scholarly journals Tuning of in vivo cognate B-T cell interactions by Intersectin 2 is required for effective anti-viral B cell immunity

eLife ◽  
2018 ◽  
Vol 7 ◽  
Author(s):  
Marianne Burbage ◽  
Francesca Gasparrini ◽  
Shweta Aggarwal ◽  
Mauro Gaya ◽  
Johan Arnold ◽  
...  
Keyword(s):  
B Cell ◽  
Adaptive Immune Response ◽  
Adaptor Protein ◽  
Small Gtpase ◽  
Germinal Centre ◽  
Nucleotide Exchange ◽  
Interacting Protein ◽  
Flu Vaccination ◽  
Cell Immunity

Wiskott-Aldrich syndrome (WAS) is an immune pathology associated with mutations in WAS protein (WASp) or in WASp interacting protein (WIP). Together with the small GTPase Cdc42 and other effectors, these proteins participate in the remodelling of the actin network downstream of BCR engagement. Here we show that mice lacking the adaptor protein ITSN2, a G-nucleotide exchange factor (GEF) for Cdc42 that also interacts with WASp and WIP, exhibited increased mortality during primary infection, incomplete protection after Flu vaccination, reduced germinal centre formation and impaired antibody responses to vaccination. These defects were found, at least in part, to be intrinsic to the B cell compartment. In vivo, ITSN2 deficient B cells show a reduction in the expression of SLAM, CD84 or ICOSL that correlates with a diminished ability to form long term conjugates with T cells, to proliferate in vivo, and to differentiate into germinal centre cells. In conclusion, our study not only revealed a key role for ITSN2 as an important regulator of adaptive immune-response during vaccination and viral infection but it is also likely to contribute to a better understanding of human immune pathologies.

scholarly journals P66Shc: A Pleiotropic Regulator of B Cell Trafficking and a Gatekeeper in Chronic Lymphocytic Leukemia

Cancers ◽  
2020 ◽  
Vol 12 (4) ◽  
pp. 1006 ◽  
Author(s):  
Laura Patrussi ◽  
Nagaja Capitani ◽  
Cosima T. Baldari
Keyword(s):  
Chronic Lymphocytic Leukemia ◽  
B Cell ◽  
Chemokine Receptors ◽  
Adaptor Protein ◽  
Leukemic Cell ◽  
Lymphocytic Leukemia ◽  
Cell Trafficking ◽  
Gene Encoding ◽  
Cell Invasiveness

Neoplastic B cells from chronic lymphocytic leukemia patients (CLL) have a profound deficiency in the expression of p66Shc, an adaptor protein with pro-apoptotic and pro-oxidant activities. This defect results in leukemic B cell resistance to apoptosis and additionally impinges on the balance between chemokine receptors that control B cell homing to secondary lymphoid organs and the sphingosine phosphate receptor S1PR1 that controls their egress therefrom, thereby favoring leukemic B cell accumulation in the pro-survival lymphoid niche. Ablation of the gene encoding p66Shc in the Eµ-TCL1 mouse model of human CLL enhances leukemogenesis and promotes leukemic cell invasiveness in both nodal and extranodal organs, providing in vivo evidence of the pathogenic role of the p66Shc defect in CLL pathogenesis. Here we present an overview of the functions of p66Shc in B lymphocytes, with a specific focus on the multiple mechanisms exploited by p66Shc to control B cell trafficking and the abnormalities in this process caused by p66Shc deficiency in CLL.

scholarly journals Molecular characterization of centerin, a germinal centre cell serpin

2007 ◽  
Vol 405 (3) ◽  
pp. 489-494 ◽  
Author(s):  
Melinda A. Paterson ◽  
Anita J. Horvath ◽  
Robert N. Pike ◽  
Paul B. Coughlin
Keyword(s):  
B Cell ◽  
Serine Protease ◽  
Molecular Characterization ◽  
Good Prognosis ◽  
Germinal Centre ◽  
Lymphoid Malignancies ◽  
B Cell Maturation ◽  
Absolute Requirement

