RESUMEN
B cells constitute an essential line of defense from pathogenic infections through the generation of class-switched antibody-secreting cells (ASCs) in germinal centers. Although this process is known to be regulated by follicular helper T (TfH) cells, the mechanism by which B cells initially seed germinal center reactions remains elusive. We found that NKT cells, a population of innate-like T lymphocytes, are critical for the induction of B cell immunity upon viral infection. The positioning of NKT cells at the interfollicular areas of lymph nodes facilitates both their direct priming by resident macrophages and the localized delivery of innate signals to antigen-experienced B cells. Indeed, NKT cells secrete an early wave of IL-4 and constitute up to 70% of the total IL-4-producing cells during the initial stages of infection. Importantly, the requirement of this innate immunity arm appears to be evolutionarily conserved because early NKT and IL-4 gene signatures also positively correlate with the levels of neutralizing antibodies in Zika-virus-infected macaques. In conclusion, our data support a model wherein a pre-TfH wave of IL-4 secreted by interfollicular NKT cells triggers the seeding of germinal center cells and serves as an innate link between viral infection and B cell immunity.
Asunto(s)
Linfocitos B/inmunología , Centro Germinal/inmunología , Inmunidad Innata , Gripe Humana/inmunología , Interleucina-4/genética , Células Asesinas Naturales/inmunología , Infección por el Virus Zika/inmunología , Animales , Pollos , Perros , Centro Germinal/citología , Humanos , Interleucina-4/metabolismo , Macaca , Macrófagos/inmunología , Células de Riñón Canino Madin Darby , Ratones , Ratones Endogámicos C57BLRESUMEN
PKCß-null (Prkcb-/-) mice are severely immunodeficient. Here we show that mice whose B cells lack PKCß failed to form germinal centers and plasma cells, which undermined affinity maturation and antibody production in response to immunization. Moreover, these mice failed to develop plasma cells in response to viral infection. At the cellular level, we have shown that Prkcb-/- B cells exhibited defective antigen polarization and mTORC1 signaling. While altered antigen polarization impaired antigen presentation and likely restricted the potential of GC development, defective mTORC1 signaling impaired metabolic reprogramming, mitochondrial remodeling, and heme biosynthesis in these cells, which altogether overwhelmingly opposed plasma cell differentiation. Taken together, our study reveals mechanistic insights into the function of PKCß as a key regulator of B cell polarity and metabolic reprogramming that instructs B cell fate.
Asunto(s)
Linfocitos B/inmunología , Diferenciación Celular/inmunología , Activación de Linfocitos/inmunología , Células Plasmáticas/inmunología , Proteína Quinasa C beta/inmunología , Animales , Hemo/biosíntesis , Ratones , Ratones Noqueados , Mitocondrias/inmunología , Mitocondrias/metabolismo , Células Plasmáticas/citologíaRESUMEN
Humans with Wiskott-Aldrich syndrome display a progressive immunological disorder associated with compromised Wiskott-Aldrich Syndrome Interacting Protein (WIP) function. Mice deficient in WIP recapitulate such an immunodeficiency that has been attributed to T cell dysfunction; however, any contribution of B cells is as yet undefined. Here we have shown that WIP deficiency resulted in defects in B cell homing, chemotaxis, survival, and differentiation, ultimately leading to diminished germinal center formation and antibody production. Furthermore, in the absence of WIP, several receptors, namely the BCR, BAFFR, CXCR4, CXCR5, CD40, and TLR4, were impaired in promoting CD19 co-receptor activation and subsequent PI3 kinase (PI3K) signaling. The underlying mechanism was due to a distortion in the actin and tetraspanin networks that lead to altered CD19 cell surface dynamics. In conclusion, our findings suggest that, by regulating the cortical actin cytoskeleton, WIP influences the function of CD19 as a general hub for PI3K signaling.
