RESUMO
B cells predominate in a quiescent state until an antigen is encountered, which results in rapid growth, proliferation and differentiation of the B cells. These distinct cell states are probably accompanied by differing metabolic needs, yet little is known about the metabolic control of B cell fate. Here we show that glycogen synthase kinase 3 (Gsk3) is a metabolic sensor that promotes the survival of naive recirculating B cells by restricting cell mass accumulation. In antigen-driven responses, Gsk3 was selectively required for regulation of B cell size, mitochondrial biogenesis, glycolysis and production of reactive oxygen species (ROS), in a manner mediated by the co-stimulatory receptor CD40. Gsk3 was required to prevent metabolic collapse and ROS-induced apoptosis after glucose became limiting, functioning in part by repressing growth dependent on the myelocytomatosis oncoprotein c-Myc. Notably, we found that Gsk3 was required for the generation and maintenance of germinal center B cells, which require high glycolytic activity to support growth and proliferation in a hypoxic microenvironment.
Assuntos
Linfócitos B/fisiologia , Centro Germinativo/imunologia , Glicogênio Sintase Quinase 3 beta/metabolismo , Animais , Antígenos CD19/genética , Antígenos CD19/metabolismo , Apoptose/genética , Ligante de CD40/metabolismo , Diferenciação Celular , Proliferação de Células , Células Cultivadas , Glicogênio Sintase Quinase 3 beta/genética , Glicólise , Interleucina-4/metabolismo , Camundongos , Camundongos Knockout , Estresse Oxidativo , Espécies Reativas de Oxigênio/metabolismo , Transdução de SinaisRESUMO
Despite the essential role of plasma cells in health and disease, the cellular mechanisms controlling their survival and secretory capacity are still poorly understood. Here, we identified the soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) Sec22b as a unique and critical regulator of plasma cell maintenance and function. In the absence of Sec22b, plasma cells were hardly detectable and serum antibody titers were dramatically reduced. Accordingly, Sec22b-deficient mice fail to mount a protective immune response. At the mechanistic level, we demonstrated that Sec22b contributes to efficient antibody secretion and is a central regulator of plasma cell maintenance through the regulation of their transcriptional identity and of the morphology of the endoplasmic reticulum and mitochondria. Altogether, our results unveil an essential and nonredundant role for Sec22b as a regulator of plasma cell fitness and of the humoral immune response.
Assuntos
Plasmócitos , Proteínas SNARE , Camundongos , Animais , Plasmócitos/metabolismo , Proteínas R-SNARE/metabolismo , Proteínas SNARE/metabolismo , Retículo Endoplasmático/metabolismo , Transporte BiológicoRESUMO
CD20 is a B cell-specific membrane protein and represents an attractive target for therapeutic antibodies. Despite widespread usage of anti-CD20 antibodies for B cell depletion therapies, the biological function of their target remains unclear. Here, we demonstrate that CD20 controls the nanoscale organization of receptors on the surface of resting B lymphocytes. CRISPR/Cas9-mediated ablation of CD20 in resting B cells resulted in relocalization and interaction of the IgM-class B cell antigen receptor with the coreceptor CD19. This receptor rearrangement led to a transient activation of B cells, accompanied by the internalization of many B cell surface marker proteins. Reexpression of CD20 restored the expression of the B cell surface proteins and the resting state of Ramos B cells. Similarly, treatment of Ramos or naive human B cells with the anti-CD20 antibody rituximab induced nanoscale receptor rearrangements and transient B cell activation in vitro and in vivo. A departure from the resting B cell state followed by the loss of B cell identity of CD20-deficient Ramos B cells was accompanied by a PAX5 to BLIMP-1 transcriptional switch, metabolic reprogramming toward oxidative phosphorylation, and a shift toward plasma cell development. Thus, anti-CD20 engagement or the loss of CD20 disrupts membrane organization, profoundly altering the fate of human B cells.
