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PNAS. 2018 Aug. 蘇郁文 副研究員

DUSP6 mediates T cell receptor-engaged glycolysis and restrains TFH cell differentiation.

Activated T cells undergo metabolic reprogramming and effector-cell differentiation but the factors involved are unclear. Utilizing mice lacking DUSP6 (DUSP6−/−), we show that this phosphatase regulates T cell receptor (TCR) signaling to influence follicular helper T (TFH) cell differentiation and T cell metabolism. In vitro, DUSP6−/− CD4+ TFH cells produced elevated IL-21. In vivo, TFH cells were increased in DUSP6−/− mice and in transgenic OTII-DUSP6−/− mice at steady state. After immunization, DUSP6−/− and OTII-DUSP6−/− mice generated more TFH cells and produced more antigen-specific IgG2 than controls. Activated DUSP6−/− T cells showed enhanced JNK and p38 phosphorylation but impaired glycolysis. JNK or p38 inhibitors significantly reduced IL-21 production but did not restore glycolysis. TCR-stimulated DUSP6−/− T cells could not induce phosphofructokinase activity and relied on glucose-independent fueling of mitochondrial respiration. Upon CD28 costimulation, activated DUSP6−/− T cells did not undergo the metabolic commitment to glycolysis pathway to maintain viability. Unexpectedly, inhibition of fatty acid oxidation drastically lowered IL-21 production in DUSP6−/− TFH cells. Our findings suggest that DUSP6 connects TCR signaling to activation-induced metabolic commitment toward glycolysis and restrains TFH cell differentiation via inhibiting IL-21 production....detail

 

FASEB Journal. 2018 Jun. 譚澤華 特聘研究員、莊懷佳助研究員

Induction of DUSP14 ubiquitination by PRMT5-mediated arginine methylation.

Dual-specificity phosphatase (DUSP)14 (also known as MAP-kinase phosphatase 6) inhibits T-cell receptor (TCR) signaling and T-cell-mediated immune responses by inactivation of the TGF-β activated kinase 1 binding protein (TAB1)-TGF-β activated kinase 1 (TAK1) complex and ERK. DUSP14 phosphatase activity is induced by the E3 ligase TNF receptor associated factor (TRAF)2-mediated Lys63-linked ubiquitination. Here we report an interaction between DUSP14 and protein arginine methyltransferase (PRMT)5 by proximity ligation assay; similarly, DUSP14 directly interacted with TAB1 but not TAK1. DUSP14 is methylated by PRMT5 at arginine 17, 38, and 45 residues. The DUSP14 triple-methylation mutant was impaired in PRMT5-mediated arginine methylation, TRAF2-mediated lysine ubiquitination, and DUSP14 phosphatase activity. Consistently, DUSP14 methylation, TRAF2 binding, and DUSP14 ubiquitination were attenuated by PRMT5 short hairpin RNA knockdown. Furthermore, DUSP14 was inducibly interacted with PRMT5 and was methylated during TCR signaling in T cells. Together, these findings reveal a novel regulatory mechanism of DUSP14 by which PRMT5-mediated arginine methylation may sequentially stimulate TRAF2-mediated DUSP14 ubiquitination and phosphatase activity, leading to inhibition of TCR signaling.-Yang, C.-Y., Chiu, L.-L., Chang, C.-C., Chuang, H.-C., Tan, T.-H. Induction of DUSP14 ubiquitination by PRMT5-mediated arginine methylation.....detail

 

Oncotarget. 2016 Aug. 7:57593-57605譚澤華 特聘研究員

Downregulation of the phosphatase JKAP/DUSP22 in T Cells as a potential new biomarker of systemic lupus erythematosus nephritis.

