Cyclin B2 impairs the p53 signaling in nasopharyngeal carcinoma.

BACKGROUND: Cyclin B2 (CCNB2), a member of the cyclin family, is an oncogene in multiple cancers, including nasopharyngeal carcinoma (NPC). However, the epigenetics mechanism for CCNB2 overexpression in NPC remains unclear. This study dissects the regulatory role of CCNB2 in NPC and the molecular mechanism. METHODS: Differentially methylated genes (DMG) and differentially expressed genes (DEG) were screened out in GSE52068 and GSE13597 databases, respectively, and candidate targets were identified by the Venn diagram. GO annotation and pathway enrichment analyses were performed on selected DMG and DEG, and a PPI network was constructed to pinpoint hub genes. PCR and qMSP were conducted to detect the expression and methylation of CCNB2 in cells. The siRNA targeting CCNB2 was transfected into NPC cells, and the migration, proliferation, cell cycle, epithelial-mesenchymal transition (EMT), tumorigenesis, and metastasis were examined. The upstream factor responsible for CCNB2 overexpression in NPC was explored. The p53 activity in NPC cells was assessed using western blot analysis. RESULTS: CCNB2 showed hypomethylation and overexpression in NPC. CCNB2 silencing inhibited cell migration, proliferation, cell cycle entry, and EMT. JMJD6 was overexpressed in NPC and upregulated CCNB2 through demethylation. JMJD6 reversed the effects of CCNB2 downregulation, resulting in elevated cellular activity in vitro and tumorigenic and metastatic activities in vivo. CCNB2 blocked the p53 pathway, while the p53 pathway inhibitor reversed the effect of CCNB2 silencing to increase the activity of NPC cells. CONCLUSIONS: JMJD6 enhanced CCNB2 transcription by demethylating CCNB2, thereby repressing the p53 pathway and promoting NPC progression.

Bacteria Exploit Autophagy for Proteasome Degradation and Enhanced Virulence in Plants.

Autophagy and the ubiquitin-proteasome system (UPS) are two major protein degradation pathways implicated in the response to microbial infections in eukaryotes. In animals, the contribution of autophagy and the UPS to antibacterial immunity is well documented and several bacteria have evolved measures to target and exploit these systems to the benefit of infection. In plants, the UPS has been established as a hub for immune responses and is targeted by bacteria to enhance virulence. However, the role of autophagy during plant-bacterial interactions is less understood. Here, we have identified both pro- and antibacterial functions of autophagy mechanisms upon infection of Arabidopsis thaliana with virulent Pseudomonas syringae pv tomato DC3000 (Pst). We show that Pst activates autophagy in a type III effector (T3E)-dependent manner and stimulates the autophagic removal of proteasomes (proteaphagy) to support bacterial proliferation. We further identify the T3E Hrp outer protein M1 (HopM1) as a principle mediator of autophagy-inducing activities during infection. In contrast to the probacterial effects of Pst-induced proteaphagy, NEIGHBOR OF BRCA1-dependent selective autophagy counteracts disease progression and limits the formation of HopM1-mediated water-soaked lesions. Together, we demonstrate that distinct autophagy pathways contribute to host immunity and bacterial pathogenesis during Pst infection and provide evidence for an intimate crosstalk between proteasome and autophagy in plant-bacterial interactions.

Transcriptional stimulation of rate-limiting components of the autophagic pathway improves plant fitness.