Assuntos
Antígenos CD20/metabolismo , Linfócitos B/imunologia , Antígenos CD19/metabolismo , Linhagem Celular Tumoral , Membrana Celular/metabolismo , Células Cultivadas , Humanos , Ativação Linfocitária , Receptores de Antígenos de Linfócitos B/metabolismoRESUMO
In response to mitogenic stimulation, B cells activate different pro-anabolic signaling pathways such as c-Myc- and mTORC1-dependent networks to satisfy the energetic demands of biomass synthesis and proliferation. In order to preserve viability and function, cell growth cannot progress unchecked and must be adjusted according to the availability of nutrients. Nutrient-sensing proteins such as AMPK antagonize mTORC1 activity in response to starvation. If pro-anabolic signaling pathways are aberrantly activated, B cells may lack the metabolic capacity to accommodate their energetic needs, which can lead to cell death. On the other hand, metabolic hyperactivation is a salient feature of cancer cells, suggesting that mechanisms exist, which allow B cells to cope with metabolic stress. The aim of this review is to discuss how B cells respond to a mismatch between energy supply and demand and what the consequences are of metabolic dysregulation in normal and malignant B cells.
Assuntos
Linfócitos B/imunologia , Linfócitos B/metabolismo , Transformação Celular Neoplásica , Metabolismo Energético , Proteínas de Checkpoint Imunológico/metabolismo , Ativação Linfocitária/imunologia , Animais , Linfócitos B/citologia , Biomarcadores , Diferenciação Celular/imunologia , Sobrevivência Celular , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/imunologia , Transformação Celular Neoplásica/metabolismo , Senescência Celular/genética , Senescência Celular/imunologia , Centro Germinativo/imunologia , Centro Germinativo/metabolismo , Homeostase , Humanos , Proteínas de Checkpoint Imunológico/genética , Memória Imunológica , Leucemia de Células B/etiologia , Leucemia de Células B/metabolismo , Leucemia de Células B/patologia , Ativação Linfocitária/genética , Plasmócitos/imunologia , Plasmócitos/metabolismo , Células Precursoras de Linfócitos B/citologia , Células Precursoras de Linfócitos B/metabolismoRESUMO
In the original published version of the article, the red squares in the figures which indicated the corrections.
RESUMO
As an important chemokine receptor, the role of CX3CR1 has been studied extensively on the migration of lymphocytes including T and B cells. Although CX3CR1+ B cells have immune suppressor properties, little is known about its role on the regulation of BCR signaling and B cell differentiation as well as the underlying molecular mechanism. We have used CX3CR1 KO mice to study the effect of CX3CR1 deficiency on BCR signaling and B cell differentiation. Interestingly, we found that proximal BCR signaling, such as the activation of CD19, BTK and SHIP was reduced in CX3CR1 KO B cells upon antigenic stimulation. However, the activation of mTORC signaling was enhanced. Mechanistically, we found that the reduced BCR signaling in CX3CR1 KO B cells was due to reduced BCR clustering, which is caused by the enhanced actin accumulation by the plasma membrane via increased activation of WASP. This caused an increased differentiation of MZ B cells in CX3CR1 KO mice and an enhanced generation of plasma cells (PC) and antibodies. Our study shows that CX3CR1 regulates BCR signaling via actin remodeling and affects B cell differentiation and the humoral immune response.
Assuntos
Actinas/metabolismo , Linfócitos B/citologia , Receptor 1 de Quimiocina CX3C/metabolismo , Receptores de Antígenos de Linfócitos B/metabolismo , Animais , Linfócitos B/metabolismo , Receptor 1 de Quimiocina CX3C/genética , Diferenciação Celular , Camundongos Endogâmicos C57BL , Camundongos Knockout , Transdução de SinaisRESUMO
B cells contribute critically to an effective immune response by producing Ag-specific Abs. During the immune response to so-called "thymus-dependent Ags," activated B cells seek T cell help and form germinal centers. In contrast, thymus-independent Ags generally do not induce germinal center formation. In the germinal center, B cells undergo somatic hypermutation, affinity-based clonal expansion, and differentiation to produce plasma cells and memory B cells. Valuable insight into these processes has been gained by using model hapten-carrier complexes or SRBCs. SRBCs induce robust germinal center formation in mice. Therefore, this Ag is commonly used to study germinal center responses. In contrast to haptenated Ags, thus far it has been difficult to measure the titer of Ag-specific Abs or the expansion of Ag-specific B cells after immunization with SRBCs. We have developed new, simple methods to access these parameters, thus providing new tools to study germinal center and Ab responses.
