating MHCIIB diversity in the Grey partridge; two recombination breakpoints and several gene conversion events were detected. In phylogenetic analysis of galliform MHCIIB, the Grey partridge alleles do not cluster together, but are scattered through the tree instead. Thus, our results indicate that the Grey partridge MHCIIB is comparable to most other galliforms in terms of copy number and population polymorphism. PMID:23935938

  16. Oxazines: A New Class of Second-Order Nonlinear Optical Switches.

    PubMed

    Beaujean, Pierre; Bondu, Flavie; Plaquet, Aurélie; Garcia-Amorós, Jaume; Cusido, Janet; Raymo, Françisco M; Castet, Frédéric; Rodriguez, Vincent; Champagne, Benoît

    2016-04-20

    A combined experimental-theoretical investigation has revealed that oxazine-based compounds are multiaddressable, multistate, and multifunctional molecular switches exhibiting contrasts of both linear and second-order nonlinear optical properties. The switching properties are particularly large when the substituent is a donor group. In this study, the cleavage of the C-O bond at the junction of the indole and oxazine cycles (of the closed a forms) is acido-triggered, leading to an open form (b(+)) characterized by larger first hyperpolarizabilities (βHRS) and smaller excitation energies than in the closed form. These results are confirmed and interpreted utilizing ab initio calculations that have been carried out on a broad set of compounds to unravel the role of the substituent. With respect to acceptor groups, oxazines bearing donor groups are characterized not only by larger βHRS and βHRS contrast ratios but also by smaller excitation energies, larger opening-induced charge transfer, and reduction of the bond length alternation, as well as smaller Gibbs energies of the opening reaction. Compared to protonated open forms (b(+)), calculations on the zwitterionic open forms (b) have pointed out similarities in the long-wavelength UV/vis absorption spectra, whereas their βHRS values might differ strongly as a function of the substituent. Indeed, the open forms present two NLOphores, the indoleninium-substituent entity and the nitrophenol (present in the protonated open form, b(+)) or nitrophenolate (present in the zwitterionic open form, b) moiety. Then, nitrophenolate displays a larger first hyperpolarizability than nitrophenol and the β tensor of the two entities might reinforce or cancel each other. PMID:26996994

  17. Hierarchical multiobjective routing model in Multiprotocol Label Switching networks with two service classes - a Pareto archive strategy

    NASA Astrophysics Data System (ADS)

    Girão-Silva, Rita; Craveirinha, José; Clímaco, João

    2012-05-01

    The article begins by reviewing a two-level hierarchical multicriteria routing model for Multiprotocol Label Switching networks with two service classes (QoS, i.e. with Quality of Service requirements, and Best Effort services) and alternative routing, as well as the foundations of a heuristic resolution approach, previously proposed by the authors. Afterwards a new variant of this heuristic approach, which includes a Pareto archive strategy, is described. In this archive, non-dominated solutions obtained throughout the heuristic are kept. At the end of the main procedure of the heuristic, these solutions are evaluated and a final solution for the routing problem is chosen using a reference point-based approach. The application of this procedure to two test networks will show, with analytic and discrete-event simulation models, that, in certain initial conditions, this approach provides improvements in the final results concerning the top-level objective functions, especially in more 'difficult' situations detected through sensitivity analysis.

  18. A recombinant, soluble, single-chain class I major histocompatibility complex molecule with biological activity.

    PubMed Central

    Mage, M G; Lee, L; Ribaudo, R K; Corr, M; Kozlowski, S; McHugh, L; Margulies, D H

    1992-01-01

    Heterodimeric class I major histocompatibility complex molecules, which consist of a 45-kDa heavy-chain and a 12-kDa beta 2-microglobulin (beta 2m) light chain, bind endogenously synthesized peptides for presentation to antigen-specific T cells. We have synthesized a gene encoding a single-chain, soluble class I molecule derived from mouse H-2Dd, in which the carboxyl terminus of beta 2m is linked via a peptide spacer to the amino terminus of the heavy chain. The chimeric protein is secreted efficiently from transfected L cells, is thermostable, and when loaded with an appropriate antigenic peptide, stimulates an H-2Dd-restricted antigen-specific T-cell hybridoma. Thus, functional binding of peptide does not require the complete dissociation of beta 2m, implying that a heavy chain/peptide complex is not an obligate intermediate in the assembly of the heavy-chain/beta 2m/peptide heterotrimer. Single-chain major histocompatibility complex molecules uniformly loaded with peptide have potential uses for structural studies, toxin or fluor conjugates, and vaccines. Images PMID:1438262

  19. Specific features of switch-on processes in high-voltage (18 kV class) optically triggered 4H-SiC thyristors

    NASA Astrophysics Data System (ADS)

    Mnatsakanov, T. T.; Yurkov, S. N.; Levinshtein, M. E.; Cheng, L.; Palmour, J. W.

    2014-05-01

    A computer simulation has been carried out to analyze and explain the specific features of the switch-on process in super-high-voltage (18 kV class) optically triggered 4H-SiC thyristors. It is shown that the previously experimentally observed two-stage character of the switch-on processes can be understood in a one-dimensional approach. The two-stage turn-on of the 18 kV structures is accounted for by the rather slow current rise in the structure with a very wide (160 µm) blocking base. In full agreement with the experimental results, a simulation in the frame of the same approximations demonstrated that, in the 12 kV class thyristor structures with a blocking base thickness of 90 µm, the current increases monotonically during the switch-on process.

  20. The Molecular Switching Mechanism at the Conserved D(E)RY Motif in Class-A GPCRs.

    PubMed

    Sandoval, Angelica; Eichler, Stefanie; Madathil, Sineej; Reeves, Philip J; Fahmy, Karim; Böckmann, Rainer A

    2016-07-12

    The disruption of ionic and H-bond interactions between the cytosolic ends of transmembrane helices TM3 and TM6 of class-A (rhodopsin-like) G protein-coupled receptors (GPCRs) is a hallmark for their activation by chemical or physical stimuli. In the bovine photoreceptor rhodopsin, this is accompanied by proton uptake at Glu(134) in the class-conserved D(E)RY motif. Studies on TM3 model peptides proposed a crucial role of the lipid bilayer in linking protonation to stabilization of an active state-like conformation. However, the molecular details of this linkage could not be resolved and have been addressed in this study by molecular dynamics (MD) simulations on TM3 model peptides in a bilayer of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC). We show that protonation of the conserved glutamic acid alters the peptide insertion depth in the membrane, its side-chain rotamer preferences, and stabilizes the C-terminal helical structure. These factors contribute to the rise of the side-chain pKa (> 6) and to reduced polarity around the TM3 C terminus as confirmed by fluorescence spectroscopy. Helix stabilization requires the protonated carboxyl group; unexpectedly, this stabilization could not be evoked with an amide in MD simulations. Additionally, time-resolved Fourier transform infrared (FTIR) spectroscopy of TM3 model peptides revealed a different kinetics for lipid ester carbonyl hydration, suggesting that the carboxyl is linked to more extended H-bond clusters than an amide. Remarkably, this was seen as well in DOPC-reconstituted Glu(134)- and Gln(134)-containing bovine opsin mutants and demonstrates that the D(E)RY motif is a hydrated microdomain. The function of the D(E)RY motif as a proton switch is suggested to be based on the reorganization of the H-bond network at the membrane interface. PMID:27410736

  1. Distinct class of DNA-binding domains is exemplified by a master regulator of phenotypic switching in Candida albicans

    PubMed Central

    Lohse, Matthew B.; Zordan, Rebecca E.; Cain, Christopher W.; Johnson, Alexander D.

    2010-01-01

    Among the most important classes of regulatory proteins are the sequence-specific DNA-binding proteins that control transcription through the occupancy of discrete DNA sequences within genomes. Currently, this class of proteins encompasses at least 37 distinct structural superfamilies and more than 100 distinct structural motifs. In this paper, we examine the transcriptional regulator Wor1, a master regulator of white-opaque switching in the human fungal pathogen Candida albicans. As assessed by a variety of algorithms, this protein has no sequence or structural similarity to any known DNA-binding protein. It is, however, conserved across the vast fungal lineage, with a 300aa region of sequence conservation. Here, we show that this 300aa region of Wor1 exhibits sequence-specific DNA binding and therefore represents a new superfamily of DNA-binding proteins. We identify the 14-nucleotide-pair DNA sequence recognized by Wor1, characterize the site through mutational analysis, and demonstrate that this sequence is sufficient for the Wor1-dependent activation of transcription in vivo. Within the 300aa DNA-binding conserved region, which we have termed the WOPR box, are two domains (WOPRa and WOPRb), dissimilar to each other but especially well-conserved across the fungal lineage. We show that the WOPR box binds DNA as a monomer and that neither domain, when expressed and purified separately, exhibits sequence-specific binding. DNA binding is restored, however, when the two isolated domains are added together. These results indicate that the WOPR family of DNA-binding proteins involves an unusual coupling between two dissimilar, covalently linked domains. PMID:20660774

  2. C-type lectin-like receptor LOX-1 promotes dendritic cell-mediated class-switched B cell responses.

    PubMed

    Joo, HyeMee; Li, Dapeng; Dullaers, Melissa; Kim, Tae-Whan; Duluc, Dorothee; Upchurch, Katherine; Xue, Yaming; Zurawski, Sandy; Le Grand, Roger; Liu, Yong-Jun; Kuroda, Marcelo; Zurawski, Gerard; Oh, SangKon

    2014-10-16

    Lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) is a pattern-recognition receptor for a variety of endogenous and exogenous ligands. However, LOX-1 function in the host immune response is not fully understood. Here, we report that LOX-1 expressed on dendritic cells (DCs) and B cells promotes humoral responses. On B cells LOX-1 signaling upregulated CCR7, promoting cellular migration toward lymphoid tissues. LOX-1 signaling on DCs licensed the cells to promote B cell differentiation into class-switched plasmablasts and led to downregulation of chemokine receptor CXCR5 and upregulation of chemokine receptor CCR10 on plasmablasts, enabling their exit from germinal centers and migration toward local mucosa and skin. Finally, we found that targeting influenza hemagglutinin 1 (HA1) subunit to LOX-1 elicited HA1-specific protective antibody responses in rhesus macaques. Thus, LOX-1 expressed on B cells and DC cells has complementary functions to promote humoral immune responses. PMID:25308333

  3. Immunoglobulin class-switched B cells provide an active immune axis between CNS and periphery in multiple sclerosis

    PubMed Central

    Wang, Shengzhi; Pitts, Steven J.; Sundar, Purnima D.; Telman, Dilduz; Zhao, Lora Z.; Derstine, Mia; Abounasr, Aya; Hauser, Stephen L.; von Büdingen, H.-Christian

    2014-01-01

    In multiple sclerosis (MS), an exchange of lymphocytes, in particular B cells, between the central nervous system (CNS) and periphery is believed to be required for the maintenance of active disease. Therapeutic monoclonal antibodies that prevent lymphocytes from crossing the blood-brain barrier (BBB) or induce near-complete peripheral B cell depletion rapidly mitigate MS disease activity. Using next-generation sequencing technology, we recently found that clonally related B cells exist in the cerebrospinal fluid (CSF) and peripheral blood (PB) of MS patients, establishing the existence of an immune axis across the BBB. However, it remains unclear which subpopulations of the highly diverse peripheral B cell compartment share antigen-specificity with intrathecal B cell repertoires, and whether their antigen stimulation occurs on both sides of the BBB. To address these questions, we combined flow cytometry sorting of PB B cell subsets with deep immune repertoire sequencing of CSF and PB B cells. Immunoglobulin (IgM and IgG) heavy chain variable (VH) region repertoires of five PB B cell subsets from MS patients (n=8) were compared with their CSF Ig-VH transcriptomes. In 6 of 8 patients, we identified peripheral CD27+IgD−memory B cells, CD27hiCD38hi plasma cells/plasmablasts, or CD27−IgD− B cells providing an immune connection to the CNS compartment. Pinpointing Ig class-switched B cells as key component of the immune axis thought to contribute to ongoing MS disease activity strengthens the rationale of current therapeutic strategies and may lead to more targeted approaches. PMID:25100740

  4. Cytokine switch and bystander suppression of autoimmune responses to multiple antigens in experimental autoimmune encephalomyelitis by a single recombinant T-cell receptor ligand.

    PubMed

    Sinha, Sushmita; Subramanian, Sandhya; Miller, Lisa; Proctor, Thomas M; Roberts, Chris; Burrows, Gregory G; Vandenbark, Arthur A; Offner, Halina

    2009-03-25

    Recombinant T-cell receptor ligands (RTLs) can reverse clinical and histological signs of experimental autoimmune encephalomyelitis (EAE) in an antigen-specific manner, and are currently in clinical trials for treatment of subjects with multiple sclerosis (MS). Antigen specificity of RTL raises the question as to whether this treatment would be successful in MS patients where target antigens are unknown. Using spinal cord homogenate or combinations of two different peptides to induce disease, we found that treatment with single RTL could reverse EAE as long as targeted T-cells were present. Therapy with three different RTLs each caused a significant reduction in IL-17 and increases in IL-10 and IL-13 in peptide-activated splenocytes, reduced proliferation of both cognate and bystander specificities of lymph node cells, and reduced inflammatory lesions and secreted IL-17 and IL-2 from peptide-activated spinal cord cells. These results show that treatment with single RTLs can induce a cytokine switch in cognate T-cells that inhibits both the target and bystander T-cells, providing new evidence for the potential applicability of RTL therapy in MS. PMID:19321778

  5. Cytokine Switch and Bystander Suppression of Autoimmune Responses to Multiple Antigens in Experimental Autoimmune Encephalomyelitis by a Single Recombinant T-Cell Receptor Ligand

    PubMed Central

    Sinha, Sushmita; Subramanian, Sandhya; Miller, Lisa; Proctor, Thomas M.; Roberts, Chris; Burrows, Gregory G.; Vandenbark, Arthur A.; Offner, Halina

    2009-01-01

    Recombinant T-cell receptor ligands (RTLs) can reverse clinical and histological signs of experimental autoimmune encephalomyelitis (EAE) in an antigen-specific manner, and are currently in clinical trials for treatment of subjects with multiple sclerosis (MS). Antigen specificity of RTL raises the question as to whether this treatment would be successful in MS patients where target antigens are unknown. Using spinal cord homogenate or combinations of two different peptides to induce disease,we found that treatment with single RTL could reverse EAE as long as targeted T-cells were present. Therapy with three different RTLs each caused a significant reduction in IL-17 and increases in IL-10 and IL-13 in peptide-activated splenocytes, reduced proliferation of both cognate and bystander specificities of lymph node cells, and reduced inflammatory lesions and secreted IL-17 and IL-2 from peptide-activated spinal cord cells. These results show that treatment with single RTLs can induce a cytokine switch in cognate T-cells that inhibits both the target and bystander T-cells, providing new evidence for the potential applicability of RTL therapy in MS. PMID:19321778

  6. The Carboxy-Terminal αN Helix of the Archaeal XerA Tyrosine Recombinase Is a Molecular Switch to Control Site-Specific Recombination

    PubMed Central

    Serre, Marie-Claude; El Arnaout, Toufic; Brooks, Mark A.; Durand, Dominique; Lisboa, Johnny; Lazar, Noureddine; Raynal, Bertrand; van Tilbeurgh, Herman; Quevillon-Cheruel, Sophie

    2013-01-01

    Tyrosine recombinases are conserved in the three kingdoms of life. Here we present the first crystal structure of a full-length archaeal tyrosine recombinase, XerA from Pyrococcus abyssi, at 3.0 Å resolution. In the absence of DNA substrate XerA crystallizes as a dimer where each monomer displays a tertiary structure similar to that of DNA-bound Tyr-recombinases. Active sites are assembled in the absence of dif except for the catalytic Tyr, which is extruded and located equidistant from each active site within the dimer. Using XerA active site mutants we demonstrate that XerA follows the classical cis-cleavage reaction, suggesting rearrangements of the C-terminal domain upon DNA binding. Surprisingly, XerA C-terminal αN helices dock in cis in a groove that, in bacterial tyrosine recombinases, accommodates in trans αN helices of neighbour monomers in the Holliday junction intermediates. Deletion of the XerA C-terminal αN helix does not impair cleavage of suicide substrates but prevents recombination catalysis. We propose that the enzymatic cycle of XerA involves the switch of the αN helix from cis to trans packing, leading to (i) repositioning of the catalytic Tyr in the active site in cis and (ii) dimer stabilisation via αN contacts in trans between monomers. PMID:23667562

  7. Spectrum Recombination.

    ERIC Educational Resources Information Center

    Greenslade, Thomas B., Jr.

    1984-01-01

    Describes several methods of executing lecture demonstrations involving the recombination of the spectrum. Groups the techniques into two general classes: bringing selected portions of the spectrum together using lenses or mirrors and blurring the colors by rapid movement or foreshortening. (JM)

  8. The Arabidopsis MutS homolog AtMSH4 functions at an early step in recombination: evidence for two classes of recombination in Arabidopsis

    PubMed Central

    Higgins, James D.; Armstrong, Susan J.; Franklin, F. Christopher H.; Jones, Gareth H.

