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Viruses often usurp host machineries for their amplification, but it remains unclear if hosts may subvert virus proteins to regulate viral proliferation. Here, we show that the 17K protein, an important virulence factor conserved in barley yellow dwarf viruses (BYDVs) and related poleroviruses, is phosphorylated by host GRIK1-SnRK1 kinases, with the phosphorylated 17K (P17K) capable of enhancing the abundance of virus-derived small interfering RNAs (vsiRNAs) and thus antiviral RNAi. Furthermore, P17K interacts with barley small RNA-degrading nuclease 1 (HvSDN1) and impedes HvSDN1-catalyzed vsiRNA degradation. Additionally, P17K weakens the HvSDN1-HvAGO1 interaction, thus hindering HvSDN1 from accessing and degrading HvAGO1-carried vsiRNAs. Importantly, transgenic expression of 17K phosphomimetics (17K

Summary The LRK10‐like receptor kinases (LRK10L‐RLKs) are ubiquitously present in higher plants, but knowledge of their expression and function is still limited. Here, we report expression and functional analysis of TtdLRK10L‐1, a typical LRK10L‐RLK in durum wheat (Triticum turgidum L. ssp. durum). The introns of TtdLRK10L‐1 contained multiple kinds of predicted cis‐elements. To investigate the potential effect of these cis‐elements on TtdLRK10L‐1 expression and function, two types of transgenic wheat lines were prepared, which expressed a GFP‐tagged TtdLRK10L‐1 protein (TtdLRK10L‐1:GFP) from the cDNA or genomic DNA (gDNA) sequence of TtdLRK10L‐1 under the native promoter. TtdLRK10L‐1:GFP expression was up‐regulated by the powdery mildew pathogen Blumeria graminis f. sp. tritici (Bgt) in both types of transgenic plants, with the scale of the elevation being much stronger in the gDNA lines. Both types of transgenic plants exhibited enhanced resistance to Bgt infection relative to wild type control. Notably, the Bgt defence activated in the gDNA lines was significantly stronger than that in the cDNA lines. Further analysis revealed that a putative MYB transcription factor binding site (MYB‐BS, CAGTTA) located in TtdLRK10L‐1 intron I was critical for the efficient expression and function of TtdLRK10L‐1 in Bgt defence. This MYB‐BS could also increase the activity of a superpromoter widely used in ectopic gene expression studies in plants. Together, our results deepen the understanding of the expression and functional characteristics of LRK10L‐RLKs. TtdLRK10L‐1 is likely useful for further dissecting the molecular processes underlying wheat defence against Bgt and for developing Bgt resistant wheat crops.

Purpose We aimed to identify the anterolateral ligament (ALL) tears in anterior cruciate ligament (ACL)-deficient knees using standard 1.5-Tesla magnetic resonance imaging (MRI). Methods We included all patients who underwent primary ACL reconstruction at our center between 2012 and 2015. Exclusion criteria included patients with multiple ligament injuries, lateral collateral ligament, posterolateral corner, and infections, and patients who underwent MRI more than 2 months after their injury. All patients (n = 148) had ACL tears that were subsequently arthroscopically reconstructed. The magnetic resonance (MR) images of the injured knees performed within 2 months of injury were reviewed by a musculoskeletal radiologist and an orthopedic surgeon. The patients were divided into two groups. The first group of patients had MRI performed within 1 month of injury. The second group of patients had MRI performed 1–2 months after the index injury. Both assessors were blinded and the MR mages were read separately to assess the presence of ALL, presence of a tear and the location of the tear. Based on their readings, interobserver agreement (kappa statistic (K)), sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), and accuracy were compared. Results The ALL was identified in 100% of the patients. However, there was a discrepancy of up to 15% in the identification of tear of the ALL. In the first group in which MRI scans were performed within 1 month of injury, the ALL tear was identified by the radiologist in 92% of patients and by the surgeon in 90% of patients (Κ = 0.86). In the second group in which MRI scans were performed within 1–2 months of the injury, the ALL tear was identified by the radiologist in 78% of patients and by the surgeon in 93% of patients (K = 0.62). Conclusion The ALL can be accurately identified on MRI, but the presence and location of ALL tear and its location cannot be reliably identified on MRI. The accuracy in identification and characterization of a tear was affected by the interval between the time of injury and the time when the MRI was performed. Level of evidence Diagnostic, level IIIb, retrospective.

