RESUMO
Ciliogenesis precedes lineage-determining signaling in skin development. To understand why, we performed shRNA-mediated knockdown of seven intraflagellar transport proteins (IFTs) and conditional ablation of Ift-88 and Kif3a during embryogenesis. In both cultured keratinocytes and embryonic epidermis, all of these eliminated cilia, and many (not Kif3a) caused hyperproliferation. Surprisingly and independent of proliferation, ciliary mutants displayed defects in Notch signaling and commitment of progenitors to differentiate. Notch receptors and Notch-processing enzymes colocalized with cilia in wild-type epidermal cells. Moreover, differentiation defects in ciliary mutants were cell autonomous and rescued by activated Notch (NICD). By contrast, Shh signaling was neither operative nor required for epidermal ciliogenesis, Notch signaling, or differentiation. Rather, Shh signaling defects in ciliary mutants occurred later, arresting hair follicle morphogenesis in the skin. These findings unveil temporally and spatially distinct functions for primary cilia at the nexus of signaling, proliferation, and differentiation.
Assuntos
Diferenciação Celular , Cílios/metabolismo , Epiderme/embriologia , Epiderme/metabolismo , Receptores Notch/metabolismo , Transdução de Sinais , Animais , Proteínas de Transporte/genética , Polaridade Celular , Proliferação de Células , Proteínas do Citoesqueleto/metabolismo , Células Epidérmicas , Técnicas de Silenciamento de Genes , Folículo Piloso/citologia , Proteínas Hedgehog/metabolismo , Cinese , Camundongos , Proteínas Supressoras de Tumor/metabolismoRESUMO
In Fig. 2g of this Article, a panel was inadvertently duplicated. The 'D30 IMQ' image was a duplicate of the 'D6 Ctrl' image. Fig. 2g has been corrected online to show the correct 'D30 IMQ' image (showing skin inflammation induced by the NALP3 agonist imiquimod, IMQ). The Supplementary Information to this Amendment contains the old, incorrect Fig. 2 for transparency.
RESUMO
The skin barrier is the body's first line of defence against environmental assaults, and is maintained by epithelial stem cells (EpSCs). Despite the vulnerability of EpSCs to inflammatory pressures, neither the primary response to inflammation nor its enduring consequences are well understood. Here we report a prolonged memory to acute inflammation that enables mouse EpSCs to hasten barrier restoration after subsequent tissue damage. This functional adaptation does not require skin-resident macrophages or T cells. Instead, EpSCs maintain chromosomal accessibility at key stress response genes that are activated by the primary stimulus. Upon a secondary challenge, genes governed by these domains are transcribed rapidly. Fuelling this memory is Aim2, which encodes an activator of the inflammasome. The absence of AIM2 or its downstream effectors, caspase-1 and interleukin-1ß, erases the ability of EpSCs to recollect inflammation. Although EpSCs benefit from inflammatory tuning by heightening their responsiveness to subsequent stressors, this enhanced sensitivity probably increases their susceptibility to autoimmune and hyperproliferative disorders, including cancer.
