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Rice OsFLS2-Mediated Perception of Bacterial flagellins Is Evaded by Xanthomonas oryzae pvs. oryzae and oryzicola

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Molecular Plant 2015年第7期8卷 1024-1037页

作者： Shanzhi Wang Zhe Sun Huiqin Wang Lijuan Liu Fen Lu Jun Yang Min Zhang Shiyong Zhang Zejian Guo Andrew F. Bent Wenxian Sun Department of Plant Pathology China Agricultural University 2 West Yuanmingyuan Road Haidian District Beijing 100193 China Key Laboratory of Plant Pathology Ministry of Agriculture China Agricultural University Beijing 100193 China Rice Research Institute Shandong Academy of Agricultural Science Jinan 250100 Shandong Province China College of Bioscience and Biotechnology Hunan Agricultural University Changsha 410128 Hunan Province China Department of Plant Pathology University of Wisconsin Madison WI 53706 USA These authors contributed equally to this article.

Bacterial flagellins are often recognized by the receptor kinase flagellin SENSITIVE2 （FLS2） and activate MAMP-triggered immunity in dicotyledonous plants. However, the capacity of monocotyledonous rice to recognize flagellins of key rice pathogens and its biological relevance remain poorly understood. We demonstrate that ectopically expressed OsFLS2 in Arabidopsis senses the eliciting fig22 peptide and in vitro purified Acidovorax avenae （Aa） flagellin in an expression level-dependent manner, but does not recognize purified flagellins or derivative fig22x~ peptides ofXanthomonas oryzae pvs. oryzae （Xoo） and or- yzicola （Xoc）. Consistently, the fig22 peptide and purified Aa flageUin, but not Xoo/Xoc flagellins, induce various immune responses such as defense gene induction and MAPK activation in rice. Perception of flagellin by rice does induce strong resistance to Xoo infection, as shown after pre-treatment of rice leaves with Aa flagellin. OsFLS2 was found to differ from AtFLS2 in its perception specificities or sensitivities to different fig22 sequences. In addition, post-translational modification of Xoc flagellin was altered by dele- tion of glycosyltransferase-encoding rbfC, but this had little effect on Xoc motility and rpfC mutation did not detectably reduce Xoc virulence on rice. Deletion of flagellin-encoding fliC from Xoo/Xoc blocked swim- ming motility but also did not significantly alter Xoo/Xoc virulence. These results suggest that Xoo/Xoc carry flg22-region amino acid changes that allow motility while evading the ancient flagellin detection sys- tem in rice, which retains recognition capacity for other bacterial pathogens.

关键词： OsFLS2 flagellin perception specificity Xanthomonas oryzae

Sensing bacterial infections by NAIP receptors in NLRC4 inflammasome activation

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Protein & Cell 2012年第2期3卷 98-105页

作者： Yi-Nan Gong Feng Shao National Institute of Biological Sciences Beijing 102206China Howard Hughes Medical Institute National Institute of Biological SciencesBeijing 102206China

The inflammasome is an emerging new pathway in innate immune defense against microbial infection or endogenous danger *** inflammasome stimulates activation of inflammatory caspases,mainly ***-1 activation is responsible for processing and secretion of IL-1βand IL-18 as well as for inducing macrophage pyroptotic *** of the large cytoplasmic inflammasome complex is thought to be mediated by members of NOD-like receptor(NLR)*** functions of most of the NLR proteins remain to be defined,several NLR proteins including NLRC4 have been shown to assemble distinct inflammasome *** inflammasome pathways,particularly the NLRC4 inflammasome,play a critical role in sensing and restricting diverse types of bacterial *** we review recent advances in defining the exact bacterial ligands and the ligand-binding receptors involved in NLRC4 inflammasome *** of the discovery of the NAIP family of inflammasome receptors for bacterial flagellin and type III secretion apparatus on future inflammasome and bacterial infection studies are also discussed.