Asunto(s)
Antígenos CD19/fisiología , Linfocitos B/inmunología , Proteínas Portadoras/fisiología , Fosfatidilinositol 3-Quinasas/fisiología , Transducción de Señal/inmunología , Citoesqueleto de Actina/ultraestructura , Actinas/análisis , Animales , Formación de Anticuerpos , Linfocitos B/efectos de los fármacos , Linfocitos B/enzimología , Linfocitos B/ultraestructura , Proteínas Portadoras/genética , Células Cultivadas , Quimiocinas/farmacología , Quimiocinas/fisiología , Quimiotaxis/efectos de los fármacos , Proteínas del Citoesqueleto , Centro Germinal/inmunología , Centro Germinal/patología , Haptenos , Hemocianinas/farmacología , Activación de Linfocitos/efectos de los fármacos , Linfopoyesis , Proteínas de la Membrana/inmunología , Ratones , Fosforilación , Células Plasmáticas/inmunología , Procesamiento Proteico-Postraduccional , Quimera por Radiación , Receptores de Antígenos de Linfocitos B/inmunología , Receptores de Quimiocina/fisiología , Tetraspaninas/análisis , Vaccinia/inmunología , Vaccinia/patologíaRESUMEN
Invariant natural killer T cells (iNKT cells) are involved in the host defense against microbial infection. Although it is known that iNKT cells recognize glycolipids presented by CD1d, how and where they encounter antigen in vivo remains unclear. Here we used multiphoton microscopy to visualize the dynamics and activation of iNKT cells in lymph nodes. After antigen administration, iNKT cells became confined in a CD1d-dependent manner in close proximity to subcapsular sinus CD169(+) macrophages. These macrophages retained, internalized and presented lipid antigen and were required for iNKT cell activation, cytokine production and population expansion. Thus, CD169(+) macrophages can act as true antigen-presenting cells controlling early iNKT cell activation and favoring the fast initiation of immune responses.
Asunto(s)
Presentación de Antígeno/inmunología , Glucolípidos/inmunología , Ganglios Linfáticos/inmunología , Activación de Linfocitos/inmunología , Macrófagos/inmunología , Células T Asesinas Naturales/inmunología , Animales , Antígenos/inmunología , Separación Celular , Ensayo de Inmunoadsorción Enzimática , Citometría de Flujo , Ganglios Linfáticos/citología , Macrófagos/metabolismo , Glicoproteínas de Membrana/inmunología , Glicoproteínas de Membrana/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Microscopía Confocal , Microscopía de Fluorescencia por Excitación Multifotónica , Receptores Inmunológicos/inmunología , Receptores Inmunológicos/metabolismo , Lectina 1 Similar a Ig de Unión al Ácido SiálicoRESUMEN
A key role is emerging for the cytoskeleton in coordinating receptor signaling, although the underlying molecular requirements remain unclear. Here we show that cytoskeleton disruption triggered signaling requiring not only the B cell receptor (BCR), but also the coreceptor CD19 and tetraspanin CD81, thus providing a mechanism for signal amplification upon surface-bound antigen stimulation. By using superresolution microscopy, we demonstrated that endogenous IgM, IgD, and CD19 exhibited distinct nanoscale organization within the plasma membrane of primary B cells. Upon stimulation, we detect a local convergence of receptors, although their global organization was not dramatically altered. Thus, we postulate that cytoskeleton reorganization releases BCR nanoclusters, which can interact with CD19 held in place by the tetraspanin network. These results not only suggest that receptor compartmentalization regulates antigen-induced activation but also imply a potential role for CD19 in mediating ligand-independent "tonic" BCR signaling necessary for B cell survival.
