scholarly journals Endoplasmic reticulum-resident E3 ubiquitin ligase Hrd1 controls B-cell immunity through degradation of the death receptor CD95/Fas

2016 ◽  
Vol 113 (37) ◽  
pp. 10394-10399 ◽  
Author(s):  
Sinyi Kong ◽  
Yi Yang ◽  
Yuanming Xu ◽  
Yajun Wang ◽  
Yusi Zhang ◽  
...  
Keyword(s):  
Cell Death ◽  
B Cells ◽  
B Cell ◽  
Ubiquitin Ligase ◽  
Molecular Mechanisms ◽  
Critical Role ◽  
E3 Ubiquitin Ligase ◽  
Death Receptor ◽  
Cell Immunity ◽  
B Cell Immunity

Humoral immunity involves multiple checkpoints during B-cell development, maturation, and activation. The cell death receptor CD95/Fas-mediated apoptosis plays a critical role in eliminating the unwanted activation of B cells by self-reactive antigens and in maintaining B-cell homeostasis through activation-induced B-cell death (AICD). The molecular mechanisms controlling AICD remain largely undefined. Herein, we show that the E3 ubiquitin ligase Hrd1 protected B cells from activation-induced cell death by degrading the death receptor Fas. Hrd1-null B cells exhibited high Fas expression during activation and rapidly underwent Fas-mediated apoptosis, which could be largely inhibited by FasL neutralization. Fas mutation in Hrd1 KO mice abrogated the increase in B-cell AICD. We identified Hrd1 as the first E3 ubiquitin ligase of the death receptor Fas and Hrd1-mediated Fas destruction as a molecular mechanism in regulating B-cell immunity.

scholarly journals Human neonatal B cell immunity differs from the adult version by conserved Ig repertoires and rapid, but transient response dynamics

2020 ◽  
Author(s):  
Bettina Budeus ◽  
Artur Kibler ◽  
Martina Brauser ◽  
Ekaterina Homp ◽  
Kevin Bronischewski ◽  
...  
Keyword(s):  
T Cell ◽  
B Cells ◽  
Cord Blood ◽  
B Cell ◽  
Transient Response ◽  
Cell System ◽  
Response Dynamics ◽  
Vaccination Strategies ◽  
Cell Immunity ◽  
B Cell Immunity

AbstractThe human infant B cell system is considered premature or impaired. Here we show that most cord blood B cells are mature and functional as seen in adults, albeit with distinct transcriptional programs providing accelerated responsiveness to T cell-independent and T cell-dependent stimulation and facilitated IgA class switching. Stimulation drives extensive differentiation into antibody-secreting cells, thereby presumably limiting memory B cell formation. The neonatal Ig-repertoire is highly variable, but conserved, showing recurrent B cell receptor (BCR) clonotypes frequently shared between neonates. Our study demonstrates that cord blood B cells are not impaired but differ from their adult counterpart in a conserved BCR repertoire and rapid but transient response dynamics. These properties may account for the sensitivity of neonates to infections and limited effectivity of vaccination strategies. Humanized mice suggest that the distinctness of cord blood versus adult B cells is already reflected by the developmental program of hematopoietic precursors, arguing for a layered B-1/B-2 lineage system as in mice. Still, our findings reveal overall limited comparability of human cord blood B cells and murine B-1 cells.Significance StatementNeonates and infants suffer from enhanced susceptibility to infections. Our study contrasts with the current concept of a premature or impaired B cell system in neonates, by showing that most cord blood B cells are mature and functional. However, their responses are rapid but provide only short-term protection, which may help to improve infant vaccination strategies. We propose an altered perspective on the early human B cell system, which looks similar to but functions differently from the adult counterpart. Finally, our analysis indicates that cord blood- and adult B cell development occur layered as in mice, but certain mouse models still may offer a limited view on human neonatal B cell immunity.