Assuntos
Células Epiteliais/citologia , Inflamação/genética , Inflamação/patologia , Pele/citologia , Pele/patologia , Células-Tronco/citologia , Cicatrização/fisiologia , Aminoquinolinas/farmacologia , Animais , Doenças Autoimunes/patologia , Caspase 1/metabolismo , Linhagem da Célula , Cromatina/genética , Proteínas de Ligação a DNA/deficiência , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Epigênese Genética/efeitos dos fármacos , Epigênese Genética/genética , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Feminino , Imiquimode , Inflamassomos/metabolismo , Inflamação/induzido quimicamente , Inflamação/imunologia , Interleucina-1beta/metabolismo , Macrófagos , Camundongos , Neoplasias/patologia , Regeneração/efeitos dos fármacos , Regeneração/genética , Pele/efeitos dos fármacos , Pele/imunologia , Células-Tronco/efeitos dos fármacos , Células-Tronco/metabolismo , Estresse Fisiológico/genética , Linfócitos T , Cicatrização/efeitos dos fármacos , Cicatrização/genéticaRESUMO
Proteins that directly regulate tumour necrosis factor receptor (TNFR) signalling have critical roles in regulating cellular activation and survival. ABIN-1 (A20 binding and inhibitor of NF-kappaB) is a novel protein that is thought to inhibit NF-kappaB signalling. Here we show that mice deficient for ABIN-1 die during embryogenesis with fetal liver apoptosis, anaemia and hypoplasia. ABIN-1 deficient cells are hypersensitive to tumour necrosis factor (TNF)-induced programmed cell death, and TNF deficiency rescues ABIN-1 deficient embryos. ABIN-1 inhibits caspase 8 recruitment to FADD (Fas-associated death domain-containing protein) in TNF-induced signalling complexes, preventing caspase 8 cleavage and programmed cell death. Moreover, ABIN-1 directly binds polyubiquitin chains and this ubiquitin sensing activity is required for ABIN-1's anti-apoptotic activity. These studies provide insights into how ubiquitination and ubiquitin sensing proteins regulate cellular and organismal survival.
Assuntos
Apoptose/fisiologia , Proteínas de Ligação a DNA/metabolismo , Desenvolvimento Embrionário/fisiologia , Ubiquitina/metabolismo , Proteínas Adaptadoras de Transdução de Sinal , Sequência de Aminoácidos , Animais , Linhagem Celular , Proteínas de Ligação a DNA/química , Desenvolvimento Embrionário/genética , Regulação da Expressão Gênica no Desenvolvimento , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/química , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Células Jurkat , Camundongos , Camundongos Knockout , Dados de Sequência Molecular , Alinhamento de Sequência , Fator de Necrose Tumoral alfa/metabolismoRESUMO
The high affinity interleukin (IL)-15 receptor, IL-15Ralpha, is essential for supporting lymphoid homeostasis. To assess whether IL-15Ralpha's role in vivo is to trans present IL-15, we generated mixed bone marrow chimera from IL-15Ralpha- and IL-2/15Rbeta-deficient mice. We find that IL-15Ralpha-competent, IL-2/15Rbeta-deficient cells are able to support IL-15Ralpha-deficient natural killer (NK) and memory CD8+ T cells, thus ruling out secondary signals on these cells and demonstrating that IL-15Ralpha-mediated presentation of IL-15 in trans is the primary mechanism by which IL-15Ralpha functions in vivo. Surprisingly, using IL-15- and IL-15Ralpha-deficient mixed chimera, we also find that IL-15 and IL-15Ralpha must be expressed by the same cells to present IL-15 in trans, indicating that IL-15Ralpha is required on a cellular level for the elaboration of IL-15. These studies indicate that IL-15Ralpha defines homeostatic niches for NK and memory CD8+ T cells by controlling both the production and the presentation of IL-15 in trans to NK and CD8+ memory T cells.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Homeostase/imunologia , Interleucina-15/metabolismo , Células Matadoras Naturais/imunologia , Receptores de Interleucina-2/metabolismo , Animais , Células da Medula Óssea , Citometria de Fluxo , Memória Imunológica/imunologia , Interleucina-15/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Modelos Imunológicos , Quimera por Radiação , Receptores de Interleucina-15 , Receptores de Interleucina-2/imunologiaRESUMO
Natural killer (NK) cells protect hosts against viral pathogens and transformed cells. IL-15 is thought to play a critical role in NK cell development, but its role in the regulation of peripheral NK cells is less well defined. We now find that adoptive transfer of normal NK cells into mice lacking the high affinity interleukin (IL)-15 receptor, IL-15Ralpha, surprisingly results in the abrupt loss of these cells. Moreover, IL-15Ralpha-deficient NK cells can differentiate successfully in radiation bone marrow chimera bearing normal cells. Finally, adoptively transferred IL-15Ralpha-deficient NK cells survive in normal but not IL-15Ralpha-deficient mice. These findings demonstrate that NK cell-independent IL-15Ralpha expression is critical for maintaining peripheral NK cells, while IL-15Ralpha expression on NK cells is not required for this function.