Asunto(s)
Actinas/inmunología , Antígenos CD19/inmunología , Receptores de Antígenos de Linfocitos B/inmunología , Transducción de Señal/inmunología , Tetraspanina 28/inmunología , Actinas/metabolismo , Animales , Antígenos CD19/genética , Antígenos CD19/metabolismo , Linfocitos B/inmunología , Linfocitos B/metabolismo , Membrana Celular/inmunología , Membrana Celular/metabolismo , Citoesqueleto/inmunología , Citoesqueleto/metabolismo , Citometría de Flujo , Immunoblotting , Inmunoglobulina D/inmunología , Inmunoglobulina D/metabolismo , Inmunoglobulina M/inmunología , Inmunoglobulina M/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Ratones Transgénicos , Microscopía Fluorescente , Modelos Inmunológicos , Nanoestructuras , Unión Proteica/inmunología , Receptores de Antígenos de Linfocitos B/metabolismo , Tetraspanina 28/genética , Tetraspanina 28/metabolismoRESUMEN
In order to mount high-affinity antibody responses, B cells internalise specific antigens and process them into peptides loaded onto MHCII for presentation to T helper cells (TH cells). While the biochemical principles of antigen processing and MHCII loading have been well dissected, how the endosomal vesicle system is wired to enable these specific functions remains much less studied. Here, we performed a systematic microscopy-based analysis of antigen trafficking in B cells to reveal its route to the MHCII peptide-loading compartment (MIIC). Surprisingly, we detected fast targeting of internalised antigen into peripheral acidic compartments that possessed the hallmarks of the MIIC and also showed degradative capacity. In these vesicles, internalised antigen converged rapidly with membrane-derived MHCII and partially overlapped with cathepsin-S and H2-M, both required for peptide loading. These early compartments appeared heterogenous and atypical as they contained a mixture of both early and late endosomal markers, indicating a specialized endosomal route. Together, our data suggest that, in addition to in the previously reported perinuclear late endosomal MIICs, antigen processing and peptide loading could have already started in these specialized early peripheral acidic vesicles (eMIIC) to support fast peptide-MHCII presentation.
Asunto(s)
Presentación de Antígeno , Linfocitos B/inmunología , Antígenos de Histocompatibilidad Clase II/inmunología , Antígenos de Histocompatibilidad Clase II/metabolismo , Traslado Adoptivo , Animales , Linfocitos B/citología , Endosomas/metabolismo , Femenino , Humanos , Lisosomas/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Transporte de Proteínas , Receptores de Antígenos de Linfocitos B/genética , Receptores de Antígenos de Linfocitos B/metabolismoRESUMEN
Receptor organization and dynamics at the cell membrane are important factors of signal transduction regulation. Using super-resolution microscopy and single-particle tracking, we show how the negative coreceptor CD22 works with the cortical cytoskeleton in restraining BCR signalling. In naïve B cells, we found endogenous CD22 to be highly mobile and organized into nanodomains. The landscape of CD22 and its lateral diffusion were perturbed either in the absence of CD45 or when the CD22 lectin domain was mutated. To understand how a relatively low number of CD22 molecules can keep BCR signalling in check, we generated Brownian dynamic simulations and supported them with ex vivo experiments. This combined approach suggests that the inhibitory function of CD22 is influenced by its nanoscale organization and is ensured by its fast diffusion enabling a "global BCR surveillance" at the plasma membrane.
Asunto(s)
Linfocitos B/fisiología , Citoesqueleto/metabolismo , Receptores de Antígenos de Linfocitos B/metabolismo , Lectina 2 Similar a Ig de Unión al Ácido Siálico/metabolismo , Transducción de Señal , Animales , Linfocitos B/citología , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Microscopía FluorescenteRESUMEN
The B cell receptor (BCR) mediates B cell antigen gathering and acquisition for presentation to T cells. Although the amount of antigen presentation to T cells determines the extent of B cell activation, the molecular mechanisms underlying antigen gathering remain unexplored. Here, through a combination of high-resolution imaging, genetics and quantitative mass spectrometry, we demonstrate that adaptors Grb2 and Dok-3, and ubiquitin ligase Cbl in signaling BCR microclusters mediate association with the microtubule motor dynein. Furthermore, we visualize the localization and movement of these microclusters on the underlying microtubule network. Importantly, disruption of this network or diminished dynein recruitment in Grb2-, Dok-3-, or Cbl-deficient B cells, does not influence microcluster formation or actin-dependent spreading, but abrogates directed movement of microclusters and antigen accumulation. Thus we identify a surprising but pivotal role for dynein and the microtubule network alongside Grb2, Dok-3, and Cbl in antigen gathering during B cell activation.
Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/inmunología , Antígenos/inmunología , Dineínas/inmunología , Proteína Adaptadora GRB2/inmunología , Proteínas Proto-Oncogénicas c-cbl/inmunología , Receptores de Antígenos de Linfocitos B/inmunología , Transducción de Señal , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Animales , Células Cultivadas , Dineínas/metabolismo , Proteína Adaptadora GRB2/metabolismo , Ratones , Microtúbulos/metabolismo , Unión Proteica , Proteínas Proto-Oncogénicas c-cbl/metabolismo , Receptores de Antígenos de Linfocitos B/metabolismo , Tubulina (Proteína)/metabolismoRESUMEN
Early events of B cell activation after B cell receptor (BCR) triggering have been well characterized. However, little is known about the steady state of the BCR on the cell surface. Here, we simultaneously visualize single BCR particles and components of the membrane skeleton. We show that an ezrin- and actin-defined network influenced steady-state BCR diffusion by creating boundaries that restrict BCR diffusion. We identified the intracellular domain of Igbeta as important in mediating this restriction in diffusion. Importantly, alteration of this network was sufficient to induce robust intracellular signaling and concomitant increase in BCR mobility. Moreover, by using B cells deficient in key signaling molecules, we show that this signaling was most probably initiated by the BCR. Thus, our results suggest the membrane skeleton plays a crucial function in controlling BCR dynamics and thereby signaling, in a way that could be important for understanding tonic signaling necessary for B cell development and survival.
Asunto(s)
Actinas/metabolismo , Linfocitos B/metabolismo , Antígenos CD79/metabolismo , Membrana Celular/inmunología , Proteínas del Citoesqueleto/metabolismo , Actinas/inmunología , Animales , Linfocitos B/efectos de los fármacos , Linfocitos B/inmunología , Linfocitos B/patología , Compuestos Bicíclicos Heterocíclicos con Puentes/farmacología , Antígenos CD79/genética , Antígenos CD79/inmunología , Línea Celular Tumoral , Membrana Celular/efectos de los fármacos , Membrana Celular/metabolismo , Proteínas del Citoesqueleto/inmunología , Citoesqueleto/efectos de los fármacos , Citoesqueleto/inmunología , Recubrimiento Inmunológico/efectos de los fármacos , Recubrimiento Inmunológico/genética , Recubrimiento Inmunológico/inmunología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Microscopía Fluorescente , Unión Proteica , Ingeniería de Proteínas , Estructura Terciaria de Proteína/genética , Transducción de Señal/efectos de los fármacos , Transducción de Señal/genética , Transducción de Señal/inmunología , Tiazolidinas/farmacologíaRESUMEN
The molecules on mammalian spermatozoa that mediate recognition and binding to the zona pellucida of the egg are still not understood. Current concepts favour their assembly into multimolecular complexes in the plasma membrane in response to cholesterol efflux, an important step during sperm capacitation. Here, we track in real time diffusion of cross-linked clusters containing zona-binding molecules and GM1 gangliosides in the plasma membrane of live boar spermatozoa before and after cholesterol reduction. Both GM1 gangliosides and zona-binding molecules partition into a low density Triton X100 resistant phase suggesting their association with lipid rafts. Initially, GM1 and zona-binding molecules localize to the apical ridge on the acrosome but following cholesterol efflux with methyl-beta-cyclodextrin, clusters containing zona-binding molecules diffuse randomly over the acrosomal domain. Diffusing clusters of either type do not access the postacrosome. Spermatozoa agglutinated head-to-head show contact-induced coalescence of GM1 gangliosides (but not zona-binding molecules) suggestive of a specific mechanosensitive response. Thus, cholesterol efflux initiates diffusion (and possibly formation) of novel lipid raft-like structures containing zona-binding molecules over the sperm acrosome. We hypothesise that in combination with contact coalescence, these mechanisms concentrate important molecules to the appropriate site on the sperm surface to mediate zona binding.
