Showing papers by "Richard A. Flavell published in 2001"
TL;DR: It is shown that mammalian TLR3 recognizes dsRNA, and that activation of the receptor induces the activation of NF-κB and the production of type I interferons (IFNs).
Abstract: Toll-like receptors (TLRs) are a family of innate immune-recognition receptors that recognize molecular patterns associated with microbial pathogens, and induce antimicrobial immune responses. Double-stranded RNA (dsRNA) is a molecular pattern associated with viral infection, because it is produced by most viruses at some point during their replication. Here we show that mammalian TLR3 recognizes dsRNA, and that activation of the receptor induces the activation of NF-kappaB and the production of type I interferons (IFNs). TLR3-deficient (TLR3-/-) mice showed reduced responses to polyinosine-polycytidylic acid (poly(I:C)), resistance to the lethal effect of poly(I:C) when sensitized with d-galactosamine (d-GalN), and reduced production of inflammatory cytokines. MyD88 is an adaptor protein that is shared by all the known TLRs. When activated by poly(I:C), TLR3 induces cytokine production through a signalling pathway dependent on MyD88. Moreover, poly(I:C) can induce activation of NF-kappaB and mitogen-activated protein (MAP) kinases independently of MyD88, and cause dendritic cells to mature.
6,066 citations
TL;DR: In mammals, BH3 domain-only molecules activate multidomain proapoptotic members to trigger a mitochondrial pathway, which both releases cytochrome c to activate caspases and initiates caspase-independent mitochondrial dysfunction.
1,708 citations
TL;DR: It is shown that T-cell activation and proliferation are defective in the absence of ICOS, and ICOS-/- mice showed greatly enhanced susceptibility to experimental autoimmune encephalomyelitis, indicating that ICOS has a protective role in inflammatory autoimmune diseases.
Abstract: T-lymphocyte activation and immune function are regulated by co-stimulatory molecules. CD28, a receptor for B7 gene products, has a chief role in initiating T-cell immune responses1,2. CTLA4, which binds B7 with a higher affinity, is induced after T-cell activation and is involved in downregulating T-cell responses3,4. The inducible co-stimulatory molecule (ICOS), a third member of the CD28/CTLA4 family, is expressed on activated T cells5,6. Its ligand B7H/B7RP-1 is expressed on B cells and in non-immune tissues after injection of lipopolysaccharide into animals6,7. To understand the role of ICOS in T-cell activation and function, we generated and analysed ICOS-deficient mice. Here we show that T-cell activation and proliferation are defective in the absence of ICOS. In addition, ICOS-/- T cells fail to produce interleukin-4 when differentiated in vitro or when primed in vivo. ICOS is required for humoral immune responses after immunization with several antigens. ICOS-/- mice showed greatly enhanced susceptibility to experimental autoimmune encephalomyelitis, indicating that ICOS has a protective role in inflammatory autoimmune diseases.
908 citations
TL;DR: It is demonstrated here that T-cell–specific blockade of TGF-β signaling allows the generation of an immune response capable of eradicating tumors in mice challenged with live tumor cells.
Abstract: Despite the existence of tumor-specific antigens and demonstrated presence of tumor-specific immune cells, the majority of tumors manage to avoid immune-mediated destruction. Various mechanisms have been suggested for tumor evasion from immune response. One such mechanism is thought to be mediated by transforming growth factor-beta (TGF-beta), an immunosuppressive cytokine found at the site of most tumors. We demonstrate here that T-cell-specific blockade of TGF-beta signaling allows the generation of an immune response capable of eradicating tumors in mice challenged with live tumor cells. In addition, we provide mechanisms through which abrogation of TGF-beta signaling leads to the enhancement of anti-tumor immunity. Our data indicate that T-cell-specific blockade of TGF-beta signaling has strong therapeutic potential to shift the balance of the immune response in favor of anti-tumor immunity.
746 citations
TL;DR: An until now unappreciated dominant role for VCAM-1 in lymphocyte homing to BM is reported, thereby overcoming the embryonic lethality seen with conventional vcam-1–deficient mice.
