RESUMO
Terminal differentiation of B cells into antibody-secreting cells is the foundation of humoral immune response. B-1 cells, which are different from B-2 cells, preferentially differentiate into plasma cells. CMTM7 is a MARVEL-domain-containing membrane protein predominantly expressed in B cells that plays an important role in B-1a cell development. The present study assessed CMTM7 function in response to antigen stimulation. Following immunization with T cell-dependent and T cell-independent antigens, Cmtm7-deficient mice exhibited decreased IgM but normal IgG responses in vivo. In vitro stimulation with LPSs induced Cmtm7-/- B-1 cell activation, whereas proliferation was marginally reduced. Notably, Cmtm7 deficiency markedly suppressed plasma cell differentiation in response to TLR agonists, accompanied by a decrease in IgM and IL-10 production. At the molecular level, loss of Cmtm7 repressed the downregulation of Pax5 and the upregulation of Xbp1, Irf4, and Prdm1. Furthermore, p38 phosphorylation was inhibited in Cmtm7-/- B-1 cells. Experiments using a p38 inhibitor revealed that p38 activation was essential for the terminal differentiation of B-1 cells, suggesting that Cmtm7 contributes to B-1 cell differentiation by maintaining p38 activation. Overall, the data reveal the crucial functions of CMTM7 in TLR-induced terminal differentiation and p38 activation in B-1 cells.
Assuntos
Subpopulações de Linfócitos B/imunologia , Diferenciação Celular/imunologia , Quimiocinas/imunologia , Sistema de Sinalização das MAP Quinases/imunologia , Proteínas com Domínio MARVEL/imunologia , Plasmócitos/imunologia , Receptores Toll-Like/imunologia , Proteínas Quinases p38 Ativadas por Mitógeno/imunologia , Animais , Subpopulações de Linfócitos B/citologia , Diferenciação Celular/genética , Quimiocinas/genética , Ativação Enzimática/genética , Ativação Enzimática/imunologia , Imunoglobulina M/genética , Imunoglobulina M/imunologia , Fatores Reguladores de Interferon/genética , Fatores Reguladores de Interferon/imunologia , Interleucina-10/genética , Interleucina-10/imunologia , Sistema de Sinalização das MAP Quinases/genética , Proteínas com Domínio MARVEL/genética , Camundongos , Camundongos Knockout , Plasmócitos/citologia , Fator 1 de Ligação ao Domínio I Regulador Positivo/genética , Fator 1 de Ligação ao Domínio I Regulador Positivo/imunologia , Receptores Toll-Like/genética , Proteínas Quinases p38 Ativadas por Mitógeno/genéticaRESUMO
BACKGROUND: Chordomas are rare, slow-growing and locally aggressive bone sarcomas. At present, chordomas are difficult to manage due to their high recurrence rate, metastasis tendency and poor prognosis. The underlying mechanisms of chordoma tumorigenesis and progression urgently need to be explored to find the effective therapeutic targets. Our previous data demonstrates that EGFR plays important roles in chordoma development and CKLF-like MARVEL transmembrane domain containing (CMTM)3 suppresses gastric cancer metastasis by inhibiting the EGFR/STAT3/EMT signaling pathway. However, the roles and mechanism of CMTM3 in chordomas remain unknown. METHODS: Primary chordoma tissues and the paired adjacent non-tumor tissues were collected to examine the expression of CMTM3 by western blot. The expression of CMTM3 in chordoma cell lines was tested by Real-time PCR and western blot. CCK-8 and colony forming unit assay were performed to delineate the roles of CMTM3 in cell proliferation. Wound healing and Transwell assays were performed to assess cell migration and invasion abilities. A xenograft model in NSG mice was used to elucidate the function of CMTM3 in vivo. Signaling pathways were analyzed by western blot and IHC. RNA-seq was performed to further explore the mechanism regulated by CMTM3 in chordoma cells. RESULTS: CMTM3 expression was downregulated in chordoma tissues compared with paired normal tissues. CMTM3 suppressed proliferation, migration and invasion of chordoma cells in vitro and inhibited tumor growth in vivo. CMTM3 accelerated EGFR degradation, suppressed EGFR/STAT3/EMT signaling pathway, upregulated TP53 expression and enriched the TP53 signaling pathway in chordoma cells. CONCLUSIONS: CMTM3 inhibited tumorigenesis and development of chordomas through activating the TP53 signaling pathway and suppressing the EGFR/STAT3 signaling pathway, which suppressed EMT progression. CMTM3 might be a potential therapeutic target for chordomas.