Centerin [SERPINA9/GCET1 (germinal centre B-cell-expressed transcript 1)] is a serpin (serine protease inhibitor) whose expression is restricted to germinal centre B-cells and lymphoid malignancies with germinal centre B-cell maturation. Expression of centerin, together with bcl-6 and GCET2, constitutes a germinal centre B-cell signature, which is associated with a good prognosis in diffuse large B-cell lymphomas, but the molecular basis for this remains to be elucidated. We report here the cloning, expression and molecular characterization of bacterial recombinant centerin. Biophysical studies demonstrated that centerin was able to undergo the ‘stressed to relaxed’ conformational change which is an absolute requirement for protease inhibitory activity. Kinetic analysis showed that centerin rapidly inhibited the serine protease trypsin (ka=1.9×105 M−1·s−1) and also demonstrated measurable inhibition of thrombin (ka=1.17×103 M−1·s−1) and plasmin (ka=1.92×103 M−1·s−1). Centerin also bound DNA and unfractionated heparin, although there was no functionally significant impact on the rate of inhibition. These results suggest that centerin is likely to function in vivo in the germinal centre as an efficient inhibitor of a trypsin-like protease.

scholarly journals Specific In Vivo Deletion of B-Cell Subpopulations Expressing Human Immunoglobulins by the B-Cell Superantigen Protein L

2004 ◽  
Vol 72 (6) ◽  
pp. 3515-3523 ◽  
Author(s):  
Muriel Viau ◽  
Nancy S. Longo ◽  
Peter E. Lipsky ◽  
Lars Björck ◽  
Moncef Zouali
Keyword(s):  
B Cells ◽  
B Cell ◽  
Immune Defense ◽  
Protein L ◽  
Variable Regions ◽  
Marginal Zone B Cells ◽  
Human Immunoglobulins ◽  
Cell Immunity ◽  
B Cell Subsets

ABSTRACT Some pathogens have evolved to produce proteins, called B-cell superantigens, that can interact with human immunoglobulin variable regions, independently of the combining site, and activate B lymphocytes that express the target immunoglobulins. However, the in vivo consequences of these interactions on human B-cell numbers and function are largely unknown. Using transgenic mice expressing fully human immunoglobulins, we studied the consequences of in vivo exposure of protein L of Peptostreptococcus magnus with human immunoglobulins. In the mature pool of B cells, protein L exposure resulted in a specific reduction of splenic marginal-zone B cells and peritoneal B-1 cells. Splenic B cells exhibited a skewed light-chain repertoire consistent with the capacity of protein L to bind specific kappa gene products. Remarkably, these two B-cell subsets are implicated in innate B-cell immunity, allowing rapid clearance of pathogens. Thus, the present study reveals a novel mechanism that may be used by some infectious agents to subvert a first line of the host's immune defense.

In vivo activity of rituximab-CpG oligodeoxynucleotide conjugate against rituximab-resistant human CD20+ B-cell lymphoma

2009 ◽  
Vol 27 (15_suppl) ◽  
pp. 8529-8529
Author(s):  
J. Timmerman ◽  
D. Betting ◽  
R. Yamada ◽  
S. Hurvitz ◽  
K. Steward ◽  
...  
Keyword(s):  
B Cell ◽  
Cell Lymphoma ◽  
Single Agent ◽  
B Cell Lymphoma ◽  
Cpg Oligodeoxynucleotides ◽  
Cell Immunity ◽  
Cpg Oligodeoxynucleotide ◽  
Immunocompetent Mice ◽  
Anti Cd20