    2004-01-01

    MSH4, a meiosis-specific member of the MutS-homolog family of genes, is required for normal levels of recombination and fertility in budding yeast, mouse, and Caenorhabditis elegans. In this paper, we report the identification and characterization of the Arabidopsis homolog of MSH4 (AtMSH4). We demonstrate that AtMSH4 expression can only be detected in floral tissues, consistent with a role in reproduction. Immunofluorescence studies indicate that its expression is limited to early meiotic prophase I, preceding the synapsis of homologous chromosomes. A T-DNA insertional mutant (Atmsh4) exhibited normal vegetative growth but a severe reduction in fertility, consistent with a meiotic defect; this was confirmed by cytological analysis of meiosis. RNAi-induced down-regulation of the MSH4 gene resulted in a similar fertility and meiotic phenotype. We demonstrate that prophase I chromosome synapsis is delayed and may be incomplete in Atmsh4, and metaphase I chiasma frequency is greatly reduced to ∼15% of wild type, leading to univalence and nondisjunction. We show that these residual chiasmata are randomly distributed among cells and chromosomes. These features of chiasma frequency and distribution in Atmsh4 show close parallels to MSH4-independent crossovers in budding yeast that have been proposed to originate by a separate pathway. Furthermore, the characteristics of the MSH4-independent chiasmata in the Atmsh4 mutant closely parallel those of second-pathway crossovers that have been postulated from Arabidopsis crossover analysis and mathematical modeling. Taken together, this evidence strongly indicates that Arabidopsis possesses two crossover pathways. PMID:15489296

  9. Markov-switching multifractal models as another class of random-energy-like models in one-dimensional space

    NASA Astrophysics Data System (ADS)

    Saakian, David B.

    2012-03-01

    We map the Markov-switching multifractal model (MSM) onto the random energy model (REM). The MSM is, like the REM, an exactly solvable model in one-dimensional space with nontrivial correlation functions. According to our results, four different statistical physics phases are possible in random walks with multifractal behavior. We also introduce the continuous branching version of the model, calculate the moments, and prove multiscaling behavior. Different phases have different multiscaling properties.

  10. The diabetogenic mouse MHC class II molecule I-A[subscript g7] is endowed with a switch that modulates TCR affinity

    SciTech Connect

    Yoshida, Kenji; Corper, Adam L.; Herro, Rana; Jabri, Bana; Wilson, Ian A.; Teyton, Luc

    2011-11-16

    Genetic susceptibility to autoimmunity is frequently associated with specific MHC alleles. Diabetogenic MHC class II molecules, such as human HLA-DQ8 and mouse I-A{sub g7}, typically have a small, uncharged amino acid residue at position 57 of their {beta} chain ({beta}57); this results in the absence of a salt bridge between {beta}57 and Arg{alpha}76, which is adjacent to the P9 pocket of the peptide-binding groove. However, the influence of Arg{alpha}76 on the selection of the TCR repertoire remains unknown, particularly when the MHC molecule binds a peptide with a neutral amino acid residue at position P9. Here, we have shown that diabetogenic MHC class II molecules bound to a peptide with a neutral P9 residue primarily selected and expanded cells expressing TCRs bearing a negatively charged residue in the first segment of their complementarity determining region 3{beta}. The crystal structure of one such TCR in complex with I-A{sub g7} bound to a peptide containing a neutral P9 residue revealed that a network of favorable long-range (greater than 4 {angstrom}) electrostatic interactions existed among Arg{alpha}76, the neutral P9 residue, and TCR, which supported the substantially increased TCR/peptide-MHC affinity. This network could be modulated or switched to a lower affinity interaction by the introduction of a negative charge at position P9 of the peptide. Our results support the existence of a switch at residue {beta}57 of the I-Ag7 and HLA-DQ8 class II molecules and potentially link normal thymic TCR selection with abnormal peripheral behavior.

  11. The diabetogenic mouse MHC class II molecule I-A[superscript g7] is endowed with a switch that modulates TCR affinity

    SciTech Connect

    Yoshida, Kenji; Corper, Adam L.; Herro, Rana; Jabri, Bana; Wilson, Ian A.; Teyton, Luc

    2010-07-22

    Genetic susceptibility to autoimmunity is frequently associated with specific MHC alleles. Diabetogenic MHC class II molecules, such as human HLA-DQ8 and mouse I-A{sup g7}, typically have a small, uncharged amino acid residue at position 57 of their {beta} chain ({beta}57); this results in the absence of a salt bridge between {beta}57 and Arg{alpha}76, which is adjacent to the P9 pocket of the peptide-binding groove. However, the influence of Arg{alpha}76 on the selection of the TCR repertoire remains unknown, particularly when the MHC molecule binds a peptide with a neutral amino acid residue at position P9. Here, we have shown that diabetogenic MHC class II molecules bound to a peptide with a neutral P9 residue primarily selected and expanded cells expressing TCRs bearing a negatively charged residue in the first segment of their complementarity determining region 3{beta}. The crystal structure of one such TCR in complex with I-Ag7 bound to a peptide containing a neutral P9 residue revealed that a network of favorable long-range (greater than 4 {angstrom}) electrostatic interactions existed among Arg{alpha}76, the neutral P9 residue, and TCR, which supported the substantially increased TCR/peptide-MHC affinity. This network could be modulated or switched to a lower affinity interaction by the introduction of a negative charge at position P9 of the peptide. Our results support the existence of a switch at residue {beta}57 of the I-A{sup g7} and HLA-DQ8 class II molecules and potentially link normal thymic TCR selection with abnormal peripheral behavior.

  12. Polymorphism, recombination and alternative unscrambling in the DNA polymerase alpha gene of the ciliate Stylonychia lemnae (Alveolata; class Spirotrichea).

    PubMed Central

    Ardell, David H; Lozupone, Catherine A; Landweber, Laura F

    2003-01-01

    DNA polymerase alpha is the most highly scrambled gene known in stichotrichous ciliates. In its hereditary micronuclear form, it is broken into >40 pieces on two loci at least 3 kb apart. Scrambled genes must be reassembled through developmental DNA rearrangements to yield functioning macronuclear genes, but the mechanism and accuracy of this process are unknown. We describe the first analysis of DNA polymorphism in the macronuclear version of any scrambled gene. Six functional haplotypes obtained from five Eurasian strains of Stylonychia lemnae were highly polymorphic compared to Drosophila genes. Another incompletely unscrambled haplotype was interrupted by frameshift and nonsense mutations but contained more silent mutations than expected by allelic inactivation. In our sample, nucleotide diversity and recombination signals were unexpectedly high within a region encompassing the boundary of the two micronuclear loci. From this and other evidence we infer that both members of a long repeat at the ends of the loci provide alternative substrates for unscrambling in this region. Incongruent genealogies and recombination patterns were also consistent with separation of the two loci by a large genetic distance. Our results suggest that ciliate developmental DNA rearrangements may be more probabilistic and error prone than previously appreciated and constitute a potential source of macronuclear variation. From this perspective we introduce the nonsense-suppression hypothesis for the evolution of ciliate altered genetic codes. We also introduce methods and software to calculate the likelihood of hemizygosity in ciliate haplotype samples and to correct for multiple comparisons in sliding-window analyses of Tajima's D. PMID:14704164

  13. Introduction of protein or DNA delivered via recombinant Salmonella typhimurium into the major histocompatibility complex class I presentation pathway of macrophages.

    PubMed

    Catic, A; Dietrich, G; Gentschev, I; Goebel, W; Kaufmann, S H; Hess, J

    1999-02-01

    Recombinant (r) Salmonella typhimurium aroA strains which display the hen egg ovalbumin OVA(257-264) peptide SIINFEKL in secreted form were constructed. In addition, attenuated rS. typhimurium pcDNA-OVA constructs harbouring a eukaryotic expression plasmid encoding complete OVA were used to introduce the immunodominant OVA(257-264) epitope into the major histocompatibility complex (MHC) class I presentation pathway. Both modes of antigen delivery (DNA and protein) by Salmonella vaccine carriers stimulated OVA(257-264)-specific CD8 T-cell hybridomas. An in vitro infection system was established that allowed both rSalmonella carrier devices to facilitate MHC class I delivery of OVA(257-264) by coexpression of listeriolysin (Hly) or by coinfection with rS. typhimurium Hlys (Hess J., Gentschev I., Miko D., Welzel M., Ladel C., Goebel W., Kaufmann S.H.E., Proc. Natl. Acad. Sci. USA 93 (1996) 1458-1463). Coexpression of Hly and coinfection with rS. typhimurium Hlys slightly improved MHC class I processing of OVA. Our data provide further evidence for the feasibility of attenuated, Hly-expressing rS. typhimurium carriers secreting heterologous antigens or harbouring heterologous DNA as effective vaccines for stimulating CD8 T cells in addition to CD4 T cells. PMID:10594975

  14. Treatment with Recombinant Trichinella spiralis Cathepsin B-like Protein Ameliorates Intestinal Ischemia/Reperfusion Injury in Mice by Promoting a Switch from M1 to M2 Macrophages.

    PubMed

    Liu, Wei-Feng; Wen, Shi-Hong; Zhan, Jian-Hua; Li, Yun-Sheng; Shen, Jian-Tong; Yang, Wen-Jing; Zhou, Xing-Wang; Liu, Ke-Xuan

    2015-07-01

    Intestinal ischemia/reperfusion (I/R) injury, in which macrophages play a key role, can cause high morbidity and mortality. The switch from classically (M1) to alternatively (M2) activated macrophages, which is dependent on the activation of STAT6 signaling, has been shown to protect organs from I/R injuries. In the current study, the effects of recombinant Trichinella spiralis cathepsin B-like protein (rTsCPB) on intestinal I/R injury and the potential mechanism related to macrophage phenotypes switch were investigated. In a mouse I/R model undergoing 60-min intestinal ischemia followed by 2-h or 7-d reperfusion, we demonstrated that intestinal I/R caused significant intestinal injury and induced a switch from M2 to M1 macrophages, evidenced by a decrease in levels of M2 markers (arginase-1 and found in inflammatory zone protein), an increase in levels of M1 markers (inducible NO synthase and CCR7), and a decrease in the ratio of M2/M1 macrophages. RTsCPB reversed intestinal I/R-induced M2-M1 transition and promoted M1-M2 phenotype switch evidenced by a significant decrease in M1 markers, an increase in M2 markers, and the ratio of M2/M1 macrophages. Meanwhile, rTsCPB significantly ameliorated intestinal injury and improved intestinal function and survival rate of animals, accompanied by a decrease in neutrophil infiltration and an increase in cell proliferation in the intestine. However, a selective STAT6 inhibitor, AS1517499, reversed the protective effects of rTsCPB by inhibiting M1 to M2 transition. These findings suggest that intestinal I/R injury causes a switch from M2 to M1 macrophages and that rTsCPB ameliorates intestinal injury by promoting STAT6-dependent M1 to M2 transition. PMID:25987744

  15. Dissection of the role of MHC class II A and E genes in autoimmune susceptibility in murine lupus models with intragenic recombination.

    PubMed

    Zhang, Danqing; Fujio, Keishi; Jiang, Yi; Zhao, Jingyuan; Tada, Norihiro; Sudo, Katsuko; Tsurui, Hiromichi; Nakamura, Kazuhiro; Yamamoto, Kazuhiko; Nishimura, Hiroyuki; Shira, Toshikazu; Hirose, Sachiko

    2004-09-21

    Systemic lupus erythematosus (SLE) is a multigenic autoimmune disease, and the major histocompatibility complex (MHC) class II polymorphism serves as a key genetic element. In SLE-prone (NZB x NZW)F(1) mice, the MHC H-2(d/z) heterozygosity (H-2(d) of NZB and H-2(z) of NZW) has a strong impact on disease; thus, congenic H-2(d/d) homozygous F(1) mice do not develop severe disease. In this study, we used Ea-deficient intra-H-2 recombination to establish A(d/d)-congenic (NZB x NZW)F(1) mice, with or without E molecule expression, and dissected the role of class II A and E molecules. Here we found that A(d/d) homozygous F(1) mice lacking E molecules developed severe SLE similar to that seen in wild-type F1 mice, including lupus nephritis, autoantibody production, and spontaneously occurring T cell activation. Additional evidence revealed that E molecules prevent the disease in a dose-dependent manner; however, the effect is greatly influenced by the haplotype of A molecules, because wild-type H-2(d/z) F(1) mice develop SLE, despite E molecule expression. Studies on the potential of dendritic cells to present a self-antigen chromatin indicated that dendritic cells from wild-type F(1) mice induced a greater response of chromatin-specific T cells than did those from A(d/d) F(1) mice, irrespective of the presence or absence of E molecules, suggesting that the self-antigen presentation is mediated by A, but not by E, molecules. Our mouse models are useful for analyzing the molecular mechanisms by which MHC class II regions regulate the process of autoimmune responses. PMID:15361580

  16. The GPS Motif Is a Molecular Switch for Bimodal Activities of Adhesion Class G Protein-Coupled Receptors

    PubMed Central

    Prömel, Simone; Frickenhaus, Marie; Hughes, Samantha; Mestek, Lamia; Staunton, David; Woollard, Alison; Vakonakis, Ioannis; Schöneberg, Torsten; Schnabel, Ralf; Russ, Andreas P.; Langenhan, Tobias

    2012-01-01

    Summary Adhesion class G protein-coupled receptors (aGPCR) form the second largest group of seven-transmembrane-spanning (7TM) receptors whose molecular layout and function differ from canonical 7TM receptors. Despite their essential roles in immunity, tumorigenesis, and development, the mechanisms of aGPCR activation and signal transduction have remained obscure to date. Here, we use a transgenic assay to define the protein domains required in vivo for the activity of the prototypical aGPCR LAT-1/Latrophilin in Caenorhabditis elegans. We show that the GPCR proteolytic site (GPS) motif, the molecular hallmark feature of the entire aGPCR class, is essential for LAT-1 signaling serving in two different activity modes of the receptor. Surprisingly, neither mode requires cleavage but presence of the GPS, which relays interactions with at least two different partners. Our work thus uncovers the versatile nature of aGPCR activity in molecular detail and places the GPS motif in a central position for diverse protein-protein interactions. PMID:22938866

  17. Biochemical characterization of a recombinant plant class III chitinase from the pitcher of the carnivorous plant Nepenthes alata.

    PubMed

    Ishisaki, Kana; Arai, Sachiko; Hamada, Tatsuro; Honda, Yuji

    2012-11-01

    A class III chitinase belonging to the GH18 family from Nepenthes alata (NaCHIT3) was expressed in Escherichia coli. The enzyme exhibited hydrolytic activity toward colloidal chitin, ethylene glycol chitin, and (GlcNAc)(n) (n=5 and 6). The enzyme hydrolyzed the fourth glycosidic linkage from the non-reducing end of (GlcNAc)(6). The anomeric form of the products indicated it was a retaining enzyme. The colloidal chitin hydrolytic reaction displayed high activity between pH 3.9 and 6.9, but the pH optimum of the (GlcNAc)(6) hydrolytic reaction was 3.9 at 37 °C. The optimal temperature for activity was 65 °C in 50 mM sodium acetate buffer (pH 3.9). The pH optima of NaCHIT3 and NaCHIT1 might be related to their roles in chitin degradation in the pitcher fluid. PMID:23026711

  18. New Class of Leukemogenic Ecotropic Recombinant Murine Leukemia Virus Isolated from Radiation-Induced Thymomas of C57BL/6 Mice

    PubMed Central

    Rassart, E.; Sankar-Mistry, P.; Lemay, G.; DesGroseillers, L.; Jolicoeur, P.

    1983-01-01

    We previously reported the establishment of several lymphoid cell lines from X-ray-induced thymomas of C57BL/Ka mice, and all, except one, produce retroviruses (P. Sankar-Mistry and P. Jolicoeur, J. Virol.35:270-275, 1980). Biological characterization of five of these new primary radiation leukemia viruses (RadLVs) indicated that they had a B-tropic, fibrotropic, and ecotropic host range and were leukemogenic when reinjected into C57BL/Ka newborn mice. The leukemogenic potential of one isolate (G6T2) was further assessed and shown to be retained after prolonged passaging on fibroblasts in vitro. Restriction endonuclease analysis of the DNA of four of our new RadLV isolates (G6T2, Ti-7, Ti-8, and Ti-9) revealed that G6T2 and Ti-7 murine leukemia virus (MuLV) genomes had identical restriction maps, whereas Ti-8 and Ti-9 genomes were different from each other and from the G6T2 and Ti-7 genomes. The physical maps of these genomes were similar to that of known ecotropic MuLV genomes (including the C57BL/Ka endogenous ecotropic MuLV) within their long terminal repeats, env, the right portion of pol, and the left portion of gag. However, a region covering the end of gag and the beginning of pol was different and showed several similarities with xenotropic MuLV genomes of BALB/c, AKR, and C58 mice previously mapped. Our results suggest that these primary RadLV genomes are recombinants between the parental ecotropic MuLV genome and a nonecotropic (xenotropic) sequence. This nonecotropic gag-pol region might be important in conferring the leukemogenic potential to these isolates. Therefore, these RadLVs appear to form a new class of leukemogenic recombinant MuLVs recovered from leukemic tissues of mice. They appear to be distinct from the recombinant AKR mink cell focus-inducing MuLVs which have a dual-tropic host range and harbor xenotropic env sequences. To further study the leukemogenic potential of these RadLVs, the genome of one of them (G6T2) was cloned in Charon 21A

  19. Increased immunogenicity and induction of class switching by conjugation of complement C3d to pneumococcal serotype 14 capsular polysaccharide.