RESUMO
B-1a cells represent a distinct B cell population with unique phenotype, self-renewing capacity and restricted Igµ repertoire. They primarily locate in body cavity and also exist in spleen. The different subpopulations of B-1a cells are heavily affected by local environment. Our previous studies revealed that MARVEL-domain-containing membrane protein, CMTM7, was involved in B-1a cell development. Here, we focused its influence on peritoneal and splenic B-1a cells. Unlike peritoneal B-1a cells, we found that splenic Cmtm7-/- B-1a cells expressed higher level of CD5, CD80 and CD86 compared with WT counterparts. They also exhibited an enhanced tonic BCR signals in steady state. Though the cell viability was unaffected in vitro, Cmtm7 knockout markedly promoted splenic B-1a cell apoptosis in situ, which was likely associated with down-regulation of Il-5rα. With regard to Igµ repertoire, peritoneal and splenic Cmtm7-/- B-1a cells exhibit similar changes exemplified by the loss of VH11 and gain of VH12, whereas an increase in VH1 usage and skewed J segments from JH1 to JH2 and JH4 families could only be detected within splenic Cmtm7-/- B-1a cells. Overall, these data indicate that Cmtm7 functions differently in peritoneal and splenic B-1a cells and plays a more important role in splenic cells.
Assuntos
Subpopulações de Linfócitos B/metabolismo , Quimiocinas/metabolismo , Proteínas com Domínio MARVEL/metabolismo , Receptores de Antígenos de Linfócitos B/metabolismo , Animais , Apoptose/imunologia , Subpopulações de Linfócitos B/imunologia , Diferenciação Celular/imunologia , Membrana Celular/metabolismo , Proliferação de Células , Quimiocinas/imunologia , Feminino , Proteínas com Domínio MARVEL/genética , Proteínas com Domínio MARVEL/imunologia , Masculino , Proteínas de Membrana/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Fenótipo , Receptores de Antígenos de Linfócitos B/imunologia , Transdução de Sinais/imunologia , Baço/imunologia , Baço/patologiaRESUMO
Innate-like B-1a cells are an important cell population for production of natural IgM and interleukin-10 (IL-10), and act as the first line against pathogens. We determined that CMTM7 is essential for B-1a cell development. Following Cmtm7 (CKLF-like MARVEL transmembrane domain-containing 7) knockout, B-1a cell numbers decreased markedly in all investigated tissues. Using a bone marrow and fetal liver adoptive transfer model and conditional knockout mice, we showed that the reduction of B-1a cells resulted from B-cell-intrinsic defects. Because of B-1a cell loss, Cmtm7-deficient mice produced less IgM and IL-10, and were more susceptible to microbial sepsis. Self-renewal and homeostasis of mature B-1a cells in Cmtm7-/- mice were not impaired, suggesting the effect of Cmtm7 on B-1a cell development. Further investigations demonstrated that the function of Cmtm7 in B-1a cell development occurred at the specific transitional B-1a (TrB-1a) stage. Cmtm7 deficiency resulted in a slow proliferation and high cell death rate of TrB-1a cells. Thus, Cmtm7 controls B-1a cell development at the transitional stage.