Vaccine-elicited CD4 T cells prevent the deletion of antiviral B cells in chronic infection

2021 ◽  
Vol 118 (46) ◽  
pp. e2108157118
Author(s):  
Kerstin Narr ◽  
Yusuf I. Ertuna ◽  
Benedict Fallet ◽  
Karen Cornille ◽  
Mirela Dimitrova ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cells ◽  
B Cell ◽  
Cd4 T Cells ◽  
Memory B Cells ◽  
Type I ◽  
Clonal Deletion ◽  
Cell Immunity ◽  
B Cell Immunity

Chronic viral infections subvert protective B cell immunity. An early type I interferon (IFN-I)–driven bias to short-lived plasmablast differentiation leads to clonal deletion, so-called “decimation,” of antiviral memory B cells. Therefore, prophylactic countermeasures against decimation remain an unmet need. We show that vaccination-induced CD4 T cells prevented the decimation of naïve and memory B cells in chronically lymphocytic choriomeningitis virus (LCMV)-infected mice. Although these B cell responses were largely T independent when IFN-I was blocked, preexisting T help assured their sustainability under conditions of IFN-I–driven inflammation by instructing a germinal center B cell transcriptional program. Prevention of decimation depended on T cell–intrinsic Bcl6 and Tfh progeny formation. Antigen presentation by B cells, interactions with antigen-specific T helper cells, and costimulation by CD40 and ICOS were also required. Importantly, B cell–mediated virus control averted Th1-driven immunopathology in LCMV-challenged animals with preexisting CD4 T cell immunity. Our findings show that vaccination-induced Tfh cells represent a cornerstone of effective B cell immunity to chronic virus challenge, pointing the way toward more effective B cell–based vaccination against persistent viral diseases.

Abstract 174: The BH3-Only Protein BNIP3 Induces Mitochondrial Clearance via Multiple Pathways

2015 ◽  
Vol 117 (suppl_1) ◽  
Author(s):  
Eileen R Gonzalez ◽  
Babette Hammerling ◽  
Rita Hanna ◽  
Dieter A Kubli ◽  
Åsa B Gustafsson
Keyword(s):  
Cell Death ◽  
Ubiquitin Ligase ◽  
Critical Role ◽  
E3 Ubiquitin Ligase ◽  
Wild Type ◽  
Potent Inducer ◽  
Autophagy Pathway ◽  
Endosomal Pathway ◽  
In Cells

Autophagy plays an important role in cellular quality control and is responsible for removing protein aggregates and dysfunctional organelles. BNIP3 is an atypical BH3-only protein which is known to cause mitochondrial dysfunction and cell death in the myocardium. Interestingly, BNIP3 can also protect against cell death by promoting removal of dysfunctional mitochondria via autophagy (mitophagy). We have previously reported that BNIP3 is a potent inducer of mitophagy in cardiac myocytes and that BNIP3 contains an LC3 Interacting Region (LIR) that binds to LC3 on the autophagosome, tethering the mitochondrion to the autophagosome for engulfment. However, the molecular mechanism(s) underlying BNIP3-mediated mitophagy are still unclear. In this study, we discovered that BNIP3 can mediate mitochondrial clearance in cells even in the absence of a functional autophagy pathway. We found that overexpression of BNIP3 led to significant clearance of mitochondria in both wild type (WT) and autophagy deficient Atg5-/- MEFs. BNIP3 caused an increase in LC3II levels in WT MEFs, indicating increased formation of autophagosomes. In contrast, LC3II was undetectable in Atg5-/- MEFs. Furthermore, we found that BNIP3-mediated clearance in WT and Atg5-/- MEFs did not require the presence of Parkin, an E3 ubiquitin ligase which plays a critical role in clearing dysfunctional mitochondria in cells. Also, overexpression of Parkin did not enhance BNIP3-mediated mitochondrial clearance. When investigating activation of alternative cellular degradation pathways, we found that BNIP3 induced activation of the endosomal-lysosomal pathway in both WT and Atg5-/- MEFs. Mutating the LC3 binding site in BNIP3 did not interfere with the activation of the endosomal pathway and clearance of mitochondria in Atg5-/- MEFs. Thus, these findings suggest that BNIP3 can promote clearance of mitochondria via multiple pathways in cells. The role of autophagy in removing mitochondria is already well established and we are currently exploring the roles of the endosomal and alternative autophagy pathways in BNIP3-mediated mitochondrial clearance in myocytes.