Abstract: We generated vascular cell adhesion molecule (VCAM)-1 "knock-in" mice and Cre recombinase transgenic mice to delete the VCAM-1 gene (vcam-1) in whole mice, thereby overcoming the embryonic lethality seen with conventional vcam-1-deficient mice. vcam-1 knock-in mice expressed normal levels of VCAM-1 but showed loss of VCAM-1 on endothelial and hematopoietic cells when interbred with a "TIE2Cre" transgene. Analysis of peripheral blood from conditional vcam-1-deficient mice revealed mild leukocytosis, including elevated immature B cell numbers. Conversely, the bone marrow (BM) had reduced immature B cell numbers, but normal numbers of pro-B cells. vcam-1-deficient mice also had reduced mature IgD+ B and T cells in BM and a greatly reduced capacity to support short-term migration of transferred B cells, CD4+ T cells, CD8+ T cells, and preactivated CD4+ T cells to the BM. Thus, we report an until now unappreciated dominant role for VCAM-1 in lymphocyte homing to BM.
497 citations
TL;DR: It is demonstrated that in cortical neurons exposed to Aβ, activated c-Jun N-terminal kinase (JNK) is required for the phosphorylation and activation of the c- Jun transcription factor, which in turn stimulates the transcription of several key target genes, including the death inducer Fas ligand.
Abstract: Elevated levels of β-Amyloid (Aβ) are present in the brains of individuals with either the sporadic or familial form of Alzheimer9s disease (AD), and the deposition of Aβ within the senile plaques that are a hallmark of AD is thought to be a primary cause of the cognitive dysfunction that occurs in AD. Recent evidence suggests that Aβ induces neuronal apoptosis in the brain and in primary neuronal cultures, and that this Aβ-induced neuronal death may be responsible in part for the cognitive decline found in AD patients. In this study we have characterized one mechanism by which Aβ induces neuronal death. We found that in cortical neurons exposed to Aβ, activated c-Jun N-terminal kinase (JNK) is required for the phosphorylation and activation of the c-Jun transcription factor, which in turn stimulates the transcription of several key target genes, including the death inducer Fas ligand. The binding of Fas ligand to its receptor Fas then induces a cascade of events that lead to caspase activation and ultimately cell death. By analyzing the effects of mutations in each of the components of the JNK–c-Jun–Fas ligand–Fas pathway, we demonstrate that this pathway plays a critical role in mediating Aβ-induced death of cultured neurons. These findings raise the possibility that the JNK pathway may also contribute to Aβ-dependent death in AD patients.
478 citations
TL;DR: It is shown that MKK4 and MKK7 serve different functions in the JNK signal transduction pathway, and disruption of the Mkk7 gene alone was sufficient to prevent JNK activation caused by proinflammatory cytokines.
Abstract: Mitogen-activated protein kinases (MAPK) are activated by phosphorylation on Thr and Tyr by MAPK kinases. Two MAPK kinases (MKK4 and MKK7) can activate the c-Jun NH(2)-terminal kinase (JNK) group of MAPK in vitro. JNK is phosphorylated preferentially on Tyr by MKK4 and on Thr by MKK7. Targeted gene-disruption studies in mice were performed to examine the role of MKK4 and MKK7 in vivo. Simultaneous disruption of the Mkk4 and Mkk7 genes was required to block JNK activation caused by exposure of cells to environmental stress. In contrast, disruption of the Mkk7 gene alone was sufficient to prevent JNK activation caused by proinflammatory cytokines. These data demonstrate that MKK4 and MKK7 serve different functions in the JNK signal transduction pathway.
371 citations
TL;DR: It is shown that the development of chronic intestinal inflammation in IL‐10‐deficient mice requires the function of TNF, indicating that theIL‐10/TNF axis regulates mucosal immunity and crosstalk is necessary for optimal TNF production and for the maintenance of immune homeostasis in the gut.
Abstract: Interleukin-10 (IL-10) is a key inhibitory signal of inflammatory responses that regulates the production of potentially pathogenic cytokines like tumor necrosis factor (TNF). We show here that the development of chronic intestinal inflammation in IL-10-deficient mice requires the function of TNF, indicating that the IL-10/TNF axis regulates mucosal immunity. We further show that IL-10 targets the 3′ AU-rich elements (ARE) of TNF mRNA to inhibit its translation. Moreover, IL-10 does not alter TNF mRNA stability, and its action does not require the presence of the stability-regulating ARE binding factor tristetraprolin, indicating a differential assembly of stability and translation determinants on the TNF ARE. Inhibition of TNF translation by IL-10 is exerted mainly by inhibition of the activating p38/MAPK-activated protein kinase-2 pathway. These results demonstrate a physiologically significant cross-talk between the IL-10 receptor and the stress-activated protein kinase modules targeting TNF mRNA translation. This cross-talk is necessary for optimal TNF production and for the maintenance of immune homeostasis in the gut.
