碟子湖磷污染的植物修复研究

磷是富营养化水体中的特征污染物,是植物和微生物的主要营养元素。而水体富营养化是水体污染中最为普遍的现象,也是国内外水环境污染治理的难题。本文以生态修复技术为切入点,研究了碟子湖湿地中酸模、蔊菜、稗草、莎草、野生胡萝卜、虉草六种优势植物的生物量和对污水中磷的净化效果,分析了根系和地上部分生物量与污水净化效果之间的关系。根据植被的生长环境条件和植被的自身特性探究出最优组合,使其适应于当地局部地区的气候及土壤环境,治理水体污染。

Rh(Ⅱ)-catalyzed intermolecular carboamination of pyridines via double Csp^(2)-H bond activations

We disclose the development of the Rh-catalyzed amine-directed remote 5,6-carboamination protocol of pyridines via dual Csp^(2)-H functionalizations.A variety of readily available 2-aminopyridines and 1,2,3-triazoles are allowed for coupling cyclization to access polyfunctionalized azaindoles.Mechanistic studies including DFT calculations unveil that relay carbenoidelectrophilic addition to pyridines and the sequential pyridyl Csp^(2)-H amination are involved in this transformation.The postsynthetic utility of this methodology is showcased by versatile and site-selective modification of azaindoles.

Tal6b/AvrXa27A,a hidden TALE targeting the susceptibility gene OsSWEET11a and the resistance gene Xa27 in rice

Xanthomonas oryzae pv.oryzae(Xoo)secretes transcription activator-like effectors(TALEs)to activate rice susceptibility(S)genes,causing bacterial blight(BB),as well as resistance(R)genes,leading to de-fense against BB.This activation follows a gene-for-gene paradigm that results in an arms race between the TALE of the pathogen and effector-binding elements(EBEs)in the promoters of host genes.In this study,we characterized a novel TALE,designated Tal6b/AvrXa27A,that activates the rice S gene OsSWEET11a and the rice R gene Xa27.Tal6b/AvrXa27A is a member of the AvrXa27/TalAO class and contains 16 repeat variable diresidues(RVDs);one RVD is altered and one is deleted in Tal6b/AvrXa27A compared with AvrXa27,a known avirulence(avr)effector of Xa27.Tal6b/AvrXa27A can transcriptionally activate the expression of Xa27 and OsSWEET11a via EBEs in their corresponding promoters,leading to effector-triggered immunity and susceptibility,respectively.The 16 RVDs in Tal6b/AvrXa27A have no obvious similarity to the 24 RVDs in the effector PthXo1,but EBETal6b and EBEPthXo1 are overlapped in the OsSWEET11a promoter.Tal6b/AvrXa27A is prevalent among Asian Xoo isolates,but PthXo1 has only been reported in the Philippine strain PXO99A.Genome editing of EBETal6b in the OsSWEET11a pro-moter further confirmed the requirement for OsSWEET11a expression in Tal6b/AvrXa27A-dependent susceptibility to Xoo.Moreover,Tal6b/AvrXa27A resulted in higher transcription of Xa27 than of OsSWEET11a,which led to a strong,rapid resistance response that blocked disease development.Thesefindings suggest that Tal6b/AvrXa27A has a dual function:triggering resistance by activating Xa27 gene expression as an avirulence factor and inducing transcription of the S gene OsSWEET11a,resulting in virulence.Intriguingly,Tal6b/AvrXa27A,but not AvrXa27,can bind to the promoter of OsSWEET11a.The underlying recognition mechanism for this binding remains unclear but appears to deviate from the currently accepted TALE code.

Engineering Broad-Spectrum Bacterial Blight Resistance by Simultaneously Disrupting Variable TALE-Binding Elements of Multiple Susceptibility Genes in Rice

Xanthomonas oryzae pv.oryzae(Xoo),the causal agent of bacterial blight of rice,employs the transcription activator-like effectors(TALEs)to induce the expression of the OsSWEET family of putative sugar transporter genes,which function in conferring disease susceptibility(S)in rice plants.To engineer broadspectrum bacterial blight resistance,we used CRISPR/Cas9-mediated gene editing to disrupt the TALEbinding elements(EBEs)of two S genes,OsSWEETH and OsSWEET14,in rice cv.Kitaake,which harbors the recessive resistance allele of Xa25/OsSWEET13.The engineered rice line MS14K exhibited broadspectrum resistance to most Xoo strains with a few exceptions,suggesting that the compatible strains may contain new TALEs.We identified two PthXo2-like TALEs,Tal5LN18 and Tal7PX061,as major virulence factors in the compatible Xoo strains LN18 and PX061,respectively,and found that Xoo encodes at least five types of PthXo2-like effectors.Given that PthXo2/PthXo2.1 target OsSlVEETf3 for transcriptional activation,the genomes of 3000 rice varieties were analyzed for EBE variationsin the OsSWEET13 promoter,and 10Xa25-like haplotypes were identified.We found that Tal5LN18 and Tal7PX〇6i bind slightly different EBE sequences in the OsSWEET13 promoter to activate its expression.CRISPR/Cas9 technology was then used to generate InDels in the EBE of the OsSWEET13 promoter in MS14K to creat a new germplasm with three edited OsSWEET EBEs and broad-spectrum resistance against all Xoo strains tested.Collectively,our findings illustrate how to disarm TALE-S co-evolved loci to generate broad-spectrum resistance through the loss of effector-triggered susceptibility in plants.