关键词： inflammasome NOD-like receptors NLRC4 caspase-1 NAIP type III secretion system flagellin Salmonella Legionella enteropathogenic E.coli Burkholderia

Antigenicity analysis of Vibrio harveyi TS-628 strain

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Frontiers in Biology 2007年第3期2卷 263-267页

作者： QIN Yingxue WANG Jun WANG Shifeng YAN Qingpi Department of Oceanography Institute of Subtropic OceanXiamen UniversityXiamen 361000China Fisheries College Jimei UniversityXiamen 361021China

Vibrio harveyi,the major causative agent of vib-riosis,affects a diverse range of marine cultured organisms over a wide geographical ***,reports about screening the effective antigen and research on vaccines of *** are ***,lipopolysaccharide(LPS)and outer membrane proteins(OMP)are major immunogenic antigens in many Gram-negative *** this study,the flagellin,OMP and LPS of the *** TS-628 strain iso-lated from infected groupers were extracted and Western blot analysis was used to detect the antigenicity of these *** of the Western blot assay reveal that there are four positive flagellin bands:35 kDa,38 kDa,43 kDa,and 52 kDa,of which the 43 kDa and 52 kDa bands displayed the strongest positive *** are five positive OMPbands about 35 kDa,38 kDa,43 kDa,47 kDa,and 52 kDa,of which the 43 kDa appeared to have the strongest positive reaction although the other four proteins also displayed strong ***,LPS is Western *** results indicate that the 43 kDa and 52 kDa flagellin and OMP of size 43 kDa,52 kDa can be candidates for developing vaccines against ***.

关键词： Vibrio harveyi flagellin LPS OMP antigenicity

rFliC prolongs allograft survival in association with the activation of recipient Tregs in a TLR5-dependent manner

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Cellular & Molecular Immunology 2014年第2期11卷 206-214页

作者： Jing Hao Chao Zhang Jing Song Guihua Hou Key Laboratory for Experimental Teratology of the Ministry of Education and Institute of Experimental Nuclear Medicine Shandong University MedicalSchool Jinan 250012 China

AIIorejection remains an obstacle for successful organ transplantation. Although different types of immunosuppressive agents are effective for controlling rejection and prolonging graft survival, drug treatment is limited because of side effects and toxicity. Therefore, it is necessary and urgent to identify new candidate drugs for inducing allotolerance. Recently, it has been reported that bacterial flagellin induces the immunosuppressive activity of regulatory T cells （Tregs） in humans in vitro. In the present study, we analyzed the effects of recombinant flagellin （rFliC） on allograft survival and explored the underlying mechanisms associated with the activation of recipient Tregs in a murine skin allotransplantation model. The results showed that rFliC administration （3 mg/kg, once per day for 3 days, i.p.） prolonged allograft survival （mean survival time： 18.4--.1.1 days） compared to the control group （10___0.7 days, P〈O.01）. Additionally, higher positive expression of Toll-like receptor 5 （TLR5） was detected within the allograft administered with rFliC. The frequency of CD4＋CD25＋Foxp3＋ Tregs; the expression of Treg-related factors TLR5, Foxp3, TGF-I^I and IL-IO; and the proliferation and suppression of Tregs were increased following rFliC administration compared to the control. Moreover, the increased expression of tolerance-related molecules and the proliferation of Tregs induced by rFliC were attenuated by an anti-TLR5 blocking antibody both in vivo and in vitro. In conclusion, rFliC administration prolongs the survival of allografts, which is associated with the activation of recipient Tregs in a TLR5-dependent manner, rFliC may be a new candidate for anti-allorejection therapy.