271 citations
TL;DR: Gata-3 mediated strong enhancement of IL-4 promoter activity in Th1 cells when the promoter was embedded in the minilocus or linked to HSS and IE, demonstrating that GATA-3 acts through these elements to regulate IL- 4 gene expression.
245 citations
TL;DR: The mouse homolog of GILT was identified and a GilT knockout mouse was generated and facilitated the processing and presentation to antigen-specific T cells of protein antigens containing disulfide bonds.
Abstract: Processing of proteins for major histocompatibility complex (MHC) class II-restricted presentation to CD4-positive T lymphocytes occurs after they are internalized by antigen-presenting cells (APCs). Antigenic proteins frequently contain disulfide bonds, and their reduction in the endocytic pathway facilitates processing. In humans, a gamma interferon-inducible lysosomal thiol reductase (GILT) is constitutively present in late endocytic compartments of APCs. Here, we identified the mouse homolog of GILT and generated a GILT knockout mouse. GILT facilitated the processing and presentation to antigen-specific T cells of protein antigens containing disulfide bonds. The response to hen egg lysozyme, a model antigen with a compact structure containing four disulfide bonds, was examined in detail.
243 citations
TL;DR: Disruption of the Jip1 gene in mice by homologous recombination prevented JNK activation caused by exposure to excitotoxic stress and anoxic stress in vivo and in vitro.
Abstract: The JNK (c-Jun N-terminal kinase) group of mitogen-activated protein (MAP) kinases phosphorylate the transcription factor c-Jun (Derijard et al. 1994; Kyriakis et al. 1994). JNK is activated in response to cellular stress and contributes to the apoptotic response (for review, see Davis 2000). Indeed, JNK is implicated in neuronal death following exposure to excitotoxins, during the progression of some neurodegenerative diseases (e.g., Huntington's), and during the recovery from stroke (Davis 2000). Therefore, JNK deficiency prevents c-Jun phosphorylation and causes resistance to stress-induced apoptosis (Yang et al. 1997b; Tournier et al. 2000). Similarly, mutation of the JNK phosphorylation sites on c-Jun (Ser63 and Ser73) prevents stress-induced neuronal apoptosis (Behrens et al. 1999). The JNK signal transduction pathway therefore contributes to physiological and pathophysiological neuronal responses to stress.
JNK is activated by dual phosphorylation on the tripeptide motif Thr–Pro–Tyr (Derijard et al. 1994). Two MAP-kinase kinases that activate JNK have been identified, MKK4 and MKK7 (for review, see Davis 2000). Disruption of the Mkk4 (Nishina et al. 1997; Yang et al. 1997a; Ganiatsas et al. 1998; Swat et al. 1998) or Mkk7 (Dong et al. 2000) genes causes defects in JNK activation in response to specific stimuli (Tournier et al. 2001). In contrast, simultaneous disruption of both Mkk4 and Mkk7 eliminates JNK activation in response to stress, indicating that the MKK4 and MKK7 protein kinases represent the major activators of JNK in vivo (Tournier et al. 2001). These MAP kinase kinases are activated, in turn, by phosphorylation by MAP kinase kinase kinases, including TAK1, TPL2, and members of the ASK, MLK, and MEKK groups (Garrington and Johnson 1999). These MAP-kinase kinase kinases serve to integrate signals mediated by upstream signaling molecules (e.g., Rho family GTPases) to the activation of the protein kinase cascade that leads to JNK activation (for review, see Davis 2000).
The protein kinases that form the JNK signal transduction pathway may be organized into modules (Whitmarsh and Davis 1998). The kinases may function as a series of sequential binary complexes (Xia et al. 1998). Alternatively, one of the protein kinases may serve to bind the other protein kinases to form a functional module (Cheng et al. 2000). It is also possible that scaffold proteins may assemble a functional signaling module (Whitmarsh and Davis 1998). Studies of yeast have established that the protein kinase components of the mating MAP-kinase pathway interact with the scaffold protein Ste5p and that this interaction is essential for the formation of a functional signaling module (Elion 2000). Recent studies of the JNK signal transduction pathway have led to the identification of two types of potential scaffold proteins, β-arrestin and JIP.