8529 Background: The anti-CD20 monoclonal antibody rituximab (R) is a mainstay in the treatment of B cell non-Hodgkin's lymphoma (NHL), exerting anti-tumor effects via antibody-dependent cellular cytotoxicity (ADCC), complement-dependent cytotoxicity, and apoptosis induction. Toll-like receptor 9 agonist CpG oligodeoxynucleotides (CpG) are potent activators of ADCC and T cell immunity, and have been studied for anti-NHL effects when administered by systemic or intratumoral routes. We sought to optimize the delivery of CpG to sites of NHL to improve R efficacy. Methods: We utilized an aggressive syngeneic human CD20-expressing murine B cell lymphoma (38C13-huCD20) to study the in vivo augmentation of R efficacy by CpG. Established tumors in immunocompetent mice were treated with R plus CpG given systemically, intratumorally, or chemically linked to antibody using maleimide-sulfhydryl chemistry (D. Betting et al, J. Immunol. 2008; 181: 4131). Results: 5- to 7-day established tumors were completely resistant to single agent R. Combining intratumoral, but not systemic administration of CpG with R resulted in tumor eradication from up to 42% of mice (p < 0.0003 vs. R alone, CpG alone, R + systemic CpG). Mechanistic studies indicated that both natural killer cells and complement participated in the cure of tumors by intratumoral CpG + R, by increasing tumor cell sensitivity to complement and ADCC lysis, and by augmenting the cytotoxicity of ADCC effectors. To overcome the need for repeated direct intratumoral injections and allow precise targeting of CpG to tumor cells, we chemically linked CpG to R using a cleavable linker. A single i.v. injection of this R-CpG conjugate achieved eradication of established tumors from 100% of mice. In contrast, equivalent doses of unlinked i.v. R + CpG, CpG alone, or R alone cured only 8% of mice. Thus, combining CpG with R was most effective using direct conjugation to the antibody. Conclusions: In conclusion, enhancement of R efficacy required sustained intratumoral delivery of CpG to maximize anti-tumor responses. R-CpG conjugate efficiently eradicated an established B cell lymphoma that is fully resistant to single-agent R. Clinical testing of anti-CD20-CpG conjugates against B cell NHL is thus warranted. [Table: see text]

scholarly journals The in vivo function of a noncanonical TRAF2-binding domain in the C-terminus of CD40 in driving B-cell growth and differentiation

Blood ◽  
2007 ◽  
Vol 110 (1) ◽  
pp. 193-200 ◽  
Author(s):  
Li-Fan Lu ◽  
Cory L. Ahonen ◽  
Evan F. Lind ◽  
Vanitha S. Raman ◽  
W. James Cook ◽  
...  
Keyword(s):  
Immune Responses ◽  
B Cell ◽  
Cell Activation ◽  
Affinity Maturation ◽  
Functional Domain ◽  
Antibody Isotype ◽  
Humoral Immune ◽  
Humoral Immune Responses ◽  
Cell Immunity

The recruitment of tumor necrosis factor receptor–associated factors (TRAFs) 1, 2, 3, 5, and 6 to the CD40 cytoplasmic tail upon CD40 trimerization results in downstream signaling events that ultimately lead to CD40-dependent, thymus-dependent (TD) humoral immune responses. Previously, we have shown signaling through the C-terminal tail of CD40 in the absence of canonical TRAF-binding sites is capable of signaling through an alternative TRAF2-binding site. Here, we demonstrate that B cells from mice harboring CD40 with only the C-terminal tail can activate both canonical and noncanonical NFκB signaling pathways. Moreover, while lacking germinal center formation, several hallmarks of humoral immune responses including clonal B-cell activation/expansion, antibody isotype switching, and affinity maturation remain normal. This study demonstrates a new functional domain in CD40 that controls critical aspects of B-cell immunity in an in vivo setting.

scholarly journals Identification of Tetratricopeptide Repeat 1 as an Adaptor Protein That Interacts with Heterotrimeric G Proteins and the Small GTPase Ras

2003 ◽  
Vol 23 (11) ◽  
pp. 3847-3858 ◽  
Author(s):  
Caroline Marty ◽  
Darren D. Browning ◽  
Richard D. Ye
Keyword(s):  
G Proteins ◽  
Mammalian Cells ◽  
Active Form ◽  
Adaptor Protein ◽  
Small Gtpases ◽  
Small Gtpase ◽  
Tetratricopeptide Repeat ◽  
Heterotrimeric G Proteins ◽  
Interacting Protein