Systemic lupus erythematosus (SLE) is a complex autoimmune disease that is characterized by systemic inflammation and multiple organ failures. Dysregulation of T cells plays a critical role in SLE pathogenesis. Our previous study indicates that JKAP (also named DUSP22) inhibits T-cell activation and that JKAP knockout mice develop spontaneous autoimmunity; therefore, we investigated whether JKAP downregulation is involved in SLE patients. JKAP protein levels in purified T cells were examined by immunoblotting using blood samples from 43 SLE patients and 32 healthy controls. SLE patients showed significantly decreased JKAP protein levels in peripheral blood T cells compared to healthy controls. JKAP protein levels in peripheral blood T cells were inversely correlated with SLE disease activity index (SLEDAI) and anti-dsDNA antibody levels. JKAP downregulation in T cells was highly correlated with daily urinary protein amounts and with poor renal outcome in lupus nephritis patients. Notably, the diagnostic power of JKAP downregulation in T cells for active lupus nephritis was higher than those of serum anti-dsDNA antibody, C3, and C4 levels. Moreover, T-cell-specific transgenic mice expressing a dominant-negative JKAP mutant developed spontaneous autoimmune nephritis. Furthermore, JKAP-deficient T cells overproduced complement components, soluble ICAM-1, and soluble VCAM-1 in the kidney; these cytokines have been reported to be involved in lupus nephritis. Taken together, JKAP downregulation in T cells is a novel diagnostic and prognostic biomarker for SLE nephritis...detail

 

Oncotarget. 2016 Jul. 7:41748-41757. 譚澤華 特聘研究員

GLK/MAP4K3 overexpression associates with recurrence risk for non-small cell lung cancer.

Lung cancer is the leading cause of cancer death worldwide. Non-small cell lung cancer (NSCLC) accounts for 85% of total lung cancers; 40% to 60% of NSCLC patients die of cancer recurrence after cancer resection. Since GLK (also named MAP4K3) induces activation of NF-κB, which contributes to tumor progression, we investigated the role of GLK in NSCLC. GLK protein levels of 190 samples from pulmonary tissue arrays and 58 pulmonary resection samples from stage I to stage III NSCLC patients were studied using immunohistochemistry or immunoblotting. High levels of GLK proteins were detected in pulmonary tissues from NSCLC patients. Elevated GLK protein levels were correlated with increased recurrence risks and poor recurrence-free survival rates in NSCLC patients after adjusting for pathologic stage, smoking status, alcohol status, and EGFR levels. Thus, GLK is a novel prognostic biomarker for NSCLC recurrence....detail

 

Cell Death and Differentiation. 2016 May. 23:841莊宗顯 副研究員

Downregulation of COMMD1 by miR-205 promotes a positive feedback loop for amplifying inflammatory- and stemness-associated properties of cancer cells.

Sustained activation of nuclear factor-kappaB (NF-kappaB) in cancer cells has been shown to promote inflammation, expansion of cancer stem cell (CSC) population, and tumor development. In contrast, recent studies reveal that CSCs exhibit increased inflammation due to constitutive NF-kappaB activation; however, the underlying molecular mechanism remains unclear. In the present study, the analysis of microarray data revealed upregulation of NF-kappaB-regulated pro-inflammatory genes and downregulation of copper metabolism MURR1 domain-containing 1 (COMMD1) during the enrichment for stemness in SAS head and neck squamous-cell carcinoma (HNSCC) cells. The 3'-UTR of COMMD1 mRNA contains microRNA (miR)-205 target site. Parallel studies with HNSCC and NSCLC cells indicated that miR-205 is upregulated upon NF-kappaB activation and suppresses COMMD1 expression in stemness-enriched cancer cells. COMMD1 negatively regulates the inflammatory responses induced by TLR agonists, IL-1beta, and TNF-alpha by targeting RelA for degradation. The shRNA-mediated downregulation of COMMD1 in cancer cells enhanced inflammatory response, generating favorable conditions for macrophage recruitment. In addition, genes associated with stemness were also upregulated in these cells, which exhibited increased potential for anchorage-independent growth. Furthermore, COMMD1 downregulation promoted in vivo tumorigenesis and tumor growth, and tumors derived from COMMD1-knockdown cells displayed elevated level of NF-kappaB activation, increased expression of inflammatory- and stemness-associated genes, and contain expanded population of tumor-associated leukocytes and stemness-enriched cancer cells. These results suggest that COMMD1 downregulation by miR-205 promotes tumor development by modulating a positive feedback loop that amplifies inflammatory- and stemness-associated properties of cancer cells...detail

 

Oncotarget. 2016 Feb. 7:10976. 譚澤華 特聘研究員

Epigenetic regulation of HGK/MAP4K4 in T cells of type 2 diabetes patients.