The invasion of osteoclasts into the cartilage via blood vessels advances the process of endochondral ossification, and dysregulation of dynamic intercellular interactions results in skeletal dysplasias. Although the regulation of osteoclasts by growth plate chondrocytes has been reported in detail, the effect of osteoclasts on chondrocytes remains to be determined. In this study, ATDC5 cells and bone marrow mesenchymal stem cells were differentiated into chondrocytes and treated with conditioned medium obtained from bone marrow macrophages differentiated to osteoclast precursors and osteoclasts. Exosomes were inhibited in conditioned medium or isolated directly from osteoclasts to further determine whether osteoclast-derived exosomes play an important role in chondrocyte hypertrophy. Additionally, exosomal miRNAs were detected, and let-7a-5p was selected as an miRNA with significantly increased expression in osteoclast-derived exosomes. Experiments were performed to verify the potential target Smad2 and investigate how let-7a-5p affected chondrocytes. The results suggest that both osteoclast precursors and osteoclasts promote chondrocyte hypertrophy and that the promotive effect of osteoclasts is more significant than that of osteoclast precursors. Osteoclast-derived exosomes promote the hypertrophic differentiation of chondrocytes. Moreover, osteoclast-derived exosomal let-7a-5p inhibits Smad2 to decrease the transforming growth factor-β-induced inhibition of chondrocyte hypertrophy. Our research reveals the role of osteoclasts in the regulation of chondrocytes and provides insights into the highly coordinated intercellular process of endochondral ossification.

Vasculogenesis (de novo formation of vessels) induced by endothelial progenitor cells (EPCs) is requisite for vascularized bone regeneration. However, there exist few available options for promoting vasculogenesis within artificial bone grafts except for exogenous EPC transplantation, which suffers from the source of EPC, safety, cost, and time concerns in clinical applications. This study aimed at endogenous EPC recruitment for vascularized bone regeneration by using a bioinspired EPC-induced graft. The EPC-induced graft was created by immobilizing two bioactive peptides, WKYMVm and YIGSR, on the surface of poly(ε-caprolactone) (PCL)/poliglecaprone (PGC) nanofibrous scaffolds via a polyglycolic acid (PGA)-binding peptide sequence. Remarkable immobilization efficacy of WKYMVm and YIGSR peptides and their sustained release (over 14 days) from scaffolds were observed. In vivo and in vitro studies showed robust recruitment of EPCs, which subsequently contributed to early vasculogenesis and ultimate bone regeneration. The dual-peptide-functionalized nanofibrous scaffolds proposed in this study provide a promising therapeutic strategy for vasculogenesis in bone defect repair.

Summary Powdery mildew disease, elicited by the obligate fungal pathogen Blumeria graminis f.sp. tritici (Bgt), causes widespread yield losses in global wheat crop. However, the molecular mechanisms governing wheat defense to Bgt are still not well understood.Here we found that TuACO3, encoding the 1‐aminocyclopropane‐1‐carboxylic acid (ACC) oxidase functioning in ethylene (ET) biosynthesis, was induced by Bgt infection of the einkorn wheat Triticum urartu, which was accompanied by increased ET content. Silencing TuACO3 decreased ET production and compromised wheat defense to Bgt, whereas both processes were enhanced in the transgenic wheat overexpressing TuACO3.TuMYB46L, phylogenetically related to Arabidopsis MYB transcription factor AtMYB46, was found to bind to the TuACO3 promoter region in yeast‐one‐hybrid and EMSA experiments. TuMYB46L expression decreased rapidly following Bgt infection. Silencing TuMYB46L promoted ET content and Bgt defense, but the reverse was observed when TuMYB46L was overexpressed.Hence, decreased expression of TuMYB46L permits elevated function of TuACO3 in ET biosynthesis in Bgt‐infected wheat. The TuMYB46L‐TuACO3 module regulates ET biosynthesis to promote einkorn wheat defense against Bgt. Furthermore, we found four chitinase genes acting downstream of the TuMYB46L‐TuACO3 module. Collectively, our data shed a new light on the molecular mechanisms underlying wheat defense to Bgt.