Asunto(s)
Membrana Celular/metabolismo , Colesterol/metabolismo , Gangliósido G(M1)/metabolismo , Espermatozoides/metabolismo , Zona Pelúcida/metabolismo , Acrosoma/metabolismo , Animales , Sitios de Unión , Masculino , PorcinosRESUMEN
We have investigated exchange of molecules between different membrane domains on a highly compartmentalized cell, the spermatozoon. Using Alexa Fluor 555-cholera toxin B-subunit we have observed clustering of preexisting GM1 gangliosides which diffused across the anterior acrosome-equatorial segment interface but did not access the postacrosome. By contrast, single lipid and protein molecules readily exchanged between all three domains, although they diffused more slowly on nearing and crossing to the postacrosome. Thus, two types of diffusion interfaces are present on sperm heads, an "open" interface and a "mass filter" interface. The latter seems to be due to a protein-cytoskeleton network.
Asunto(s)
Microdominios de Membrana/metabolismo , Espermatozoides/citología , Animales , Toxina del Cólera/metabolismo , Difusión , Cinética , Masculino , Espermatozoides/metabolismo , PorcinosRESUMEN
Although RNA aptamers can show comparable or better specificity and affinity to antibodies and have the advantage of being able to access different live cell compartments, they are often much less stable in vivo. We report here the first aptamer that binds human retinoblastoma protein (RB) and is stable in live cells. RB is both a key protein in cell cycle control and also a tumour suppressor. The aptamer was selected from an RNA library against a unique 12-residue helical peptide derived from RB rather than the whole protein molecule. It binds RB with high affinity (K d = 5.1 ± 0.1 nM) and is a putative RNA G-quadruplex structure formed by an 18-nucleotide sequence (18E16 - GGA GGG UGG AGG GAA GGG), which may account for its high stability. Confocal fluorescence microscopy of live cells transfected with the aptamer shows it is stable intracellularly and efficient in entering the nucleus where an analogous antibody was inaccessible. The findings demonstrate this aptamer is an advanced probe for RB in live cell applications.
RESUMEN
Single molecule tracking is widely used to monitor the change in position of lipids and proteins in living cells. In many experiments in which molecules are tagged with a single or small number of fluorophores, the signal/noise ratio may be limiting, the number of molecules is not known, and fluorophore blinking and photobleaching can occur. All these factors make accurate tracking over long trajectories difficult and hence there is still a pressing need to develop better algorithms to extract the maximum information from a sequence of fluorescence images. We describe here a Bayesian-based inference approach, based on a trans-dimensional sequential Monte Carlo method that utilizes both the spatial and temporal information present in the image sequences. We show, using model data, where the real trajectory of the molecule is known, that our method allows accurate tracking of molecules over long trajectories even with low signal/noise ratio and in the presence of fluorescence blinking and photobleaching. The method is then applied to real experimental data.
Asunto(s)
Inteligencia Artificial , Biopolímeros/química , Aumento de la Imagen/métodos , Interpretación de Imagen Asistida por Computador/métodos , Microscopía Fluorescente/métodos , Microscopía por Video/métodos , Reconocimiento de Normas Patrones Automatizadas/métodos , Algoritmos , Teorema de Bayes , Movimiento (Física)RESUMEN
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.