The β-arrestin scaffolds are implicated in signaling by G protein coupled receptors (GPCR). These receptors bind ligand and are phosphorylated by GPCR kinases (GRK; Pitcher et al. 1998). The phosphorylated receptors can engage the scaffold protein β-arrestin-2 (Miller and Lefkowitz 2001), which serves as a site of assembly for a functional JNK signaling module (McDonald et al. 2000). The β-arrestin-2 scaffold protein directly binds JNK3 and ASK1 at different sites and indirectly interacts with MKK4. This scaffold protein may therefore serve to recruit a JNK signaling module to activated seven transmembrane-spanning receptors.
The JIP group of scaffold proteins (also known as IB/JSAP) bind to JNK, MKK7, and to members of the mixed-lineage protein kinase (MLK) group (Dickens et al. 1997; Bonny et al. 1998; Whitmarsh et al. 1998; Ito et al. 1999; Yasuda et al. 1999; Kelkar et al. 2000; Negri et al. 2000). The Jip1 gene is expressed in many tissues, including neurons, neuroendocrine cells (e.g., the β cells of the islets of Langerhans), lung, kidney, and in several other tissues at lower levels. In contrast, the JIP2 and JIP3 proteins are selectively expressed in neurons and in neuroendocrine cells, but low levels can be detected in some other tissues. In vitro biochemical assays and transfection assays show that JNK, MKK7, and MLK bind to separate sites on JIP proteins (Whitmarsh et al. 1998). These assays also show that JIP proteins potentiate the activation of JNK (Whitmarsh et al. 1998). Therefore, JIP proteins represent putative scaffolds that may contribute to the activation of JNK in vivo (Whitmarsh and Davis 1998).
Although biochemical studies and transfection assays indicate that these putative JNK scaffold proteins can potentiate JNK activation in cultured cells, the function of these scaffold proteins in vivo has not been established (Davis 2000). The purpose of this study was to examine the role of the JIP1 scaffold protein. A null allele of the Jip1 gene in mice was created by homologous recombination. We show that JIP1 is required for stress-induced activation of JNK in hippocampal neurons in vivo and in vitro. In addition, we show that the JIP1 protein accumulates in the perinuclear region of hippocampal neurons following exposure to stress. These data show that JIP1 acts as a dynamically regulated scaffold protein for a JNK signaling module in vivo.
Journal Article•
TL;DR: It is shown that in JNK2-deficient (Jnk2(-/-)) mice, the multiplicity of papillomas induced by 12-O-tetradecanoylphorbol-13-acetate (TPA) was lower than that in wild-type mice, and data suggest that J NK2 is critical in the tumor promotion process.
Abstract: Previous studies have shown that c-Jun NH(2)-terminal kinase (JNK) belongs to the mitogen-activated protein kinase (MAPK) family of signal transduction components that are rapidly initiated and activated by many extracellular stimuli. However, the potential role of JNK in mediating tumor promotion and carcinogenesis is unclear. We show here that in JNK2-deficient (Jnk2(-/-)) mice, the multiplicity of papillomas induced by 12-O-tetradecanoylphorbol-13-acetate (TPA) was lower than that in wild-type mice. Papillomas on wild-type mice grew rapidly and were well vascularized compared with Jnk2(-/-) mice. After the 12th week of TPA treatment, the mean number of tumors per mouse was 4.13-4.86 in wild-type mice but only 1.13-2.5 in Jnk2(-/-) mice. TPA induced phosphorylation of extracellular signal-regulated kinases and activator protein-1 DNA binding activity in wild-type mice, but the phosphorylation of extracellular signal-regulated kinases and activator protein-1 DNA binding were inhibited in Jnk2(-/-) mice. These data suggest that JNK2 is critical in the tumor promotion process.
TL;DR: It was found that ICOS−/− mice, when immunized with SRBC, had smaller GCs and IgG1 class switching in the GCs, and GC formation in response to a secondary recall challenge was completely absent in ICOS knockout mice, establishing a critical role of ICOS in regulation of humoral immunity.