Angiogenesis is essential for successful bone defect repair. In normal tissue repair, the physiological inflammatory response is the main regulator of angiogenesis through the activity of macrophages and the cytokines secreted by them. In particular, M2 macrophages which secrete high levels of PDGF-BB are typically considered to promote angiogenesis. A hexapeptide [WKYMVm, (Trp-Lys-Tyr-Met-Val-D-Met-NH2)] has been reported to modulate inflammatory activities. However, the underlying mechanisms by which WKYMVm regulates macrophages remain unclear. In this study, the possible involvement by which WKYMVm induces the polarization of macrophages and affects their behaviors was evaluated. In vitro results showed that macrophages were induced to an M2 rather than M1 phenotype and the M2 phenotype was enhanced by WKYMVm through activation of the JAK1/STAT6 signaling pathway. It was also found that WKYMVm played an important role in the PDGF-BB production increase and proangiogenic abilities in M2 macrophages. Consistent with the results in vitro, the elevated M2/M0 ratio induced by WKYMVm enhanced the formation of new blood vessels in a femoral defect mouse model. In summary, these findings suggest that WKYMVm could be a promising alternative strategy for angiogenesis in bone repair by inducing M2 macrophage polarization.

Triticum urartu (AA, 2n = 2x = 14), a wild grass endemic to the Fertile Crescent (FC), is the progenitor of the A subgenome in common wheat. It belongs to the primary gene pool for wheat improvement. Here, we evaluated the yellow rust (caused by Puccinia striiformis f. sp. tritici, Pst) reactions of 147 T. urartu accessions collected from different parts of the FC. The reactions varied from susceptibility to strong resistance. In general, there were more accessions with stronger resistance to race CYR33 than to CYR 32. In most cases the main form of defense was a moderate resistance characterized by the presence of necrotic/chlorotic lesions with fewer Pst uredinia on the leaves. Forty two accessions displayed resistance to both races. Histological analysis showed that Pst growth was abundant in the compatible interaction but significantly suppressed by the resistant response. Gene silencing mediated by Barley stripe mosaic virus was effective in two T. urartu accessions with different resistance responses, indicating that this method can expedite future functional analysis of resistance genes. Our data suggest that T. urartu is a valuable source of resistance to yellow rust, and represents a model for studying the genetic, genomic and molecular basis underlying interaction between wheat and Pst. Keywords: Common wheat, Disease resistance, Gene silencing, Puccinia striiformis

A distinct class of mitosis regulators, conserved between plant and animal viruses, play active roles in viral pathogenesis., Many animal viral proteins, e.g., Vpr of HIV-1, disrupt host mitosis by directly interrupting the mitotic entry switch Wee1-Cdc25-Cdk1. However, it is unknown whether plant viruses may use this mechanism in their pathogenesis. Here, we report that the 17K protein, encoded by barley yellow dwarf viruses and related poleroviruses, delays G2/M transition and disrupts mitosis in both host (barley) and nonhost (fission yeast, Arabidopsis thaliana, and tobacco) cells through interrupting the function of Wee1-Cdc25-CDKA/Cdc2 via direct protein-protein interactions and alteration of CDKA/Cdc2 phosphorylation. When ectopically expressed, 17K disrupts the mitosis of cultured human cells, and HIV-1 Vpr inhibits plant cell growth. Furthermore, 17K and Vpr share similar secondary structural feature and common amino acid residues required for interacting with plant CDKA. Thus, our work reveals a distinct class of mitosis regulators that are conserved between plant and animal viruses and play active roles in viral pathogenesis.

