HvWRKY2 acts as an immunity suppressor and targets HvCEBiP to regulate powdery mildew resistance in barley

Plants use a sophisticated immune system to perceive pathogen infection and activate immune responses in a tightly controlled manner.In barley,Hv WRKY2 acts as a repressor in barley disease resistance to the powdery mildew fungus,Blumeria graminis f.sp.hordei(Bgh).However,the molecular features of Hv WRKY2 in its DNA-binding and repressor functions,as well as its target genes,are uncharacterized.We show that the W-box binding of Hv WRKY2 requires an intact WRKY domain and an upstream sequence of~75 amino acids,and the Hv WRKY2 W-box binding activity is linked to its repressor function in disease resistance.Chromatin immunoprecipitation(ChIP)-seq analysis identified HvCEBiP,a putative chitin receptor gene,as a target gene of Hv WRKY2 in overexpressing transgenic barley plants.ChIP-qPCR and Electrophoretic Mobility Shift Assay(EMSA)verified the direct binding of Hv WRKY2 to a W-boxcontaining sequence in the HvCEBiP promoter.Hv CEBiP positively regulates resistance against Bgh in barley.Our findings suggest that Hv WRKY2 represses barley basal immunity by directly targeting pathogen-associated molecular pattern(PAMP)recognition receptor genes,suggesting that Hv CEBiP and likely chitin signaling function in barley PAMP-triggered immune responses to Bgh infection.

HvMPK4 phosphorylates HvWRKY1 to enhance its suppression of barley immunity to powdery mildew fungus

Mitogen-activated protein kinase(MAPK)cascades play important roles in disease resistance in model plant species.However,the functions of MAPK signaling pathways in crop disease resistance are largely unknown.Here we report the function of HvMKK1-HvMPK4-HvWRKY1 module in barley immune system.HvMPK4 is identified to play a negative role in barley immune response against Bgh,as virus-induced gene silencing of HvMPK4 results in enhanced disease resistance whilst stably overexpressing HvMPK4 leads to super-susceptibility to Bgh infection.Furthermore,the barley MAPK kinase HvMKK1 is found to specifically interact with HvMPK4,and the activated HvMKK1^(DD) variant specifically phosphorylates HvMPK4 in vitro.Moreover,the transcription factor HvWRKY1 is identified to be a downstream target of HvMPK4 and phosphorylated by HvMPK4 in vitro in the presence of HvMKK1^(DD).Phosphorylation assay coupled with mutagenesis analyses identifies S122,T284,and S347 in HvWRKY1 as the major residues phosphorylated by HvMPK4.HvWRKY1 is phosphorylated in barley at the early stages of Bgh infection,which enhances its suppression on barley immunity likely due to enhanced DNA-binding and transcriptional repression activity.Our data suggest that the HvMKK1-HvMPK4 kinase pair acts upstream of HvWRKY1 to negatively regulate barley immunity against powdery mildew.

Orthologous genesPm12andPm21from twowild relatives of wheat show evolutionary conservation but divergent powdery mildew resistance

Wheat powdery mildew,caused by Blumeria graminis f.sp.tritici(Bgt),is a devastating disease that threatens wheat production worldwide.Pm12,which originated from Aegilops speltoides,a wild relative of wheat,confers strong resistance to powdery mildew and therefore has potential use in wheat breeding.Using susceptible mutants induced by gamma irradiation,we physically mapped and isolated Pm12 and showed it to be orthologous to Pm21 from Dasypyrum villosum,also a wild relative of wheat.The resistance function of Pm12 was validated via ethyl methanesulfonatemutagenesis,virus-induced gene silencing,and stable genetic transformation.Evolutionary analysis indicates that the Pm12/Pm21 loci in wheat species are relatively conserved but dynamic.Here,we demonstrated that the two orthologous genes,Pm12 and Pm21,possess differential resistance against the same set of Bgt isolates.Overexpression of the coiledcoil domains of both PM12 and PM21 induces cell death in Nicotiana benthamiana leaves.However,their full-length forms display different cell death-inducing activities caused by their distinct intramolecular interactions.Cloning of Pm12 will facilitate its application in wheat breeding programs.This study also gives new insight into two orthologous resistance genes,Pm12 and Pm21,which show different race specificities and intramolecular interaction patterns.

Development, identification and utilization of introgression lines using Chinese endemic and synthetic wheat as donors

Chromosome segmental introgression lines（ILs）are an effective way to utilize germplasm resources in crops.To improve agronomic traits of wheat cultivar（Triticum aestivum） Shi 4185, four sets of ILs were developed. The donors were Chinese endemic subspecies accessions Yunnan wheat（T. aestivum ssp. yunnanense） YN3, Tibetan semiwild wheat（T. aestivum ssp. tibetanum） XZ-ZM19450, and Xinjiang wheat（T. aestivum ssp. petropavlovskyi） XJ5, and synthetic wheat HC-XM1620 derived from a cross between T. durum acc. D67.2/P66.270 with Aegilops tauschii acc. 218.Totals of 356, 366, 445 and 457 simple sequence repeat（SSR）markers were polymorphic between Shi 4185 and YN3, XZZM19450, XJ5 and HC-XM1620, respectively. In total, 991 ILs were identified, including 300 derived from YN3, covering 95%of the genome of Shi 4185, 218 from XZ-ZM19450（79%）, 279 from XJ5（97%）, and 194 from HC-ZX1620（84%）. The sizes and locations of each introgression were determined from a consensus SSR linkage map. Using the ILs, 11 putative Resear quantitative trait loci（QTLs） were identified for plant height（PH）, spike length（SL） and grain number per spike（GNS）.Comparative analyses of 24 elite ILs with the parents revealed that the four donor parents could be important resources to improve wheat SL and GNS. Our work offers a case for utilizing endemic landraces for QTL mapping and improvement of wheat cultivars using introgression lines.

Development of Triticum aestivume——Leymus mollis Translocation Lines and Identification of Resistance to Stripe Rust

Wheat stripe rust, caused by Puccinia striiformis f. sp. tritici, is one of the most widely distributed and destructive fungal diseases worldwide. Since 1995, most Chinese wheat cultivars have lost their stripe rust resistance due to the subsequent emergence of the new races CYR30, CYR31, CYR32, and CYR33 （Han et al., 2010）. Therefore, it is necessary to seek effective resistance genes and develop new resistance germ- plasm for wheat resistance breeding.

Dissecting and Enhancing the Contributions of High-Molecular-Weight Glutenin Subunits to Dough Functionality and Bread Quality

Dear Editor,Seed storage proteins （SSPs） are frequently important determinants of crop quality traits （Shewry and Casey, 1999）. Dissecting and enhancing the genetic contributions of individual SSPs to their target traits are essential for effectively improving crop quality attributes. However, such a task is often difficult to accomplish, because SSPs are frequently expressed from multigene families and exhibit strong allelic variation. Consequently, detailed knowledge of the function of individual SSPs in crop quality trait is still limited. This scenario is well illustrated by high-molecular-weight glutenin subunits （HMW- GSs）, a complex family of SSPs that are involved in wheat enduse quality through affecting dough functionality （Bek6s, 2012; Rasheed et al., 2014）.