Assuntos
Sobrevivência Celular/fisiologia , Células Matadoras Naturais/metabolismo , Receptores de Interleucina-2/metabolismo , Transferência Adotiva , Animais , Homeostase , Interleucina-15/genética , Interleucina-15/metabolismo , Células Matadoras Naturais/citologia , Camundongos , Camundongos Endogâmicos , Camundongos Knockout , Quimera por Radiação/fisiologia , Receptores de Interleucina-15 , Receptores de Interleucina-2/genética , Transdução de Sinais/fisiologia , Baço/citologiaRESUMO
Interleukin-15 (IL-15) is a cytokine that plays unique roles in both innate and adaptive immune cell homeostasis. While early studies suggested that IL-15 resembled IL-2, more recent work suggests that IL-15 may play multiple unique roles in immune homeostasis befitting its pleiotropic expression pattern. This review will focus on recent studies that highlight some of these functions.
Assuntos
Interleucina-15/fisiologia , Linfócitos/imunologia , Animais , Homeostase , Memória Imunológica , Interleucina-2/fisiologia , Ativação Linfocitária , Receptores de Interleucina-15 , Receptores de Interleucina-2/metabolismo , Transdução de Sinais , Linfócitos T/imunologiaRESUMO
During mouse development, core planar cell polarity (PCP) proteins become polarized in the epidermal plane to guide angling/morphogenesis of hair follicles. How PCP is established is poorly understood. Here, we identify a key role for Wdr1 (also known as Aip1), an F-actin-binding protein that enhances cofilin/destrin-mediated F-actin disassembly. We show that cofilin and destrin function redundantly in developing epidermis, but their combined depletion perturbs cell adhesion, cytokinesis, apicobasal polarity and PCP. Although Wdr1 depletion accentuates single-loss-of-cofilin/destrin phenotypes, alone it resembles core PCP mutations. Seeking a mechanism, we find that Wdr1 and cofilin/destrin-mediated actomyosin remodelling are essential for generating or maintaining cortical tension within the developing epidermal sheet and driving the cell shape and planar orientation changes that accompany establishment of PCP in mammalian epidermis. Our findings suggest intriguing evolutionary parallels but mechanistic modifications to the distal wing hinge-mediated mechanical forces that drive cell shape change and orient PCP in the Drosophila wing disc.
Assuntos
Polaridade Celular , Forma Celular , Epiderme/metabolismo , Queratinócitos/metabolismo , Mecanotransdução Celular , Proteínas dos Microfilamentos/metabolismo , Fatores de Despolimerização de Actina/metabolismo , Actomiosina/metabolismo , Animais , Adesão Celular , Linhagem Celular , Citocinese , Citoesqueleto/metabolismo , Destrina/deficiência , Destrina/genética , Células Epidérmicas , Evolução Molecular , Genótipo , Terapia a Laser , Camundongos Knockout , Proteínas dos Microfilamentos/genética , Microscopia de Vídeo , Fenótipo , Transporte Proteico , Interferência de RNA , Estresse Mecânico , Fatores de Tempo , TransfecçãoRESUMO
The intestinal lamina propria contains lymphocytes that are chronically activated by exposure to luminal antigens. Dysregulation of these cells is thought to be central to the pathogenesis of bowel inflammation in experimental models of inflammatory bowel disease. CD28 signals on peripheral T cells provide important costimulatory signals that enhance T-cell proliferation and activation responses to antigens. However, the role of CD28 signals in lamina propria T cells or models of inflammatory bowel disease have not been determined. Accordingly, we examined T lymphocyte activation and proliferation in CD28-deficient (CD28-/-) mice to examine the in vivo roles of CD28 signals in lamina propria T-cell homeostasis. We further generated CD28-/- interleukin (IL)-2-/- double mutant mice to assess the role of CD28 signals in supporting the spontaneously activated and pathogenic T cells that accumulate in IL-2-/- mice. CD28-/- lamina propria T cells displayed reduced activation markers, but were present in normal numbers and proliferated normally. IL-2-/- lymphocytes expressed high levels of bcl-xL, whereas CD28-/- IL-2-/- cells had substantially less bcl-xL. However, lymphadenopathy and ulcerative colitis-like disease occurred in both IL-2-/- and CD28-/- IL-2-/- mice. Thus, CD28 provides a functional costimulatory signal to lamina propria T cells but is not required for homeostasis of these cells. In addition, neither CD28 nor bcl-xL appears to be required for the spontaneous accumulation of T cells in IL-2-/- mice. This suggests that other costimulatory molecules or T-cell receptor ligation alone drive lymphocyte expansion in IL-2-deficient mice.