Asunto(s)
Proteínas Adaptadoras del Transporte Vesicular/metabolismo , Linfocitos B/inmunología , Vacunas contra la Influenza/inmunología , Infecciones por Orthomyxoviridae/patología , Orthomyxoviridae/inmunología , Linfocitos T/inmunología , Proteínas Adaptadoras del Transporte Vesicular/deficiencia , Animales , Adhesión Celular , Proliferación Celular , Vacunas contra la Influenza/administración & dosificación , Ratones , Análisis de SupervivenciaRESUMEN
During B-cell activation, the dynamic reorganisation of the cytoskeleton is crucial for multiple cellular responses, such as receptor signalling, cell spreading, antigen internalisation, intracellular trafficking, and antigen presentation. However, the role of intermediate filaments (IFs), which represent a major component of the mammalian cytoskeleton, is not well defined. Here, by using multiple super-resolution microscopy techniques, including direct stochastic optical reconstruction microscopy, we show that IFs in B cells undergo drastic reorganisation immediately upon antigen stimulation and that this reorganisation requires actin and microtubules. Although the loss of vimentin in B cells did not impair B-cell development, receptor signalling, and differentiation, vimentin-deficient B cells exhibit altered positioning of antigen-containing and lysosomal associated membrane protein 1 (LAMP1+) compartments, implying that vimentin may play a role in the fine-tuning of intracellular trafficking. Indeed, vimentin-deficient B cells exhibit impaired antigen presentation and delayed antibody responses in vivo. Thus, our study presents a new perspective on the role of IFs in B-cell activation.
RESUMEN
Wiskott-Aldrich syndrome protein (WASp) is a main cytoskeletal regulator in B cells. WASp-interacting protein (WIP) binds to and stabilizes WASp but also interacts with actin. Using mice with a mutated actin binding domain of WIP (WIPΔABD), we here investigated the role of WIP binding to actin during B cell activation. We found an altered differentiation of WIPΔABD B cells and diminished antibody affinity maturation after immunization. Mechanistically, WIPΔABD B cells showed impaired B cell receptor (BCR)-induced PI3K signaling and actin reorganization, likely caused by diminished CD81 expression and altered CD19 dynamics on the B cell surface. WIPΔABD B cells displayed reduced in vivo motility, concomitantly with impaired chemotaxis and defective F-actin polarization, HS1 phosphorylation, and polarization of HS1 to F-actin-rich structures after CXCL12 stimulation in vitro. We thus concluded that WIP binding to actin, independent of its binding to WASp, is critical for actin cytoskeleton plasticity in B cells.
Asunto(s)
Actinas/metabolismo , Linfocitos B/citología , Linfocitos B/metabolismo , Movimiento Celular , Inmunidad Humoral , Animales , Afinidad de Anticuerpos , Antígenos CD/metabolismo , Proteínas Portadoras/metabolismo , Membrana Celular/metabolismo , Polaridad Celular , Quimiotaxis , Proteínas del Citoesqueleto , Difusión , Centro Germinal/metabolismo , Factor Estimulante de Colonias de Granulocitos/metabolismo , Ratones , Fosfatidilinositol 3-Quinasas/metabolismo , Unión Proteica , Receptores de Antígenos de Linfocitos B/metabolismo , Transducción de SeñalRESUMEN
AIM: In the present study, a variety of high resolution microscopy techniques were used to visualize the organization and motion of lipids and proteins in the sperm's plasma membrane. We have addressed questions such as the presence of diffusion barriers, confinement of molecules to specific surface domains, polarized diffusion and the role of cholesterol in regulating lipid rafts and signal transduction during capacitation. METHODS: Atomic force microscopy identified a novel region (EqSS) within the equatorial segment of bovine, porcine and ovine spermatozoa that was enriched in constitutively phosphorylated proteins. The EqSS was assembled during epididymal maturation. Fluorescence imaging techniques were then used to follow molecular diffusion on the sperm head. RESULTS: Single lipid molecules were freely exchangeable throughout the plasma membrane and showed no evidence for confinement within domains. Large lipid aggregates, however, did not cross over the boundary between the post-acrosome and equatorial segment suggesting the presence of a molecular filter between these two domains. CONCLUSION: A small reduction in membrane cholesterol enlarges or increases lipid rafts concomitant with phosphorylation of intracellular proteins. Excessive removal of cholesterol, however, disorganizes rafts with a cessation of phosphorylation. These techniques are forcing a revision of long-held views on how lipids and proteins in sperm membranes are assembled into larger complexes that mediate recognition and fusion with the egg.