关键词： allotransplantation flagellin regulatory T cells TLR5

TLR5 involvement in attenuated IL-8 production in nuclear decorin silenced oral mucosal dysplastic keratinocytes and squamous cell carcinoma

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American Journal of Molecular Biology 2013年第4期3卷 248-254页

作者： Nyla Dil Abhijit G. Banerjee Department of Oral Biology Faculty of Dentistry University of Manitoba Winnipeg Canada Center for Genomic Bio-Medicine and Research Institute Durg Chhattisgarh India Department of Oral Biology Faculty of Dentistry University of Manitoba Winnipeg Canada Department of Immunology and Department of Medical Microbiology and Infectious Diseases Faculty of Medicine University of Manitoba Winnipeg Canada

Head and neck cancer is one of the most prevalent cancers in the world. Roughly half of these malignancies originate from oral mucosa and constitute Oral squamous cell carcinomas. Despite many advances in diagnostic and therapeutic regimens, five year survival rate remains at roughly 50 %, indicating the need for in depth understanding of the oral squamous cell carcinoma immunobiology. We have previously shown that in human dysplastic oral keratinocytes (DOK) and malignant squamous cells carcinoma (SCC-25), multifunctional proteoglycan decorin is aberrantly expressed and localized in the nucleus bound to nuclear EGFR. In vitro nuclear decorin knockdown significantly reduced IL-8 and IL8-dependent migration, invasion and angiogenesis in these cells. Since toll-like receptor (TLR) signalling leads to IL-8 production we examined here if these receptors play a role in decorin silencing mediated reduction in IL-8 levels. We have used immunological and molecular techniques to study toll-like receptors involvement in attenuated IL-8 production in nuclear decorin silenced (stable knockdown) oral mucosal dysplastic keratinocytes and squamous carcinoma cells. We show that nuclear decorin silenced DOK and SCC-25 cells show marked diminution of TLR5 mRNA and protein expression compared with respective controls that translated into loss of function in response to appropriate TLR ligand. In these mucosal oral epithelia, decorin stable knockdown significantly down-regulated IL-8 production following activation with TLR5 ligand flagellin. These data suggest that decorin silencing interferes with IL-8 production, in part, by altering TLR5 expression and signaling in dysplastic and malignant oral epithelia. This study highlights the significance of TLR5 expression and signaling in mucosal cancers.

关键词： Oral Cancer Proteoglycans flagellin DAMP Mucosal Cancer TLR5 PAMP IL-8 SLRP Nuclear decorin