Assuntos
Antígenos CD28/genética , Antígenos CD28/fisiologia , Colite/genética , Colite/fisiopatologia , Interleucina-2/deficiência , Interleucina-2/fisiologia , Ativação Linfocitária/genética , Ativação Linfocitária/fisiologia , Animais , Modelos Animais de Doenças , Interleucina-2/genética , Mucosa Intestinal/fisiopatologia , Camundongos , Camundongos Endogâmicos C57BL , Transdução de Sinais/genética , Transdução de Sinais/fisiologia , Linfócitos T/fisiologiaRESUMO
NK cells protect hosts against viral pathogens and transformed cells, and dendritic cells (DCs) play important roles in activating NK cells. We now find that murine IL-15Ralpha-deficient DCs fail to support NK cell cytolytic activity and elaboration of IFN-gamma, despite the fact that these DCs express normal levels of costimulatory molecules and IL-12. By contrast, IL-15Ralpha expression on NK cells is entirely dispensable for their activation by DCs. In addition, blockade with anti-IL-15Ralpha and anti-IL-2Rbeta but not anti-IL-2Ralpha-specific Abs prevents NK cell activation by wild-type DCs. Finally, presentation of IL-15 by purified IL-15Ralpha/Fc in trans synergizes with IL-12 to support NK cell priming. These findings suggest that murine DCs require IL-15Ralpha to present IL-15 in trans to NK cells during NK cell priming.
Assuntos
Células Matadoras Naturais/imunologia , Células Matadoras Naturais/metabolismo , Ativação Linfocitária/imunologia , Subunidades Proteicas/fisiologia , Receptores de Interleucina-2/fisiologia , Animais , Anticorpos Bloqueadores/farmacologia , Células da Medula Óssea/citologia , Células da Medula Óssea/imunologia , Células da Medula Óssea/metabolismo , Diferenciação Celular/genética , Diferenciação Celular/imunologia , Sistema Livre de Células/imunologia , Células Cultivadas , Técnicas de Cocultura , Células Dendríticas/citologia , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Humanos , Imunossupressores/farmacologia , Interleucina-15/metabolismo , Interleucina-15/fisiologia , Ativação Linfocitária/genética , Camundongos , Camundongos Congênicos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Subunidades Proteicas/antagonistas & inibidores , Subunidades Proteicas/imunologia , Subunidades Proteicas/metabolismo , Receptores de Interleucina-15 , Receptores de Interleucina-2/antagonistas & inibidores , Receptores de Interleucina-2/imunologia , Receptores de Interleucina-2/metabolismoRESUMO
IL-2 receptor alpha-deficient (IL2Ralpha-/-) mice spontaneously accumulate vast numbers of intestinal lamina propria (LP) T cells and develop bowel inflammation. The accumulation of T cells in IL2Ralpha-/- mice is thought to result, in part, from defective Fas-induced cell death. To understand the role of cell proliferation and death in regulating LP T cells in IL2Ralpha-/- mice, we have directly examined the proliferation and Fas sensitivity of wild-type, lpr/lpr, and IL2Ralpha-/- LP T cells. In wild-type mice, 5'-bromodeoxyuridine (BrdU) labeling and Fas susceptibility are greatest in CD44Hi LP T cells. Fas-deficient lpr/lpr mice have normal total numbers of LP T cells, despite an increased proportion of BrdU+ T cells. By contrast, IL2Ralpha-/- mice possess increased total numbers of LP T cells, despite normal proportions of BrdU+ LP T cells. Finally, wild-type and IL2Ralpha-/- LP T cells are equivalently Fas sensitive. These results demonstrate that LP T cells proliferate and are Fas-sensitive cells. IL2Ralpha-/- mice accumulate a large number of these Fas-sensitive LP T cells and clearly differ from Fas-deficient lpr/lpr mice in this regard. Thus our studies reveal that Fas is dispensable for LP T cell homeostasis and suggest that the intestinal inflammation observed in IL2Ralpha-/- mice is independent of defective Fas-induced cell death.
Assuntos
Intestinos/patologia , Linfócitos T/fisiologia , Receptor fas/fisiologia , Animais , Apoptose , Bromodesoxiuridina/metabolismo , Divisão Celular , Homeostase , Receptores de Hialuronatos/análise , Hipertrofia , Doenças Inflamatórias Intestinais/etiologia , Doenças Inflamatórias Intestinais/patologia , Mucosa Intestinal/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptores de Interleucina-2/deficiência , Receptores de Interleucina-2/fisiologia , Linfócitos T/imunologia , Linfócitos T/patologia , Timectomia , Receptor fas/genéticaRESUMO
The generation and maintenance of immunological memory requires the activation, expansion, and persistent proliferation of antigen-specific T cells. Recent work suggests that IL-15 may be important for this process. Surprisingly, we now find that expression of the high-affinity receptor for IL-15, IL-15R alpha, on T cells is dispensable for the generation or maintenance of memory CD8(+) T cells. By contrast, IL-15R alpha expression on cells other than T cells is absolutely critical for this function. These findings may be related to IL-15R alpha's ability to present IL-15 in trans to low-affinity IL-15R beta gamma(c) receptors on memory CD8(+) T cells. These unexpected results provide insights into how IL-15R alpha supports memory CD8(+) T cells.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Memória Imunológica , Receptores de Interleucina-2/metabolismo , Adjuvantes Imunológicos/farmacologia , Transferência Adotiva , Animais , Linfócitos T CD8-Positivos/citologia , Linfócitos T CD8-Positivos/efeitos dos fármacos , Divisão Celular , Camundongos , Camundongos Congênicos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Poli I-C/farmacologia , Quimera por Radiação , Tolerância a Radiação/imunologia , Receptores de Interleucina-15 , Receptores de Interleucina-2/deficiência , Receptores de Interleucina-2/genética , Transdução de SinaisRESUMO
A20 is a cytoplasmic protein required for the termination of tumor necrosis factor (TNF)-induced signals. We show here that mice doubly deficient in either A20 and TNF or A20 and TNF receptor 1 developed spontaneous inflammation, indicating that A20 is also critical for the regulation of TNF-independent signals in vivo. A20 was required for the termination of Toll-like receptor-induced activity of the transcription factor NF-kappaB and proinflammatory gene expression in macrophages, and this function protected mice from endotoxic shock. A20 accomplished this biochemically by directly removing ubiquitin moieties from the signaling molecule TRAF6. The critical function of this deubiquitinating enzyme in the restriction of TLR signals emphasizes the importance of the regulation of ubiquitin conjugation in innate immune cells.