Type 2 diabetes (T2D) is a complex and heterogeneous disease. Obesity increases the risk of obese T2D; but in Asia non-obese T2D is prevalent. The cause of non-obese T2D has remained elusive. We studied the potential involvement of HGK/MAP4K4 in T2D using clinical samples from newly diagnosed, drug-naïve patients and healthy controls. HGK levels fell and IL-6 levels increased in T cells from T2D patients. Frequencies of IL-6-producing T cells were correlated with glucose levels after glucose-tolerance tests (but not body mass index and waist circumference) and inversely correlated with HGK expression levels. Moreover, methylation frequencies of the HGK promoter were increased in T2D patients and correlated with glucose levels after glucose-tolerance tests. The correlation was independent of body mass index. Demethylation treatment increased HGKexpression levels and reduced IL-6 production in T2D T cells. This report identifies HGK methylation/downregulation in T cells as a potential biomarker for non-obese T2D...detail

 

Oncotarget. 2016 Feb. 7:8389. 林文傑 助研究員

Serum chemokine (CC motif) ligand 2 level as a diagnostic, predictive, and prognostic biomarker for prostate cancer.

Prostate-specific antigen (PSA) is regarded as the most sensitive biomarker for prostate cancer. Although androgen/androgen receptor (AR) signaling promotes prostate cancer progression, suppression of AR signaling induces chemokine (CC motif) ligand 2 (CCL2), which enables prostate cancer cells to gain metastatic potential. AR-controlled PSA alone may be an unreliable biomarker for patients receiving androgen deprivation therapy. Therefore, we investigated the validity of CCL2 as a complementary biomarker to PSA for prostate cancer. Our in vitro approach of enriching for prostate cancer cells with higher migration potential showed that CCL2 activated cellular migration. Importantly, we found that CCL2 levels were significantly different between men (n = 379) with and without prostate cancer. Patients with CCL2 ≥ 320 pg/mL had worse overall survival and prostate cancer -specific survival than those with CCL2 < 320 pg/mL. A novel risk classification was developed according to the risk factors CCL2 ≥ 320 pg/mL and PSA ≥ 100 ng/mL, and scores of 2, 1, and 0 were defined as poor, intermediate, and good risk, respectively, and clearly distinguished patient outcomes. CCL2 may serve as a novel biomarker for prostate cancer. The novel risk classification based on combining CCL2 and PSA is more reliable than using either alone...detail

 

Journal of Immunology. 2015 Oct. 195:3912. 莊宗顯 副研究員

Identification of Thiostrepton as a Novel Inhibitor for Psoriasis-like Inflammation Induced by TLR7-9.

Activation of TLR7-9 has been linked to the pathogenesis of autoimmune diseases such as rheumatoid arthritis, systemic lupus erythematosus, and psoriasis. Thus, therapeutic applications of antagonists of these TLRs for such disorders are being investigated. Bortezomib (Velcade) is a proteasome inhibitor known to suppress activation of these TLRs. To identify novel TLR7-9 inhibitors, we searched the Gene Expression Omnibus database for gene expression profiles of bortezomib-treated cells. These profiles were then used to screen the Connectivity Map database for chemical compounds with similar functions as bortezomib. A natural antibiotic, thiostrepton, was identified for study. Similar to bortezomib,thiostrepton effectively inhibits TLR7-9 activation in cell-based assays and in dendritic cells. In contrast to bortezomib, thiostrepton does not inhibit NF-κB activation induced by TNF-α, IL-1, and other TLRs, and it is less cytotoxic to dendritic cells. Thiostrepton inhibits TLR9 localization in endosomes for activation via two mechanisms, which distinguish it from currently used TLR7-9 inhibitors. One mechanism is similar to the proteasome inhibitory function of bortezomib, whereas the other is through inhibition of endosomal acidification. Accordingly, in different animal models, thiostrepton attenuated LL37- and imiquimod-induced psoriasis-like inflammation. These results indicated that thiostrepton is a novel TLR7-9inhibitor, and compared with bortezomib, its inhibitory effect is more specific to these TLRs, suggesting the potential therapeutic applications ofthiostrepton on immunologic disorders elicited by inappropriate activation of TLR7-9...detail