scholarly journals The E3 ubiquitin ligase Itch restricts antigen-driven B cell responses

2019 ◽  
Vol 216 (9) ◽  
pp. 2170-2183 ◽  
Author(s):  
Emily K. Moser ◽  
Jennifer Roof ◽  
Joseph M. Dybas ◽  
Lynn A. Spruce ◽  
Steven H. Seeholzer ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Cell Fate ◽  
Ubiquitin Ligase ◽  
Cell Activity ◽  
E3 Ubiquitin Ligase ◽  
Normal Numbers ◽  
Cell Responses ◽  
B Cell Responses

The E3 ubiquitin ligase Itch regulates antibody levels and prevents autoimmune disease in humans and mice, yet how Itch regulates B cell fate or function is unknown. We now show that Itch directly limits B cell activity. While Itch-deficient mice displayed normal numbers of preimmune B cell populations, they showed elevated numbers of antigen-experienced B cells. Mixed bone marrow chimeras revealed that Itch acts within B cells to limit naive and, to a greater extent, germinal center (GC) B cell numbers. B cells lacking Itch exhibited increased proliferation, glycolytic capacity, and mTORC1 activation. Moreover, stimulation of these cells in vivo by WT T cells resulted in elevated numbers of GC B cells, PCs, and serum IgG. These results support a novel role for Itch in limiting B cell metabolism and proliferation to suppress antigen-driven B cell responses.

scholarly journals Latency-Associated Nuclear Antigen E3 Ubiquitin Ligase Activity Impacts Gammaherpesvirus-Driven Germinal Center B Cell Proliferation

2016 ◽  
Vol 90 (17) ◽  
pp. 7667-7683 ◽  
Author(s):  
Sofia A. Cerqueira ◽  
Min Tan ◽  
Shijun Li ◽  
Franceline Juillard ◽  
Colin E. McVey ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Ubiquitin Ligase ◽  
Germinal Center ◽  
E3 Ubiquitin Ligase ◽  
E3 Ligase ◽  
Nuclear Antigen ◽  
Ubiquitin Ligase Activity ◽  
Socs Box

ABSTRACTViruses have evolved mechanisms to hijack components of cellular E3 ubiquitin ligases, thus modulating the ubiquitination pathway. However, the biological relevance of such mechanisms for viral pathogenesisin vivoremains largely unknown. Here, we utilized murid herpesvirus 4 (MuHV-4) infection of mice as a model system to address the role of MuHV-4 latency-associated nuclear antigen (mLANA) E3 ligase activity in gammaherpesvirus latent infection. We show that specific mutations in the mLANA SOCS box (V199A, V199A/L202A, or P203A/P206A) disrupted mLANA's ability to recruit Elongin C and Cullin 5, thereby impairing the formation of the Elongin BC/Cullin 5/SOCS (EC5SmLANA) complex and mLANA's E3 ligase activity on host NF-κB and Myc. Although these mutations resulted in considerably reduced mLANA binding to viral terminal repeat DNA as assessed by electrophoretic mobility shift assay (EMSA), the mutations did not disrupt mLANA's ability to mediate episome persistence.In vivo, MuHV-4 recombinant viruses bearing these mLANA SOCS box mutations exhibited a deficit in latency amplification in germinal center (GC) B cells. These findings demonstrate that the E3 ligase activity of mLANA contributes to gammaherpesvirus-driven GC B cell proliferation. Hence, pharmacological inhibition of viral E3 ligase activity through targeting SOCS box motifs is a putative strategy to control gammaherpesvirus-driven lymphoproliferation and associated disease.IMPORTANCEThe gammaherpesviruses Epstein-Barr virus (EBV) and Kaposi's sarcoma-associated herpesvirus (KSHV) cause lifelong persistent infection and play causative roles in several human malignancies. Colonization of B cells is crucial for virus persistence, and access to the B cell compartment is gained by virus-driven proliferation in germinal center (GC) B cells. Infection of B cells is predominantly latent, with the viral genome persisting as a multicopy episome and expressing only a small subset of viral genes. Here, we focused on latency-associated nuclear antigen (mLANA) encoded by murid herpesvirus-4 (MuHV-4), which exhibits homology in sequence, structure, and function to KSHV LANA (kLANA), thereby allowing the study of LANA-mediated pathogenesis in mice. Our experiments show that mLANA's E3 ubiquitin ligase activity is necessary for efficient expansion of latency in GC B cells, suggesting that the development of pharmacological inhibitors of LANA E3 ubiquitin ligase activity may allow strategies to interfere with gammaherpesvirus-driven lymphoproliferation and associated disease.

