AURKA inhibitor-induced PD-L1 upregulation impairs antitumor immune responses.

Introduction: Tumor immunotherapy targeting PD-L1 has emerged as one of the powerful tools for tumor therapy. Numerous studies indicate that tumor-targeted drugs critically have an influence on the interaction between the immune system and tumors by changing the expression of PD-L1, which is beneficial for immunotherapy. Our study provided novel evidence for improving the drug regimen in tumor targeted therapy and immunotherapy. Methods: The expression of PD-L1 on SKBR3, MDA-MB-231, MCF7, 4T1, MC38 and B16 cells was evaluated by flow cytometry after treatment with six preclinical targeted drugs (ARN-509, AZD3514, Galeterone, Neratinib, MLN8237 and LGK974). AURKA was knockdowned by using the specific siRNA or CRISPR-Cas9 technology. In the 4T1-breast tumor and colorectal cancer xenograft tumor models, we determined the number of infiltrated CD3+ and CD8+ T cells in tumor tissues by IHC. Results: We found that AURKA inhibitor MLN8237 promoted the expression of PD-L1 in a time- and concentration-dependent manner while exerted its antitumor effect. Knockdown of AURKA could induce the upregulation of PD-L1 on SKBR3 cells. MLN8237-induced PD-L1 upregulation was mainly associated with the phosphorylation of STAT3. In the 4T1-breast tumor xenograft model, the infiltrated CD3+ and CD8+ T cells decreased after treatment with MLN8237. When treated with MLN8237 in combination with anti-PD-L1 antibody, the volumes of tumor were significantly reduced and accompanied by increasing the infiltration of CD3+ and CD8+ T cells in colorectal cancer xenograft tumor model. Discussion: Our data demonstrated that MLN8237 improved the effect of immunology-related therapy on tumor cells by interacting with anti-PD-L1 antibody, which contributed to producing creative sparks for exploring the possible solutions to overcoming drug resistance to tumor targeted therapy.

Direct Evidence Demonstrates that Puccinia striiformis f. sp. tritici Infects Susceptible Barberry to Complete Sexual Cycle in Autumn.

Wheat stripe rust is an airborne and destructive disease caused by a heteroecious rust fungus Puccinia striiformis f. sp. tritici (Pst). Studies have demonstrated that the rust pathogen accomplishes sexual reproduction on susceptible barberry under natural conditions in spring, whereas Pst infection on barberry is still in blank in other seasons. In late October 2016, aecial production on barberry shrubs were observed in Linzhi, Tibet, China. Therefore, experimental tests were conducted to verify the existence of sexual cycles of Pst in this season. By inoculating 52 aecial clusters from 30 rusted barberry leaves, four Pst samples, T1 to T4, were successfully recovered from the rusted barberry shrubs. Sixty-five single uredinium (SU) isolates were derived from the four Pst samples. Based on virulence tests on the Chinese differential hosts, T1 to T4 samples were unknown races and showed mixed reactions on some differentials. Twenty-one known races and 44 unknown races belonging to five race groups were identified among the 65 SU isolates. Meanwhile, the 65 SU isolates produced 26 various virulence patterns (VPs; called VP1-VP26) on 25 single Yr gene lines and 15 multilocus genotypes (MLGs) at nine simple sequence repeat marker loci. Clustering analysis showed similar lineage among subpopulations and different lineage between subpopulations. Linkage disequilibrium analysis indicated that the SU population was produced sexually. This study first reported that Pst infects susceptible barberry to complete sexual reproduction in autumn. The results update the knowledge of disease cycle and management of wheat stripe rust and contribute to the understanding of rust genetic diversity in Tibet.

Molecular Characterization of Novel Totivirus-Like Double-Stranded RNAs from Puccinia striiformis f. sp. tritici, the Causal Agent of Wheat Stripe Rust.

Characterization of newly isolated mycoviruses may contribute to understanding of the evolution and diversity of viruses. Here, a deep sequencing approach was used to analyze the double-stranded RNA (dsRNA) mycoviruses isolated from field-collected P. striiformis samples in China. Database searches showed the presence of at least four totivirus-like sequences, termed Puccinia striiformis virus 1 to 4 (PsV1 to 4). All of these identified sequences contained two overlapping open reading frames (ORFs) which encode a putative coat protein (CP) and an RNA-dependent RNA polymerase (RdRp) showing similar structures to members of the genus Totivirus. Each PsV contained a -1 ribosomal frameshifting region with a slippery site and a pseudoknot structure in the overlapped regions of these ORFs, indicating that the RdRp is translated as a CP-RdRp fusion. Phylogenetic analyses based on RdRp and CP suggested that these novel viruses belong to the genus Totivirus in the family Totiviridae. The presences of these PsVs were further validated by transmission electron microscope (TEM) and RT-PCR. Taken together, our results demonstrate the presence of diverse, novel totiviruses in the P. striiformis field populations.

Identification of a Novel Alternaria alternata Strain Able to Hyperparasitize Puccinia striiformis f. sp. tritici, the Causal Agent of Wheat Stripe Rust.

The obligate bitrophic fungus Puccinia striiformis f. sp. tritici (Pst) causes stripe (yellow) rust on wheat worldwide. Here, we report a novel fungal strain able to hyperparasitize Pst. The strain was isolated from gray-colored rust pustules, and was identified as Alternaria alternata (Fr.: Fr.) keissler based on a combination of morphological characteristics and multi-locus (ITS, GAPDH, and RPB2) phylogeny. Upon artificial inoculation, the hyperparasite reduced the production and viability of urediniospores, and produced a typical gray-colored rust pustule symptom. Scanning electron microscopy demonstrated that the strain could efficiently penetrate and colonize Pst urediniospores. This study first demonstrates that A. alternata could parasitize Pst and indicates its potential application in the biological control of wheat stripe rust disease.

Screening for simple sequence repeat markers in Puccinia striiformis tritici based on genomic sequence.

Puccinia striiformis f. sp. tritici (Pst) is the obligate biotrophic fungus responsible for stripe rust wheat. In this study, we developed and characterized 20 polymorphic microsatellite markers from the genomic sequence of an isolate of Chinese Pst race CY32. Polymorphism at each simple sequence repeat (SSR) locus was determined using 32 Pst isolates from 7 countries. The number of alleles varied from 2 to 7 across isolates, and the observed and expected heterozygosities ranged from 0.33 to 0.97 (mean 0.62) and 0.23 to 0.73 (mean 0.51), respectively. As expected the genomic SSR markers were more polymorphic than the expressed sequence tag (EST)-SSR markers developed previously. These markers will be more useful for population genetics and molecular genetics studies in Pst.