 

 PLoS One. 2014 Sep. 12:9. 蘇郁文 助研究員

PP4 is essential for germinal center formation and class switch recombination in mice.

PP4 is a serine/threonine phosphatase required for immunoglobulin (Ig) VDJ recombination and pro-B/pre-B cell development in mice. To elucidate the role of PP4 in mature B cells, we ablated the catalytic subunit of murine PP4 in vivo utilizing the CD23 promoter and cre-loxP recombination and generated CD23(cre)PP4(F/F) mice. The development of follicular and marginal zone B cells was unaffected in these mutants, but the proliferation of mature PP4-deficient B cells stimulated by in vitro treatment with either anti-IgM antibody (Ab) or LPS was partially impaired. Interestingly, the induction of CD80 and CD86 expression on these stimulated B cells was normal. Basal levels of serum Igs of all isotypes were strongly reduced in CD23(cre)PP4(F/F) mice, and their B cells showed a reduced efficiency of class switch recombination (CSR) in vitro upon stimulation by LPS or LPS plus IL-4. When CD23(cre)PP4(F/F) mice were challenged with either the T cell-dependent antigen TNP-KLH or the T cell-independent antigen TNP-Ficoll, or by H1N1 virus infection, the mutant animals failed to form germinal centers (GCs) in the spleen and the draining mediastinal lymph nodes, and did not efficiently mount antigen-specific humoral responses. In the resting state, PP4-deficient B cells exhibited pre-existing DNA fragmentation. Upon stimulation by DNA-damaging drug etoposide in vitro, mutant B cells showed increased cleavage of caspase 3. In addition, the mutant B cells displayed impaired CD40-mediated MAPK activation, abnormal IgM-mediated NF-κB activation, and reduced S phase entry upon IgM/CD40-stimulation. Taken together, our results establish a novel role for PP4 in CSR, and reveal crucial functions for PP4 in the maintenance of genomic stability, GC formation, and B cell-mediated immune responsesdetail

 

 Nat. Commun. 2014 Aug. 5:4602. 譚澤華 特聘研究員

HGK/MAP4K4 deficiency induces TRAF2 stabilization and Th17 differentiation leading to insulin resistance.

Proinflammatory cytokines play important roles in insulin resistance. Here we report that mice with a T-cell-specific conditional knockout of HGK (T-HGK cKO) develop systemic inflammation and insulin resistance. This condition is ameliorated by either IL-6 or IL-17 neutralization. HGK directly phosphorylates TRAF2, leading to its lysosomal degradation and subsequent inhibition of IL-6 production. IL-6-overproducing HGK-deficient T cells accumulate in adipose tissue and further differentiate into IL-6/IL-17 double-positive cells. Moreover, CCL20 neutralization or CCR6 deficiency reduces the Th17 population or insulin resistance in T-HGK cKO mice. In addition, leptin receptor deficiency in T cells inhibits Th17 differentiation and improves the insulin sensitivity in T-HGK cKO mice, which suggests that leptin cooperates with IL-6 to promote Th17 differentiation. Thus, HGK deficiency induces TRAF2/IL-6 upregulation, leading to IL-6/leptin-induced Th17 differentiation in adipose tissue and subsequent insulin resistance. These findings provide insight into the reciprocal regulation between the immune system and the metabolismdetail

 

 Nat. Commun. 2014 Apr. 5:3618. 譚澤華 特聘研究員

The phosphatase JKAP/DUSP22 inhibits T-cell receptor signalling and autoimmunity by inactivating Lck.