    PubMed

    Test, S T; Mitsuyoshi, J; Connolly, C C; Lucas, A H

    2001-05-01

    Previous studies have demonstrated an adjuvant effect for the C3d fragment of complement C3 when coupled to T-dependent protein antigens. In this study, we examined the antibody response to covalent conjugates of C3d and a T-independent antigen, the capsular polysaccharide of serotype 14 Streptococcus pneumoniae (PPS14). We prepared a conjugate of mouse C3d and PPS14 and compared its immunogenicity with that of a conjugate of PPS14 and ovalbumin (OVA). When BALB/c mice were immunized with PPS14-C3d, there was a significant increase in serum anti-PPS14 concentrations compared with either native PPS14 or control PPS14-glycine conjugates. This was accompanied by a switch in anti-PPS14 from predominantly immunoglobulin M (IgM) to IgG1 by day 25 following primary immunization. Following secondary immunization with PPS14-C3d, there was a marked booster response and a further increase in the ratio of IgG1 to IgM anti-PPS14. Although the primary antibody response to the PPS14-OVA conjugate exceeded that induced by immunization with PPS14-C3d, serum anti-PPS14 concentrations after a second injection of PPS14-C3d were nearly identical to those induced by secondary immunization with PPS14-OVA. Experiments with athymic nude mice suggested that T cells were not required for the adjuvant effect of C3d on the primary immune response to PPS14 but were necessary for enhancement of the memory response after a second injection of PPS14-C3d. These studies show that the adjuvant effects of C3d extend to T-independent antigens as well as T-dependent antigens. As a means of harnessing the adjuvant potential of the innate immune system, C3d conjugates may prove useful as a component of vaccines against encapsulated bacteria. PMID:11292721

  20. Increased Immunogenicity and Induction of Class Switching by Conjugation of Complement C3d to Pneumococcal Serotype 14 Capsular Polysaccharide

    PubMed Central

    Test, Samuel T.; Mitsuyoshi, Joyce; Connolly, Charles C.; Lucas, Alexander H.

    2001-01-01

    Previous studies have demonstrated an adjuvant effect for the C3d fragment of complement C3 when coupled to T-dependent protein antigens. In this study, we examined the antibody response to covalent conjugates of C3d and a T-independent antigen, the capsular polysaccharide of serotype 14 Streptococcus pneumoniae (PPS14). We prepared a conjugate of mouse C3d and PPS14 and compared its immunogenicity with that of a conjugate of PPS14 and ovalbumin (OVA). When BALB/c mice were immunized with PPS14-C3d, there was a significant increase in serum anti-PPS14 concentrations compared with either native PPS14 or control PPS14-glycine conjugates. This was accompanied by a switch in anti-PPS14 from predominantly immunoglobulin M (IgM) to IgG1 by day 25 following primary immunization. Following secondary immunization with PPS14-C3d, there was a marked booster response and a further increase in the ratio of IgG1 to IgM anti-PPS14. Although the primary antibody response to the PPS14-OVA conjugate exceeded that induced by immunization with PPS14-C3d, serum anti-PPS14 concentrations after a second injection of PPS14-C3d were nearly identical to those induced by secondary immunization with PPS14-OVA. Experiments with athymic nude mice suggested that T cells were not required for the adjuvant effect of C3d on the primary immune response to PPS14 but were necessary for enhancement of the memory response after a second injection of PPS14-C3d. These studies show that the adjuvant effects of C3d extend to T-independent antigens as well as T-dependent antigens. As a means of harnessing the adjuvant potential of the innate immune system, C3d conjugates may prove useful as a component of vaccines against encapsulated bacteria. PMID:11292721

  1. Evolutionary genetics and vector adaptation of recombinant viruses of the western equine encephalitis antigenic complex provides new insights into alphavirus diversity and host switching

    PubMed Central

    Allison, Andrew B.; Stallknecht, David E.; Holmes, Edward C.

    2014-01-01

    Western equine encephalitis virus (WEEV), Highlands J virus (HJV), and Fort Morgan virus (FMV) are the sole representatives of the WEE antigenic complex of the genus Alphavirus, family Togaviridae, that are endemic to North America. All three viruses have their ancestry in a recombination event involving eastern equine encephalitis virus (EEEV) and a Sindbis (SIN)-like virus that gave rise to a chimeric alphavirus that subsequently diversified into the present-day WEEV, HJV, and FMV. Here, we present a comparative analysis of the genetic, ecological, and evolutionary relationships among these recombinant-origin viruses, including the description of a nsP4 polymerase mutation in FMV that allows it to circumvent the host range barrier to Asian tiger mosquito cells, a vector species that is normally refractory to infection. Notably, we also provide evidence that the recombination event that gave rise to these three WEEV antigenic complex viruses may have occurred in North America. PMID:25463613

  2. MHC class II derived recombinant T cell receptor ligands protect DBA/1LacJ mice from collagen-induced arthritis.

    PubMed

    Huan, Jianya; Kaler, Laurie J; Mooney, Jeffery L; Subramanian, Sandhya; Hopke, Corwyn; Vandenbark, Arthur A; Rosloniec, Edward F; Burrows, Gregory G; Offner, Halina

    2008-01-15

    We previously demonstrated the therapeutic effects of MHC class II derived recombinant T cell receptor ligands (RTL), single-chain two domain complexes of the alpha1 and beta1 domains of MHC class II molecules genetically linked with an immunodominant peptide, in experimental autoimmune encephalomyelitis. In the current study, we produced a monomeric murine I-Aq-derived RTL construct covalently linked with bovine collagen type II peptide (bCII257-270) suitable for use in DBA/1LacJ mice that develop collagen-induced arthritis (CIA), an animal model of human rheumatoid arthritis, after immunization with bCII protein in CFA. In this study, we demonstrate that the I-Aq-derived RTLs reduced the incidence of the disease, suppressed the clinical and histological signs of CIA and induced long-term modulation of T cells specific for arthritogenic Ags. Our results showed that the I-Aq/bCII257-270 molecule could systemically reduce proinflammatory IL-17 and IFN-gamma production and significantly increase anti-inflammatory IL-10, IL-13, and FoxP3 gene expression in splenocytes. Moreover, I-Aq/bCII257-270 molecule could also selectively inhibit IL-1beta, IL-6, and IL-23 expression in local joint tissue. This is the first report demonstrating effective prevention of joint inflammation and clinical signs of CIA with an I-Aq-derived RTL, thus supporting the possible clinical use of this approach for treating rheumatoid arthritis in humans. PMID:18178865

  3. MHC Class II Derived Recombinant T Cell Receptor Ligands Protect DBA/1LacJ Mice from Collagen-Induced Arthritis1

    PubMed Central

    Huan, Jianya; Kaler, Laurie J.; Mooney, Jeffery L.; Subramanian, Sandhya; Hopke, Corwyn; Vandenbark, Arthur A.; Rosloniec, Edward F.; Burrows, Gregory G.; Offner, Halina

    2012-01-01

    We previously demonstrated the therapeutic effects of MHC class II derived recombinant T cell receptor ligands (RTL), single-chain two domain complexes of the α1 and β1 domains of MHC class II molecules genetically linked with an immunodominant peptide, in experimental autoimmune encephalomyelitis. In the current study, we produced a monomeric murine I-Aq-derived RTL construct covalently linked with bovine collagen type II peptide (bCII257–270) suitable for use in DBA/1LacJ mice that develop collagen-induced arthritis (CIA), an animal model of human rheumatoid arthritis, after immunization with bCII protein in CFA. In this study, we demonstrate that the I-Aq-derived RTLs reduced the incidence of the disease, suppressed the clinical and histological signs of CIA and induced long-term modulation of T cells specific for arthritogenic Ags. Our results showed that the I-Aq/bCII257–270 molecule could systemically reduce proinflammatory IL-17 and IFN-γ production and significantly increase anti-inflammatory IL-10, IL-13, and FoxP3 gene expression in splenocytes. Moreover, I-Aq/bCII257–270 molecule could also selectively inhibit IL-1β, IL-6, and IL-23 expression in local joint tissue. This is the first report demonstrating effective prevention of joint inflammation and clinical signs of CIA with an I-Aq-derived RTL, thus supporting the possible clinical use of this approach for treating rheumatoid arthritis in humans. PMID:18178865

  4. Integration-dependent bacteriophage immunity provides insights into the evolution of genetic switches.

    PubMed

    Broussard, Gregory W; Oldfield, Lauren M; Villanueva, Valerie M; Lunt, Bryce L; Shine, Emilee E; Hatfull, Graham F

    2013-01-24

    Genetic switches are critical components of developmental circuits. Because temperate bacteriophages are vastly abundant and greatly diverse, they are rich resources for understanding the mechanisms and evolution of switches and the molecular control of genetic circuitry. Here, we describe a new class of small, compact, and simple switches that use site-specific recombination as the key decision point. The phage attachment site attP is located within the phage repressor gene such that chromosomal integration results in removal of a C-terminal tag that destabilizes the virally encoded form of the repressor. Integration thus not only confers prophage stability but also is a requirement for lysogenic establishment. The variety of these self-contained integration-dependent immunity systems in different genomic contexts suggests that these represent ancestral states in switch evolution from which more-complex switches have evolved. They also provide a powerful toolkit for building synthetic biological circuits. PMID:23246436

  5. Integration-dependent bacteriophage immunity provides insights into the evolution of genetic switches

    PubMed Central

    Broussard, Gregory W.; Oldfield, Lauren M.; Villanueva, Valerie M.; Lunt, Bryce L.; Shine, Emilee E.; Hatfull, Graham F.

    2012-01-01

    SUMMARY Genetic switches are critical components of developmental circuits. Because temperate bacteriophages are vastly abundant and greatly diverse, they are rich resources for understanding the mechanisms and evolution of switches and the molecular control of genetic circuitry. Here, we describe a new class of small, compact, and simple switches that use site-specific recombination as the key decision point. The phage attachment site attP is located within the phage repressor gene such that chromosomal integration results in removal of a C-terminal tag that destabilizes the virally-encoded form of the repressor. Integration thus not only confers prophage stability, but also is a requirement for lysogenic establishment. The variety of these self-contained integration-dependent immunity systems in different genomic contexts suggests that these represent ancestral states in switch evolution from which more complex switches have evolved. They also provide a powerful toolkit for building synthetic biological circuits. PMID:23246436

  6. New Switches for Utility-Scale Inverters: First In-Class Demonstration of a Completely New Type of SiC Bipolar Switch (15kV-20kV) for Utility-Scale Inverters

    SciTech Connect

    2011-12-31

    Solar ADEPT Project: The SiCLAB is developing a new power switch for utility-scale PV inverters that would improve the performance and significantly reduce the size, weight, and energy loss of PV systems. A power switch controls the electrical energy flowing through an inverter, which takes the electrical current from a PV solar panel and converts it into the type and amount of electricity that is compatible with the electric grid. SiCLAB is using silicon carbide (SiC) semiconductors in its new power switches, which are more efficient than the silicon semiconductors used to conduct electricity in most conventional power switches today. Switches with SiC semiconductors can operate at much higher temperatures, as well as higher voltage and power levels than silicon switches. SiC-based power switches are also smaller than those made with silicon alone, so they result in much smaller and lighter electrical devices. In addition to their use in utility-scale PV inverters, SiCLAB’s new power switches can also be used in wind turbines, railways, and other smart grid applications.

  7. λ Recombination and Recombineering.

    PubMed

    Murphy, Kenan C

    2016-05-01

    The bacteriophage λ Red homologous recombination system has been studied over the past 50 years as a model system to define the mechanistic details of how organisms exchange DNA segments that share extended regions of homology. The λ Red system proved useful as a system to study because recombinants could be easily generated by co-infection of genetically marked phages. What emerged from these studies was the recognition that replication of phage DNA was required for substantial Red-promoted recombination in vivo, and the critical role that double-stranded DNA ends play in allowing the Red proteins access to the phage DNA chromosomes. In the past 16 years, however, the λ Red recombination system has gained a new notoriety. When expressed independently of other λ functions, the Red system is able to promote recombination of linear DNA containing limited regions of homology (∼50 bp) with the Escherichia coli chromosome, a process known as recombineering. This review explains how the Red system works during a phage infection, and how it is utilized to make chromosomal modifications of E. coli with such efficiency that it changed the nature and number of genetic manipulations possible, leading to advances in bacterial genomics, metabolic engineering, and eukaryotic genetics. PMID:27223821

  8. Scintigraphic detection of overexpressed c-erbB-2 protooncogene products by a class-switched murine anti-c-erbB-2 protein monoclonal antibody

    SciTech Connect

    Saga, T.; Endo, K.; Akiyama, T.; Sakahara, H.; Koizumi, M.; Watanabe, Y.; Nakai, T.; Hosono, M.; Yamamoto, T.; Toyoshima, K. )

    1991-02-01

    Class-switched monoclonal antibody SV2-61r recognized the extracellular domain of c-erbB-2 protooncogene products separate from the epidermal growth factor receptor. We studied the potential of SV2-61r for evaluating the amplification of c-erbB-2 protooncogene on cancer cells, which has been reported to have prognostic value in adenocarcinoma patients. Radiolabeled SV2-61r specifically bound to various adenocarcinoma cells in addition to c-erbB-2-transfected NIH-3T3 cells (A4) with the affinity constant of 4.4 x 10(8) M-1. SV2-61r injected i.v. localized well to A4 cells xenografted in nude mice. Tumor uptake and localization index of radioiodinated SV2-61r were lower than those of 111In-labeled SV2-61r, probably due to the internalization and dehalogenation of formed antibody-antigen complexes. Biodistribution and specificity of targeting were assessed by comparison among three cells, A4, lung cancer SBC-3 (c-erbB-2 weakly positive) and B-lymphoblastoid Manca cells (c-erbB-2 negative). Tumor:blood ratios, obtained 48 h after injection, were 5.63, 1.45, and 0.68, respectively, indicating the potential of 111In-labeled SV2-61r for evaluating the amplification of c-erbB-2 protooncogene on cancer cells. Because of its close relationship with carcinogenesis and the uniform expression, c-erbB-2 protooncogene products seem to be the optimal target of imaging and therapy of adenocarcinoma patients.

  9. Plasmonic enhanced ultrafast switch.

    SciTech Connect

    Subramania,Ganapathi Subramanian; Reno, John Louis; Passmore, Brandon Scott; Harris, Tom.; Shaner, Eric Arthur; Barrick, Todd A.

    2009-09-01

    Ultrafast electronic switches fabricated from defective material have been used for several decades in order to produce picosecond electrical transients and TeraHertz radiation. Due to the ultrashort recombination time in the photoconductor materials used, these switches are inefficient and are ultimately limited by the amount of optical power that can be applied to the switch before self-destruction. The goal of this work is to create ultrafast (sub-picosecond response) photoconductive switches on GaAs that are enhanced through plasmonic coupling structures. Here, the plasmonic coupler primarily plays the role of being a radiation condenser which will cause carriers to be generated adjacent to metallic electrodes where they can more efficiently be collected.

  10. Effect of gamma radiation on retroviral recombination.

    PubMed

    Hu, W S; Temin, H M

    1992-07-01

    To elucidate the mechanism(s) of retroviral recombination, we exposed virions to gamma radiation prior to infecting target cells. By using previously described spleen necrosis virus-based vectors containing multiple markers, recombinant proviruses were studied after a single round of retrovirus replication. The current models of retroviral recombination predict that breaking virion RNA should promote minus-strand recombination (forced copy-choice model), decrease or not affect plus-strand recombination (strand displacement/assimilation model), and shift plus-strand recombination towards the 3' end of the genome. However, we found that while gamma irradiation of virions reduced the amount of recoverable viral RNA, it did not primarily cause breaks. Thus, the frequency of selected recombinants was not significantly altered with greater doses of radiation. In spite of this, the irradiation did decrease the number of recombinants with only one internal template switch. As a result, the average number of additional internal template switches in the recombinant proviruses increased from 0.7 to 1.4 as infectivity decreased to 6%. The unselected internal template switches tended to be 5' of the selected crossover even in the recombinants from irradiated viruses, inconsistent with a plus-strand recombination mechanism. PMID:1602553

  11. Integrated network analysis identifies fight-club nodes as a class of hubs encompassing key putative switch genes that induce major transcriptome reprogramming during grapevine development.