Assuntos
Quimiocinas/imunologia , Proteínas com Domínio MARVEL/imunologia , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Animais , Subpopulações de Linfócitos B/imunologia , Morte Celular , Proliferação de Células , Quimiocinas/deficiência , Proteínas com Domínio MARVEL/deficiência , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas Serina-Treonina Quinases/imunologiaRESUMO
BACKGROUND: Obesity plays crucial roles in the development of cardiovascular diseases. However, the mechanisms that link obesity and cardiovascular diseases remain elusive. Compelling evidence indicates that adipokines play an important role in obesity-related cardiovascular diseases. Here, we found a new adipokine-named family with sequence similarity 19, member A5 (FAM19A5), a protein with unknown function that was predicted to be distantly related to the CC-chemokine family. We aimed to test whether adipose-derived FAM19A5 regulates vascular pathology on injury. METHODS: DNA cloning, protein expression, purification, and N-terminal sequencing were applied to characterize FAM19A5. Adenovirus infection and siRNA transfection were performed to regulate FAM19A5 expression. Balloon and wire injury were performed in vivo on the rat carotid arteries and mouse femoral arteries, respectively. Bioinformatics analysis, radioactive ligand-receptor binding assays, receptor internalization, and calcium mobilization assays were used to identify the functional receptor for FAM19A5. RESULTS: We first characterized FAM19A5 as a secreted protein, and the first 43 N-terminal amino acids were the signal peptides. Both FAM19A5 mRNA and protein were abundantly expressed in the adipose tissue but were downregulated in obese mice. Overexpression of FAM19A5 markedly inhibited vascular smooth muscle cell proliferation and migration and neointima formation in the carotid arteries of balloon-injured rats. Accordingly, FAM19A5 silencing in adipocytes significantly promoted vascular smooth muscle cell activation. Adipose-specific FAM19A5 transgenic mice showed greater attenuation of neointima formation compared with wild-type littermates fed with or without Western-style diet. We further revealed that sphingosine-1-phosphate receptor 2 was the functional receptor for FAM19A5, with a dissociation constant (Kd) of 0.634 nmol/L. Inhibition of sphingosine-1-phosphate receptor 2 or its downstream G12/13-RhoA signaling circumvented the suppressive effects of FAM19A5 on vascular smooth muscle cell proliferation and migration. CONCLUSIONS: We revealed that a novel adipokine, FAM19A5, was capable of inhibiting postinjury neointima formation via sphingosine-1-phosphate receptor 2-G12/13-RhoA signaling. Downregulation of FAM19A5 during obesity may trigger cardiometabolic diseases.
Assuntos
Tecido Adiposo/metabolismo , Citocinas/metabolismo , Músculo Liso Vascular/metabolismo , Neointima , Receptores de Lisoesfingolipídeo/metabolismo , Lesões do Sistema Vascular/metabolismo , Adipócitos/metabolismo , Animais , Sinalização do Cálcio , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Citocinas/genética , Modelos Animais de Doenças , Subunidades alfa G12-G13 de Proteínas de Ligação ao GTP/metabolismo , Células HEK293 , Humanos , Masculino , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Músculo Liso Vascular/lesões , Músculo Liso Vascular/patologia , Miócitos de Músculo Liso/metabolismo , Miócitos de Músculo Liso/patologia , Obesidade/genética , Obesidade/metabolismo , Comunicação Parácrina , Ratos Sprague-Dawley , Receptores de Lisoesfingolipídeo/genética , Receptores de Esfingosina-1-Fosfato , Técnicas de Cultura de Tecidos , Lesões do Sistema Vascular/genética , Lesões do Sistema Vascular/patologia , Proteína rhoA de Ligação ao GTP/metabolismoRESUMO
FAM19A1 is a member of the family with sequence similarity 19 with unknown function. FAM19A1 mRNA expression is restricted to the CNS. Here, we report that FAM19A1 is a classic secretory protein, and expression levels correlate with brain development, increasing from embryonic d 12.5, peaking between postnatal d (P)1 and P7 and decreasing at wk 8. The adult hippocampus is a region of FAM19A1 high expression. Recombinant FAM19A1 suppressed the proliferation and self-renewal of neural stem cells (NSCs) and altered the lineage progression of NSCs with promoted neuron differentiation and suppressed astrocyte differentiation. Although GPCR 1 (GPR1) has been reported to be expressed in the CNS, its functions in the brain remain unclear. We identified GPR1 to be a functional receptor for FAM19A1. FAM19A1 interacted with GPR1 via the N-terminal domain (GPR1-ND), and its NSC modulatory functions required the Rho-associated protein kinase (ROCK) /ERK1/2 and ROCK/signal transducer and activator of transcription 3 signaling pathways. GPR1-ND that selectively bound to FAM19A1 neutralized the effects of FAM19A1 on NSC functions. Taken together, our results show, for the first time to our knowledge, that FAM19A1 is a novel regulatory factor of the proliferation and differentiation of NSCs, and identified a novel mechanism by which GPCR mediates the effects of FAM19A1 on NSC functions that may be important for brain development and neurogenesis. Additional exploration of the functions of FAM19A1 and GPR1 in the CNS may broaden the range of therapeutic options available for major brain disorders.-Zheng, C., Chen, D., Zhang, Y., Bai, Y., Huang, S., Zheng, D., Liang, W., She, S., Peng, X., Wang, P., Mo, X., Song, Q., Lv, P., Huang, J., Ye, R. D., Wang, Y. FAM19A1 is a new ligand for GPR1 that modulates neural stem-cell proliferation and differentiation.