JNK pathway-associated phosphatase (JKAP, also known as DUSP22 or JSP-1) is a JNK activator. The in vivo role of JKAP in immune regulation remains unclear. Here we report that JKAP directly inactivates Lck by dephosphorylating tyrosine-394 residue during T-cell receptor (TCR) signalling. JKAP-knockout T cells display enhanced cell proliferation and cytokine production. JKAP-knockout mice show enhanced T-cell-mediated immune responses and are more susceptible to experimental autoimmune encephalomyelitis (EAE). In addition, the recipient mice that are adoptively transferred with JKAP-knockout T cells show exacerbated EAE symptoms. Aged JKAP-knockout mice spontaneously develop inflammation and autoimmunity. Thus, our results indicate that JKAP is an important phosphatase that inactivates Lck in the TCR signalling turn-off stage, leading to suppression of T-cell-mediated immunity and autoimmunity.detail

 

 J. Immunol. 2014 Feb. 192:1547-57. 譚澤華 特聘研究員

Dual-specificity phosphatase 14 (DUSP14/MKP6) negatively regulates TCR signaling by inhibiting TAB1.

T cell activation is dependent upon phosphorylation of MAPKs, which play a critical role in the regulation of immune responses. Dual-specificity phosphatase 14 (DUSP14; also known as MKP6) is classified as a MAPK phosphatase. The in vivo functions of DUSP14 remain unclear. Thus, we generated DUSP14-deficient mice and characterized the roles of DUSP14 in T cell activation and immune responses. DUSP14 deficiency in T cells resulted in enhanced T cell proliferation and increased cytokine production upon T cell activation. DUSP14 directly interacted with TGF-β-activated kinase 1 (TAK1)-binding protein 1 (TAB1) and dephosphorylated TAB1 at Ser(438), leading to TAB1-TAK1 complex inactivation in T cells. The phosphorylation levels of the TAB1-TAK1 complex and its downstream molecules, including JNK and IκB kinase, were enhanced in DUSP14-deficient T cells upon stimulation. The enhanced JNK and IκB kinase activation in DUSP14-deficient T cells was attenuated by TAB1 short hairpin RNA knockdown. Consistent with that, DUSP14-deficient mice exhibited enhanced immune responses and were more susceptible to experimental autoimmune encephalomyelitis induction. Thus, DUSP14 negatively regulates TCR signaling and immune responses by inhibiting TAB1 activation…detail

 

 PNAS. 2013 Dec. 110:20711-6. 莊宗顯 副研究員

Toll-like receptor 9 and 21 have different ligand recognition profiles and cooperatively mediate activity of CpG-oligodeoxynucleotides in zebrafish.

CpG-oligodeoxynucleotides (CpG-ODNs) are potent immune stimuli currently under investigation as antimicrobial agents for different species. Toll-like receptor (TLR) 9 and TLR21 are the cellular receptors of CpG-ODN in mammals and chickens, respectively. The avian genomes lack TLR9, whereas mammalian genomes lack TLR21. Although fish contain both of these genes, the biological functions of fish TLR9 and TLR21 have not been investigated previously. In this study, we comparatively investigated zebrafish TLR9 (zebTLR9) and TLR21 (zebTLR21). The two TLRs have similar expression profiles in zebrafish. They are expressed during early development stages and are preferentially expressed in innate immune function-related organs in adult fish. Results from cell-based activation assays indicate that these two zebrafish TLRs are functional, responding to CpG-ODN but not to other TLR ligands. zebTLR9 broadly recognized CpG-ODN with different CpG motifs, but CpG-ODN with GACGTT or AACGTT had better activity to this TLR. In contrast, zebTLR21 responded preferentially to CpG-ODN with GTCGTT motifs. The distinctive ligand recognition profiles of these two TLRs were determined by their ectodomains. Activation of these two TLRs by CpG-ODN occurred inside the cells and was modulated by UNC93B1. The biological functions of these two TLRs were further investigated. The CpG-ODNs that activate both zebTLR9 and zebTLR21 were more potent than others that activate only zebTLR9 in the activation of cytokine productions and were more bactericidal in zebrafish. These results suggest that zebTLR9 and zebTLR21 cooperatively mediate the antimicrobial activities of CpG-ODN. Overall, this study provides a molecular basis for the activities of CpG-ODN in fishdetail