Assuntos
Linfócitos B/fisiologia , Transfusão de Eritrócitos/métodos , Eritrócitos/imunologia , Centro Germinativo/imunologia , Imunidade Humoral , Técnicas Imunológicas/métodos , Linfócitos T Auxiliares-Indutores/imunologia , Animais , Bovinos , Diferenciação Celular , Células Cultivadas , Feminino , Imunização , Memória Imunológica , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Hipermutação Somática de ImunoglobulinaRESUMO
B cell activation is accompanied by metabolic adaptations to meet the increased energetic demands of proliferation. The metabolic composition of the microenvironment is known to change during a germinal center response, in inflamed tissue and to vary significantly between different organs. To sustain cellular homeostasis B cells need to be able to dynamically adapt to changes in their environment. An inability to take up and process available nutrients can result in impaired B cell growth and a diminished humoral immune response. Furthermore, the metabolic microenvironment can affect B cell signaling and provide a means to avoid aberrant proliferation or modulate B cell function. Thus, a better understanding of the intricate interplay between cell signaling and metabolism could provide novel insight into how B cell function is regulated and have implications for the development of vaccines or treatment of autoimmune disorders and B cell derived malignancies.
Assuntos
Linfócitos B/fisiologia , Proliferação de Células , Homeostase , Transdução de Sinais , Estresse Fisiológico , AnimaisRESUMO
B cell growth and proliferation is tightly regulated by signaling through the B cell receptor and by other membrane bound receptors responding to different cytokines. The PI3K signaling pathway has been shown to play a crucial role in B cell activation, differentiation and survival. Activated B cells undergo metabolic reprograming in response to changing energetic and biosynthetic demands. B cells also need to be able to coordinate metabolic activity and proliferation with nutrient availability. The PI3K signaling network has been implicated in regulating nutrient acquisition, utilization and biosynthesis, thus integrating receptor-mediated signaling with cell metabolism. In this review, we discuss the current knowledge about metabolic changes induced in activated B cells, strategies to adapt to metabolic stress and the role of PI3K signaling in these processes.
Assuntos
Linfócitos B/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Transdução de Sinais , Animais , Autofagia , Linfócitos B/citologia , Diferenciação Celular , Proliferação de Células , Humanos , Redes e Vias Metabólicas , Mitocôndrias/metabolismo , Neoplasias/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Serina-Treonina Quinases TOR/metabolismoRESUMO
Survivin is a member of the inhibitor of apoptosis family of proteins and a biomarker of poor prognosis in aggressive B cell non-Hodgkin's lymphoma. In addition to its role in inhibition of apoptosis, survivin also regulates mitosis. In this article, we show that deletion of survivin during early B cell development results in a complete block at the cycling pre-B stage. In the periphery, B cell homeostasis is not affected, but survivin-deficient B cells are unable to mount humoral responses. Correspondingly, we show that survivin is required for cell division in response to mitogenic stimulation. Thus, survivin is essential for proliferation of B cell progenitors and activated mature B cells, but is dispensable for B cell survival. Moreover, a small-molecule inhibitor of survivin strongly impaired the growth of representative B lymphoma lines in vitro, supporting the validity of survivin as an attractive therapeutic target for high-grade B cell non-Hodgkin's lymphoma.