RESUMO
CMTM4 (CKLF-like MARVEL transmembrane domain containing 4), a potential tumor suppressor gene, is involved in several types of malignancies. It has been reported to be downregulated and exhibit anti-tumorigenic activities by regulating cell growth and cell cycle in clear cell renal cell carcinoma. It has also been identified as a tumor suppressor in hepatocellular carcinoma (HCC), and its negative expression is a risk factor for poor prognosis of HCC patients. In the present study, an integrated bioinformatics analysis based on The Cancer Genome Atlas (TCGA) database showed that CMTM4 was frequently reduced in colorectal cancer (CRC) and high expression of CMTM4 was associated with increased overall survival rates. Based on these findings, we adopted gain-of-function and lost-of-function strategies using SW480 and HT29 CRC cell lines which have relatively low and high endogenous CMTM4 levels, respectively. We observed impeded cell proliferation and migration upon overexpression of CMTM4 in SW480 cells, and the opposite effects were observed upon knockdown of CMTM4 in HT-29 cells. Cell signaling pathways essential for CRC progression were then examined, and the phosphorylation levels of AKT, ERK1/2, and STAT3 were found to be decreased by CMTM4 overexpression in SW480 cells and elevated by CMTM4 silencing in HT29 cells. Their inhibitors were used to validate that the three signaling pathways contributed to the inhibitory effects of CMTM4 on CRC cells. Taken together, our results suggest that CMTM4 plays a tumor suppressive role in CRC.
Assuntos
Adenocarcinoma/metabolismo , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Proteínas com Domínio MARVEL/fisiologia , Adenocarcinoma/patologia , Movimento Celular , Proliferação de Células , Células HT29 , Humanos , Sistema de Sinalização das MAP Quinases , Proteínas com Domínio MARVEL/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Fator de Transcrição STAT3/metabolismoRESUMO
The family with sequence similarity 3 (FAM3) gene family is a cytokine-like gene family with four members FAM3A, FAM3B, FAM3C and FAM3D. In this study, we found that FAM3D strongly chemoattracted human peripheral blood neutrophils and monocytes. To identify the FAM3D receptor, we used chemotaxis, receptor internalization, Ca(2+) flux and radioligand-binding assays in FAM3D-stimulated HEK293 cells that transiently expressed formyl peptide receptor (FPR)1 or FPR2 to show that FAM3D was a high affinity ligand of these receptors, both of which were highly expressed on the surface of neutrophils, and monocytes and macrophages. After being injected into the mouse peritoneal cavity, FAM3D chemoattracted CD11b+ Ly6G+ neutrophils in a short time. In response to FAM3D stimulation, phosphorylated ERK1/2 and phosphorylated p38 MAPK family proteins were upregulated in the mouse neutrophils, and this increase was inhibited upon treatment with an inhibitor of FPR1 or FPR2. FAM3D has been reported to be constitutively expressed in the gastrointestinal tract. We found that FAM3D expression increased significantly during colitis induced by dextran sulfate sodium. Taken together, we propose that FAM3D plays a role in gastrointestinal homeostasis and inflammation through its receptors FPR1 and FPR2.
Assuntos
Quimiotaxia , Colite/metabolismo , Citocinas/metabolismo , Sistema de Sinalização das MAP Quinases , Monócitos/metabolismo , Neutrófilos/metabolismo , Receptores de Formil Peptídeo , Receptores de Lipoxinas , Animais , Colite/genética , Colite/patologia , Citocinas/genética , Sulfato de Dextrana/toxicidade , MAP Quinases Reguladas por Sinal Extracelular/genética , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Células HEK293 , Humanos , Camundongos , Monócitos/patologia , Neutrófilos/patologia , Receptores de Formil Peptídeo/agonistas , Receptores de Formil Peptídeo/genética , Receptores de Formil Peptídeo/metabolismo , Receptores de Lipoxinas/agonistas , Receptores de Lipoxinas/genética , Receptores de Lipoxinas/metabolismoRESUMO
CCDC134 (coiled-coil domain containing 134), a cytokine-like molecule, was previously reported to exert antitumor effects by augmenting CD8+ T-cell mediated immunity. However, the dynamic changes in CCDC134 expression patterns in the spinal cord that may be involved in the progression of experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis, remains unclear. In this study, we found that CCDC134 expression was markedly increased in the spinal cord during the progression of EAE. Furthermore, we demonstrated that CCDC134 significantly reduced the severity and slowed the progression of EAE, which correlated with reduced spinal cord inflammation and demyelination. The underlying mechanism of CCDC134-induced effects involved inhibition of T helper (Th)-1 and Th17 cell differentiation and secretion of its key effector molecules IFN-γ and IL-17A via regulation of JAK/STAT signaling. These findings indicate that CCDC134 exerts potent anti-inflammatory effects through the selective modulation of pathogenic Th1 and Th17 cells by targeting critical signaling pathways. The study provides insights into the role of CCDC134 as a unique therapeutic agent for the treatment of autoimmune diseases.