Abstract: Inducible costimulator (ICOS) is a new member of the CD28/CTLA-4 family that is expressed on activated and germinal center (GC) T cells. Recently, we reported that ICOS-deficient mice exhibited profound defects in T cell activation and effector function. Ab responses in a T-dependent primary reaction and in a murine asthma model were also diminished. In the current study, we investigate the mechanism by which ICOS regulates humoral immunity and examine B cell GC reactions in the absence of ICOS. We found that ICOS(-/-) mice, when immunized with SRBC, had smaller GCs. Furthermore, IgG1 class switching in the GCs was impaired. Remarkably, GC formation in response to a secondary recall challenge was completely absent in ICOS knockout mice. These data establish a critical role of ICOS in regulation of humoral immunity.
TL;DR: Results are consistent with the growing idea that the involvement of specific caspases and the occurrence of caspase-independent programmed cell death may be dependent on brain region, cell type, age, and species or may be the result of specific perturbations or pathology.
Abstract: An analysis of programmed cell death of several populations of developing postmitotic neurons after genetic deletion of two key members of the caspase family of pro-apoptotic proteases, caspase-3 and caspase-9, indicates that normal neuronal loss occurs. Although the amount of cell death is not altered, the death process may be delayed, and the cells appear to use a nonapoptotic pathway of degeneration. The neuronal populations examined include spinal interneurons and motor, sensory, and autonomic neurons. When examined at both the light and electron microscopic levels, the caspase-deficient neurons exhibit a nonapoptotic morphology in which nuclear changes such as chromatin condensation are absent or reduced; in addition, this morphology is characterized by extensive cytoplasmic vacuolization that is rarely observed in degenerating control neurons. There is also reduced terminal deoxynucleotidyl transferase-mediated biotinylated UTP nick end labeling in dying caspase-deficient neurons. Despite the altered morphology and apparent temporal delay in cell death, the number of neurons that are ultimately lost is indistinguishable from that seen in control animals. In contrast to the striking perturbations in the morphology of the forebrain of caspase-deficient embryos, the spinal cord and brainstem appear normal. These results are consistent with the growing idea that the involvement of specific caspases and the occurrence of caspase-independent programmed cell death may be dependent on brain region, cell type, age, and species or may be the result of specific perturbations or pathology.
[...]
TL;DR: This review summarizes recent progress on different features of co-stimulatory regulation and chemokine-mediated homing of Th1 and Th2 cells.
Abstract: After activation, CD4 helper T (Th) cells differentiate into Th1or Th2 effector subsets. These two types of cells produce distinct profiles of cytokines and regulate different immune responses. This review summarizes recent progress on different features of co-stimulatory regulation and chemokine-mediated homing of Th1 and Th2 cells. Transcription factors and signaling pathways that are selectively expressed or activated in Th1 and Th2 cells to regulate cytokine gene expression are discussed.
TL;DR: It is determined that oocyte death initiated as a result of either developmental cues or pathological insults was unaffected by the absence of caspase-3, and that the enzyme is dispensable for germ cell apoptosis in the female.
Abstract: Previous studies have proposed the involvement of caspase-3, a downstream executioner enzyme common to many paradigms of programmed cell death (PCD), in mediating the apoptosis of both germ and somatic cells in the ovary. Herein we used caspase-3 gene knockout mice to directly test for the functional requirement of this protease in oocyte and/or granulosa cell demise. Using both in vivo and in vitro approaches, we determined that oocyte death initiated as a result of either developmental cues or pathological insults was unaffected by the absence of caspase-3. However, granulosa cells of degenerating antral follicles in both mouse and human ovaries showed a strong immunoreaction using an antibody raised against the cleaved (activated) form of caspase-3. Furthermore, caspase-3 mutant female mice possessed aberrant atretic follicles containing granulosa cells that failed to be eliminated by apoptosis, as confirmed by TUNEL (terminal deoxynucleotidyl transferase-mediated deoxy-UTP nick end labeling) analysis ...
TL;DR: GADD45gamma mediates activation of the p38 and JNK pathways and effector function of TH1 cells and causes reduced contact hypersensitivity in mice.
TL;DR: This work shows that Stra13-deficient mice exhibit defects in several phases of CD4+ T cell activation, and identifies Stra13 as a key regulator of lymphocyte activation that is vital for maintenance of self-tolerance and for constraint of autoimmunity.