水稻条斑病菌pilT基因在致病性中的功能分析

【目的】本实验室前期研究发现水稻条斑病菌(Xanthomonas oryzae pv.oryzicola,Xoc)RS105菌株中pil T基因Tn5转座子插入突变体在寄主水稻上致病性明显降低,在非寄主烟草上激发过敏反应(hypersensitive response,HR)的能力也明显减弱。为了揭示pilT基因在Xoc菌株中的功能,本文进行了深入研究。【方法】本实验通过无标记双交换敲除的方法获得Xoc RS105菌株pil T基因缺失突变体RΔpilT,并对该突变体的游动性以及生物膜等表型进行了检测。【结果】与野生型RS105菌株相比,敲除突变体RΔpilT不仅在感病水稻IR24上的致病性显著降低,在非寄主烟草上产生过敏反应的能力减弱,而且突变体游动性降低,生物膜含量增加,互补子能够恢复上述缺陷至野生型水平。qRT-PCR结果显示,在RΔpil T中,hrpG、hrpX、hrcC、clp、rpfG、pilA、pilC基因表达量明显降低。【结论】Xoc中pilT为重要的毒性相关基因,其致病性与游动性和生物膜含量变化相关,并且受clp、rpfG、pilA、pilC等基因调控。pil T基因编码的Pil T蛋白是构成IV型菌毛的亚基之一,为菌体运动提供能量。本文对pilT基因的功能研究,为进一步分析IV型菌毛在Xoc中的功能提供了线索。

The Xanthomonas typeⅢeffector Xop AP prevents stomatal closure by interfering with vacuolar acidification

Plant stomata close rapidly in response to a rise in the plant hormone abscisic acid(ABA)or salicylic acid(SA)and after recognition of pathogenassociated molecular patterns(PAMPs).Stomatal closure is the result of vacuolar convolution,ion efflux,and changes in turgor pressure in guard cells.Phytopathogenic bacteria secrete typeⅢeffectors(T3Es)that interfere with plant defense mechanisms,causing severe plant disease symptoms.Here,we show that the virulence and infection of Xanthomonas oryzae pv.oryzicola(Xoc),which is the causal agent of rice bacterial leaf streak disease,drastically increased in transgenic rice(Oryza sativa L.)plants overexpressing the Xoc T3E gene Xop AP,which encodes a protein annotated as a lipase.We discovered that Xop AP binds to phosphatidylinositol 3,5-bisphosphate(Ptd Ins(3,5)P_(2)),a memb rane phospholipid that functions in p H control in lysosomes,membrane dynamics,and protein trafficking.Xop AP inhibited the acidification of vacuoles by competing with vacuolar H^(+)-pyrophosphatase(V-PPase)for binding to Ptd Ins(3,5)P_(2),leading to stomatal opening.Transgenic rice overexpressing Xop AP also showed inhibition of stomatal closure when challenged by Xoc infection and treatment with the PAMP flg22.Moreover,Xop AP suppressed flg22-induced gene expression,reactive oxygen species burst and callose deposition in host plants,demonstrating that Xop AP subverts PAMP-triggered immunity during Xoc infection.Taken together,these findings demonstrate that Xop AP overcomes stomatal immunity in plants by binding to lipids.

A Xanthomonas transcription activator-like effector is trapped in nonhost plants for immunity

Xanthomonas oryzae pv.oryzae(Xoo),the causal agent of bacterial leaf blight in rice,delivers transcription activator-like effector(TALE)proteins into host cells to activate susceptibility or resistance(R)genes that promote disease or immunity,respectively.Nonhost plants serve as potential reservoirs of R genes;consequently,nonhost R genes may trap TALEs to trigger an immune response.In this study,we screened 17 Xoo TALEs for their ability to induce a hypersensitive response(HR)in the nonhost plant Nicotiana benthamiana(Nb);only AvrXa10 elicited an HR when transiently expressed in Nb.The HR generated by AvrXa10 required both the central repeat region and the activation domain,suggesting a specific interaction between AvrXa10 and a potential R-like gene in nonhost plants.Evans blue staining and ion leakage measurements confirmed that the AvrXa10-triggered HR was a form of cell death,and the transient expression of AvrXa10 in Nb induced immune responses.Genes targeted by AvrXa10 in the Nb genome were identified by transcriptome profiling and prediction of effector binding sites.Using several approaches(in vivo reporter assays,electrophoretic mobility-shift assays,targeted designer TALEs,and on-spot gene silencing),we confirmed that AvrXa10 targets NbZnFP1,a C2H2-type zinc finger protein that resides in the nucleus.Functional analysis indicated that overexpression of NbZnFP1 and its rice orthologs triggered cell death in rice protoplasts.An NbZnFP1 ortholog was also identified in tomato and was specifically activated by AvrXa10.These results demonstrate that NbZnFP1 is a nonhost R gene that traps AvrXa10 to promote plant immunity in Nb.

A Potential Disease Susceptibility Gene CsLOB of Citrus Is Targeted by a Major Virulence Effector PthA of Xanthomonas citri subsp, citri

Dear Editor, Citrus bacterial canker （CBC）, one of the most destructive diseases of citrus, has a severe economic impact on citrus industries in tropical and subtropical countries （Das, 2003）. It has been widely known that Xanthomonas citri subsp, citri （Xcc） causes the CBC by transferring the type-Ill effectors （T3SEs） to the interior of the plant cell through type-Ill secretion system （T3S） to modulate transcription of host plants to trigger CBC disease （Schaad et al., 2006）. Pathotype A,