    PubMed

    Palumbo, Maria Concetta; Zenoni, Sara; Fasoli, Marianna; Massonnet, Mélanie; Farina, Lorenzo; Castiglione, Filippo; Pezzotti, Mario; Paci, Paola

    2014-12-01

    We developed an approach that integrates different network-based methods to analyze the correlation network arising from large-scale gene expression data. By studying grapevine (Vitis vinifera) and tomato (Solanum lycopersicum) gene expression atlases and a grapevine berry transcriptomic data set during the transition from immature to mature growth, we identified a category named "fight-club hubs" characterized by a marked negative correlation with the expression profiles of neighboring genes in the network. A special subset named "switch genes" was identified, with the additional property of many significant negative correlations outside their own group in the network. Switch genes are involved in multiple processes and include transcription factors that may be considered master regulators of the previously reported transcriptome remodeling that marks the developmental shift from immature to mature growth. All switch genes, expressed at low levels in vegetative/green tissues, showed a significant increase in mature/woody organs, suggesting a potential regulatory role during the developmental transition. Finally, our analysis of tomato gene expression data sets showed that wild-type switch genes are downregulated in ripening-deficient mutants. The identification of known master regulators of tomato fruit maturation suggests our method is suitable for the detection of key regulators of organ development in different fleshy fruit crops. PMID:25490918

  12. Evolution of genetic switch complexity

    PubMed Central

    Broussard, Gregory W.; Hatfull, Graham F.

    2013-01-01

    The circuitry of the phage λ genetic switch determining the outcome of lytic or lysogenic growth is well-integrated and complex, raising the question as to how it evolved. It is plausible that it arose from a simpler ancestral switch with fewer components that underwent various additions and refinements, as it adapted to vast numbers of different hosts and conditions. We have recently identified a new class of genetic switches found in mycobacteriophages and other prophages, in which immunity is dependent on integration. These switches contain only three genes (integrase, repressor and cro) and represent a major departure from the λ-like circuitry, lacking many features such as xis, cII and cIII. These small self-contained switches represent an unrealized, elegant circuitry for controlling infection outcome. In this addendum, we propose a model of possible events in the evolution of a complex λ-like switch from a simpler integration-dependent switch. PMID:23819104

  13. Polymorphisms in HLA Class I Genes Associated with both Favorable Prognosis of Human Immunodeficiency Virus (HIV) Type 1 Infection and Positive Cytotoxic T-Lymphocyte Responses to ALVAC-HIV Recombinant Canarypox Vaccines

    PubMed Central

    Kaslow, Richard A.; Rivers, Charles; Tang, Jianming; Bender, Thomas J.; Goepfert, Paul A.; El Habib, Raphaelle; Weinhold, Kent; Mulligan, Mark J.

    2001-01-01

    Carriers of certain human leukocyte antigen class I alleles show favorable prognosis of human immunodeficiency virus type 1 (HIV-1) infection, presumably due to effective CD8+ cytotoxic T-lymphocyte responses, but close relationships between class I variants mediating such responses to natural and to vaccine HIV-1 antigen have not been established. During 6 to 30 months of administration and follow-up in trials of ALVAC-HIV recombinant canarypox vaccines, cells from 42% of 291 HIV-1-negative vaccinated subjects typed at class I loci responded to an HIV-1 protein in a lytic bulk CD8+ cytotoxic T-lymphocyte assay. By 2 weeks after the second dose, higher proportions of vaccinees carrying one of two alleles consistently associated with slower progression of natural HIV-1 infection reacted at least once: B∗27 carriers reacted to Gag (64%; odds ratio [OR] = 10.3, P = 0.001) and Env (36%; OR = 4.6, P = 0.04), and B∗57 carriers reacted to Env (44%; OR = 6.6, P < 0.05). By 2 weeks after the third or fourth dose, B∗27 carriers had responded (two or more reactions) to Gag (33%; OR = 4.4, P < 0.05) and B∗57 carriers had responded to both Gag (39%; OR = 5.3, P = 0.013) and Env (39%; OR = 9.5, P = 0.002). Homozygosity at class I loci, although conferring an unfavorable prognosis following natural infection, showed no such disadvantage for vaccine response. Individual class I alleles have not previously demonstrated such clear and consistent relationship with both the clinical course of an infection and cellular immunity to a vaccine against the infectious agent. This proof of principle that class I an alleles modulate both processes has implications for development of HIV-1 and presumably other vaccines. PMID:11507213

  14. DNA secondary structures are associated with recombination in major Plasmodium falciparum variable surface antigen gene families

    PubMed Central

    Sander, Adam F.; Lavstsen, Thomas; Rask, Thomas S.; Lisby, Michael; Salanti, Ali; Fordyce, Sarah L.; Jespersen, Jakob S.; Carter, Richard; Deitsch, Kirk W.; Theander, Thor G.; Pedersen, Anders Gorm; Arnot, David E.

    2014-01-01

    Many bacterial, viral and parasitic pathogens undergo antigenic variation to counter host immune defense mechanisms. In Plasmodium falciparum, the most lethal of human malaria parasites, switching of var gene expression results in alternating expression of the adhesion proteins of the Plasmodium falciparum-erythrocyte membrane protein 1 class on the infected erythrocyte surface. Recombination clearly generates var diversity, but the nature and control of the genetic exchanges involved remain unclear. By experimental and bioinformatic identification of recombination events and genome-wide recombination hotspots in var genes, we show that during the parasite’s sexual stages, ectopic recombination between isogenous var paralogs occurs near low folding free energy DNA 50-mers and that these sequences are heavily concentrated at the boundaries of regions encoding individual Plasmodium falciparum-erythrocyte membrane protein 1 structural domains. The recombinogenic potential of these 50-mers is not parasite-specific because these sequences also induce recombination when transferred to the yeast Saccharomyces cerevisiae. Genetic cross data suggest that DNA secondary structures (DSS) act as inducers of recombination during DNA replication in P. falciparum sexual stages, and that these DSS-regulated genetic exchanges generate functional and diverse P. falciparum adhesion antigens. DSS-induced recombination may represent a common mechanism for optimizing the evolvability of virulence gene families in pathogens. PMID:24253306

  15. Gain of function mutation in tobacco MADS box promoter switch on the expression of flowering class B genes converting sepals to petals.

    PubMed

    Mahajan, Monika; Yadav, Sudesh Kumar

    2014-02-01

    One mutant transgenic line displaying homeotic conversion of sepals to petals with other phenotypic aberrations was selected and characterized at molecular level. The increased transcript level of gene encoding anthocyanidin synthase and petal specific class B genes, GLOBOSA and DEFECIENS in sepals of mutant line may be responsible for its homeotic conversion to petaloid organs. While characterizing this mutant line for locus identification, T-DNA was found to be inserted in 3' untranslated region of promoter of class B MADS box gene, GLOBOSA. Here, CaMV 35S promoter of T-DNA might be deriving the expression of class B genes. PMID:24362510

  16. Switch wear leveling

    SciTech Connect

    Wu, Hunter; Sealy, Kylee; Gilchrist, Aaron

    2015-09-01

    An apparatus for switch wear leveling includes a switching module that controls switching for two or more pairs of switches in a switching power converter. The switching module controls switches based on a duty cycle control technique and closes and opens each switch in a switching sequence. The pairs of switches connect to a positive and negative terminal of a DC voltage source. For a first switching sequence a first switch of a pair of switches has a higher switching power loss than a second switch of the pair of switches. The apparatus includes a switch rotation module that changes the switching sequence of the two or more pairs of switches from the first switching sequence to a second switching sequence. The second switch of a pair of switches has a higher switching power loss than the first switch of the pair of switches during the second switching sequence.

  17. Condition known to be sufficient for rearrangeability of the benes class of interconnection networks with 2*2 switches is also necessary

    SciTech Connect

    Kothari, S.C.; Lakshmivarahan, S.

    1983-01-01

    An n*n (n inputs and outputs) multistage interconnection network is said to be rearrangeable if it can realise all the possible connections of the n input terminals to the n output terminals in a one-to-one fashion. Starting with the pioneering works of Clos (1953) and Benes (1968) to this date a variety of sufficient conditions for rearrangeability are known in the literature. It is shown that the well known sufficient condition due to Benes on the link permutation is also necessary for rearrangeability if the network is made up of 2*2 switches. 15 references.

  18. Independent Roles of Switching and Hypermutation in the Development and Persistence of B Lymphocyte Memory.

    PubMed

    Gitlin, Alexander D; von Boehmer, Lotta; Gazumyan, Anna; Shulman, Ziv; Oliveira, Thiago Y; Nussenzweig, Michel C

    2016-04-19

    Somatic hypermutation (SHM) and class-switch recombination (CSR) increase the affinity and diversify the effector functions of antibodies during immune responses. Although SHM and CSR are fundamentally different, their independent roles in regulating B cell fate have been difficult to uncouple because a single enzyme, activation-induced cytidine deaminase (encoded by Aicda), initiates both reactions. Here, we used a combination of Aicda and antibody mutant alleles that separate the effects of CSR and SHM on polyclonal immune responses. We found that class-switching to IgG1 biased the fate choice made by B cells, favoring the plasma cell over memory cell fate without significantly affecting clonal expansion in the germinal center (GC). In contrast, SHM reduced the longevity of memory B cells by creating polyreactive specificities that were selected against over time. Our data define the independent contributions of SHM and CSR to the generation and persistence of memory in the antibody system. PMID:26944202

  19. Classifications and comparisons of multilocus recombination distributions

    PubMed Central

    Karlin, Samuel; Liberman, Uri

    1978-01-01

    Various classifications and representations of multilocus recombination structures are delineated based on generalized notions of linkage values and recombination rates. An important class of recombination distributions (called the count-location chiasma process) is parameterized by a distribution of the number of crossover events and, for each such crossover count, by a conditional distribution of crossover locations. A number of properties of this recombination structure are developed. A multilocus definition of a “natural” recombination range is set forth. Orderings among recombination distributions in the multilocus setting are also discussed. Comparisons are made in terms of complete linkage, free assortment and noninterference schemes serving as standards. PMID:16592601

  20. Genetic Recombination

    ERIC Educational Resources Information Center

    Whitehouse, H. L. K.

    1973-01-01

    Discusses the mechanisms of genetic recombination with particular emphasis on the study of the fungus Sordaria brevicollis. The study of recombination is facilitated by the use of mutants of this fungus in which the color of the ascospores is affected. (JR)

  1. Optical switches and switching methods

    DOEpatents

    Doty, Michael

    2008-03-04

    A device and method for collecting subject responses, particularly during magnetic imaging experiments and testing using a method such as functional MRI. The device comprises a non-metallic input device which is coupled via fiber optic cables to a computer or other data collection device. One or more optical switches transmit the subject's responses. The input device keeps the subject's fingers comfortably aligned with the switches by partially immobilizing the forearm, wrist, and/or hand of the subject. Also a robust nonmetallic switch, particularly for use with the input device and methods for optical switching.

  2. 47 CFR 32.2210 - Central office-switching.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... Central office—switching. This account shall be used by Class B companies to record the original cost of switching assets of the type and character required of Class A companies in Accounts 2211 through 2212. ... 47 Telecommunication 2 2010-10-01 2010-10-01 false Central office-switching. 32.2210 Section...

  3. New class of leukemogenic ecotropic recombinant murine leukemia virus isolated from radiation-induced thymomas of C57BL/6 mice

    SciTech Connect

    Rassart, E.; Sankar-Mistry, P.; Lemay, G.; DesGroseillers, L.; Jalicoeur, P.

    1983-02-01

    We previously reported the establishment of several lymphoid cell lines from X-ray-induced thymomas of C57BL/Ka mice, and all, except one, produce retroviruses. Biological characterization of five of these new primary radiation leukemia viruses (RadLVs) indicated that they had a B-tropic, fibrotropic, and ecotropic host range and were leukemogenic when reinjected into C57BL/Ka newborn mice. The leukemogenic potential of one isolate (G/sub 6/T/sub 2/) was further assessed and shown to be retained after prolonged passaging on fibroblasts in vitro. Restriction endonuclease analysis of the DNA of four of our new RadLV isolates (G/sub 6/T/sub 2/, Ti-7, Ti-8, and Ti-9) revealed that G/sub 6/T/sub 2/ and Ti-7 murine leukemia virus (MuLV) genomes had identical restriction maps, whereas Ti-8 and Ti-9 genomes were different from each other and from the G/sub 6/T/sub 2/ and Ti-7 genomes. The physical maps of these genomes were similar to that of known ecotropic MuLV genomes (including the C57BL/Ka endogenous ecotropic MuLV) within their long terminal repeats, env, the right portion of pol, and the left portion of gag. However, a region covering the end of gag and the beginning of pol was different and showed several similarities with xenotropic MuLV genomes of BALB/c, AKR, and C58 mice previously mapped. Our results suggest that these primary RadLV genomes are recombinants between the parental ecotropic MuLV genome and a nonecotropic (xenotropic) sequence. To further study the leukemic potential of these RadLVs, the genome of one of them (G/sub 6/T/sub 2/) was cloned in Charon 21A as an infectious molecule.

  4. Nondisjunction of chromosome 15: Origin and recombination

    SciTech Connect

    Robinson, W.P.; Bernasconi, F.; Schinzel, A.A.; Mutirangura, A.; Ledbetter, D.H. ); Langlois, S. ); Morris, M.A.; Malcolm, S.

    1993-09-01

    Thirty-two cases of uniparental disomy (UPD), ascertained from Prader-Willi syndrome patients (N=27) and Angelman syndrome patients (N-5), are used to investigate the pattern of recombination associated with nondisjunction of chromosome 15. In addition, the meiotic stage of nondisjunction is inferred by using markers mapping near the centromere. Two basic approaches to the analysis of recombination in specific pairwise intervals along the chromosome. This method shows a significant reduction in recombination for two of five intervals examined. Second, the observed frequency of each recombinant class (i.e., zero, one, two, three, or more observable crossovers) is compared with expected values. This is useful for testing whether the reduction in recombination can be attributed solely to a proportion of cases with no recombination at all (because of asynapsis), with the remaining groups showing normal recombination (or even excess recombination), or whether recombination is uniformly reduced. Analysis of maternal UPD(15) data shows a slight reduction in the multiple-recombinant classes, with a corresponding increase in both the zero- and one-recombinant classes over expected values. The majority, more than 82%, of the extra chromosomes in maternal UPD(15) cases are due to meiotic I nondisjunction events. In contrast, more paternal UPD(15) cases so far examined appear to have a postzygotic origin of the extra paternal chromosome. 33 refs., 1 fig., 7 tabs.

  5. ION SWITCH

    DOEpatents

    Cook, B.

    1959-02-10

    An ion switch capable of transferring large magnitudes of power is described. An ion switch constructed in accordance with the invention includes a pair of spaced control electrodes disposed in a highly evacuated region for connection in a conventional circuit to control the passing of power therethrough. A controllable ionic conduction path is provided directiy between the control electrodes by a source unit to close the ion switch. Conventional power supply means are provided to trigger the source unit and control the magnitude, durations and pulse repetition rate of the aforementioned ionic conduction path.

  6. Acceleration switch

    DOEpatents

    Abbin, J.P. Jr.; Devaney, H.F.; Hake, L.W.

    1979-08-29

    The disclosure relates to an improved integrating acceleration switch of the type having a mass suspended within a fluid filled chamber, with the motion of the mass initially opposed by a spring and subsequently not so opposed.

  7. Acceleration switch

    DOEpatents

    Abbin, Jr., Joseph P.; Devaney, Howard F.; Hake, Lewis W.

    1982-08-17

    The disclosure relates to an improved integrating acceleration switch of the type having a mass suspended within a fluid filled chamber, with the motion of the mass initially opposed by a spring and subsequently not so opposed.

  8. Cosmological Recombination

    NASA Astrophysics Data System (ADS)

    Wong, Wan Yan

    2008-11-01

    In this thesis we focus on studying the physics of cosmological recombination and how the details of recombination affect the Cosmic Microwave Background (CMB) anisotropies. We present a detailed calculation of the spectral line distortions on the CMB spectrum arising from the Lyman-alpha and the lowest two-photon transitions in the recombination of hydrogen (H), and the corresponding lines from helium (He). The peak of these distortions mainly comes from the Lyman-alpha transition and occurs at about 170 microns, which is the Wien part of the CMB. The major theoretical limitation for extracting cosmological parameters from the CMB sky lies in the precision with which we can calculate the cosmological recombination process. With this motivation, we perform a multi-level calculation of the recombination of H and He with the addition of the spin-forbidden transition for neutral helium (He I), plus the higher order two-photon transitions for H and among singlet states of He I. We find that the inclusion of the spin-forbidden transition results in more than a percent change in the ionization fraction, while the other transitions give much smaller effects. Last we modify RECFAST by introducing one more parameter to reproduce recent numerical results for the speed-up of helium recombination. Together with the existing hydrogen `fudge factor', we vary these two parameters to account for the remaining dominant uncertainties in cosmological recombination. By using a Markov Chain Monte Carlo method with Planck forecast data, we find that we need to determine the parameters to better than 10% for He I and 1% for H, in order to obtain negligible effects on the cosmological parameters.

  9. Activation-Induced Cytidine Deaminase Does Not Impact Murine Meiotic Recombination

    PubMed Central

    Cortesao, Catarina S.; Freitas, Raquel F.; Barreto, Vasco M.