Abstract: Stra13, a basic helix-loop-helix transcription factor, is up-regulated upon activation of CD4+ T cells. Here we show that Stra13-deficient mice exhibit defects in several phases of CD4+ T cell activation. In vivo, Stra13 deficiency results in ineffective elimination of activated T and B cells, which accumulate progressively, leading to lymphoid organ hyperplasia. Consequently, aging Stra13-/- mice develop autoimmune disease characterized by accumulation of spontaneously activated T and B cells, circulating autoantibodies, infiltration of T and B lymphocytes in several organs and immune complex deposition in glomeruli. Our studies identify Stra13 as a key regulator of lymphocyte activation that is vital for maintenance of self-tolerance and for constraint of autoimmunity.
TL;DR: It is reported here that islet-specific expression of TNF-α can play a dual role in autoimmune diabetes, depending on its precise timing in relation to the ongoing autoimmune process, and the fine-tuned kinetics of an autoreactive process undergo distinct stages that respond in a differential way to the presence of T NF-α.
Abstract: We report here that islet-specific expression of TNF-alpha can play a dual role in autoimmune diabetes, depending on its precise timing in relation to the ongoing autoimmune process. In a transgenic model (rat insulin promoter-lymphocytic choriomeningitis virus) of virally induced diabetes, TNF-alpha enhanced disease incidence when induced through an islet-specific tetracycline-dependent promoter system early during pathogenesis. Blockade of TNF-alpha during this phase prevented diabetes completely, suggesting its pathogenetic importance early in disease development. In contrast, TNF-alpha expression abrogated the autoimmune process when induced late, which was associated with a reduction of autoreactive CD8 lymphocytes in islets and their lytic activities. Thus, the fine-tuned kinetics of an autoreactive process undergo distinct stages that respond in a differential way to the presence of TNF-alpha. This observation has importance for understanding the complex role of inflammatory cytokines in autoimmunity.
TL;DR: The results demonstrate that PP are not an absolute requirement for the induction of either high‐ or low‐dose oral tolerance, although oral tolerance could not be induced in animals devoid of both PP and MLN.
Abstract: Systemic hyporesponsiveness occurs following oral administration of antigen (oral tolerance) and involves the uptake and processing of antigen by the gut-associated lymphoid tissue (GALT), which includes Peyer's patches (PP) lamina propria lymphocytes and mesenteric lymph nodes (MLN). Animals with targeted mutations of genes in the tumor necrosis factor (TNF) family have differential defects in the development of peripheral lymphoid organs including PP and MLN, and provide a unique opportunity to investigate the role of GALT structures in the induction of oral tolerance. Oral tolerance could not be induced in TNF/lymphotoxin (LT) α–/– mice, which are devoid of both PP and MLN, although these animals could be tolerized by intraperitoneal administration of antigen, demonstrating the requirement for GALT for oral tolerance induction. LTβ–/– mice and LTα/LTβ+/– animals do not have PP but could be orally tolerized, as measured by IFN-γ production and delayed-type hypersensitivity responses by administration of both low or high doses of ovalbumin. To further investigate the requirement for PP, we tested the progeny of LTβ-receptor-IgG-fusion-protein (LTβRigG)-treated mice, which do not form PP but have an otherwise intact immune system. Although these animals had decreased fecal IgA production, they could be orally tolerized. Our results demonstrate that PP are not an absolute requirement for the induction of either high- or low-dose oral tolerance, although oral tolerance could not be induced in animals devoid of both PP and MLN.
TL;DR: Comparing signaling characteristics of 9 single-chain receptors consisting of the H-2K(b) extracellular and transmembrane domains and various combinations of T cell signal transduction domains shows that it is possible to link TCR, coreceptor, and costimulatory activities in a single functional entity using modular domains.
TL;DR: This work presents a novel probabilistic construct called a “spatially aggregating immune checkpoints” that are able to be distinguished between the immune checkpoints of the immune system and other immune checkpoints.
Abstract: Immunobiology Program, Department of Medicine, University of Vermont, Burlington, Vermont, VT 05405, USA; Howard Hughes Medical Institute and Section of Immunobiology, Yale University School of Medicine, New Haven, Connecticut CT 06520, USA; Howard Hughes Medical Institute and Program in Molecular Medicine, Department of Biochemistry & Molecular Biology, University of Massachussetts Medical School, Worcester, Massachusetts, MA 01605, USA
TL;DR: The results indicate that Rac2 is important in mediating both transcriptional and cytoskeletal changes during T cell activation, and the phenotypic similarity of Rac2−/− to Vav −/− cells implicates Rac2 as a downstream mediator of Vav signaling.