 

 Arthritis Rheum. 2013 Oct. 65:2573-82. 譚澤華 特聘研究員

Germinal center kinase-like kinase overexpression in T cells as a novel biomarker in rheumatoid arthritis.

Objective - Germinal center kinase–like kinase (GLK; also called MAPKKKK-3) activates protein kinase Cθ (PKCθ) during T cell activation and controls autoimmunity in lupus patients. Intracellular kinases are involved in the pathogenesis of rheumatoid arthritis (RA). We undertook this study to determine the role of GLK in RA. Methods - The severity of collagen-induced arthritis (CIA) was studied in GLK-deficient mice. Expression levels of GLK from RA patients were determined by Western blotting, flow cytometry, real-time polymerase chain reaction, and immunohistochemical staining. Localization of GLK in T cells was identified by confocal microscopy. RA disease activity was assessed using the Disease Activity Score in 28 joints…detail

 

 PLoS One. 2013 Jul. 8:e68804. 蘇郁文 助研究員

The serine/threonine phosphatase PP4 is required for pro-B cell development through its promotion of immunoglobulin VDJ recombination.

PP4 phosphatase regulates a number of crucial processes but the role of PP4 in B cells has never been reported. We generated B cell-specific pp4 knockout mice and have identified an essential role for PP4 in B cell development. Deficiency of PP4 in B lineage cells leads to a strong reduction in pre-B cell numbers, an absence in immature B cells, and a complete loss of mature B cells. In PP4-deficient pro-B cells, immunoglobulin (Ig) DJ(H) recombination is impaired and Ig µ heavy chain expression is greatly decreased. In addition, PP4-deficient pro-B cells show an increase of DNA double-strand breaks at Ig loci. Consistent with their reduced numbers, residual PP4-deficient pre-B cells accumulate in the G1 phase, exhibit excessive DNA damage, and undergo increased apoptosis. Overexpression of transgenic Ig in PP4-deficient mice rescues the defect in B cell development such that the animals have normal numbers of IgM(+) B cells. Our study therefore reveals a novel function for PP4 in pro-B cell development through its promotion of V(H)DJ(H) recombination.…detail

 

 J. Biol. Chem. 2012 Oct. 287:34091-100. 譚澤華 特聘研究員

Attenuation of T cell receptor signaling by serine phosphorylation-mediated lysine 30 ubiquitination of SLP-76 protein.

SLP-76 (SH2 domain-containing leukocyte protein of 76 kDa) is an adaptor protein that is essential for T cell development and T cell receptor (TCR) signaling activation. Previous studies have identified an important negative feedback regulation of SLP-76 by HPK1 (hematopoietic progenitor kinase 1; MAP4K1)-induced Ser-376 phosphorylation. Ser-376 phosphorylation of SLP-76 mediates 14-3-3 binding, resulting in the attenuation of SLP-76 activation and downstream signaling; however, the underlying mechanism of this action remains unknown. Here, we report that phosphorylated SLP-76 is ubiquitinated and targeted for proteasomal degradation during TCR signaling. SLP-76 ubiquitination is mediated by Ser-376 phosphorylation. Furthermore, Lys-30 is identified as a ubiquitination site of SLP-76. Loss of Lys-30 ubiquitination of SLP-76 results in enhanced anti-CD3 antibody-induced ERK and JNK activation. These results reveal a novel regulation mechanism of SLP-76 by ubiquitination and proteasomal degradation of activated SLP-76, which is mediated by Ser-376 phosphorylation, leading to down-regulation of TCR signaling…detail