    2013-01-01

    Activation-induced cytidine deaminase (AID) was first described as the triggering enzyme of the B-cell−specific reactions that edit the immunoglobulin genes, namely somatic hypermutation, gene conversion, and class switch recombination. Over the years, AID was also detected in cells other than lymphocytes, and it has been assigned additional roles in the innate defense against transforming retroviruses, in retrotransposition restriction and in DNA demethylation. Notably, AID expression was found in germline tissues, and in heterologous systems it can induce the double-strand breaks required for the initiation of meiotic recombination and proper gamete formation. However, because AID-deficient mice are fully fertile, the molecule is not essential for meiosis. Thus, the remaining question that we addressed here is whether AID influences the frequency of meiotic recombination in mice. We measured the recombination events in the meiosis of male and female mice F1 hybrids of C57BL/6J and BALB/c, in Aicda+/+ and Aicda−/− background by using a panel of single-nucleotide polymorphisms that distinguishes C57BL/6J from BALB/c genome across the 19 autosomes. In agreement with the literature, we found that the frequency of recombination in the female germline was greater than in male germline, both in the Aicda+/+ and Aicda−/− backgrounds. No statistical difference was found in the average recombination events between Aicda+/+ and Aidca−/− animals, either in females or males. In addition, the recombination frequencies between single-nucleotide polymorphisms flanking the immunoglobulin heavy and immunoglobulin kappa loci was also not different. We conclude that AID has a minor impact, if any, on the overall frequency of meiotic recombination. PMID:23550130

  10. Switched steerable multiple beam antenna system

    NASA Astrophysics Data System (ADS)

    Iwasaki, Richard S.

    1988-09-01

    A steerable multibeam five element cross-feed cluster antenna system is described. The feed power is divided into five branches. Each branch includes a switching network comprised of a plurality of time delay elements each individually controlled by a respective electromagnetic latching switch. Frequency independent individual two-dimensional beam steering at intermediate (IF) scanning frequencies is thereby provided wherein discrete incremental time delays are introduced by the switching networks into each branch and the signals recombined thereafter to form each beam. The electromagnetic latched switching reduces power consumption and permits higher power switching and reciprocal coincident tranmsit and receive operation. Frequency independence due to incremental time delay switching permits coincident reciprocal operation and steering for transmit-receive signal paths carrying different transmit-receive frequencies. Diagonal quarter wave plates in the waveguides alter polarization from the circular to orthogonal linear to provide transmitter-receiver isolation.

  11. Optical switch

    DOEpatents

    Reedy, Robert P.

    1987-01-01

    An optical switching device (10) is provided whereby light from a first glass fiber (16) or a second glass fiber (14) may be selectively transmitted into a third glass fiber (18). Each glass fiber is provided with a focusing and collimating lens system (26, 28, 30). In one mode of operation, light from the first glass fiber (16) is reflected by a planar mirror (36) into the third glass fiber (18). In another mode of operation, light from the second glass fiber (14) passes directly into the third glass fiber (18). The planar mirror (36) is attached to a rotatable table (32) which is rotated to provide the optical switching.

  12. Learner Code-Switching versus English Only

    ERIC Educational Resources Information Center

    Sampson, Andrew

    2012-01-01

    This article describes a study into the functions of code-switching in EFL classes at a Colombian language school. It was undertaken to decide whether the official "English-only" policy in place in this and other classrooms is pedagogically justified. The results suggest that code-switching may not necessarily be connected to ability level and…

  13. Controlled Release from Recombinant Polymers

    PubMed Central

    Price, Robert; Poursaid, Azadeh; Ghandehari, Hamidreza

    2014-01-01

    Recombinant polymers provide a high degree of molecular definition for correlating structure with function in controlled release. The wide array of amino acids available as building blocks for these materials lend many advantages including biorecognition, biodegradability, potential biocompatibility, and control over mechanical properties among other attributes. Genetic engineering and DNA manipulation techniques enable the optimization of structure for precise control over spatial and temporal release. Unlike the majority of chemical synthetic strategies used, recombinant DNA technology has allowed for the production of monodisperse polymers with specifically defined sequences. Several classes of recombinant polymers have been used for controlled drug delivery. These include, but are not limited to, elastin-like, silk-like, and silk-elastinlike proteins, as well as emerging cationic polymers for gene delivery. In this article, progress and prospects of recombinant polymers used in controlled release will be reviewed. PMID:24956486

  14. Switching Transistor

    NASA Technical Reports Server (NTRS)

    1981-01-01

    Westinghouse Electric Corporation's D60T transistors are used primarily as switching devices for controlling high power in electrical circuits. It enables reduction in the number and size of circuit components and promotes more efficient use of energy. Wide range of application from a popcorn popper to a radio frequency generator for solar cell production.

  15. The Strength of an Ig Switch Region is Determined by its Ability to Drive R-loop Formation and its Number of WGCW Sites

    PubMed Central

    Zhang, Zheng Z.; Pannunzio, Nicholas R.; Han, Li; Hsieh, Chih-Lin; Yu, Kefei; Lieber, Michael R.

    2014-01-01

    SUMMARY R-loops exist at the murine IgH switch regions and possibly other locations, but their functional importance is unclear. In biochemical systems, R-loop initiation requires DNA sequence regions containing clusters of G nucleotides, but cellular studies have not been done. Here, we vary the G-clustering, total switch region length, and the number of target sites (WGCW sites for the activation-induced deaminase) at synthetic switch regions in a murine B cell line to determine the effect on class switch recombination (CSR). G-clusters increase CSR, regardless of their immediate proximity to the WGCW sites. This increase is accompanied by an increase in R-loop formation. CSR efficiency correlates better with the absolute number of WGCW sites in the switch region rather than the total switch region length or density of WGCW sites. Thus, the overall strength of the switch region depends on G-clusters, which initiate R-loop formation, and on the number of WGCW sites. PMID:25017067

  16. State-dependent switching control of switched positive fractional-order systems.

    PubMed

    Zhao, Xudong; Yin, Yunfei; Zheng, Xiaolong

    2016-05-01

    In this paper, the problem of switching stabilization for a class of continuous-time switched positive fractional-order systems is studied by using state-dependent switching. First, the asymptotic stability condition of switched positive fractional-order systems with state-dependent switching is given, which is based on the fractional co-positive Lyapunov method. Moreover, by the sliding sector method, the stability condition of switched positive fractional-order systems whose subsystems are possibly all unstable is obtained. A variable structure (VS) switching law with sliding sector is also proposed to guarantee the switched positive fractional-order system to be asymptotically stable. Finally, two numerical examples are given to demonstrate the advantages and effectiveness of our developed results. PMID:26922493

  17. Ligand-responsive RNA mechanical switches.

    PubMed

    Boerneke, Mark A; Hermann, Thomas

    2015-01-01

    Ligand-responsive RNA mechanical switches represent a new class of simple switching modules that adopt well-defined ligand-free and bound conformational states, distinguishing them from metabolite-sensing riboswitches. Initially discovered in the internal ribosome entry site (IRES) of hepatitis C virus (HCV), these RNA switch motifs were found in the genome of diverse other viruses. Although large variations are seen in sequence and local secondary structure of the switches, their function in viral translation initiation that requires selective ligand recognition is conserved. We recently determined the crystal structure of an RNA switch from Seneca Valley virus (SVV) which is able to functionally replace the switch of HCV. The switches from both viruses recognize identical cognate ligands despite their sequence dissimilarity. Here, we describe the discovery of 7 new switches in addition to the previously established 5 examples. We highlight structural and functional features unique to this class of ligand-responsive RNA mechanical switches and discuss implications for therapeutic development and the construction of RNA nanostructures. PMID:26158858

  18. Molecular Switches and Motors on Surfaces

    NASA Astrophysics Data System (ADS)

    Pathem, Bala Krishna; Claridge, Shelley A.; Zheng, Yue Bing; Weiss, Paul S.

    2013-04-01

    Molecular switches and motors respond structurally, electronically, optically, and/or mechanically to external stimuli, testing and potentially enabling extreme miniaturization of optoelectronic devices, nanoelectromechanical systems, and medical devices. The assembly of motors and switches on surfaces makes it possible both to measure the properties of individual molecules as they relate to their environment and to couple function between assembled molecules. In this review, we discuss recent progress in assembling molecular switches and motors on surfaces, measuring static and dynamic structures, understanding switching mechanisms, and constructing functional molecular materials and devices. As demonstrative examples, we choose a representative molecule from three commonly studied classes including molecular switches, photochromic molecules, and mechanically interlocked molecules. We conclude by offering perspectives on the future of molecular switches and motors on surfaces.

  19. Wide Bandgap Extrinsic Photoconductive Switches

    NASA Astrophysics Data System (ADS)

    Sullivan, James Stephen

    Wide Bandgap Extrinsic Photoconductive Switches Semi-insulating Gallium Nitride, 4H and 6H Silicon Carbide are attractive materials for compact, high voltage, extrinsic, photoconductive switches due to their wide bandgap, high dark resistance, high critical electric field strength and high electron saturation velocity. These wide bandgap semiconductors are made semi-insulating by the addition of vanadium (4H and 6H-SiC) and iron (2H-GaN) impurities that form deep acceptors. These deep acceptors trap electrons donated from shallow donor impurities. The electrons can be optically excited from these deep acceptor levels into the conduction band to transition the wide bandgap semiconductor materials from a semi-insulating to a conducting state. Extrinsic photoconductive switches with opposing electrodes have been constructed using vanadium compensated 6H-SiC and iron compensated 2H-GaN. These extrinsic photoconductive switches were tested at high voltage and high power to determine if they could be successfully used as the closing switch in compact medical accelerators. The successful development of a vanadium compensated, 6H-SiC extrinsic photoconductive switch for use as a closing switch for compact accelerator applications was realized by improvements made to the vanadium, nitrogen and boron impurity densities. The changes made to the impurity densities were based on the physical intuition outlined and simple rate equation models. The final 6H-SiC impurity 'recipe' calls for vanadium, nitrogen and boron densities of 2.5 e17 cm-3, 1.25e17 cm-3 and ≤ 1e16 cm-3, respectively. This recipe was originally developed to maximize the quantum efficiency of the vanadium compensated 6H-SiC, while maintaining a thermally stable semi-insulating material. The rate equation models indicate that, besides increasing the quantum efficiency, the impurity recipe should be expected to also increase the carrier recombination time. Three generations of 6H-SiC materials were tested. The

  20. Lymphocyte-specific compensation for XLF/cernunnos end-joining functions in V(D)J recombination.

    PubMed

    Li, Gang; Alt, Frederick W; Cheng, Hwei-Ling; Brush, James W; Goff, Peter H; Murphy, Mike M; Franco, Sonia; Zhang, Yu; Zha, Shan

    2008-09-01

    Mutations in XLF/Cernunnos (XLF) cause lymphocytopenia in humans, and various studies suggest an XLF role in classical nonhomologous end joining (C-NHEJ). We now find that XLF-deficient mouse embryonic fibroblasts are ionizing radiation (IR) sensitive and severely impaired for ability to support V(D)J recombination. Yet mature lymphocyte numbers in XLF-deficient mice are only modestly decreased. Moreover, XLF-deficient pro-B lines, while IR-sensitive, perform V(D)J recombination at nearly wild-type levels. Correspondingly, XLF/p53-double-deficient mice are not markedly prone to the pro-B lymphomas that occur in previously characterized C-NHEJ/p53-deficient mice; however, like other C-NHEJ/p53-deficient mice, they still develop medulloblastomas. Despite nearly normal V(D)J recombination in developing B cells, XLF-deficient mature B cells are moderately defective for immunoglobulin heavy-chain class switch recombination. Together, our results implicate XLF as a C-NHEJ factor but also indicate that developing mouse lymphocytes harbor cell-type-specific factors/pathways that compensate for the absence of XLF function during V(D)J recombination. PMID:18775323

  1. Attenuating homologous recombination stimulates an AID-induced antileukemic effect

    PubMed Central

    Lamont, Kristin R.; Hasham, Muneer G.; Donghia, Nina M.; Branca, Jane; Chavaree, Margaret; Chase, Betsy; Breggia, Anne; Hedlund, Jacquelyn; Emery, Ivette; Cavallo, Francesca; Jasin, Maria; Rüter, Jens

    2013-01-01

    Activation-induced cytidine deaminase (AID) is critical in normal B cells to initiate somatic hypermutation and immunoglobulin class switch recombination. Accumulating evidence suggests that AID is also prooncogenic, inducing cancer-promoting mutations or chromosome rearrangements. In this context, we find that AID is expressed in >40% of primary human chronic lymphocytic leukemia (CLL) cases, consistent with other reports. Using a combination of human B lymphoid leukemia cells and mouse models, we now show that AID expression can be harnessed for antileukemic effect, after inhibition of the RAD51 homologous recombination (HR) factor with 4,4′-diisothiocyanatostilbene-2-2′-disulfonic acid (DIDS). As a proof of principle, we show that DIDS treatment inhibits repair of AID-initiated DNA breaks, induces apoptosis, and promotes cytotoxicity preferentially in AID-expressing human CLL. This reveals a novel antineoplastic role of AID that can be triggered by inhibition of HR, suggesting a potential new paradigm to treat AID-expressing tumors. Given the growing list of tumor types with aberrant AID expression, this novel therapeutic approach has potential to impact a significant patient population. PMID:23589568

  2. Nanoelectromechanical contact switches

    NASA Astrophysics Data System (ADS)

    Loh, Owen Y.; Espinosa, Horacio D.

    2012-05-01

    Nanoelectromechanical (NEM) switches are similar to conventional semiconductor switches in that they can be used as relays, transistors, logic devices and sensors. However, the operating principles of NEM switches and semiconductor switches are fundamentally different. These differences give NEM switches an advantage over semiconductor switches in some applications -- for example, NEM switches perform much better in extreme environments -- but semiconductor switches benefit from a much superior manufacturing infrastructure. Here we review the potential of NEM-switch technologies to complement or selectively replace conventional complementary metal-oxide semiconductor technology, and identify the challenges involved in the large-scale manufacture of a representative set of NEM-based devices.

  3. An evaluation of Access Tier local area network switches.

    SciTech Connect

    Eldridge, John M.; Olsberg, Ronald R.

    2004-06-01

    This reports tabulates the Test and Evaluation results of the Access Class Switch tests conducted by members of Department 9336. About 15 switches were reviewed for use in the enterprise network as access tier switches as defined in a three tier architecture. The Access Switch Tier has several functions including: aggregate customer desktop ports, preserve and apply QoS tags, provide switched LAN access, provide VLAN assignment, as well as others. The typical switch size is 48 or less user ports. The evaluation team reviewed network switch evaluation reports from the Tolly Group as well as other sources. We then used these reports as a starting point to identify particular switches for evaluation. In general we reviewed the products of dominant equipment manufacturers. Also, based on architectural design requirements, the majority of the switches tested were of relatively small monolithic unit variety.

  4. Mechanisms and Factors that Influence High Frequency Retroviral Recombination

    PubMed Central

    Delviks-Frankenberry, Krista; Galli, Andrea; Nikolaitchik, Olga; Mens, Helene; Pathak, Vinay K.; Hu, Wei-Shau

    2011-01-01

    With constantly changing environmental selection pressures, retroviruses rely upon recombination to reassort polymorphisms in their genomes and increase genetic diversity, which improves the chances for the survival of their population. Recombination occurs during DNA synthesis, whereby reverse transcriptase undergoes template switching events between the two copackaged RNAs, resulting in a viral recombinant with portions of the genetic information from each parental RNA. This review summarizes our current understanding of the factors and mechanisms influencing retroviral recombination, fidelity of the recombination process, and evaluates the subsequent viral diversity and fitness of the progeny recombinant. Specifically, the high mutation rates and high recombination frequencies of HIV-1 will be analyzed for their roles in influencing HIV-1 global diversity, as well as HIV-1 diagnosis, drug treatment, and vaccine development. PMID:21994801

  5. Optical switching technologies and their applications

    NASA Astrophysics Data System (ADS)

    Ciminelli, Caterina; Meli, F.; Grasso, Giorgio

    2001-05-01

    In this paper system applications, geometry, physical effects and materials of optical switching devices are reviewed. Main system scenarios are presented and some key features such as size, loss, speed, scalability and granularity are highlighted. Two principal categories of optical switches are considered, i.e. guided-wave switches and free-space switches. In the first category some sub- classes have been identified according to their geometrical configuration, principle of operation and, then physical mechanism and materials. As for the geometry, the most frequently used configurations are briefly described together with their advantages and disadvantages. Different physical effects suitable to obtain the index change, which the switching function is based on, are also described with reference to the material substrates. Switches based on semiconductor optical amplifier gate are also analyzed. In the free-space category the main sub-classes are represented by the opto-mechanical devices and micro-opto-electro- mechanical systems switches. The last technology combines the free-space interconnecting with the integration capability on a single silica chip. The main advantages such as the ability to scaling up to large switch fabric and some issues such as packaging and reliability are analyzed. Finally, devices based on polarization change, acousto- optics interaction, total internal reflection and holography are illustrated.