Assuntos
Encefalomielite Autoimune Experimental/tratamento farmacológico , Encefalomielite Autoimune Experimental/imunologia , Proteínas de Membrana/farmacologia , Animais , Diferenciação Celular/efeitos dos fármacos , Citocinas/metabolismo , Modelos Animais de Doenças , Encefalomielite Autoimune Experimental/metabolismo , Feminino , Proteínas de Membrana/metabolismo , Proteínas de Membrana/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Esclerose Múltipla/metabolismo , Transdução de Sinais/efeitos dos fármacos , Medula Espinal/metabolismo , Células Th1/efeitos dos fármacos , Células Th1/imunologia , Células Th1/fisiologia , Células Th17/efeitos dos fármacos , Células Th17/imunologia , Células Th17/fisiologiaRESUMO
Previous research revealed that CMTM8 acts as a tumor suppressor gene in variety cancers. However, the role of CMTM8 in bladder cancer has never been reported. In this study, the expression profile of CMTM8 was examined in bladder cancer tissues and bladder cancer cell lines. The effects of CMTM8 on bladder cancer cell proliferation, apoptosis, migration, and invasion were examined. Bladder tumor tissues from 84 cases were examined for CMTM8 expression by immunohistochemistry. Disease-specific survival was investigated using a Kaplan-Meier analysis, and Cox proportional hazards analysis was assessed. Our results showed that upregulation of CMTM8 in the T24 cell line could suppress T24 cells proliferation, migration and invasion and enhance the sensitivity to Epirubicin. Kaplan-Meier analysis revealed that the expression of CMTM8 was correlated with the survival time of bladder cancer patients. Altogether, our data suggested that CMTM8 is an important tumor suppressor gene in human bladder cancer and qualified as a useful prognostic indicator for patients with bladder cancer.
Assuntos
Biomarcadores Tumorais/genética , Quimiocinas/genética , Proteínas com Domínio MARVEL/genética , Prognóstico , Neoplasias da Bexiga Urinária/genética , Adulto , Idoso , Apoptose/genética , Biomarcadores Tumorais/biossíntese , Linhagem Celular Tumoral , Proliferação de Células/genética , Quimiocinas/biossíntese , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Estimativa de Kaplan-Meier , Proteínas com Domínio MARVEL/biossíntese , Masculino , Pessoa de Meia-Idade , Proteínas Supressoras de Tumor/genética , Neoplasias da Bexiga Urinária/patologiaRESUMO
PC3-secreted microprotein (PSMP) or microseminoprotein is a newly discovered secreted protein whose function is currently unknown. In this study, PSMP was found to possess chemotactic ability toward monocytes and lymphocytes, and its functional receptor was identified as CCR2B. PSMP was identified as a chemoattractant protein from a PBMC chemoattractant platform screen that we established. The mature secreted PSMP was able to chemoattract human peripheral blood monocytes, PBLs, and CCR2B-expressing THP-1 cells, but not peripheral blood neutrophils, even though it does not contain the classical structure of chemokines. CCR2B was identified as one receptor for PSMP-mediated chemotaxis by screening HEK293 cells that transiently expressed classical chemokine receptors; results obtained from the chemotaxis, calcium flux, receptor internalization, and radioligand-binding assays all confirmed this finding. To further identify the major function of PSMP, we analyzed its expression profile in tissues. PSMP is highly expressed in benign prostatic hyperplasia and in some prostate cancers, and can also be detected in breast tumor tissue. In response to PSMP stimulation, phosphorylated ERK levels downstream of CCR2B signaling were upregulated in the PC3 cell line. Taken together, our data collectively suggest that PSMP is a chemoattractant protein acting as a novel CCR2 ligand that may influence inflammation and cancer development.
