High-density mapping of durable and broad-spectrum stripe rust resistance gene Yr30 in wheat.

KEY MESSAGE: The durable stripe rust resistance gene Yr30 was fine-mapped to a 610-kb region in which five candidate genes were identified by expression analysis and sequence polymorphisms. The emergence of genetically diverse and more aggressive races of Puccinia striiformis f. sp. tritici (Pst) in the past twenty years has resulted in global stripe rust outbreaks and the rapid breakdown of resistance genes. Yr30 is an adult plant resistance (APR) gene with broad-spectrum effectiveness and its durability. Here, we fine-mapped the YR30 locus to a 0.52-cM interval using 1629 individuals derived from residual heterozygous F5:6 plants in a Yaco"S"/Mingxian169 recombinant inbred line population. This interval corresponded to a 610-kb region in the International Wheat Genome Sequencing Consortium (IWGSC) RefSeq version 2.1 on chromosome arm 3BS harboring 30 high-confidence genes. Five genes were identified as candidate genes based on functional annotation, expression analysis by RNA-seq and sequence polymorphisms between cultivars with and without Yr30 based on resequencing. Haplotype analysis of the target region identified six haplotypes (YR30_h1-YR30_h6) in a panel of 1215 wheat accessions based on the 660K feature genotyping array. Lines with YR30_h6 displayed more resistance to stripe rust than the other five haplotypes. Near-isogenic lines (NILs) with Yr30 showed a 32.94% higher grain yield than susceptible counterparts when grown in a stripe rust nursery, whereas there was no difference in grain yield under rust-free conditions. These results lay a foundation for map-based cloning Yr30.

Himalayan mountains imposing a barrier on gene flow of wheat yellow rust pathogen in the bordering regions of Pakistan and China.

The wheat yellow rust pathogen has been shown to be diverse and potentially originated in the Himalayan region. Although Himalayan populations of Pakistan, Nepal and Bhutan have been previously compared, little is known about the relative divergence and diversity in Puccinia striiformis populations in the bordering regions of Pakistan and China. To assess the relative diversity and divergence in these regions of Pakistan (Gilgit-Baltistan, Hazara and Azad Jammu Kashmir) and China (Xinjiang, Qinghai, Tibet, Sichuan, Guizhou and Yunnan), a total of 1245 samples were genotyped using 17 microsatellite SSR markers. A clear divergence was observed between the bordering regions of Pakistan and China (FST = 0.28) without any resampling of genetic groups and multilocus genotypes across two sides of the Himalayan mountains. The closest subpopulations across the two countries were Xinjiang and Gilgit-Baltistan (Nei's distance = 0.147), which were close geographically. A very high diversity and recombinant population structure was observed in both populations, though slightly higher in China (Genotypic diversity = 0.970; r¯d = 0.000) than in Pakistan (Genotypic diversity = 0.902; r¯d = 0.065). The distribution of genetic groups and resampling of MLGs revealed more gene flow across Yunnan, Guizhou and Sichuan regions in China, while between Hazara and Azad-Jammu Kashmir in Pakistan. The lack of gene flow between Pakistan and China populations is due to geographical barriers and a large patch of land without wheat. The information on the relative diversity and divergence in different geographical zones of the pathogen center of diversity and neighboring region should be considered in resistant wheat deployment while considering the invasion potential of the pathogen at regional and global contexts.

Identification of Mahonia Species as Alternate Hosts for Puccinia striiformis f. sp. tritici and Determination of Existence of Sexual Propagation of the Rust Pathogen on Mahonia Under Natural Conditions in China.

Many Berberis species have been identified as alternate hosts for Puccinia striiformis f. sp. tritici. Importantly, susceptible Berberis species are determined to play an important role in the occurrence of sexual reproduction, generation of new races of the rust pathogen. However, little is known about Mahonia serving as alternate hosts for P. striiformis f. sp. tritici and their role to commence sexual reproduction of the rust fungus under natural conditions. Herein, three Mahonia species or subspecies, Mahonia fortunei, M. eurybracteata subsp. ganpinensis, and M. sheridaniana, were identified as alternate hosts for P. striiformis f. sp. tritici, and seven Mahonia species were highly resistant to the rust pathogen. We recovered seven samples of P. striiformis f. sp. tritici from naturally rusted Mahonia cardiophylla plants. Totally, 54 single uredinium (SU) isolates, derived from the seven samples, generated 20 different race types, including one known race type, and 19 new race types. SNP markers analysis showed that all SU isolates displayed high phenotype diversity (H = 0.32) with a high Shannon's information index (I = 0.49). Analysis of linkage disequilibrium indicated an insignificant rbarD value (rbarD = 0.003, P < 0.1). As a result, all SU isolates are sexually produced, suggesting that P. striiformis f. sp. tritici parasitizes susceptible Mahonia to complete sexual reproduction under natural conditions. The role of Mahonia in occurrence of wheat stripe rust are needed to study for management of the disease.

Evaluation of the Potential Risk of the Emerging Yr5-Virulent Races of Puccinia striiformis f. sp. tritici to 165 Chinese Wheat Cultivars.