 

 BMC Med. 2012 Aug. 10:84.  譚澤華 特聘研究員

Germinal center kinase-like kinase (GLK/MAP4K3) expression is increased in adult-onset Still's disease and may act as an activity marker

Background - Germinal center kinase-like kinase (GLK, also termed MAP4K3), a member of the MAP4K family, may regulate gene transcription, apoptosis and immune inflammation in response to extracellular signals. The enhanced expression of GLK has been shown to correspond with disease severity in patients with systemic lupus erythematosus. We investigated the role of GLK in the pathogenesis of adult-onset Still's disease, which shares some similar clinical characteristics with systemic lupus erythematosus. Methods - The frequencies of circulating GLK-expressing T-cells in 24 patients with active adult-onset Still's disease and 12 healthy controls were determined by flow cytometry analysis. The expression levels of GLK proteins and transcripts were evaluated in peripheral blood mononuclear cells by immunoblotting and quantitative PCR. Serum levels of T helper (Th)17-related cytokines, including IL-1β, IL-6, IL-17 and TNF-α, were measured by ELISA…detail

 

 J. Biol. Chem. 2012 Feb. 287:11037-48. 譚澤華 特聘研究員

Downregulation of B-cell receptor signaling by hematopoietic progenitor kinase 1 (HPK1)-mediated phosphorylation and ubiquitination of the activated BLNK

Hematopoietic progenitor kinase 1 (HPK1) is a Ste20-like serine/threonine kinase that suppresses immune responses and autoimmunity. B-cell receptor (BCR) signaling activates HPK1 by inducing BLNK/HPK1 interaction. Whether HPK1 can reciprocally regulate BLNK during BCR signaling is unknown. Here we show that HPK1-deficient B cells display hyper-proliferation and hyper-activation of IKK and MAPKs (ERK, p38, and JNK) upon the ligation of BCR. HPK1 attenuates BCR-induced cell activation via inducing BLNK threonine152 phosphorylation, which mediates BLNK/14-3-3 binding. Furthermore, threonine152-phosphorylated BLNK is ubiquitinated at lysine37, 38 and 42 residues, leading to attenuation of MAPK and IKK activation in B cells during BCR signaling. These results reveal a novel negative feedback regulation of BCR signaling by HPK1-mediated phosphorylation, ubiquitination, and subsequent degradation of the activated BLNK…detail

 

 Eur. J. Immunol. 2012 Feb. 42:476-88. 黃慶裕 助研究員

DUSP4 deficiency enhances CD25 expression and CD4+ T-cell proliferation without impeding T-cell development.

The differentiation and activation of T cells are critically modulated by MAP kinases, which are in turn feed-back regulated by dual-specificity phosphatases (DUSPs) to determine the duration and magnitude of MAP kinase activation. DUSP4 (also known as MKP2) is a MAP kinase-induced DUSP member that is dynamically expressed during thymocyte differentiation. We generated DUSP4-deficient mice to study the function of DUSP4 in T-cell development and activation. Our results show that thymocyte differentiation and activation-induced MAP kinase phosphorylation were comparable between DUSP4-deficient and WT mice. Interestingly, activated DUSP4-/- CD4+ T cells were hyperproliferative while DUSP4-/- CD8+ T cells proliferated normally. Further mechanistic studies suggested that the hyperproliferation of DUSP4-/- CD4+ T cells resulted from enhanced CD25 expression and IL-2 signaling through increased STAT5 phosphorylationdetail

 

 Nat. Immunol. 2011 Nov.12:1113-8. 譚澤華 特聘研究員

GLK controls autoimmunity and NF-kB signaling by activating PKCθ in T cells.