scholarly journals Transcription factors IRF8 and PU.1 are required for follicular B cell development and BCL6-driven germinal center responses

2019 ◽  
Vol 116 (19) ◽  
pp. 9511-9520 ◽  
Author(s):  
Hongsheng Wang ◽  
Shweta Jain ◽  
Peng Li ◽  
Jian-Xin Lin ◽  
Jangsuk Oh ◽  
...  
Keyword(s):  
Transcription Factors ◽  
B Cells ◽  
B Cell ◽  
Germinal Center ◽  
Molecular Mechanisms ◽  
Target Genes ◽  
Critical Role ◽  
B Cell Lymphoma ◽  
Cell Development ◽  
Affinity Maturation

The IRF and Ets families of transcription factors regulate the expression of a range of genes involved in immune cell development and function. However, the understanding of the molecular mechanisms of each family member has been limited due to their redundancy and broad effects on multiple lineages of cells. Here, we report that double deletion of floxed Irf8 and Spi1 (encoding PU.1) by Mb1-Cre (designated DKO mice) in the B cell lineage resulted in severe defects in the development of follicular and germinal center (GC) B cells. Class-switch recombination and antibody affinity maturation were also compromised in DKO mice. RNA-seq (sequencing) and ChIP-seq analyses revealed distinct IRF8 and PU.1 target genes in follicular and activated B cells. DKO B cells had diminished expression of target genes vital for maintaining follicular B cell identity and GC development. Moreover, our findings reveal that expression of B-cell lymphoma protein 6 (BCL6), which is critical for development of germinal center B cells, is dependent on IRF8 and PU.1 in vivo, providing a mechanism for the critical role for IRF8 and PU.1 in the development of GC B cells.

scholarly journals The E3 Ubiquitin Ligase ATL9 Affects Expression of Defense Related Genes, Cell Death and Callose Deposition in Response to Fungal Infection

Pathogens ◽  
2022 ◽  
Vol 11 (1) ◽  
pp. 68
Author(s):  
Tingwei Guo ◽  
Feng Kong ◽  
Carter Burton ◽  
Steven Scaglione ◽  
Blake Beagles ◽  
...  
Keyword(s):  
Cell Death ◽  
Fungal Infection ◽  
Ubiquitin Ligase ◽  
Critical Role ◽  
E3 Ubiquitin Ligase ◽  
Defense Responses ◽  
Callose Deposition ◽  
Expression Levels ◽  
Insertional Mutants ◽  
Protein Ubiquitination