同种排斥过程中TLR5及Foxp3的表达、调节及意义

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同种排斥过程中TLR5及Foxp3的表达、调节及意义

作者：陈富强山东大学

学位级别：硕士

研究目的近年来Toll样受体家族(Toll-like Receptor,TLRs)在机体免疫应答中的意义受到高度关注。TLRs家族成员作为一种模式识别受体(pattern recognitionreceptor,PRR)可特异性识别病原相关分子模式(pathogen-associated molecularpatterns,PAMPs),不仅在激活天然免疫中发挥重要作用,而且还在诱导和调节获得性免疫中具有重要意义,是连接天然免疫和获得性免疫的重要桥梁。TLRs信号可以在自身抗原,同种抗原和异种抗原的刺激下激活。2008年最新研究发现小鼠心脏皮肤同种移植物的长期生存需要天然Tregs的存在,TLR9的特异激活剂CpG可以通过抑制Treg的功能和促进Th1效应性T细胞的分化,抑制同种移植物的存活。TLR7的激活可以促进皮肤同种移植物的排斥;TLR4的激活阻断了利用抗CD154单抗建立的心脏同种移植物的长期存活,阻止了CD4/FoxpTreg在移植物的聚集,因此TLRs信号的激活状态对移植物的耐受或排斥非常关键。研究显示,CD4 CD25调节性T细胞(Treg)参与多种途径诱导的移植免疫耐受,Treg不仅在维持自身耐受方面发挥重要作用,而且在防治同种移植物排斥方面也有重要意义。因此如何有效扩增Treg细胞是诱导移植耐受的关键因素之一。近来研究发现TLR5是唯一具有免疫负调节作用的分子,可直接间接地参与自身免疫病的发病过程。细菌鞭毛蛋白(flagellin)是TLR5的唯一配体,flagellin可特异识别结合TLR5,通过激活核转录因子NF-κB刺激TNF-α、IL-1β、IL-8等前炎症细胞因子的产生,介导机体天然免疫。以往研究表明Treg功能的调节主要是通过细胞因子或者是通过APCs细胞上共刺激分子来实现的,然而新的证据表明Treg可以通过TLRs直接感受外来抗原的刺激,TLR5选择性高表达在Treg细胞上,TLR5的特异激活剂flagellin可增强Treg对效应性T细胞的抑制作用。2008年有研究报告体内给予TLR5特异激活剂flagellin,可以保护机体免受化学、细菌病毒和辐射等损害。因此如何通过TLRs调控Treg细胞活性,对于诱导机体免疫耐受具有重要意义。免疫抑制药物雷帕霉素(Rapamycin,Rapa)和环孢素A(Cyclosporin A,CsA)广泛用于器官移植后的抗移植物排斥反应,并且在实验模型中作为诱导免疫耐受的工具。近来研究报道Rapa在体外可以选择性地扩增正常小鼠的天然CD4CD25调节性T细胞,而不阻断该小鼠CD4T细胞的扩增和活化诱导细胞凋亡,但其确切机制尚不清楚,而CsA则可抑制TCR介导的T细胞激活和IL-2的产生。提示CsA和Rapa对Treg可能有不同的作用。我们前期的研究证实Rapa和CsA体内处理可以明显增加同种移植受体的Treg比例,增强Foxp3的表达。但是即使体内使用Rapa,同种移植个体中的Treg比例仍然处于较低水平。因此如何通过调节Treg细胞活性,体内扩增Treg细胞,更好地诱导同种移植耐受仍是目前亟待解决的问题。综上所述,迄今为止的研究表明Treg细胞具有免疫负调节作用,TLR5的激活对某些损伤因素具有免疫保护作用,Treg细胞上TLR5/TLR7/8高表达,但是同种移植状态下,TLR5的活化状态和活化的调节及对Treg的影响途径、机理和信号传导途径尚不清楚。我们推测利用flagellin刺激同种移植TLR5活化,可能在体内扩增Treg的比例,增强Foxp3的表达,从而达到形成移植耐受的目的。本课题首先分析了同种皮肤移植急性排斥过程中TLR5和Foxp3基因的动态表达状况,然后应用Rapa和CsA两种不同的免疫抑制剂对小鼠同种皮肤移植模型进行耐受初步诱导,研究了Rapa和CsA体内应用对TLR5和Foxp3基因表达的影响及与移植物生存的关系:研究了flagellin体外处理对移植耐受模型中T细胞及相关分子TLR5、Foxp3表达影响及与Rapa和CsA是否与协同作用;探讨了同种移植状态下TLR5表达的变化及意义,探讨TLR5和Foxp3表达的相关性,旨在阐明Rapa、CsA及flagellin在同种移植耐受状态下,是否能够通过激活TLR5,增强Treg的活性,强化免疫耐受的诱导,为利用TLR5特异配体-flagellin和抗排斥药物Rapa、CsA联合进行同种移植耐受诱导奠定实验基础,为CD4CD25Treg体外扩增和应用于临床防治同种移植排斥反应探索新的途径。实验方法 ***的提取和鉴定:利用LB营养培养基扩增甲型副伤寒杆菌,用PBS离心后洗涤按照文献[25]方法分离粗制鞭毛蛋白(flagellin),然后通过Sephacryl S-20

关键词： flagellin Rapamycin Cyclosporin A Treg TLR5 Foxp3 同种移植免疫耐受

三种佐剂对防龋亚单位疫苗黏膜免疫增强效应比较

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三种佐剂对防龋亚单位疫苗黏膜免疫增强效应比较