Assuntos
Linfócitos B/metabolismo , Proteínas Inibidoras de Apoptose/genética , Células Precursoras de Linfócitos B/metabolismo , Proteínas Repressoras/genética , Alelos , Animais , Formação de Anticorpos/genética , Formação de Anticorpos/imunologia , Apoptose/genética , Linfócitos B/citologia , Linfócitos B/efeitos dos fármacos , Biomarcadores , Diferenciação Celular/genética , Sobrevivência Celular/genética , Dano ao DNA , Expressão Gênica , Genótipo , Imunidade Humoral/genética , Imunidade Humoral/imunologia , Imunofenotipagem , Proteínas Inibidoras de Apoptose/antagonistas & inibidores , Proteínas Inibidoras de Apoptose/deficiência , Ativação Linfocitária/genética , Camundongos , Camundongos Transgênicos , Células Precursoras de Linfócitos B/citologia , Células Precursoras de Linfócitos B/efeitos dos fármacos , Proteínas Repressoras/antagonistas & inibidores , Proteínas Repressoras/deficiência , SurvivinaRESUMO
The mammalian target of rapamycin (mTOR), an essential serine/threonine kinase, functions in biochemically distinct multiprotein complexes, but little is known about roles of the complexes in B cells. The acutely rapamycin-sensitive mTOR complex 1 (mTORC1) is defined by a core subunit Raptor, whereas mTORC2 lacks Raptor and, instead, has Rictor and SIN1 as distinct essential components. We now show that homeostasis and function of B cells require Rictor. Conditional deletion of Rictor before lymphoid specification impaired generation of mature follicular, marginal zone, and B1a B lymphocytes. Induced inactivation in adult mice caused cell-autonomous defects in B lymphoid homeostasis and antibody responses in vivo, along with affecting plasma cells in bone marrow. Survival of B lymphocytes depended on Rictor, which was vital for normal induction of prosurvival genes, suppression of proapoptotic genes, nuclear factor κB induction after B-cell receptor stimulation, and B-cell activating factor-induced nuclear factor κB2/p52 generation. Collectively, the findings provide evidence that mTOR signaling affects survival and proliferation of mature B lymphocytes, and establish Rictor as an important signal relay in B-cell homeostasis, fate, and functions.
Assuntos
Linfócitos B/citologia , Linfócitos B/imunologia , Proteínas de Transporte/imunologia , Diferenciação Celular/imunologia , Homeostase/imunologia , Transferência Adotiva , Animais , Western Blotting , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Proliferação de Células , Sobrevivência Celular/imunologia , Ensaio de Desvio de Mobilidade Eletroforética , Ensaio de Imunoadsorção Enzimática , Citometria de Fluxo , Imuno-Histoquímica , Camundongos , Camundongos Endogâmicos C57BL , Proteína Companheira de mTOR Insensível à Rapamicina , Transdução de Sinais/imunologia , Serina-Treonina Quinases TOR/genética , Serina-Treonina Quinases TOR/imunologia , Serina-Treonina Quinases TOR/metabolismoRESUMO
In the secondary lymphoid organs, intimate contact with follicular dendritic cells (FDCs) is required for B cell retention and Ag-driven selection during the germinal center response. However, selection of self-reactive B cells by Ag on FDCs has not been addressed. To this end, we generated a mouse model to conditionally express a membrane-bound self-antigen on FDCs and to monitor the fate of developing self-reactive B cells. In this article, we show that self-antigen displayed on FDCs mediates effective elimination of self-reactive B cells at the transitional stage. Notwithstanding, some self-reactive B cells persist beyond this checkpoint, showing evidence of Ag experience and intact proximal BCR signaling, but they are short-lived and unable to elicit T cell help. These results implicate FDCs as an important component of peripheral B cell tolerance that prevents the emergence of naive B cells capable of responding to sequestered self-antigens.