Plants recognize pathogens and activate immune responses, which usually involve massive transcriptional reprogramming. The evolutionarily conserved kinase, Sucrose non-fermenting-related kinase 1 (SnRK1), functions as a metabolic regulator that is essential for plant growth and stress responses. Here, we identify barley SnRK1 and a WRKY3 transcription factor by screening a cDNA library. SnRK1 interacts with WRKY3 in yeast, as confirmed by pull-down and luciferase complementation assays. Förster resonance energy transfer combined with noninvasive fluorescence lifetime imaging analysis indicates that the interaction occurs in the barley nucleus. Transient expression and virus-induced gene silencing analyses indicate that WRKY3 acts as a repressor of disease resistance to the Bgh fungus. Barley plants overexpressing WRKY3 have enhanced fungal microcolony formation and sporulation. Phosphorylation assays show that SnRK1 phosphorylates WRKY3 mainly at Ser83 and Ser112 to destabilize the repressor, and WRKY3 non-phosphorylation-null mutants at these two sites are more stable than the wild-type protein. SnRK1-overexpressing barley plants display enhanced disease resistance to Bgh. Transient expression of SnRK1 reduces fungal haustorium formation in barley cells, which probably requires SnRK1 nuclear localization and kinase activity. Together, these findings suggest that SnRK1 is directly involved in plant immunity through phosphorylation and destabilization of the WRKY3 repressor, revealing a new regulatory mechanism of immune derepression in plants., Plants activate immune responses that usually involve massive transcriptional reprogramming. The metabolic sensor SnRK1 interacts with and phosphorylates the repressor WRKY3 in barley, leading to the degradation of WRKY3 and enhanced barley immunity against powdery mildew. This study data reveals a new regulatory mechanism of immune derepression in plants.

The balance between bone formation and bone resorption is closely related to bone homeostasis. Osteoclasts, originating from the monocyte/macrophage lineage, are the only cell type possessing bone resorption ability. Osteoclast overactivity is thought to be the major reason underlying osteoclast‐related osteolytic problems, such as Paget's disease, aseptic loosening of prostheses and inflammatory osteolysis; therefore, disruption of osteoclastogenesis is considered a crucial treatment option for these issues. WKYMVm, a synthetic peptide, which is a potent FPR2 agonist, exerts an immunoregulatory effect. This peptide inhibits the production of inflammatory cytokines, such as (IL)‐1β and TNF‐α, thus regulating inflammation. However, there are only few reports on the role of WKYMVm and FPR2 in osteoclast cytology. In the current study, we found that WKYMVm negatively regulates RANKL‐ and lipopolysaccharide (LPS)‐induced osteoclast differentiation and maturation in vitro and alleviates LPS‐induced osteolysis in animal models. WKYMVm down‐regulated the expression of osteoclast marker genes and resorption activity. Furthermore, WKYMVm inhibited osteoclastogenesis directly through reducing the phosphorylation of STAT3 and NF‐kB and indirectly through the CD9/gp130/STAT3 pathway. In conclusion, our findings demonstrated the potential medicinal value of WKYMVm for the treatment of inflammatory osteolysis.

The accuracy of magnetic resonance (MR) imaging in assessing meniscal and cartilage injuries in anterior cruciate ligament (ACL)-deficient knees as compared to arthroscopy was evaluated in the present study. The results of all preoperative MR imaging performed within 3 months prior to the ACL reconstruction were compared against intraoperative arthroscopic findings. A total of 206 patients were identified. The location and type of meniscal injuries as well as the location and grade of the cartilage injuries were studied. The negative predictive value, positive predictive value, sensitivity, specificity and accuracy of MR imaging for these 206 cases were calculated and analysed. In patients with an ACL injury, the highest incidence of concomitant injury was that of medial meniscus tears, 124 (60.2 %), followed by lateral meniscus tears, 105 (51.0 %), and cartilage injuries, 66 (32.0 %). Twenty-three (11.2 %) patients sustained injuries to all of the previously named structures. MR imaging was most accurate in detecting medial meniscus tears (85.9 %). MR imaging for medial meniscus tears also had the highest sensitivity (88.0 %) and positive predictive value (88.7 %), while MR imaging for cartilage injuries had the largest specificity (84.1 %) and negative predictive value (87.1 %). It was least accurate in evaluating lateral meniscus tears (74.3 %). The diagnostic accuracy of medial meniscus imaging is significantly influenced by age and the presence of lateral meniscus tears, while the duration between MR imaging and surgery has greater impact on the likelihood of lateral meniscus and cartilage injuries actually being present during surgery. The majority of meniscus tears missed by MR imaging affected the posterior horn and were complex in nature. Cartilage injuries affecting the medial femoral condyle or medial patella facet were also often missed by MR imaging. MR imaging remains a reliable tool for assessing meniscus tears and cartilage defects preoperatively. It is most accurate when evaluating medial meniscus tears. However, MR imaging should be used with discretion especially if there is a high index of suspicion of lateral meniscus tears. IV.