In 2017, a new race (TSA-6) of the wheat stripe rust pathogen, Puccinia striiformis f. sp. tritici, virulent to resistance gene Yr5, was detected in China. However, whether Chinese wheat cultivars are resistant to the new race was unknown. In this study, two isolates (TSA-6 and TSA-9) with virulence to Yr5 were tested on other wheat Yr gene lines for their avirulence and virulence patterns and used, together with prevalent races CYR32 and CYR34 without the Yr5 virulence, to evaluate 165 major Chinese wheat cultivars for their reactions. Isolates TSA-6 and TSA-9 had similar but different virulence spectra and therefore should be considered two different races. Their avirulence and virulence patterns were remarkably different from that of CYR34 but quite similar to that of CYR32. Of the 165 wheat cultivars, 21 had all-stage resistance to TSA-6, 34 to TSA-9, and 20 to both races. Adult plant resistance (APR) was detected in 35 cultivars to TSA-6 and 27 to TSA-9, but only three cultivars showed APR to both new races. Slow rusting resistance was observed in 24 cultivars to TSA-6 and 33 to TSA-9. Analysis of variance of disease index indicated a significant difference between cultivars but not between the four races. Based on the molecular marker data, a low percentage of wheat cultivars carried Yr5, Yr7, Yr10, Yr15, Yr26, or YrSP. Because TSA-6 and TSA-9 can be a serious threat to wheat production in China, continual monitoring of TSA-6, TSA-9, and other races is needed.

Evidence of Occurrence of Barley Crown Rust Caused by Puccinia coronata var. hordei and Sexual Reproduction of the Pathogen Under Field Conditions in China.

Crown rust of barley, caused by Puccinia coronata var. hordei, was first reported by Jin and Steffenson in 1992, and the fungus has been reported only in the United States and Hungary. In China, stripe, stem, and leaf rusts have been reported on barley, but not for crown rust. Recently, a sample (HZJ0004) of rust collected from barley in Qilian county in Qinghai, China, appeared different from the three rusts based on color, size, arrangements of uredinia and/or telia. Teliospores had crown-shaped appendages on the top. Based on the disease symptoms and morphology of urediniospores and teliospores, the fungus was identified as P. coronata var. hordei. Using the internal transcribed spacer sequences, the isolates HZJ0004 from barley and POR3 from buckthorn (Rhamnus sp.) were clustered in one clade with P. coronata var. hordei isolates from barley and Elymus repens but in a different clade from the isolate POC8 from wild oat and the varieties of P. coronata from oats and grasses. At the seedling stage, most of the tested cultivars of barley and rye were susceptible to P. coronata var. hordei isolates HZJ0004 and POR3, but the cultivars of oats, triticale, wheat, and most grasses of genera Aegilops, Brachypodium, Bromus, Calamagrostis, Deschampsia, Elymus, Festuca, and Phleum were resistant, indicating their host specialization on barley. To our knowledge, this is the first report of crown rust on barley in China.

Repositioning liothyronine for cancer immunotherapy by blocking the interaction of immune checkpoint TIGIT/PVR.

BACKGROUND: Inhibitors targeting immune checkpoint were proved effective in cancer immunotherapy, such as PD-1/PD-L1 blockade. The novel immune checkpoint TIGIT/PVR plays critical roles in suppressing the anti-tumor effects of CD8+ T and NK cells, and dual blockade of TIGIT/PVR and PD-1/PD-L1 by antibody can elicit synergistic effects in tumor models and clinical trials. However, small molecules for TIGIT/PVR blockade have not been investigated. METHODS: The expression of PVR in tumors were analyzed by using TCGA, Oncomine and GEO database, and in cancer cell lines examined by flow cytometry. Natural product compounds were docked to PVR for virtual screening by using the software Molecular Operating Environment (MOE). Candidate compounds were further tested by biolayer interferometry-based binding assay, microscale thermophoresis assay and cell based blocking assay. The in vitro activity of the candidate compound was determined by MTT, peripheral blood mononuclear cells (PBMCs) activation assay and coculture assay. The anti-tumor effects and mechanism were also investigated by using MC38 tumor-bearing mice model and immune cell depletion tumor model. RESULTS: PVR was over-expressed in many tumor tissues and cancer cell lines, making it a promising therapeutic target. Through virtual screening, binding, and blocking assay, liothyronine was discovered to bind PVR and block the interaction of TIGIT/PVR. Liothyronine could enhance the function of CD4+ and CD8+ T cells in PBMCs. Besides, in the Jurkat-hTIGIT and CHOK1-hPVR coculture assay, liothyronine could reverse the IL-2 secretion inhibition resulted by TIGIT/PVR ligation. Although had no influence on the proliferation of tumor cells in vitro, liothyronine could significantly inhibit tumor growth when administrated in vivo, by enhancing CD8+ T cell infiltration and immune responses in the tumor bearing mice. The immune cell depletion model showed that the anti-tumor effects of liothyronine depends on CD4+ T cells, CD8+ T cells and NK cells. CONCLUSIONS: A small molecule liothyronine was discovered to serve as a potential candidate for cancer immunotherapy by blocking the immune checkpoint TIGIT/PVR. Video abstract.