  6. THYRATRON SWITCH

    DOEpatents

    Creveling, R.; Bourgeois, N.A. Jr.

    1959-04-21

    An arrangement for utilizing a thyratron as a noise free switch is described. It has been discovered that the voltage between plate and cathode of a thyratron will oscillate, producing voltage spikes, if the tube carries only a fraction of its maximum rated current. These voltage spikes can produce detrimental effects where the thyratron is used in critical timing circuits. To alleviate this problem the disclosed circuit provides a charged capacitor and a resistor in parallel with the tube and of such value that the maximum current will flow from the capacitor through the thyratron when it is triggered. During this time the signal current is conducted through the tube, before the thyratron voltage starts to oscillate, and the signal current output is free of noise spikes.

  7. Expression of Recombinant Antibodies

    PubMed Central

    Frenzel, André; Hust, Michael; Schirrmann, Thomas

    2013-01-01

    Recombinant antibodies are highly specific detection probes in research, diagnostics, and have emerged over the last two decades as the fastest growing class of therapeutic proteins. Antibody generation has been dramatically accelerated by in vitro selection systems, particularly phage display. An increasing variety of recombinant production systems have been developed, ranging from Gram-negative and positive bacteria, yeasts and filamentous fungi, insect cell lines, mammalian cells to transgenic plants and animals. Currently, almost all therapeutic antibodies are still produced in mammalian cell lines in order to reduce the risk of immunogenicity due to altered, non-human glycosylation patterns. However, recent developments of glycosylation-engineered yeast, insect cell lines, and transgenic plants are promising to obtain antibodies with “human-like” post-translational modifications. Furthermore, smaller antibody fragments including bispecific antibodies without any glycosylation are successfully produced in bacteria and have advanced to clinical testing. The first therapeutic antibody products from a non-mammalian source can be expected in coming next years. In this review, we focus on current antibody production systems including their usability for different applications. PMID:23908655

  8. Exponential stability and robust H∞ control of a class of discrete-time switched non-linear systems with time-varying delays via T-S fuzzy model

    NASA Astrophysics Data System (ADS)

    Mao, Yanbing; Zhang, Hongbin

    2014-05-01

    This paper deals with stability and robust H∞ control of discrete-time switched non-linear systems with time-varying delays. The T-S fuzzy models are utilised to represent each sub-non-linear system. Thus, with two level functions, namely, crisp switching functions and local fuzzy weighting functions, we introduce a discrete-time switched fuzzy systems, which inherently contain the features of the switched hybrid systems and T-S fuzzy systems. Piecewise fuzzy weighting-dependent Lyapunov-Krasovskii functionals (PFLKFs) and average dwell-time approach are utilised in this paper for the exponentially stability analysis and controller design, and with free fuzzy weighting matrix scheme, switching control laws are obtained such that H∞ performance is satisfied. The conditions of stability and the control laws are given in the form of linear matrix inequalities (LMIs) that are numerically feasible. The state decay estimate is explicitly given. A numerical example and the control of delayed single link robot arm with uncertain part are given to demonstrate the efficiency of the proposed method.

  9. Passivity of switched recurrent neural networks with time-varying delays.

    PubMed

    Lian, Jie; Wang, Jun

    2015-02-01

    This paper is concerned with the passivity analysis for switched neural networks subject to stochastic disturbances and time-varying delays. First, using the multiple Lyapunov functions method, a state-dependent switching law is designed to present a stochastic passivity condition. Second, a hysteresis switching law involving both the current state and the previous value of the switching signal are presented to avoid chattering resulted from the state-dependent switching. Third, based on the average dwell-time approach, a class of switching signals is determined to guarantee the switched neural network stochastically passive. Finally, three numerical examples are provided to illustrate the characteristics of three proposed switching laws. PMID:25576577

  10. Phenotypic switching in bacteria

    NASA Astrophysics Data System (ADS)

    Merrin, Jack

    genetic networks that would implement a more general theoretical model of phenotypic switching. We will use a new cloning strategy in order to systematically assemble a large number of genetic features, such as site-specific recombination components from the R64 plasmid, which invert several coexisting DNA segments. The inversion of these segments would lead to discrete phenotypic transitions inside a living cell. These artificial phenotypic switches can be controlled precisely in experiments and may serve as a benchmark for their natural counterparts.

  11. Integrated Network Analysis Identifies Fight-Club Nodes as a Class of Hubs Encompassing Key Putative Switch Genes That Induce Major Transcriptome Reprogramming during Grapevine Development[W][OPEN

    PubMed Central

    Palumbo, Maria Concetta; Zenoni, Sara; Fasoli, Marianna; Massonnet, Mélanie; Farina, Lorenzo; Castiglione, Filippo; Pezzotti, Mario; Paci, Paola

    2014-01-01

    We developed an approach that integrates different network-based methods to analyze the correlation network arising from large-scale gene expression data. By studying grapevine (Vitis vinifera) and tomato (Solanum lycopersicum) gene expression atlases and a grapevine berry transcriptomic data set during the transition from immature to mature growth, we identified a category named “fight-club hubs” characterized by a marked negative correlation with the expression profiles of neighboring genes in the network. A special subset named “switch genes” was identified, with the additional property of many significant negative correlations outside their own group in the network. Switch genes are involved in multiple processes and include transcription factors that may be considered master regulators of the previously reported transcriptome remodeling that marks the developmental shift from immature to mature growth. All switch genes, expressed at low levels in vegetative/green tissues, showed a significant increase in mature/woody organs, suggesting a potential regulatory role during the developmental transition. Finally, our analysis of tomato gene expression data sets showed that wild-type switch genes are downregulated in ripening-deficient mutants. The identification of known master regulators of tomato fruit maturation suggests our method is suitable for the detection of key regulators of organ development in different fleshy fruit crops. PMID:25490918

  12. Latching relay switch assembly

    DOEpatents

    Duimstra, Frederick A.

    1991-01-01

    A latching relay switch assembly which includes a coil section and a switch or contact section. The coil section includes at least one permanent magnet and at least one electromagnet. The respective sections are, generally, arranged in separate locations or cavities in the assembly. The switch is latched by a permanent magnet assembly and selectively switched by an overriding electromagnetic assembly.

  13. Miniature intermittent contact switch

    NASA Technical Reports Server (NTRS)

    Sword, A.

    1972-01-01

    Design of electric switch for providing intermittent contact is presented. Switch consists of flexible conductor surrounding, but separated from, fixed conductor. Flexing of outside conductor to contact fixed conductor completes circuit. Advantage is small size of switch compared to standard switches.

  14. Radiation hard vacuum switch

    DOEpatents

    Boettcher, Gordon E.

    1990-03-06

    A vacuum switch with an isolated trigger probe which is not directly connected to the switching electrodes. The vacuum switch within the plasmatron is triggered by plasma expansion initiated by the trigger probe which travels through an opening to reach the vacuum switch elements. The plasma arc created is directed by the opening to the space between the anode and cathode of the vacuum switch to cause conduction.

  15. Radiation hard vacuum switch

    DOEpatents

    Boettcher, Gordon E.

    1990-01-01

    A vacuum switch with an isolated trigger probe which is not directly connected to the switching electrodes. The vacuum switch within the plasmatron is triggered by plasma expansion initiated by the trigger probe which travels through an opening to reach the vacuum switch elements. The plasma arc created is directed by the opening to the space between the anode and cathode of the vacuum switch to cause conduction.

  16. Characteristics of current filamentation in high gain photoconductive semiconductor switching

    SciTech Connect

    Zutavern, F J; Loubriel, G M; O'Malley, M W; Helgeson, W D; McLaughlin, D L; Denison, G J

    1992-01-01

    Characteristics of current filamentation are reported for high gain photoconductive semiconductor switches (PCSS). Infrared photoluminescence is used to monitor carrier recombination radiation during fast initiation of high gain switching in large (1.5 cm gap) lateral GaAs PCSS. Spatial modulation of the optical trigger, a 200--300 ps pulse width laser, is examined. Effects on the location and number of current filaments, rise time, and delay to high gain switching, minimum trigger energy, and degradation of switch contacts are presented. Implications of these measurements for the theoretical understanding and practical development of these switches are discussed. Efforts to increase current density and reduce switch size and optical trigger energy requirements are described. Results from contact development and device lifetime testing are presented and the impact of these results on practical device applications is discussed.

  17. Isotype-switched follicular lymphoma displays dissociation between activation-induced cytidine deaminase expression and somatic hypermutation.

    PubMed

    Scherer, Florian; Navarrete, Marcelo A; Bertinetti-Lapatki, Cristina; Boehm, Joachim; Schmitt-Graeff, Annette; Veelken, Hendrik

    2016-01-01

    In B-cells, activation-induced cytidine deaminase (AID) is required for somatic hypermutation (SHM) and class switch recombination (CSR) of immunoglobulin genes. AID introduces mutations in immunoglobulin variable regions (IGV) during B-cell receptor affinity maturation, but may also introduce aberrant mutations into non-immunoglobulin genes, most commonly BCL6. Follicular lymphoma (FL) B-cells constitutively express AID and undergo CSR, SHM and aberrant SHM. We have studied AID expression, the presence of SHM mutations, CSR, and aberrant SHM in BCL6 in a cohort of 75 FL patients. Whereas IgM-expressing (non-switched) FL were characterized by an expected positive correlation between AID and IGV and BCL6 mutations, isotype-switched FL showed dissociation between AID expression and aberrant SHM, and inverse correlation between SHM and AID expression. Our results unveil two manifest biological subgroups of FL and indicate that the specific dissociation between AID and SHM after isotype switch may correlate with the clinical outcome of this heterogeneous disease. PMID:25860234

  18. "Racializing" Class

    ERIC Educational Resources Information Center

    Hatt-Echeverria, Beth; Urrieta, Luis, Jr.

    2003-01-01

    In an effort to explore how racial and class oppressions intersect, the authors use their autobiographical narratives to depict cultural and experiential continuity and discontinuity in growing up white working class versus Chicano working class. They specifically focus on "racializing class" due to the ways class is often used as a copout by…

  19. Dysregulation of CD30+ T cells by leukemia impairs isotype switching in normal B cells

    PubMed Central

    Cerutti, Andrea; Kim, Edmund C.; Shah, Shefali; Schattner, Elaine J.; Zan, Hong; Schaffer, András; Casali, Paolo

    2015-01-01

    Chronic lymphocytic leukemia (CLL) is associated with impaired immunoglobulin (Ig) class-switching from IgM to IgG and IgA, a defect that leads to recurrent infections. When activated in the presence of leukemic CLL B cells, T cells rapidly up-regulate CD30 through an OX40 ligand and interleukin 4 (IL-4)–dependent mechanism. These leukemia-induced CD30+ T cells inhibit CD40 ligand (CD40L)-mediated Sµ→Sγ and Sµ→Sα class-switch DNA recombination (CSR) by engaging CD30 ligand (CD30L), a molecule that interferes with the assembly of the CD40–tumor necrosis factor receptor–associated factor (TRAF) complex in nonmalignant IgD+ B cells. In addition, engagement of T cell CD30 by CD30L on neoplastic CLL B cells down-regulates the CD3-induced expression of CD40L. These findings indicate that, in CLL, abnormal CD30-CD30L interaction impairs IgG and IgA production by interfering with the CD40-mediated differentiation of nonmalignant B cells. PMID:11175813

  20. Switching-on quantum size effects in silicon nanocrystals.

    PubMed

    Sun, Wei; Qian, Chenxi; Wang, Liwei; Wei, Muan; Mastronardi, Melanie L; Casillas, Gilberto; Breu, Josef; Ozin, Geoffrey A

    2015-01-27

    The size-dependence of the absolute luminescence quantum yield of size-separated silicon nanocrystals reveals a "volcano" behavior, which switches on around 5 nm, peaks at near 3.7-3.9 nm, and decreases thereafter. These three regions respectively define: i) the transition from bulk to strongly quantum confined emissive silicon, ii) increasing confinement enhancing radiative recombination, and iii) increasing contributions favoring non-radiative recombination. PMID:25472530

  1. Latching micro optical switch

    DOEpatents

    Garcia, Ernest J; Polosky, Marc A

    2013-05-21

    An optical switch reliably maintains its on or off state even when subjected to environments where the switch is bumped or otherwise moved. In addition, the optical switch maintains its on or off state indefinitely without requiring external power. External power is used only to transition the switch from one state to the other. The optical switch is configured with a fixed optical fiber and a movable optical fiber. The movable optical fiber is guided by various actuators in conjunction with a latching mechanism that configure the switch in one position that corresponds to the on state and in another position that corresponds to the off state.

  2. Recombinant protein production technology

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Recombinant protein production is an important technology for antibody production, biochemical activity study, and structural determination during the post-genomic era. Limiting factors in recombinant protein production include low-level protein expression, protein precipitation, and loss of protein...

  3. Comprehensive analysis of sequences of a protein switch.

    PubMed

    Chen, Szu-Hua; Meller, Jaroslaw; Elber, Ron

    2016-01-01

    Switches form a special class of proteins that dramatically change their three-dimensional structures upon a small perturbation. One possible perturbation that we explore is that of a single point mutation. Building on the pioneering experimental work of Alexander et al. (Alexander et al. PNAS, 2007; 104,11963-11968) that determines switch sequences between α and α+β folds we conduct a comprehensive sequence sampling by a Markov Chain with multiple fitness criteria to identify new switches given the experimental folds. We screen for switch sequences using a combination of contact potential, secondary structure prediction, and finally molecular dynamics simulations. Statistical properties of switch sequences are discussed and illustrated to be most sensitive to mutation at the N- and C- termini of the switch protein. Based on this analysis, a particularly stable putative switch pair is identified and proposed for further experimental analysis. PMID:26073558

  4. Therapeutic Recombinant Monoclonal Antibodies

    ERIC Educational Resources Information Center

    Bakhtiar, Ray

    2012-01-01

    During the last two decades, the rapid growth of biotechnology-derived techniques has led to a myriad of therapeutic recombinant monoclonal antibodies with significant clinical benefits. Recombinant monoclonal antibodies can be obtained from a number of natural sources such as animal cell cultures using recombinant DNA engineering. In contrast to…

  5. Recombineering homologous recombination constructs in Drosophila.

    PubMed

    Carreira-Rosario, Arnaldo; Scoggin, Shane; Shalaby, Nevine A; Williams, Nathan David; Hiesinger, P Robin; Buszczak, Michael

    2013-01-01

    The continued development of techniques for fast, large-scale manipulation of endogenous gene loci will broaden the use of Drosophila melanogaster as a genetic model organism for human-disease related research. Recent years have seen technical advancements like homologous recombination and recombineering. However, generating unequivocal null mutations or tagging endogenous proteins remains a substantial effort for most genes. Here, we describe and demonstrate techniques for using recombineering-based cloning methods to generate vectors that can be used to target and manipulate endogenous loci in vivo. Specifically, we have established a combination of three technologies: (1) BAC transgenesis/recombineering, (2) ends-out homologous recombination and (3) Gateway technology to provide a robust, efficient and flexible method for manipulating endogenous genomic loci. In this protocol, we provide step-by-step details about how to (1) design individual vectors, (2) how to clone large fragments of genomic DNA into the homologous recombination vector using gap repair, and (3) how to replace or tag genes of interest within these vectors using a second round of recombineering. Finally, we will also provide a protocol for how to mobilize these cassettes in vivo to generate a knockout, or a tagged gene via knock-in. These methods can easily be adopted for multiple targets in parallel and provide a means for manipulating the Drosophila genome in a timely and efficient manner. PMID:23893070

  6. Heat switches for ADRs

    NASA Astrophysics Data System (ADS)

    DiPirro, M. J.; Shirron, P. J.

    2014-07-01

    Heat switches are key elements in the cyclic operation of Adiabatic Demagnetization Refrigerators (ADRs). Several of the types of heat switches that have been used for ADRs are described in this paper. Key elements in selection and design of these switches include not only ON/OFF switching ratio, but also method of actuation, size, weight, and structural soundness. Some of the trade-off are detailed in this paper.

  7. Heat Switches for ADRs

    NASA Technical Reports Server (NTRS)

    DiPirro, M. J.; Shirron, P. J.

    2014-01-01

    Heat switches are key elements in the cyclic operation of Adiabatic Demagnetization Refrigerators (ADRs). Several of the types of heat switches that have been used for ADRs are described in this paper. Key elements in selection and design of these switches include not only ON/OFF switching ratio, but also method of actuation, size, weight, and structural soundness. Some of the trade-off are detailed in this paper.

  8. Robust stabilisation and L2 -gain analysis for switched systems with actuator saturation under asynchronous switching

    NASA Astrophysics Data System (ADS)

    Wang, Juan; Zhao, Jun

    2016-09-01

    Robust stabilisation and L2-gain analysis for a class of switched systems with actuator saturation are studied in this paper. The switching signal of the controllers lags behind that of the system modes, which leads to the asynchronous switching between the candidate controllers and the subsystems. By combining the piecewise Lyapunov function method with the convex hull technique, sufficient conditions in terms of LMIs for the solvability of the robust stabilisation and weighted L2-gain problems are presented respectively under the dwell time scheme. Finally, a numerical example is given to demonstrate the feasibility and effectiveness of the proposed results.