Assuntos
Fatores Quimiotáticos/metabolismo , Proteínas de Neoplasias/metabolismo , Receptores CCR2/metabolismo , Neoplasias da Mama/metabolismo , Linhagem Celular , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Feminino , Expressão Gênica , Células HEK293 , Humanos , Inflamação/metabolismo , Ligantes , Linfócitos/metabolismo , Masculino , Monócitos/metabolismo , Neutrófilos/metabolismo , Fosforilação , Hiperplasia Prostática/metabolismo , Neoplasias da Próstata/metabolismo , Ligação ProteicaRESUMO
The CKLF-like MARVEL transmembrane domain containing 3 (CMTM3) gene is a novel tumor suppressor with frequent epigenetic inactivation. In this study, we showed the role played by CMTM3 in gastric cancer cells as a tumor suppressor gene, and examined the correlation between CMTM3 expression and clinicopathological parameters using immunohistochemistry in gastric cancer patients with different pathological stages (n = 350). We found that CMTM3 expression was reduced or silenced by epigenetic regulation in gastric cell lines, and dramatically downregulated in primary gastric cancer tissues. Restoration of CMTM3 significantly affected migration and invasion of AGS and SGC-7901 cells (P < 0.001). In vivo experiments showed that peritoneal disseminated metastases were significantly suppressed by CMTM3 (P < 0.001). We further showed that the expression of MMP2 and the phosphorylation of Erk1/2 were decreased when CMTM3 was restored. In addition, by immunohistochemical staining, we found that the expression of CMTM3 was remarkably weaker in gastric cancer tissues than in normal mucosae (P = 0.008), and was significantly correlated with gender (P = 0.033), tumor depth (P = 0.049), stage (P = 0.021), and histological grade (P = 0.022). More importantly, CMTM3 expression was associated with prognosis in gastric cancer patients (P = 0.041), and was a significant independent prognostic indicator (hazard ratio = 0.704, 95% confidence interval, 0.498-0.994; P = 0.046). Our findings indicate that CMTM3 regulates migration and invasion of gastric cancer cells. Moreover, CMTM3 is a candidate marker for prognosis of gastric cancer in the clinic.
Assuntos
Movimento Celular/genética , Quimiocinas/genética , Proteínas com Domínio MARVEL/genética , Neoplasias Gástricas/genética , Neoplasias Gástricas/patologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Linhagem Celular Tumoral , Estudos de Coortes , Regulação para Baixo , Feminino , Mucosa Gástrica/patologia , Genes Supressores de Tumor , Humanos , Sistema de Sinalização das MAP Quinases , Masculino , Metaloproteinase 2 da Matriz/genética , Pessoa de Meia-Idade , Invasividade Neoplásica , Fosforilação , Prognóstico , Adulto JovemRESUMO
Dehydrogenase/reductase (SDR family) X-linked (DHRSX) is a novel human gene without any substantial functional annotation and was initially cloned and identified in our laboratory. In this study, we present evidence that it encodes a non-classical secretory protein and promotes starvation induced autophagy. Using the Baf.A1 assay and N-terminal sequencing, we showed that DHRSX is secreted in a non-classical form. We expressed and purified a recombinant human GST-DHRSX fusion protein. Functional studies revealed that HeLa and U2OS cells overexpressing DHRSX or treated with the GST-DHRSX fusion protein exhibited higher levels of starvation-induced autophagy, resulting in increased endogenous LC3-II levels, a punctate GFP-LC3 distribution, and structures associated with autophagy, with a lower accumulation of autophagy substrates such as p62 and polyQ80. Accordingly, knockdown of endogenous DHRSX through specific siRNAs reduced LC3-II levels obviously in U2OS cells induced by starvation. Collectively, these results demonstrate that DHRSX is a novel non-classical secretory protein involved in the positive regulation of starvation induced autophagy and provide a new avenue for research on this protein family and autophagy regulation.