Abstract: Rac2 is a hematopoietic-specific GTPase acting as a molecular switch to mediate both transcriptional activation and cell morphological changes. We have examined the effect of Rac2 deficiency during T cell activation. In Rac2−/− T cells, proliferation was reduced upon stimulation with either plate-bound anti-CD3 or T cell receptor–specific antigen. This defect is accompanied with decreased activation of mitogen activated protein kinase extracellular signal–regulated kinase (ERK)1/2 and p38, and reduced Ca2+ mobilization. TCR stimulation–induced actin polymerization is also reduced. In addition, anti-CD3 cross-linking–induced T cell capping is reduced compared with wild-type T cells. These results indicate that Rac2 is important in mediating both transcriptional and cytoskeletal changes during T cell activation. The phenotypic similarity of Rac2−/− to Vav−/− cells implicates Rac2 as a downstream mediator of Vav signaling.
TL;DR: These studies indicate that chloroquine activates a regulated cell death pathway that partially overlaps with the apoptotic cascade, and is used as a model of lysosomal-dependent cell death.
Abstract: Chloroquine is a lysosomotropic agent that causes marked changes in intracellular protein processing and traf- ficking and extensive autophagic vacuole formation. Chloroquine may be cytotoxic and has been used as a model of lysosomal- dependent cell death. Recent studies indicate that autophagic cell death may involve Bcl-2 family members and share some features with caspase-dependent apoptotic death. To determine the molecular pathway of chloroquine-induced neuronal cell death, we examined the effects of chloroquine on primary telencephalic neuronal cultures derived from mice with targeted gene disruptions in p53, and various caspase and bcl-2 family members. In wild-type neurons, chloroquine produced con- centration- and time-dependent accumulation of autophagosomes, caspase-3 activation, and cell death. Cell death was inhibited by 3-methyladenine, an inhibitor of autophagic vacuole formation, but not by Boc-Asp-FMK (BAF), a broad caspase inhibitor. Targeted gene disruptions of p53 and bax inhibited and bcl-x potentiated chloroquine-induced neuron death. Caspase-9- and caspase-3-deficient neurons were not protected from chloroquine cytotoxicity. These studies indicate that chloroquine activates a regulated cell death pathway that partially overlaps with the apoptotic cascade.
TL;DR: Overall, these results indicate that JNK3 plays a critical role in activation of c‐Jun and apoptosis in a classic model of cell‐autonomous programmed neuron death.
Abstract: The stress activated protein kinase pathway culminates in c-Jun phosphorylation mediated by the Jun Kinases (JNKs). The role of the JNK pathway in sympathetic neuronal death is unclear in that apoptosis is not inhibited by a dominant negative protein of one JNK kinase, SEK1, but is inhibited by CEP-1347, a compound known to inhibit this overall pathway but not JNKs per se. To evaluate directly the apoptotic role of the JNK isoform that is selectively expressed in neurons, JNK3, we isolated sympathetic neurons from JNK3-deficient mice and quantified nerve growth factor (NGF) deprivation-induced neuronal death, oxidative stress, c-Jun phosphorylation, and c-jun induction. Here, we report that oxidative stress in neurons from JNK3-deficient mice is normal after NGF deprivation. In contrast, NGF-deprivation-induced increases in the levels of phosphorylated c-Jun, c-jun, and apoptosis are each inhibited in JNK3-deficient mice. Overall, these results indicate that JNK3 plays a critical role in activation of c-Jun and apoptosis in a classic model of cell-autonomous programmed neuron death.
TL;DR: Results indicate a transition from Caspase-9- to Bax- and Bcl-XL-mediated neuronal apoptosis, which contrast with the previous investigation of AraC-induced apoptosis of telencephalic neural precursor cells in which death was completely blocked by p53 or Casp enzyme-9 deficiency but not Bax deficiency.