Plants use diverse strategies to defend themselves from biotic stresses in nature, which include the activation of defense gene expression and a variety of signal transduction pathways. Previous studies have shown that protein ubiquitination plays a critical role in plant defense responses, however the details of its function remain unclear. Our previous work has shown that increasing expression levels of ATL9, an E3 ubiquitin ligase in Arabidopsis thaliana, increased resistance to infection by the fungal pathogen, Golovinomyces cichoracearum. In this study, we demonstrate that the defense-related proteins PDF1.2, PCC1 and FBS1 directly interact with ATL9 and are targeted for degradation to the proteasome by ATL9. The expression levels of PDF1.2, PCC1 and FBS1 are decreased in T-DNA insertional mutants of atl9 and T-DNA insertional mutants of pdf1.2, pcc1 and fbs1 are more susceptible to fungal infection. In addition, callose is more heavily deposited at infection sites in the mutants of atl9, fbs1, pcc1 and pdf1.2. Overexpression of ATL9 and of mutants in fbs1, pcc1 and pdf1.2 showed increased levels of cell death during infection. Together these results indicate that ubiquitination, cell death and callose deposition may work together to enhance defense responses to fungal pathogens.

scholarly journals The E3 ubiquitin ligase Mule acts through the ATM–p53 axis to maintain B lymphocyte homeostasis

2012 ◽  
Vol 209 (1) ◽  
pp. 173-186 ◽  
Author(s):  
Zhenyue Hao ◽  
Gordon S. Duncan ◽  
Yu-Wen Su ◽  
Wanda Y. Li ◽  
Jennifer Silvester ◽  
...  
Keyword(s):  
Steady State ◽  
B Cells ◽  
B Cell ◽  
Ubiquitin Ligase ◽  
B Lymphocyte ◽  
E3 Ubiquitin Ligase ◽  
Genotoxic Stress ◽  
Cell Development ◽  
B Cell Development ◽  
Lymphocyte Homeostasis

Cellular homeostasis is controlled by pathways that balance cell death with survival. Mcl-1 ubiquitin ligase E3 (Mule) is an E3 ubiquitin ligase that targets the proapoptotic molecule p53 for polyubiquitination and degradation. To elucidate the role of Mule in B lymphocyte homeostasis, B cell–specific Mule knockout (BMKO) mice were generated using the Cre–LoxP recombination system. Analysis of BMKO mice showed that Mule was essential for B cell development, proliferation, homeostasis, and humoral immune responses. p53 transactivation was increased by two- to fourfold in Mule-deficient B cells at steady state. Genetic ablation of p53 in BMKO mice restored B cell development, proliferation, and homeostasis. p53 protein was increased in resting Mule-deficient mouse embryonic fibroblasts (MEFs) and embryonic stem (ES) cells. Loss of Mule in both MEFs and B cells at steady state resulted in increased levels of phospho–ataxia telangiectasia mutated (ATM) and the ATM substrate p53. Under genotoxic stress, BMKO B cells were resistant to apoptosis, and control MEFs exhibited evidence of a physical interaction between Mule and phospho-ATM. Phospho-ATM, phospho-p53, and Brca1 levels were reduced in Mule-deficient B cells and MEFs subjected to genotoxic stress. Thus, Mule regulates the ATM–p53 axis to maintain B cell homeostasis under both steady-state and stress conditions.

scholarly journals Regulatory T Cell Adoptive Immunotherapy Promotes B Cell Immunity after Haploidentical Transplantation

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 1917-1917
Author(s):  
Sara Piccinelli ◽  
Antonella Mancusi ◽  
Eni Hoxha ◽  
Loredana Ruggeri ◽  
Franca Falzetti ◽  
...  
Keyword(s):  
T Cell ◽  
B Cells ◽  
Mouse Model ◽  
B Cell ◽  
Adoptive Transfer ◽  
Post Transplant ◽  
Cell Immunity ◽  
Human Hscs ◽  
Cmv Reactivation ◽  
B Cell Immunity