Assuntos
Autofagia/genética , Oxirredutases/biossíntese , Oxirredutases/genética , Inanição/enzimologia , Regulação da Expressão Gênica , Proteínas de Fluorescência Verde , Células HEK293 , Células HeLa , Humanos , Oxirredutases/metabolismo , Análise de Sequência de DNA , Inanição/genéticaRESUMO
Modulation of surface T cell antigen receptor (TCR) expression is crucial for proper T cell development and maintenance of mature T cell function at steady state and upon stimulation. We previously determined that CCDC134 (coiled-coil domain containing 134), a cytokine-like molecule that served as a potential member of the γc cytokine family, contributes to antitumor responses by augmenting CD8+ T cell-mediated immunity. Here we show that T cell-specific deletion of Ccdc134 decreased peripheral mature CD4+ and CD8+ T cells, which resulted in impaired T cell homeostasis. Moreover, Ccdc134-deficient T cells exhibited an attenuated response to TCR stimulation in vitro, showing lower activation and proliferative capacity. This was further reflected in vivo, rendering mice refractory to T cell-mediated inflammatory and antitumor responses. More importantly, CCDC134 is associated with TCR signaling components, including CD3ϵ, and attenuated TCR signaling in Ccdc134-deficient T cells via altered CD3ϵ ubiquitination and degradation. Taken together, these findings suggest a role for CCDC134 as a positive regulator of TCR-proximal signaling and provide insight into the cell-intrinsic functional consequences of Ccdc134 deficiency in the attenuation of T cell-mediated inflammatory and antitumor responses.
Assuntos
Linfócitos T CD8-Positivos , Transdução de Sinais , Camundongos , Animais , Receptores de Antígenos de Linfócitos T/metabolismo , Ativação Linfocitária , Citocinas/metabolismoRESUMO
Cytokines are soluble proteins that mediate immune reactions and are responsible for communication among immune cells. CD4(+) T cells are the principle sources of cytokines of adaptive immunity. Cytokines play critical roles in the differentiation and effector function of CD4(+) T cells. They also play key roles in diseases, and some of them have been developed into drugs in the forms of recombinant cytokines, soluble receptors and neutralizing antibodies. Therefore, identifying novel potential cytokines is necessary and beneficial for better understanding immunology and enhancing human health. To find novel potential cytokines, we carried out an integrated bioinformatics analysis on the whole human genome. Cytokine candidates were selected for cDNA cloning, sub-cloning, secretion verification, expression profile analysis and functional study. Here, we report a novel soluble protein, VSTM1-v2, which is a classical secretory glycoprotein mainly expressed in immune tissues, and can promote the differentiation and activation of Th17 cells.
Assuntos
Citocinas/isolamento & purificação , Glicoproteínas/isolamento & purificação , Receptores Imunológicos/imunologia , Células Th17/efeitos dos fármacos , Processamento Alternativo , Sequência de Aminoácidos , Animais , Diferenciação Celular/efeitos dos fármacos , Diferenciação Celular/imunologia , Clonagem Molecular , Citocinas/genética , Citocinas/imunologia , Citocinas/farmacologia , Expressão Gênica , Biblioteca Gênica , Glicoproteínas/genética , Glicoproteínas/imunologia , Glicoproteínas/farmacologia , Células HEK293 , Humanos , Interleucina-17/imunologia , Interleucina-17/metabolismo , Ativação Linfocitária/efeitos dos fármacos , Dados de Sequência Molecular , Isoformas de Proteínas/genética , Isoformas de Proteínas/imunologia , Isoformas de Proteínas/isolamento & purificação , Coelhos , Receptores Imunológicos/genética , Receptores Imunológicos/isolamento & purificação , Solubilidade , Células Th17/citologia , Células Th17/imunologia , TransfecçãoRESUMO
Most studies have shown that improper disposal of e-waste can accelerate the release of high concentrations of polybrominated diphenyl ethers (PBDEs), and this situation causes environmental pollution and human health risks. The recycling technology of waste electronic plastics based on solvent processes can reduce environmental pollution and health risks from PBDEs. In this study, high impact polystyrene (HIPS) from waste TV sets was taken as the research object, and d-limonene and n-propanol were used as solvent and precipitant, respectively. We studied the relationship between the precipitation conditions and the size of precipitate particles, and the effect laws of precipitation conditions on the removal percentage of PBDEs were discussed. Transferring behavior of PBDEs during precipitation was investigated, and the parameters suitable for removing PBDEs from HIPS solution were confirmed. Results showed that lower HIPS concentration in d-limonene, lower precipitation temperature, higher mass ratio of n-propanol to HIPS solution, and greater stirring speed were conducive to form smaller and more uniform precipitate particles. All conditions (concentration, temperature, mass ratio, and stirring rate) that could increase the solubility of PBDEs in the mixed solvent of limonene and n-propanol or decrease the swelling degree of HIPS precipitate particles, or reduce the size of particles could improve the removal percentage of PBDEs. The investigated results indicated that insoluble PBDEs (e.g., decabromodiphenyl ether) transferred into the HIPS precipitate mainly through the generated crystals and then precipitated together with the HIPS particles, and soluble PBDEs (e.g., octabromodiphenyl ether) migrated into the precipitate by the solution entrained. The precipitate particles, which measured approximately 1.0 mm (on average), were obtained when the solution containing 10% of HIPS from waste TV shell was precipitated by adding n-propanol equivalent to twice the mass of the solution at 40 °C and 3000 r/min stirring speed. The total concentration of PBDEs in the precipitate particles (dried) was reduced to 2369 mg/kg, and 88.06% of the PBDEs in the original plastic solution was successfully removed by this process.