Assuntos
Autoantígenos/imunologia , Autoimunidade , Linfócitos B/imunologia , Células Dendríticas Foliculares/imunologia , Transferência Adotiva , Animais , Complexo Antígeno-Anticorpo/imunologia , Apoptose/imunologia , Proliferação de Células , Células Cultivadas , Ativação Linfocitária , Camundongos , Camundongos Transgênicos , Receptores de Antígenos de Linfócitos B/metabolismo , Transdução de Sinais/imunologiaRESUMO
Members of the tumor necrosis factor receptor superfamily (TNFRSF) participate prominently in B-cell maturation and function. In particular, B-cell activating factor belonging to the TNF family receptor (BAFF-R), B-cell maturation antigen (BCMA), and transmembrane activator and calcium modulator and cyclophilin ligand interactor (TACI) play critical roles in promoting B-cell survival at distinct stages of development by engaging a proliferation-inducing ligand (APRIL) and/or BAFF. CD40 is also essential for directing the humoral response to T-cell-dependent antigens. Signaling by the TNFRSF is mediated primarily, albeit not exclusively, via the TNFR-associated factor (TRAF) proteins and activation of the canonical and/or non-canonical nuclear factor-κB (NF-κB) pathways. Dysregulated signaling by TNFRSF members can promote B-cell survival and proliferation, causing autoimmunity and neoplasia. In this review, we present a current understanding of the functions of and distinctions between APRIL/BAFF signaling by their respective receptors expressed on particular B-cell subsets. These findings are compared and contrasted with CD40 signaling, which employs similar signaling conduits to achieve distinct cellular outcomes in the context of the germinal center response. We also underscore how new findings and conceptual insights into TNFRSF signaling are facilitating the understanding of B-cell malignancies and autoimmune diseases.
Assuntos
Fator Ativador de Células B/imunologia , Linfócitos B/imunologia , Antígenos CD40/imunologia , Imunidade Inata , NF-kappa B/imunologia , Transdução de Sinais/imunologia , Membro 13 da Superfamília de Ligantes de Fatores de Necrose Tumoral/imunologia , Animais , Doenças Autoimunes , Fator Ativador de Células B/genética , Fator Ativador de Células B/metabolismo , Receptor do Fator Ativador de Células B/genética , Receptor do Fator Ativador de Células B/imunologia , Receptor do Fator Ativador de Células B/metabolismo , Antígeno de Maturação de Linfócitos B/genética , Antígeno de Maturação de Linfócitos B/imunologia , Antígeno de Maturação de Linfócitos B/metabolismo , Subpopulações de Linfócitos B/imunologia , Subpopulações de Linfócitos B/metabolismo , Linfócitos B/metabolismo , Antígenos CD40/genética , Antígenos CD40/metabolismo , Proliferação de Células , Sobrevivência Celular/imunologia , Expressão Gênica , Humanos , Camundongos , Camundongos Transgênicos , NF-kappa B/genética , NF-kappa B/metabolismo , Neoplasias/imunologia , Proteína Transmembrana Ativadora e Interagente do CAML/genética , Proteína Transmembrana Ativadora e Interagente do CAML/imunologia , Proteína Transmembrana Ativadora e Interagente do CAML/metabolismo , Membro 13 da Superfamília de Ligantes de Fatores de Necrose Tumoral/genética , Membro 13 da Superfamília de Ligantes de Fatores de Necrose Tumoral/metabolismoRESUMO
Jun activation domain-binding protein 1 (JAB1) regulates ubiquitin-dependent protein degradation by deneddylation of cullin-based ubiquitin ligases and, therefore, plays a central role in regulating proliferation and apoptosis. Because these processes are decisive for B cell development, we investigated JAB1 functions in B cells by establishing a mouse strain with a B cell-specific JAB1 deletion. We show that JAB1 is essential for early B cell development, because the ablation of JAB1 expression blocks B cell development between the pro-B and pre-B cell stages. Furthermore, JAB1 deletion leads to aberrant expression of the apoptosis-triggering protein Fas ligand in pro-B cells. Concomitant B cell-specific overexpression of the antiapoptotic protein Bcl2 partially reverses the block in B cell development; rescued JAB1-deficient B cells reach the periphery and produce protective class-switched Abs after Borrelia burgdorferi infection. Interestingly, B cell-rescued mice exhibit no germinal centers but a striking extrafollicular plasma cell accumulation. In addition, JAB1 is essential for Bcl6 expression, a transcriptional repressor required for germinal center formation. These findings identify JAB1 as an important factor in checkpoint control during early B cell development, as well as in fate decisions in mature Ag-primed B cells.