  9. On Input-to-State Stability of Switched Stochastic Nonlinear Systems Under Extended Asynchronous Switching.

    PubMed

    Kang, Yu; Zhai, Di-Hua; Liu, Guo-Ping; Zhao, Yun-Bo

    2016-05-01

    An extended asynchronous switching model is investigated for a class of switched stochastic nonlinear retarded systems in the presence of both detection delay and false alarm, where the extended asynchronous switching is described by two independent and exponentially distributed stochastic processes, and further simplified as Markovian. Based on the Razumikhin-type theorem incorporated with average dwell-time approach, the sufficient criteria for global asymptotic stability in probability and stochastic input-to-state stability are given, whose importance and effectiveness are finally verified by numerical examples. PMID:26068932

  10. Multilingual Switch in Peer Classroom Interaction

    ERIC Educational Resources Information Center

    Unamuno, Virginia

    2008-01-01

    Focusing on code-switching from conversation analysis and sociolinguistic perspectives, this paper examines interactions between 10-12-year-old language learners of immigrant origin and locally born students as they are engaging in verbal pair work. All are students attending language classes in state primary schools in Barcelona in which Catalan…

  11. Design of microwave beam-switching networks

    NASA Astrophysics Data System (ADS)

    Burrows, M. L.

    1983-12-01

    An investigation of RF beam-switching networks for creating, with a multiple-beam antenna (MBA), a set of electronically steerable antenna beams, shows that there are three main classes of network. Taking the case of a 61-beam MBA with 8 simultaneously steered beams as an example, we can describe these classes as: (1) networks which can connect any one of the 8 output ports to any one of the 61 beam feeds, (2) networks which can connect the 8 output ports to any set of 8 beam feeds selected from the 61, but with a constraint imposed on the order in which the 8 ports are connected to the 8 selected feeds, and (3) networks in which the set of 8 beam feeds cannot be selected arbitrarily - some fraction of the total number of conceivable interconnections cannot be completed. Networks exist in each of the three classes having very similar traffic handling performance and yet requiring a total number of switches which is very different from one class to the next. Their number is 907, 387, and 175 for the unconstrained, order-constrained and selection-constrained networks, respectively, needed to do the 61-to-8 switching job described above. The report examines the design, performance and complexity of such networks for general N and M. Included are two further measures of complexity, as well as the switching algorithm and the effect of non-uniform traffic. The results are presented as graphs, tables and formulas.

  12. Switching from Computer to Microcomputer Architecture Education

    ERIC Educational Resources Information Center

    Bolanakis, Dimosthenis E.; Kotsis, Konstantinos T.; Laopoulos, Theodore

    2010-01-01

    In the last decades, the technological and scientific evolution of the computing discipline has been widely affecting research in software engineering education, which nowadays advocates more enlightened and liberal ideas. This article reviews cross-disciplinary research on a computer architecture class in consideration of its switching to…

  13. Q-Switching in a Neodymium Laser

    ERIC Educational Resources Information Center

    Holgado, Warein; Sola, Inigo J.; Jarque, Enrique Conejero; Jarabo, Sebastian; Roso, Luis

    2012-01-01

    We present a laboratory experiment for advanced undergraduate or graduate laser-related classes to study the performance of a neodymium laser. In the experiment, the student has to build the neodymium laser using an open cavity. After that, the cavity losses are modulated with an optical chopper located inside, so the Q-switching regime is…

  14. Opc expression, LPS immunotype switch and pilin conversion contribute to serum resistance of unencapsulated meningococci.

    PubMed

    Hubert, Kerstin; Pawlik, Marie-Christin; Claus, Heike; Jarva, Hanna; Meri, Seppo; Vogel, Ulrich

    2012-01-01

    Neisseria meningitidis employs polysaccharides and outer membrane proteins to cope with human serum complement attack. To screen for factors influencing serum resistance, an assay was developed based on a colorimetric serum bactericidal assay. The screening used a genetically modified sequence type (ST)-41/44 clonal complex (cc) strain lacking LPS sialylation, polysaccharide capsule, the factor H binding protein (fHbp) and MutS, a protein of the DNA repair mechanism. After killing of >99.9% of the bacterial cells by serum treatment, the colorimetric assay was used to screen 1000 colonies, of which 35 showed enhanced serum resistance. Three mutant classes were identified. In the first class of mutants, enhanced expression of Opc was identified. Opc expression was associated with vitronectin binding and reduced membrane attack complex deposition confirming recent observations. Lipopolysaccharide (LPS) immunotype switch from immunotype L3 to L8/L1 by lgtA and lgtC phase variation represented the second class. Isogenic mutant analysis demonstrated that in ST-41/44 cc strains the L8/L1 immunotype was more serum resistant than the L3 immunotype. Consecutive analysis revealed that the immunotypes L8 and L1 were frequently observed in ST-41/44 cc isolates from both carriage and disease. Immunotype switch to L8/L1 is therefore suggested to contribute to the adaptive capacity of this meningococcal lineage. The third mutant class displayed a pilE allelic exchange associated with enhanced autoaggregation. The mutation of the C terminal hypervariable region D of PilE included a residue previously associated with increased pilus bundle formation. We suggest that autoaggregation reduced the surface area accessible to serum complement and protected from killing. The study highlights the ability of meningococci to adapt to environmental stress by phase variation and intrachromosomal recombination affecting subcapsular antigens. PMID:23028802

  15. Apollo Ring Optical Switch

    SciTech Connect

    Maestas, J.H.

    1987-03-01

    An optical switch was designed, built, and installed at Sandia National Laboratories in Albuquerque, New Mexico, to facilitate the integration of two Apollo computer networks into a single network. This report presents an overview of the optical switch as well as its layout, switch testing procedure and test data, and installation.

  16. Triggered plasma opening switch

    SciTech Connect

    Mendel, C W

    1988-02-23

    A triggerable opening switch for a very high voltage and current pulse includes a transmission line extending from a source to a load and having an intermediate switch section including a plasma for conducting electrons between transmission line conductors and a magnetic field for breaking the plasma conduction path and magnetically insulating the electrons when it is desired to open the switch.

  17. Triggered plasma opening switch

    DOEpatents

    Mendel, Clifford W.

    1988-01-01

    A triggerable opening switch for a very high voltage and current pulse includes a transmission line extending from a source to a load and having an intermediate switch section including a plasma for conducting electrons between transmission line conductors and a magnetic field for breaking the plasma conduction path and magnetically insulating the electrons when it is desired to open the switch.

  18. Photoionization and Recombination

    NASA Technical Reports Server (NTRS)

    Nahar, Sultana N.

    2000-01-01

    Theoretically self-consistent calculations for photoionization and (e + ion) recombination are described. The same eigenfunction expansion for the ion is employed in coupled channel calculations for both processes, thus ensuring consistency between cross sections and rates. The theoretical treatment of (e + ion) recombination subsumes both the non-resonant recombination ("radiative recombination"), and the resonant recombination ("di-electronic recombination") processes in a unified scheme. In addition to the total, unified recombination rates, level-specific recombination rates and photoionization cross sections are obtained for a large number of atomic levels. Both relativistic Breit-Pauli, and non-relativistic LS coupling, calculations are carried out in the close coupling approximation using the R-matrix method. Although the calculations are computationally intensive, they yield nearly all photoionization and recombination parameters needed for astrophysical photoionization models with higher precision than hitherto possible, estimated at about 10-20% from comparison with experimentally available data (including experimentally derived DR rates). Results are electronically available for over 40 atoms and ions. Photoionization and recombination of He-, and Li-like C and Fe are described for X-ray modeling. The unified method yields total and complete (e+ion) recombination rate coefficients, that can not otherwise be obtained theoretically or experimentally.

  19. Genetically encoded optical activation of DNA recombination in human cells.

    PubMed

    Luo, J; Arbely, E; Zhang, J; Chou, C; Uprety, R; Chin, J W; Deiters, A

    2016-06-30

    We developed two tightly regulated, light-activated Cre recombinase enzymes through site-specific incorporation of two genetically-encoded photocaged amino acids in human cells. Excellent optical off to on switching of DNA recombination was achieved. Furthermore, we demonstrated precise spatial control of Cre recombinase through patterned illumination. PMID:27277957

  20. REMOTE CONTROLLED SWITCHING DEVICE

    DOEpatents

    Hobbs, J.C.

    1959-02-01

    An electrical switching device which can be remotely controlled and in which one or more switches may be accurately operated at predetermined times or with predetermined intervening time intervals is described. The switching device consists essentially of a deck, a post projecting from the deck at right angles thereto, cam means mounted for rotation around said posts and a switch connected to said deck and actuated by said cam means. Means is provided for rotating the cam means at a constant speed and the switching apparatus is enclosed in a sealed container with external adjusting means and electrical connection elements.

  1. Serial Recombination during Circulation of Type 1 Wild-Vaccine Recombinant Polioviruses in China

    PubMed Central

    Liu, Hong-Mei; Zheng, Du-Ping; Zhang, Li-Bi; Oberste, M. Steven; Kew, Olen M.; Pallansch, Mark A.

    2003-01-01

    Type 1 wild-vaccine recombinant polioviruses sharing a 367-nucleotide (nt) block of Sabin 1-derived sequence spanning the VP1 and 2A genes circulated widely in China from 1991 to 1993. We surveyed the sequence relationships among 34 wild-vaccine recombinants by comparing six genomic intervals: the conserved 5′-untranslated region (5′-UTR) (nt 186 to 639), the hypervariable portion of the 5′-UTR (nt 640 to 742), the VP4 and partial VP2 genes (nt 743 to 1176), the VP1 gene (nt 2480 to 3385), the 2A gene (nt 3386 to 3832), and the partial 3D gene (nt 6011 to 6544). The 5′-UTR, capsid (VP4-VP2 and VP1), and 2A sequence intervals had similar phylogenies. By contrast, the partial 3D sequences could be distributed into five divergent genetic classes. Most (25 of 34) of the wild-vaccine recombinant isolates showed no evidence of additional recombination beyond the initial wild-Sabin recombination event. Eight isolates from 1992 to 1993, however, appear to be derived from three independent additional recombination events, and one 1993 isolate was derived from two consecutive events. Complete genomic sequences of a representative isolate for each 3D sequence class demonstrated that these exchanges had occurred in the 2B, 2C, and 3D genes. The 3D gene sequences were not closely related to those of the Sabin strains or 53 diverse contemporary wild poliovirus isolates from China, but all were related to the 3D genes of species C enteroviruses. The appearance within approximately 2.5 years of five recombinant classes derived from a single ancestral infection illustrates the rapid emergence of new recombinants among circulating wild polioviruses. PMID:14512548

  2. Recombination of cluster ions

    NASA Technical Reports Server (NTRS)

    Johnsen, Rainer

    1993-01-01

    Some of our recent work on molecular band emissions from recombination of molecular dimer ions (N4(+) and CO(+) CO) is discussed. Much of the experimental work was done by Y. S. Cao; the results on N4(+) recombination have been published. A brief progress report is given on our ongoing measurements of neutral products of recombination using the flowing-afterglow Langmuir-probe technique in conjunction with laser-induced fluorescence.

  3. Class Size.

    ERIC Educational Resources Information Center

    Johnston, Holly R.

    Exploring the class-size issue, this paper focuses on the primary grades and asks questions such as "does a reduction in class size promote an increase in academic achievement?" and "how substantial does the reduction in numbers have to be in order for a significant increase to occur?" The paper surveys debates on class size and the social factors…

  4. Class Size.

    ERIC Educational Resources Information Center

    Underwood, Siobhan; Lumsden, Linda S.

    1994-01-01

    The items featured in this annotated bibliography touch on several aspects of the multifaceted class-size debate. Allen Odden reviews the literature and contends that class-size reduction should be used "sparingly and strategically." C. M. Achilles and colleagues examines two different class-size situations and find student test performance in the…

  5. 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,…

  6. Recombination in electron coolers

    NASA Astrophysics Data System (ADS)

    Wolf, A.; Gwinner, G.; Linkemann, J.; Saghiri, A. A.; Schmitt, M.; Schwalm, D.; Grieser, M.; Beutelspacher, M.; Bartsch, T.; Brandau, C.; Hoffknecht, A.; Müller, A.; Schippers, S.; Uwira, O.; Savin, D. W.

    2000-02-01

    An introduction to electron-ion recombination processes is given and recent measurements are described as examples, focusing on low collision energies. Discussed in particular are fine-structure-mediated dielectronic recombination of fluorine-like ions, the moderate recombination enhancement by factors of typically 1.5-4 found for most ion species at relative electron-ion energies below about 10 meV, and the much larger enhancement occurring for specific highly charged ions of complex electronic structure, apparently caused by low-energy dielectronic recombination resonances. Recent experiments revealing dielectronic resonances with very large natural width are also described.

  7. Effective switching frequency multiplier inverter

    SciTech Connect

    Su, Gui-Jia; Peng, Fang Z.

    2007-08-07

    A switching frequency multiplier inverter for low inductance machines that uses parallel connection of switches and each switch is independently controlled according to a pulse width modulation scheme. The effective switching frequency is multiplied by the number of switches connected in parallel while each individual switch operates within its limit of switching frequency. This technique can also be used for other power converters such as DC/DC, AC/DC converters.

  8. Molecular Characterization of IgA- and/or IgG-Switched Chronic Lymphocytic Leukemia B Cells

    PubMed Central

    Matolcsy, András; Casali, Paolo; Nádor, Roland G.; Liu, Yi-Fang; Knowles, Daniel M.

    2015-01-01

    The immunoglobulin (Ig) variable region (V) genes expressed by IgM chronic lymphocytic leukemia (CLL) B cells display little or no somatic mutations. However, preliminary findings have shown that Ig V genes of IgA and IgG CLLs may be somatically mutated, suggesting that isotype-switched CLLs may represent a “subtype” of the disease. To investigate the degree and nature of somatic mutations and the role of antigen (Ag) in the clonal selection and expansion of isotype-switched CLLs, and to determine whether specific oncogene or tumor suppressor gene mutations are associated with isotype-switched CLLs, we analyzed the expressed Ig VH gene, bcl-1 and bcl-2 proto-oncogene, and p53 tumor suppressor gene configurations of 3 IgA-, 1 IgG-, and 1 IgA/IgG-expressing CLLs. These isotype-switched CLL B cells expressed surface HLA-DR, CD19, CD23, and CD5, and dis played no alterations of the bcl-1 and bcl-2 oncogenes and the p53 tumor-suppressor gene. The cDNA VH-D-JH gene sequence was joined with that of the Cα gene in the B cells of the three IgA CLLs, and with that of the Cγ gene in the IgG CLL B cells. In the IgA/IgG-coexpressing CLL B cells, identical VH-D-JH cDNA sequences were spliced to either Cα or Cγ genes. In all five CLLs, the pattern of Cμ DNA probe hybridization to the digested genomic DNAs was consistent with deletion of the Cm exon from the rearranged Ig gene locus, suggesting that these CLL B cells had undergone DNA switch recombination. In one IgA CLL, the expressed VH gene was unmutated. In all other class-switched CLLs, the Ig VH segment gene was mutated, but the point mutations were not associated with intraclonal diversification. In one IgA and in the IgA/IgG-coexpressing CLL, the nature and distribution of the mutations were consistent with Ag selection. These findings suggest that IgA- and/or IgG-expressing CLLs represent, in their VH gene structure, transformants of B cells at different stages of ontogeny. They also suggest that Ag may play a

  9. Thermally actuated thermionic switch

    DOEpatents

    Barrus, D.M.; Shires, C.D.

    1982-09-30

    A thermally actuated thermionic switch which responds to an increase of temperature by changing from a high impedance to a low impedance at a predictable temperature set point. The switch has a bistable operation mode switching only on temperature increases. The thermionic material may be a metal which is liquid at the desired operation temperature and held in matrix in a graphite block reservoir, and which changes state (ionizes, for example) so as to be electrically conductive at a desired temperature.

  10. Thermally actuated thermionic switch

    DOEpatents

    Barrus, Donald M.; Shires, Charles D.

    1988-01-01

    A thermally actuated thermionic switch which responds to an increase of temperature by changing from a high impedance to a low impedance at a predictable temperature set point. The switch has a bistable operation mode switching only on temperature increases. The thermionic material may be a metal which is liquid at the desired operation temperature and held in matrix in a graphite block reservoir, and which changes state (ionizes, for example) so as to be electrically conductive at a desired temperature.

  11. AC magnetohydrodynamic microfluidic switch

    SciTech Connect

    Lemoff, A V; Lee, A P

    2000-03-02

    A microfluidic switch has been demonstrated using an AC Magnetohydrodynamic (MHD) pumping mechanism in which the Lorentz force is used to pump an electrolytic solution. By integrating two AC MHD pumps into different arms of a Y-shaped fluidic circuit, flow can be switched between the two arms. This type of switch can be used to produce complex fluidic routing, which may have multiple applications in {micro}TAS.