Abstract: Programmed cell death is critical for normal nervous system development and is regulated by Bcl-2 and Caspase family members. Targeted disruption of bcl-x(L), an antiapoptotic bcl-2 gene family member, causes massive death of immature neurons in the developing nervous system whereas disruption of caspase-9, a proapoptotic caspase gene family member, leads to decreased neuronal apoptosis and neurodevelopmental abnormalities. To determine whether Bcl-X(L) and Caspase-9 interact in an obligate pathway of neuronal apoptosis, bcl-x/caspase-9 double homozygous mutants were generated. The increased apoptosis of immature neurons observed in Bcl-X(L)-deficient embryos was completely prevented by concomitant Caspase-9 deficiency. In contrast, bcl-x(-/-)/caspase-9(-/-) embryonic mice exhibited an expanded ventricular zone and neuronal malformations identical to that observed in mice lacking only Caspase-9. These results indicate both epistatic and independent actions of Bcl-X(L) and Caspase-9 in neuronal programmed cell death. To examine Bcl-2 and Caspase family-dependent apoptotic pathways in telencephalic neurons, we compared the effects of cytosine arabinoside (AraC), a known neuronal apoptosis inducer, on wild-type, Bcl-X(L)-, Bax-, Caspase-9-, Caspase-3-, and p53-deficient telencephalic neurons in vitro. AraC caused extensive apoptosis of wild-type and Bcl-X(L)-deficient neurons. p53- and Bax-deficient neurons showed marked protection from AraC-induced death, whereas Caspase-9- and Caspase-3-deficient neurons showed minimal or no protection, respectively. These findings contrast with our previous investigation of AraC-induced apoptosis of telencephalic neural precursor cells in which death was completely blocked by p53 or Caspase-9 deficiency but not Bax deficiency. In total, these results indicate a transition from Caspase-9- to Bax- and Bcl-X(L)-mediated neuronal apoptosis.
TL;DR: Advances in understanding the expression, function, and regulation of the JNK pathway in T-lymphocyte activation and differentiation are summarized.
Abstract: c-Jun N-terminal kinase (JNK) is one of the several main MAP kinase groups identified in mammals. Original studies by use of Jurkat T cells implicated JNK in T cell activation and interleukin (IL-2) expression. Recent advances using mouse genetic approaches have revealed novel functions of this pathway in primary mouse T cells. JNK is not essential for T-cell activation; instead, it is required for helper T differentiation into effector cells and their cytokine production. In this review, we summarize these advances in understanding the expression, function, and regulation of the JNK pathway in T-lymphocyte activation and differentiation.
TL;DR: The results suggest that CD62L is not required for activation of autoimmune CD4 T cells but is important for the final destructive function of effector cells in the CNS and support a novel mechanism wherebyCD62L expressed on effector Cells is important in mediating myelin damage.
[...]
TL;DR: Cavitation, an early developmental process in the mouse embryo that requires cell death, has now been suggested to occur through a caspase-independent pathway that instead uses a mitochondrial protein called AIF (apoptosis-inducing factor).
Abstract: Hunot and Flavell
discuss recent findings in
Nature
by Joza
et al. that point to the existence of a new sort of cell death. Cell death is required during the development of an organism to remove portions of the body that will not be needed; this cell death usually requires destructive enzymes called caspases to perform the final demolition of the cell. But cavitation, an early developmental process in the mouse embryo that requires cell death, has now been suggested to occur through a caspase-independent pathway that instead uses a mitochondrial protein called AIF (apoptosis-inducing factor).
TL;DR: It is reported that LNs are absolutely required for generating contact hypersensitivity, a T cell–dependent cellular immune response induced by epicutaneous hapten, and reversed the LN defect in lymphotoxin-α−/− mice, thereby restoring the capacity for contact hypers sensitivity.
Abstract: Lymph nodes (LNs) are important sentinal organs, populated by circulating lymphocytes and antigen-bearing cells exiting the tissue beds. Although cellular and humoral immune responses are induced in LNs by antigenic challenge, it is not known if LNs are essential for acquired immunity. We examined immune responses in mice that lack LNs due to genetic deletion of lymphotoxin ligands or in utero blockade of membrane lymphotoxin. We report that LNs are absolutely required for generating contact hypersensitivity, a T cell–dependent cellular immune response induced by epicutaneous hapten. We show that the homing of epidermal Langerhans cells in response to hapten application is specifically directed to LNs, providing a cellular basis for this unique LN function. In contrast, the spleen cannot mediate contact hypersensitivity because antigen-bearing epidermal Langerhans cells do not access splenic white pulp. Finally, we formally demonstrate that LNs provide a unique environment essential for generating this acquired immune response by reversing the LN defect in lymphotoxin-α−/− mice, thereby restoring the capacity for contact hypersensitivity.