Assuntos
Linfócitos B/citologia , Diferenciação Celular/imunologia , Proteínas de Ligação a DNA/biossíntese , Proteína Ligante Fas/biossíntese , Centro Germinativo/imunologia , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Peptídeo Hidrolases/metabolismo , Animais , Linfócitos B/imunologia , Complexo do Signalossomo COP9 , Separação Celular , Proteínas de Ligação a DNA/imunologia , Ensaio de Imunoadsorção Enzimática , Proteína Ligante Fas/imunologia , Citometria de Fluxo , Centro Germinativo/citologia , Immunoblotting , Peptídeos e Proteínas de Sinalização Intracelular/imunologia , Ativação Linfocitária/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Peptídeo Hidrolases/imunologia , Células Precursoras de Linfócitos B/imunologia , Células Precursoras de Linfócitos B/metabolismo , Proteínas Proto-Oncogênicas c-bcl-6 , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
The therapeutic humanized monoclonal antibody IgG1 known as Herceptin® has shown remarkable antitumor effects. Although this type of therapy has increased the cancer-free survival of patients, not all tumors respond to this treatment and cancers often develop resistance to the antibody. Despite the fact that Herceptin function has been extensively studied, the precise mechanism underlying its antitumor activity still remains incompletely defined. We previously demonstrated on human breast MCF-7 carcinoma and T-lymphoblastoid CEM cells that monoclonal antibody in combination with Lipoplex consisting of Lipofectamine mixed with plasmid DNA showed a more profound effect on cancer cell viability than antibody alone. The analyses of N-glycans isolated from cancer cells showed dramatic differences in profiles when cells were exposed to Herceptin. Moreover, the investigation of glycosylated peptides from the same cancer cell models after treatment revealed further alterations in the post-translational modifications. Tandem mass spectra obtained from the samples treated confirmed the presence of a series of glycopeptides bearing characteristic oligosaccharides as described in IgG1. However some of them differed by mass differences that corresponded to peptide backbones not described previously and more of them were detected from Herceptin treated samples than from cells transfected with Heceptin/Lipoplex. The results indicate that the presence of Lipoplex prevents antibody transformation and elongates its proper function. The better understanding of the multipart changes described in the glycoconjugates could provide new insights into the mechanism by which antibody induces regression in cancers.
Assuntos
Anticorpos Monoclonais Humanizados , Antineoplásicos , Neoplasias da Mama/metabolismo , Glicômica/métodos , Glicopeptídeos , Leucemia-Linfoma Linfoblástico de Células Precursoras/metabolismo , Proteômica/métodos , Anticorpos Monoclonais Humanizados/química , Anticorpos Monoclonais Humanizados/farmacologia , Anticorpos Monoclonais Humanizados/uso terapêutico , Antineoplásicos/química , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Sequência de Carboidratos , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Cromatografia Líquida de Alta Pressão , Resistencia a Medicamentos Antineoplásicos/genética , Feminino , Glicopeptídeos/análise , Glicopeptídeos/química , Humanos , Lipídeos , Dados de Sequência Molecular , Plasmídeos , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/patologia , Processamento de Proteína Pós-Traducional , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Transfecção , Trastuzumab , Tripsina/metabolismoRESUMO
In recent years, the field of immunometabolism has solidified its position as a prominent area of investigation within the realm of immunological research. An expanding body of scientific literature has unveiled the intricate interplay between energy homeostasis, signalling molecules, and metabolites in relation to fundamental aspects of our immune cells. It is now widely accepted that disruptions in metabolic equilibrium can give rise to a myriad of pathological conditions, ranging from autoimmune disorders to cancer. Emerging evidence, although sometimes fragmented and anecdotal, has highlighted the indispensable role of lipids in modulating the behaviour of immune cells, including B cells. In light of these findings, this review aims to provide a comprehensive overview of the current state of knowledge regarding lipid metabolism in the context of B cell biology.