Assuntos
Resíduo Eletrônico , Éteres Difenil Halogenados , 1-Propanol , Resíduo Eletrônico/análise , Monitoramento Ambiental , Éteres Difenil Halogenados/análise , Humanos , Limoneno , Plásticos/química , Poliestirenos , SolventesRESUMO
Chemokine-like factor 1 (CKLF1) was the first member of the CKLF-like MARVEL transmembrane domain containing member (CMTM) family to be discovered. Its expression level is increased clearly in peripheral blood lymphocytes upon phytohemagglutinin stimulation, but little is known about the expression and function of CKLF1 in dendritic cells (DCs), which are the most potent antigen-presenting cells. In the present study, we showed that CKLF1 was highly expressed in monocytes. During differentiation from monocytes to immature DCs, CKLF1 was increased dramatically on day 2 and then decreased from day 3 to 5. Upon maturation with different stimuli, CKLF1 was down-regulated. Two peptides of CKLF1, C19 and C27, stimulated the effect of immature DCs (imDCs) on T-cell proliferation and IFN-gamma production. Further study on DC maturation showed that C19 and C27 up-regulated HLA-DR expression and IL-12 secretion, with no obvious effects on CD80, CD83 or CD86. Thus, CKLF1-C19 and -C27 can stimulate antigen-presenting capability of imDCs.
Assuntos
Diferenciação Celular , Quimiocinas/metabolismo , Células Dendríticas/citologia , Células Dendríticas/imunologia , Regulação da Expressão Gênica , Diferenciação Celular/efeitos dos fármacos , Quimiocinas/farmacologia , Células Dendríticas/efeitos dos fármacos , Regulação para Baixo , Humanos , Interleucina-12/metabolismo , Leucócitos Mononucleares/citologia , Leucócitos Mononucleares/imunologia , Proteínas com Domínio MARVELRESUMO
Antibodies have a common structure consisting of two identical heavy (H) and two identical light (L) chains. It is widely accepted that a single mature B cell produces a single antibody through restricted synthesis of only one VHDJH (encoding the H-chain variable region) and one VLJL (encoding the L-chain variable region) via recombination. Naive B cells undergo class-switch recombination (CSR) from initially producing membrane-bound IgM and IgD to expressing more effective membrane-bound IgG, IgA, or IgE when encountering antigens. To ensure the "one cell - one antibody" paradigm, only the constant region of the H chain is replaced during CSR, while the rearranged VHDJH pattern and the L chain are kept unchanged. To define those long-standing classical concepts at the single-cell transcriptome level, we applied the Chromium Single-Cell Immune Profiling Solution and Sanger sequencing to evaluate the Ig transcriptome repertoires of single B cells. Consistent with the "one cell - one antibody" rule, most of the B cells showed one V(D)J recombination pattern. Intriguingly, however, two or more VHDJH or VLJL recombination patterns of IgH chain or IgL chain were also observed in hundreds to thousands of single B cells. Moreover, each Ig class showed unique VHDJH recombination pattern in a single B-cell expressing multiple Ig classes. Together, our findings reveal an unprecedented presence of multi-Ig specificity in some single B cells, implying regulation of Ig gene rearrangement and class switching that differs from the classical mechanisms of both the "one cell - one antibody" rule and CSR.
RESUMO
CMTM is a novel family of proteins linking chemokines and TM4SF. Several members of this family are highly expressed in testes and regulate androgen receptor (AR) transcription activity. One member of this family, CMTM3, has the highest expression level in testes and contains one leucine zipper and two LXXLL motifs. As assessed with the dual-luciferase reporter assay, overexpression of CMTM3 represses AR transactivation, while knocking down it can increase AR transactivation. Moreover, CMTM3 inhibits prostate-specific antigen (PSA) expression in LNCaP cells at both mRNA and protein levels with no obvious influence on AR expression. Taken together, CMTM3 may play some roles in the maturation and maintenance of the male reproduction.