Background: Adoptive transfer of CD4+CD25+FOXP3+ regulatory T cells (Tregs) effectively prevents conventional T cell (Tcons) mediated Graft versus Host Disease (GvHD) while it does not impair graft-versus-leukemia effect in haploidentical hematopoietic cell transplantation (haplo-HCT). Moreover Treg immunotherapy promotes fast donor T cell recovery after transplant. Recent studies showed that mouse bone marrow (BM) Tregs localize in the hematopoietic stem cell (HSC) niche, where they contribute to HSCs maintenance and promote donor engraftment and B cell lymphopoiesis. We are investigating if human Tregs promote B cell reconstitution and immunity in preclinical models and in haplo-HCT patients. Methods: Human sample analysis: B cell reconstitution was analysed monthly by FACS in BM and peripheral blood (PB) samples from 66 patients who underwent either Treg/Tcon haplo-HCT (45 patients), or T-cell depleted haplo-HCT (8 patients) or haplo-HCT with post-transplant cyclophosphamide (PTCy, 13 patients). Diagnosis was acute leukemia in 52 patients, lymphoma in 11 and multiple myeloma in 3. PB total immunoglobulin (Ig), anti-Cytomegalovirus (CMV) IgM and CMV viremia were monitored. Humanized mouse model: donor derived human Tregs and purified CD34+ HSCs were co-infused in sublethally irradiated (2 Gy) immune-deficient NSG mice. Donor engraftment and B cell reconstitution were analysed by FACS and histology. Results: B cell reconstitution was faster after Treg/Tcon haplo-HCT when compared to other haplo-HCT protocols. B cell counts were higher in PB of patients that received Treg/Tcon haplo-HCT (p<.05) and were comparable to those of healthy subjects by 4 months after transplant (117±148 cells/mm, Fig.1A). We could detect early frequencies of CD34+CD38+CD10+CD127+ common lymphoid progenitors, CD45+CD10+CD38+CD19- Pre/Pro-B, CD45+CD10+CD38+CD19+ Pre-B, and Pro-B cells in the BM of these patients, that resulted in an increased production of CD38+CD19+CD5-IgM+ immature B cells, CD38+CD19+CD5+IgM+ transitional B cells and CD19+CD20+ mature B cells. We used a mouse model of xenotransplantation to understand whether donor B cell reconstitution in Treg/Tcon haplo-HCT is boosted by a Treg-mediated effect on donor human HSCs. We found that infusion of human Tregs facilitated donor HSC engraftment. BM and PB human chimerism was increased (p<.05) in mice that received Treg infusion. BM histology revealed the presence of in vivo expanded human CXCR4+ Tregs in the femurs of Treg receiving mice 1 month after transplant. In the same samples we observed a lower number of CD34+ HSCs, suggesting BM Tregs facilitate donor HSC differentiation. Moreover, Treg treatment allowed human HSCs to localize preferentially in the epiphyseal areas of the femurs, where donor cells started to engraft (p<.04). When looking at donor B cell reconstitution, we found HSC-derived mature B cells rapidly abundant and easily detectable starting 30 days after HSC infusion in PB of Treg-treated animals (p<.05). To further assess whether HSC-derived B cells were functional in patients that received Treg/Tcon haplo-HCT, we analysed total Ig production and specific responses to CMV reactivation. Post-transplant hypogammaglobulinemia was rapidly corrected in Treg/Tcon haplo-HCT patients. Total IgM were higher compared to other haplo-HCT protocols and reached normal levels by 3 months after transplant (96±155 mg/dL, Fig.1B). CMV reactivation rate was lower (47% vs 79%) and it occurred later after Treg/Tcon immunotherapy (51±19 vs 40±48 days). New production of anti-CMV specific IgM was documented in 43% of CMV seropositive patients 98±48 days after Treg/Tcon haplo-HCT, while anti-CMV specific IgM were almost undetectable after other haplo-HCT protocols within the first 6 months after transplant (Fig.1C). Such potent B cell responses coupled with T cell reconstitution resulted in a reduction of CMV second reactivations (Fig.1D p<.05) with no patient that died with CMV disease. Conclusions: Adoptive transfer of human Tregs boosts donor HSC engraftment and facilitates the reconstitution of functional HSC-derived donor B cells. Such results suggest that Treg/Tcon haplo-HCT patients could be early vaccinated after transplant. Treg/Tcon immunotherapy promotes control of infections and B cell immunity in patients undergoing haplo-HCT. Disclosures No relevant conflicts of interest to declare.

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