RESUMO
Despite available targeted treatments for the disease, drug-resistant chronic lymphocytic leukemia (CLL) poses a clinical challenge. The objective of this study is to examine whether the dual-specific phosphatases DUSP1 and DUSP6 are required to negatively regulate mitogen-activated protein kinases (MAPKs) and thus counterbalance excessive MAPK activity. We show that high expression of DUSP6 in CLL correlates with poor clinical prognosis. Importantly, genetic deletion of the inhibitory phosphatase DUSP1 or DUSP6 and blocking DUSP1/6 function using a small-molecule inhibitor reduces CLL cell survival in vitro and in vivo. Using global phospho-proteome approaches, we observe acute activation of MAPK signaling by DUSP1/6 inhibition. This promotes accumulation of mitochondrial reactive oxygen species and, thereby, DNA damage and apoptotic cell death in CLL cells. Finally, we observe that DUSP1/6 inhibition is particularly effective against treatment-resistant CLL and therefore suggest transient DUSP1/6 inhibition as a promising treatment concept to eliminate drug-resistant CLL cells.
Assuntos
Leucemia Linfocítica Crônica de Células B , Humanos , Retroalimentação , Proteínas Quinases Ativadas por MitógenoRESUMO
In most cell types, nuclear ß-catenin functions as prominent oncogenic driver and pairs with TCF7-family factors for transcriptional activation of MYC. Surprisingly, B-lymphoid malignancies not only lacked expression and activating lesions of ß-catenin but critically depended on GSK3ß for effective ß-catenin degradation. Our interactome studies in B-lymphoid tumors revealed that ß-catenin formed repressive complexes with lymphoid-specific Ikaros factors at the expense of TCF7. Instead of MYC-activation, ß-catenin was essential to enable Ikaros-mediated recruitment of nucleosome remodeling and deacetylation (NuRD) complexes for transcriptional repression of MYC. To leverage this previously unrecognized vulnerability of B-cell-specific repressive ß-catenin-Ikaros-complexes in refractory B-cell malignancies, we examined GSK3ß small molecule inhibitors to subvert ß-catenin degradation. Clinically approved GSK3ß-inhibitors that achieved favorable safety prof les at micromolar concentrations in clinical trials for neurological disorders and solid tumors were effective at low nanomolar concentrations in B-cell malignancies, induced massive accumulation of ß-catenin, repression of MYC and acute cell death. Preclinical in vivo treatment experiments in patient-derived xenografts validated small molecule GSK3ß-inhibitors for targeted engagement of lymphoid-specific ß-catenin-Ikaros complexes as a novel strategy to overcome conventional mechanisms of drug-resistance in refractory malignancies. HIGHLIGHTS: Unlike other cell lineages, B-cells express nuclear ß-catenin protein at low baseline levels and depend on GSK3ß for its degradation.In B-cells, ß-catenin forms unique complexes with lymphoid-specific Ikaros factors and is required for Ikaros-mediated tumor suppression and assembly of repressive NuRD complexes. CRISPR-based knockin mutation of a single Ikaros-binding motif in a lymphoid MYC superenhancer region reversed ß-catenin-dependent Myc repression and induction of cell death. The discovery of GSK3ß-dependent degradation of ß-catenin as unique B-lymphoid vulnerability provides a rationale to repurpose clinically approved GSK3ß-inhibitors for the treatment of refractory B-cell malignancies. GRAPHICAL ABSTRACT: Abundant nuclear ß-cateninß-catenin pairs with TCF7 factors for transcriptional activation of MYCB-cells rely on efficient degradation of ß-catenin by GSK3ßB-cell-specific expression of Ikaros factors Unique vulnerability in B-cell tumors: GSK3ß-inhibitors induce nuclear accumulation of ß-catenin.ß-catenin pairs with B-cell-specific Ikaros factors for transcriptional repression of MYC.