Asunto(s)
Membrana Celular/ultraestructura , Capacitación Espermática/fisiología , Maduración del Esperma/fisiología , Espermatozoides/citología , Membrana Celular/fisiología , Humanos , Masculino , Lípidos de la Membrana/fisiología , Microscopía de Fuerza Atómica , Espermatozoides/fisiologíaRESUMEN
Autophagy is important in a variety of cellular and pathophysiological situations; however, its role in immune responses remains elusive. Here, we show that among B cells, germinal center (GC) cells exhibited the highest rate of autophagy during viral infection. In contrast to mechanistic target of rapamycin complex 1-dependent canonical autophagy, GC B cell autophagy occurred predominantly through a noncanonical pathway. B cell stimulation was sufficient to down-regulate canonical autophagy transiently while triggering noncanonical autophagy. Genetic ablation of WD repeat domain, phosphoinositide-interacting protein 2 in B cells alone enhanced this noncanonical autophagy, resulting in changes of mitochondrial homeostasis and alterations in GC and antibody-secreting cells. Thus, B cell activation prompts a temporal switch from canonical to noncanonical autophagy that is important in controlling B cell differentiation and fate.
Asunto(s)
Autofagia/inmunología , Linfocitos B/inmunología , Linfocitos B/virología , Virosis/inmunología , Animales , Regulación hacia Abajo , Centro Germinal/inmunología , Centro Germinal/virología , Activación de Linfocitos , Diana Mecanicista del Complejo 1 de la Rapamicina , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Proteínas Asociadas a Microtúbulos/genética , Proteínas Asociadas a Microtúbulos/metabolismo , Mitocondrias/metabolismo , Complejos Multiproteicos/metabolismo , Serina-Treonina Quinasas TOR/metabolismo , Repeticiones WD40/genéticaRESUMEN
Cancer-associated fibroblasts (CAFs) are non-cancerous cells found in solid tumors that remodel the tumor matrix and promote cancer invasion and angiogenesis. Here, we demonstrate that Cdc42EP3/BORG2 is required for the matrix remodeling, invasion, angiogenesis, and tumor-growth-promoting abilities of CAFs. Cdc42EP3 functions by coordinating the actin and septin networks. Furthermore, depletion of SEPT2 has similar effects to those of loss of Cdc42EP3, indicating a role for the septin network in the tumor stroma. Cdc42EP3 is upregulated early in fibroblast activation and precedes the emergence of the highly contractile phenotype characteristic of CAFs. Depletion of Cdc42EP3 in normal fibroblasts prevents their activation by cancer cells. We propose that Cdc42EP3 sensitizes fibroblasts to further cues-in particular, those activating actomyosin contractility-and thereby enables the generation of the pathological activated fibroblast state.
Asunto(s)
Fibroblastos/metabolismo , Fibroblastos/patología , Reguladores de Proteínas de Unión al GTP/metabolismo , Neoplasias/metabolismo , Neoplasias/patología , Septinas/metabolismo , Proteína de Unión al GTP cdc42/metabolismo , Animales , Línea Celular , Línea Celular Tumoral , Humanos , Ratones , Regulación hacia ArribaRESUMEN
The layer of macrophages at the subcapsular sinus (SCS) captures pathogens entering the lymph node, preventing their global dissemination and triggering an immune response. However, how infection affects SCS macrophages remains largely unexplored. Here we show that infection and inflammation disrupt the organization of SCS macrophages in a manner that involves the migration of mature dendritic cells to the lymph node. This disrupted organization reduces the capacity of SCS macrophages to retain and present antigen in a subsequent secondary infection, resulting in diminished B cell responses. Thus, the SCS macrophage layer may act as a sensor or valve during infection to temporarily shut down the lymph node to further antigenic challenge. This shutdown may increase an organism's susceptibility to secondary infections.