ABSTRACT The biological functions of heterotrimeric G proteins and small GTPases are modulated by both extracellular stimuli and intracellular regulatory proteins. Using Saccharomyces cerevisiae two-hybrid screening, we identified tetratricopeptide repeat 1 (TPR1), a 292-amino-acid protein with three TPR motifs, as a Gα16-binding protein. The interaction was confirmed both in vitro and in transfected mammalian cells, where TPR1 also binds to several other Gα proteins. TPR1 was found to interact with Ha-Ras preferentially in its active form. Overexpression of TPR1 promotes accumulation of active Ras. TPR1 was found to compete with the Ras-binding domain (RBD) of Raf-1 for binding to the active Ras, suggesting that it may also compete with Ras GTPase-activating protein, thus contributing to the accumulation of GTP-bound Ras. Expression of Gα16 strongly enhances the interaction between TPR1 and Ras. Removal of the TPR1 N-terminal 112 residues abolishes potentiation by Gα16 while maintaining the interaction with Gα16 and the ability to discriminate active Ras from wild-type Ras. We have also observed that LGN, a Gαi-interacting protein with seven TPR motifs, binds Ha-Ras. Thus, TPR1 is a novel adaptor protein for Ras and selected Gα proteins that may be involved in protein-protein interaction relating to G-protein signaling.

scholarly journals Loss of EPAC2 results in altered synaptic composition of dendritic spines and excitatory and inhibitory balance

2019 ◽  
Author(s):  
Kelly A Jones ◽  
Michiko Sumiya ◽  
Kevin M Woolfrey ◽  
Deepak P Srivastava ◽  
Peter Penzes
Keyword(s):  
Ampa Receptors ◽  
Cortical Neurons ◽  
Glutamate Transporter ◽  
Autism Spectrum ◽  
Small Gtpase ◽  
Anterior Cingulate ◽  
Synaptic Proteins ◽  
Inhibitory Synapses ◽  
Nucleotide Exchange

EPAC2 is a guanine nucleotide exchange factor that regulates GTPase activity of the small GTPase Rap and Ras) and is highly enriched at synapses. Activation of EPAC2 has been shown to induce dendritic spine shrinkage and increase spine motility, effects that are necessary for synaptic plasticity. These morphological effects are dysregulated by rare mutations of EPAC2 associated with autism spectrum disorders. In addition, EPAC2 destabilizes synapses through the removal of synaptic GluA2/3-containing AMPA receptors. Previous work has shown that Epac2 knockout mice (Epac2-/-) display abnormal social interactions, as well as gross disorganization of the frontal cortex and abnormal spine motility in vivo. In this study we sought to further understand the cellular consequences of knocking out Epac2 on the development of neuronal and synaptic structure and organization of cortical neurons. Using primary cortical neurons generated from Epac2+/+ or Epac2-/- mice, we confirm that EPAC2 is required for cAMP-dependent spine shrinkage. Neurons from Epac2-/- mice also displayed increased synaptic expression of GluA2/3-containing AMPA receptors, as well as of the adhesion protein N-cadherin. Intriguingly, analysis of excitatory and inhibitory synaptic proteins revealed that loss of EPAC2 resulted in altered of expression of vesicular glutamate transporter 1 (VGluT1) and vesicular GABA transporter (VGAT), indicating a potential imbalance in excitatory/inhibitory inputs onto neurons. Finally, examination of cortical neurons located within the anterior cingulate cortex further revealed subtle deficits in the establishment of dendritic arborization in vivo. These data provide evidence that EPAC2 is required for the correct composition of synapses and that loss of this protein could result in an imbalance of excitatory and inhibitory synapses.

The small GTPase KlRho5 responds to oxidative stress and affects cytokinesis

2021 ◽  
Vol 134 (18) ◽  
Author(s):  
Marius Musielak ◽  
Carolin C. Sterk ◽  
Felix Schubert ◽  
Christian Meyer ◽  
Achim Paululat ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Mitochondrial Dynamics ◽  
Kluyveromyces Lactis ◽  
Neck Region ◽  
Small Gtpase ◽  
Nutrient Sensing ◽  
Deletion Mutants ◽  
Nucleotide Exchange ◽  
Morphological Defects