Protein kinase C-θ (PKC-θ) is required for activation of the transcription factor NF-κB induced by signaling via the T cell antigen receptor (TCR); however, the direct activator of PKC-θ is unknown. We report that the kinase GLK (MAP4K3) directly activated PKC-θ during TCR signaling. TCR signaling activated GLK by inducing its direct interaction with the upstream adaptor SLP-76. GLK-deficient mice had impaired immune responses and were resistant to experimental autoimmune encephalomyelitis. Consistent with that, people with systemic lupus erythematosus had considerable enhanced GLK expression and activation of PKC-θ and the kinase IKK in T cells, and the frequency of GLK-overexpressing T cells was directly correlated with disease severity. Thus, GLK is a direct activator of PKC-θ, and activation of GLK-PKC-θ-IKK could be used as new diagnostic biomarkers and therapeutic targets for systemic lupus erythematosus….detail

 

 J. Biol. Chem. 2010 Feb. 285:5472-8. 譚澤華 特聘研究員

JNK pathway-associated phosphatase dephosphorylates focal adhesion kinase and suppresses cell migration.

JNK pathway-associated phosphatase (JKAP, also named DUSP22) is expressed in various tissues, indicating that JKAP may have an important biological function. We showed that JKAP localized in the actin filament-enriched region. Expression of JKAP reduced cell migration, whereas a JKAP mutant lacking catalytic activity promoted cell motility. JKAP efficiently removed tyrosine phosphorylation of several proteins. We have identified focal adhesion kinase (FAK) as a substrate of JKAP. Overexpression of JKAP, but not JKAP mutant lacking catalytic activity, decreased FAK phosphorylation at tyrosines 397, 576, and 577 in H1299 cells. Consistent with these results, decreasing JKAP expression by RNA interference promoted cell migration and Src-induced FAK phosphorylation. Taken together, this study identified a new role for JKAP in the modulation of FAK phosphorylation and cell motility...detail

 

 FASEB J. 2011 Aug. 25:2700-10. 莊宗顯 副研究員

Functional interaction of Hsp90 and Beclin 1 modulates Toll-like receptor-mediated autophagy.

Autophagy is one of the downstream effector mechanisms for elimination of intracellular microbes following activation of the Toll-like receptors (TLRs). Although the detailed molecular mechanism for this cellular process is still unclear, Beclin 1, a key molecule for autophagy, has been suggested to play a role. Heat shock protein 90 (Hsp90) is a molecular chaperone that regulates the stability of signaling proteins. Herein, we show that Hsp90 forms a complex with Beclin 1 through an evolutionarily conserved domain to maintain the stability of Beclin 1. In monocytic cells, geldanamycin (GA), an Hsp90 inhibitor, effectively promoted proteasomal degradation of Beclin 1 in a concentration-dependent (EC50 100 nM) and time-dependent (t50 2 h) manner. In contrast, KNK437/Hsp inhibitor I had no effect. Hsp90 specifically interacted with Beclin 1 but not with other adapter proteins in the TLR signalsome...detail

 PNAS. 2011 Nov. 108:18354-9.  蘇郁文 助研究員

Crucial role for TNF receptor-associated factor 2(TRAF2) in regulating NFκB2 signaling that contributes to autoimmunity.

TNF receptor-associated factor 2 (TRAF2) is a key intracellular signaling mediator that acts downstream of not only TNFα but also various members of the TNFα superfamily. Here, we report that, despite their lack of TNFα signaling, TRAF2−/−TNFα−/− mice develop an inflammatory disorder characterized by autoantibody accumulation and organ infiltration by T cells with the phenotypes of activated, effector, and memory cells. RAG1−/− mice reconstituted with TRAF2−/−TNFα−/− bone marrow cells showed increased numbers of hyperactive T cells and rapidly developed progressive and eventually lethal inflammation. No inflammation was observed in RAG1−/− mice reconstituted with TRAF2−/−TNFα−/−T-cell receptor β−/− or TRAF2−/−TNFα−/−NFκB-induced kinase+/− bone marrow cells. The pathogenic TRAF2−/−TNFα−/− T cells showed constitutive NFκB2p52 activation and produced elevated levels of T-helper 1 and T-helper 17 cytokines...detail

  

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