  12. Solid state switch

    DOEpatents

    Merritt, Bernard T.; Dreifuerst, Gary R.

    1994-01-01

    A solid state switch, with reverse conducting thyristors, is designed to operate at 20 kV hold-off voltage, 1500 A peak, 1.0 .mu.s pulsewidth, and 4500 pps, to replace thyratrons. The solid state switch is more reliable, more economical, and more easily repaired. The switch includes a stack of circuit card assemblies, a magnetic assist and a trigger chassis. Each circuit card assembly contains a reverse conducting thyristor, a resistor capacitor network, and triggering circuitry.

  13. In vitro analysis of a primary, major histocompatibility complex (MHC)-restricted, cytotoxic T-lymphocyte response to avian leukosis virus (ALV), using target cells expressing MHC class I cDNA inserted into a recombinant ALV vector.

    PubMed

    Thacker, E L; Fulton, J E; Hunt, H D

    1995-10-01

    The interaction between the major histocompatibility complex (MHC) and cytotoxic T lymphocytes (CTLs) is an important component of the host's resistance to viral infections and tumor formation. In this study, an avian leukosis virus (ALV) vector system, RCASBP, expressing MHC chicken class I (B-F) cDNA was used to develop target cells expressing the chicken class I glycoproteins complexed with ALV antigens on the cell surface. Peripheral blood from chickens inoculated with ALV was shown to contain antigen-specific, MHC-restricted, CD8+ effector CTLs, using a 51Cr release assay utilizing the RCASBP B-F target cells. The stimulated effector cells were also predominantly alpha beta T-cell receptor-positive (TCR2) T cells. The CTL response varied between two haplotypes of chickens which differed in their response to Rous sarcoma virus (RSV)-induced tumors. Chickens with the B21 haplotype which regress RSV-induced tumors showed maximal cytolytic activity, while chickens with the B13 haplotype which do not regress RSV-induced tumors had minimal to no cytolytic activity. In addition to assessing the CTL response to ALV, the creation of MHC-specific immortal target cell lines will be extremely useful in evaluating CTL responses to other viral disease in chickens. PMID:7666545

  14. Reusable fast opening switch

    DOEpatents

    Van Devender, J.P.; Emin, D.

    1983-12-21

    A reusable fast opening switch for transferring energy, in the form of a high power pulse, from an electromagnetic storage device such as an inductor into a load. The switch is efficient, compact, fast and reusable. The switch comprises a ferromagnetic semiconductor which undergoes a fast transition between conductive and metallic states at a critical temperature and which undergoes the transition without a phase change in its crystal structure. A semiconductor such as europium rich europhous oxide, which undergoes a conductor to insulator transition when it is joule heated from its conductor state, can be used to form the switch.

  15. Alarm toe switch

    DOEpatents

    Ganyard, Floyd P.

    1982-01-01

    An alarm toe switch inserted within a shoe for energizing an alarm circuit n a covert manner includes an insole mounting pad into which a miniature reed switch is fixedly molded. An elongated slot perpendicular to the reed switch is formed in the bottom surface of the mounting pad. A permanent cylindrical magnet positioned in the forward portion of the slot with a diameter greater than the pad thickness causes a bump above the pad. A foam rubber block is also positioned in the slot rearwardly of the magnet and holds the magnet in normal inoperative relation. A non-magnetic support plate covers the slot and holds the magnet and foam rubber in the slot. The plate minimizes bending and frictional forces to improve movement of the magnet for reliable switch activation. The bump occupies the knuckle space beneath the big toe. When the big toe is scrunched rearwardly the magnet is moved within the slot relative to the reed switch, thus magnetically activating the switch. When toe pressure is released the foam rubber block forces the magnet back into normal inoperative position to deactivate the reed switch. The reed switch is hermetically sealed with the magnet acting through the wall so the switch assembly S is capable of reliable operation even in wet and corrosive environments.

  16. Simplified Design Equations for Class-E Neural Prosthesis Transmitters

    PubMed Central

    Troyk, Philip; Hu, Zhe

    2013-01-01

    Extreme miniaturization of implantable electronic devices is recognized as essential for the next generation of neural prostheses, owing to the need for minimizing the damage and disruption of the surrounding neural tissue. Transcutaneous power and data transmission via a magnetic link remains the most effective means of powering and controlling implanted neural prostheses. Reduction in the size of the coil, within the neural prosthesis, demands the generation of a high-intensity radio frequency magnetic field from the extracoporeal transmitter. The Class-E power amplifier circuit topology has been recognized as a highly effective means of producing large radio frequency currents within the transmitter coil. Unfortunately, design of a Class-E circuit is most often fraught by the need to solve a complex set of equations so as to implement both the zero-voltage-switching and zero-voltage-derivative-switching conditions that are required for efficient operation. This paper presents simple explicit design equations for designing the Class-E circuit topology. Numerical design examples are presented to illustrate the design procedure. PMID:23292784

  17. Recombination between bacteriophage lambda and plasmid pBR322 in Escherichia coli.

    PubMed Central

    Pogue-Geile, K L; Dassarma, S; King, S R; Jaskunas, S R

    1980-01-01

    Recombinant lambda phages were isolated that resulted from recombination between the lambda genome and plasmid pBR322 in Escherichia coli, even though these deoxyribonucleic acids (DNAs) did not share extensive regions of homology. The characterization of these recombinant DNAs by heteroduplex analysis and restriction endonucleases is described. All but one of the recombinants appeared to have resulted from reciprocal recombination between a site on lambda DNA and a site on the plasmid. In general, there were two classes of recombinants. One class appeared to have resulted from recombination at the phage attachment site that probably resulted from lambda integration into secondary attachment sites on the plasmid. Seven different secondary attachment sites on pBR322 were found. The other class resulted from plasmid integration at other sites that were widely scattered on the lambda genome. For this second class of recombinants, more than one site on the plasmid could recombine with lambda DNA. Thus, the recombination did not appear to be site specific with respect to lambda or the plasmid. Possible mechanisms for generating these recombinants are discussed. Images PMID:6247334

  18. The mismatch repair system reduces meiotic homeologous recombination and stimulates recombination-dependent chromosome loss.

    PubMed Central

    Chambers, S R; Hunter, N; Louis, E J; Borts, R H

    1996-01-01

    Efficient genetic recombination requires near-perfect homology between participating molecules. Sequence divergence reduces the frequency of recombination, a process that is dependent on the activity of the mismatch repair system. The effects of chromosomal divergence in diploids of Saccharomyces cerevisiae in which one copy of chromosome III is derived from a closely related species, Saccharomyces paradoxus, have been examined. Meiotic recombination between the diverged chromosomes is decreased by 25-fold. Spore viability is reduced with an observable increase in the number of tetrads with only two or three viable spores. Asci with only two viable spores are disomic for chromosome III, consistent with meiosis I nondisjunction of the homeologs. Asci with three viable spores are highly enriched for recombinants relative to tetrads with four viable spores. In 96% of the class with three viable spores, only one spore possesses a recombinant chromosome III, suggesting that the recombination process itself contributes to meiotic death. This phenomenon is dependent on the activities of the mismatch repair genes PMS1 and MSH2. A model of mismatch-stimulated chromosome loss is proposed to account for this observation. As expected, crossing over is increased in pms1 and msh2 mutants. Furthermore, genetic exchange in pms1 msh2 double mutants is affected to a greater extent than in either mutant alone, suggesting that the two proteins act independently to inhibit homeologous recombination. All mismatch repair-deficient strains exhibited reductions in the rate of chromosome III nondisjunction. PMID:8887641

  19. Design and characterization of the Pegasus I plasma flow switch

    SciTech Connect

    Wysocki, F.J.; Bartsch, R.R.; Bowers, R.L.; Cochrane, J.C.; Greene, A.E.; Ladish, J.S.; Lee, P.H.Y.; Parker, J.V.; Peterson, D.L.; Benage, J.F.; Gribble, R.F.; Shlachter, J.S.; Scudder, D.W. ); Roderick, N.F. ); Turchi, P.J. )

    1992-01-01

    A plasma flow switch (PFS) based on an experimental design utilized at the Air Force Weapons Laboratory (AFWL) has been fielded on the Pegasus I system. Currents of as much as 5.8 MA have been switched on a time scale (10--90%) of 200 ns into a static load. Tests with a foil load have resulted in a less than ideal current transfer because of interaction between the dynamic load and the plasma flow switch. Two-dimensional radiation magnetohydrodynamic (RMHD) calculations have been used to characterize the behavior of the PFS and to aid in the design. These calculations have also been used to investigate several classes of perturbations in the switch-plasma, the interaction of the switch-plasma with the coaxial electrodes, and their effect on switching efficiency. The Pegasus I PFS experiments reproduce the results obtained at AFWL throughout the switch initiation and switching phases. However, early in the foil implosion the switched current drops significantly. We believe this current drop results from a small amount of residual plasma left behind in the PFS region that impedes the magnetic-flux transport, thereby preventing efficient foil implosion. The experimental results suggest that the method of PFS initiation (viz. chordal wire arrays) may be an important factor in switch operation.

  20. Genetic recombination. [Escherichia coli

    SciTech Connect

    Stahl, F.W.

    1987-02-01

    The molecular pathways of gene recombination are explored and compared in studies of the model organisms, Escherichia coli and phase lambda. In the discussion of data from these studies it seems that recombination varies with the genetic idiosyncrasies of the organism and may also vary within a single organism.

  1. Control discrete-time switched singular systems with state delays under asynchronous switching

    NASA Astrophysics Data System (ADS)

    Lin, Jinxing; Fei, Shumin; Gao, Zhifeng

    2013-06-01

    This article is concerned with the problem of state feedback control for a class of discrete-time switched singular systems with time-varying state delays under asynchronous switching. The asynchronous switching considered here means that the switching instants of the candidate controllers lag behind those of the system modes. The concept of mismatched control rate is introduced. By using the multiple Lyapunov function approach and the average dwell time technique, a sufficient condition for the existence a stabilising switching law is first derived to guarantee the regularity, causality and exponential stability of the closed-loop system in the presence of asynchronous switching. The stabilising switching law is characterised by a upper bound on the mismatched control rate and a lower bound on the average dwell time. Then, the corresponding solvability condition for a set of mode-dependent state feedback controllers is established by using the linear matrix inequality (LMI) technique. Finally, a numerical example is provided to illustrate the effectiveness of the proposed method.

  2. Asymmetrical Switch Costs in Children

    ERIC Educational Resources Information Center

    Ellefson, Michelle R.; Shapiron, Laura R.; Chater, Nick

    2006-01-01

    Switching between tasks produces decreases in performance as compared to repeating the same task. Asymmetrical switch costs occur when switching between two tasks of unequal difficulty. This asymmetry occurs because the cost is greater when switching to the less difficult task than when switching to the more difficult task. Various theories about…

  3. [Switching and combining strategies of antidepressant medications].

    PubMed

    Charpeaud, Thomas; Moliere, Fanny; Bubrovszky, Maxime; Haesebaert, Frédéric; Allaïli, Najib; Bation, Rémy; Nieto, Isabel; Richieri, Raphaëlle; Saba, Ghassen; Bellivier, Frank; Bennabi, Djamila; Holtzmann, Jérôme; Camus, Vincent; Courtet, Philippe; Courvoisier, Pierre; d'Amato, Thierry; Doumy, Olivier; Garnier, Marion; Bougerol, Thierry; Lançon, Christophe; Haffen, Emmanuel; Leboyer, Marion; Llorca, Pierre-Michel; Vaiva, Guillaume; El-Hage, Wissam; Aouizerate, Bruno

    2016-03-01

    Switching antidepressant medication may be helpful in depressed patients having no benefit from the initial antidepressant treatment. Before considering switching strategy, the initial antidepressant treatment should produce no therapeutic effect after at least 4 weeks of administration at adequate dosage. Choosing an antidepressant of pharmacologically distinct profile fails to consistently demonstrate a significant superiority in terms of effectiveness over the switching to another antidepressant within the same pharmacological class. Augmenting SSRI/SNRIs with mirtazapine/mianserin has become the most recommended strategy of antidepressant combinations. Augmenting SSRI with tricyclic drugs is now a less recommended strategy of antidepressant combinations given the increased risk for the occurrence of pharmacokinetic drug-drug interactions and adverse effects. PMID:26995511

  4. Manually operated coded switch

    DOEpatents

    Barnette, Jon H.

    1978-01-01

    The disclosure relates to a manually operated recodable coded switch in which a code may be inserted, tried and used to actuate a lever controlling an external device. After attempting a code, the switch's code wheels must be returned to their zero positions before another try is made.

  5. Reflective HTS switch

    DOEpatents

    Martens, Jon S.; Hietala, Vincent M.; Hohenwarter, Gert K. G.

    1994-01-01

    A HTS switch includes a HTS conductor for providing a superconducting path for an electrical signal and an serpentine wire actuator for controllably heating a portion of the conductor sufficiently to cause that portion to have normal, and not superconducting, resistivity. Mass of the portion is reduced to decrease switching time.

  6. Reflective HTS switch

    DOEpatents

    Martens, J.S.; Hietala, V.M.; Hohenwarter, G.K.G.

    1994-09-27

    A HTS (High Temperature Superconductor) switch includes a HTS conductor for providing a superconducting path for an electrical signal and an serpentine wire actuator for controllably heating a portion of the conductor sufficiently to cause that portion to have normal, and not superconducting, resistivity. Mass of the portion is reduced to decrease switching time. 6 figs.

  7. Multidimensional set switching.

    PubMed

    Hahn, Sowon; Andersen, George J; Kramer, Arthur F

    2003-06-01

    The present study examined the organization of preparatory processes that underlie set switching and, more specifically, switch costs. On each trial, subjects performed one of two perceptual judgment tasks, color or shape discrimination. Subjects also responded with one of two different response sets. The task set and/or the response set switched from one to the other after 2-6 repeated trials. Response set, task set, and double set switches were performed in both blocked and randomized conditions. Subjects performed with short (100-msec) and long (800-msec) preparatory intervals. Task and response set switches had an additive effect on reaction times (RTs) in the blocked condition. Such a pattern of results suggests a serial organization of preparatory processes when the nature of switches is predictable. However, task and response set switches had an underadditive effect on RTs in the random condition when subjects performed with a brief cue-to-target interval. This pattern of results suggests overlapping task and response set preparation. These findings are discussed in terms of strategic control of preparatory processes in set switching. PMID:12921431

  8. A Novel Molecular Switch

    PubMed Central

    Daber, Robert; Lewis, Mitchell

    2009-01-01

    Transcriptional regulation is a fundamental process for regulating the flux of all metabolic pathways. For the last several decades, the lac operon has served as a valuable model for studying transcription. More recently, the switch that controls the operon has also been successfully adapted to function in mammalian cells. Here we describe how, using directed evolution, we have created a novel switch that recognizes an asymmetric operator sequence. The new switch has a repressor with altered headpiece domains for operator recognition, and a redesigned dimer interface to create a heterodimeric repressor. Quite unexpectedly, the heterodimeric switch functions better than the natural system. It can repress more tightly than the naturally occurring switch of the lac operon; it is less leaky and can be induced more efficiently. Ultimately these novel repressors could be evolved to recognize eukaryotic promoters and used to regulate gene expression in mammalian systems. PMID:19540845

  9. Erected mirror optical switch

    DOEpatents

    Allen, James J.

    2005-06-07

    A microelectromechanical (MEM) optical switching apparatus is disclosed that is based on an erectable mirror which is formed on a rotatable stage using surface micromachining. An electrostatic actuator is also formed on the substrate to rotate the stage and mirror with a high angular precision. The mirror can be erected manually after fabrication of the device and used to redirect an incident light beam at an arbitrary angel and to maintain this state in the absence of any applied electrical power. A 1.times.N optical switch can be formed using a single rotatable mirror. In some embodiments of the present invention, a plurality of rotatable mirrors can be configured so that the stages and mirrors rotate in unison when driven by a single micromotor thereby forming a 2.times.2 optical switch which can be used to switch a pair of incident light beams, or as a building block to form a higher-order optical switch.

  10. Nanoscale memristive radiofrequency switches

    NASA Astrophysics Data System (ADS)

    Pi, Shuang; Ghadiri-Sadrabadi, Mohammad; Bardin, Joseph C.; Xia, Qiangfei

    2015-06-01

    Radiofrequency switches are critical components in wireless communication systems and consumer electronics. Emerging devices include switches based on microelectromechanical systems and phase-change materials. However, these devices suffer from disadvantages such as large physical dimensions and high actuation voltages. Here we propose and demonstrate a nanoscale radiofrequency switch based on a memristive device. The device can be programmed with a voltage as low as 0.4 V and has an ON/OFF conductance ratio up to 1012 with long state retention. We measure the radiofrequency performance of the switch up to 110 GHz and demonstrate low insertion loss (0.3 dB at 40 GHz), high isolation (30 dB at 40 GHz), an average cutoff frequency of 35 THz and competitive linearity and power-handling capability. Our results suggest that, in addition to their application in memory and computing, memristive devices are also a leading contender for radiofrequency switch applications.