ABSTRACT Rho5 is the yeast homolog of the human small GTPase Rac1. We characterized the genes encoding Rho5 and the subunits of its dimeric activating guanine-nucleotide-exchange factor (GEF), Dck1 and Lmo1, in the yeast Kluyveromyces lactis. Rapid translocation of the three GFP-tagged components to mitochondria upon oxidative stress and carbon starvation indicate a similar function of KlRho5 in energy metabolism and mitochondrial dynamics as described for its Saccharomyces cerevisiae homolog. Accordingly, Klrho5 deletion mutants are hyper-resistant towards hydrogen peroxide. Moreover, synthetic lethalities of rho5 deletions with key components in nutrient sensing, such as sch9 and gpr1, are not conserved in K. lactis. Instead, Klrho5 deletion mutants display morphological defects with strengthened lateral cell walls and protruding bud scars. The latter result from aberrant cytokinesis, as observed by following the budding process in vivo and by transmission electron microscopy of the bud neck region. This phenotype can be suppressed by KlCDC42G12V, which encodes a hyper-active variant. Data from live-cell fluorescence microscopy support the notion that KlRho5 interferes with the actin moiety of the contractile actomyosin ring, with consequences different from those previously reported for mutants lacking myosin.

scholarly journals Retinitis Pigmentosa GTPase Regulator (RPGR)-interacting Protein Is Stably Associated with the Photoreceptor Ciliary Axoneme and Anchors RPGR to the Connecting Cilium

2000 ◽  
Vol 276 (15) ◽  
pp. 12091-12099 ◽  
Author(s):  
Dong-Hyun Hong ◽  
Guohua Yue ◽  
Michael Adamian ◽  
Tiansen Li
Keyword(s):  
Retinitis Pigmentosa ◽  
Coiled Coil ◽  
Retinal Disease ◽  
Photoreceptor Cells ◽  
Nucleotide Exchange ◽  
Specific Expression ◽  
Interacting Protein ◽  
Nucleotide Exchange Factor ◽  
Retinitis Pigmentosa Gtpase Regulator

Retinitis pigmentosa (RP) is a blinding retinal disease in which the photoreceptor cells degenerate. Mutations in the gene for retinitis pigmentosa GTPase regulator (RPGR) are a frequent cause of RP. The function of RPGR is not well understood, but it is thought to be a putative guanine nucleotide exchange factor for an unknown G protein. Ablation of theRPGRgene in mice suggested a role in maintaining the polarized distribution of opsin across the cilia. To investigate its function, we used a protein interaction screen to identify candidate proteins that may interact physiologically with RPGR. One such protein, designated RPGR-interacting protein (RPGRIP), is expressed specifically in rod and cone photoreceptors. It consists of an N-terminal region predicted to form coiled coil structures linked to a C-terminal tail that binds RPGR.In vivo, both proteins co-localize in the photoreceptor connecting cilia. RPGRIP is stably associated with the ciliary axoneme independent of RPGR and is resistant to extraction under conditions that partially solubilized other cytoskeletal components. When over-expressed in heterologous cell lines, RPGRIP appears in insoluble punctate and filamentous structures. These data suggest that RPGRIP is a structural component of the ciliary axoneme, and one of its functions is to anchor RPGR within the cilium. RPGRIP is the only protein known to localize specifically in the photoreceptor connecting cilium. As such, it is a candidate gene for human photoreceptor disease. The tissue-specific expression of RPGRIP explains why mutations in the ubiquitously expressed RPGR confer a photoreceptor-specific phenotype.

scholarly journals The Ability of Sos1 to Oligomerize the Adaptor Protein LAT Is Separable from Its Guanine Nucleotide Exchange Activity in Vivo

2013 ◽  
Vol 6 (301) ◽  
pp. ra99-ra99 ◽  
Author(s):  
R. L. Kortum ◽  
L. Balagopalan ◽  
C. P. Alexander ◽  
J. Garcia ◽  
J. M. Pinski ◽  
...  
Keyword(s):  
Adaptor Protein ◽  
Guanine Nucleotide ◽  
Nucleotide Exchange ◽  
Guanine Nucleotide Exchange ◽  
Exchange Activity
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