Autophagy is a major catabolic process whereby autophagosomes deliver cytoplasmic content to the lytic compartment for recycling. Autophagosome formation requires two ubiquitin-like systems conjugating Atg12 with Atg5, and Atg8 with lipid phosphatidylethanolamine (PE), respectively. Genetic suppression of these systems causes autophagy-deficient phenotypes with reduced fitness and longevity. We show that Atg5 and the E1-like enzyme, Atg7, are rate-limiting components of Atg8-PE conjugation in Arabidopsis. Overexpression of ATG5 or ATG7 stimulates Atg8 lipidation, autophagosome formation, and autophagic flux. It also induces transcriptional changes opposite to those observed in atg5 and atg7 mutants, favoring stress resistance and growth. As a result, ATG5- or ATG7-overexpressing plants exhibit increased resistance to necrotrophic pathogens and oxidative stress, delayed aging and enhanced growth, seed set, and seed oil content. This work provides an experimental paradigm and mechanistic insight into genetic stimulation of autophagy in planta and shows its efficiency for improving plant productivity.

Retromer contributes to immunity-associated cell death in Arabidopsis.

Membrane trafficking is required during plant immune responses, but its contribution to the hypersensitive response (HR), a form of programmed cell death (PCD) associated with effector-triggered immunity, is not well understood. HR is induced by nucleotide binding-leucine-rich repeat (NB-LRR) immune receptors and can involve vacuole-mediated processes, including autophagy. We previously isolated lazarus (laz) suppressors of autoimmunity-triggered PCD in the Arabidopsis thaliana mutant accelerated cell death11 (acd11) and demonstrated that the cell death phenotype is due to ectopic activation of the LAZ5 NB-LRR. We report here that laz4 is mutated in one of three VACUOLAR PROTEIN SORTING35 (VPS35) genes. We verify that LAZ4/VPS35B is part of the retromer complex, which functions in endosomal protein sorting and vacuolar trafficking. We show that VPS35B acts in an endosomal trafficking pathway and plays a role in LAZ5-dependent acd11 cell death. Furthermore, we find that VPS35 homologs contribute to certain forms of NB-LRR protein-mediated autoimmunity as well as pathogen-triggered HR. Finally, we demonstrate that retromer deficiency causes defects in late endocytic/lytic compartments and impairs autophagy-associated vacuolar processes. Our findings indicate important roles of retromer-mediated trafficking during the HR; these may include endosomal sorting of immune components and targeting of vacuolar cargo.

[Effects of surgical operation on helper-inducer T-lymphocytes in sino-nasal neoplasms patients].

OBJECTIVE: To explore the effects of surgical operation on helper-inducer T-lymphocytes (Th1/Th2) in sino-nasal neoplasms. METHODS: From January 2004 to December 2011, 80 patients with malignant tumor in nasal cavities or sinuses were enrolled as experimental group and another 80 subjects with deflection of nasal septum as control group. The phorbol-12-myristate-13-acetate (PMA)-stimulated peripheral blood mononuclear cells (PBMC) were obtained from both malignant tumor and normal control tissues. And flow cytometry was used to detect the expression percentages of interleukin-4 (IL-4) in Th2 cell and interferon-gamma (IFN-γ) in Th1 cell at pre and post-operation. RESULTS: The preoperative expression percentage of CD3(+) CD8(-) IL-4(+) cell in malignant tumor patients was higher than that in controls (6.6% ± 1.7% vs 2.8% ± 1.7%, P < 0.05) while the expression percentage of CD3(+) CD8(-) IFN-γ(+) cell was lower (18.7% ± 5.7% vs 59.3% ± 1.5%, P < 0.05). The preoperative expression percentage of CD3(+) CD8(-) IL-4(+) cell was higher than that at postoperation (6.6% ± 1.7 %vs 2.8% ± 1.5%, P < 0.05). And the postoperative expression percentage of CD3(+) CD8(-) IFN-γ(+) cell was higher than that at preoperation (54.0% ± 4.0% vs 18.7% ± 5.7%, P < 0.05). CONCLUSION: Surgery may restore the immune balance in patients with malignant tumors in nasal cavities or sinuses.

Spray-Induced Gene Silencing to Study Gene Function in Phytophthora.

RNA interference (RNAi) is a conserved cellular defense mechanism mediated by double-stranded RNA (dsRNA) that can regulate gene expression through targeted destruction of mRNAs (messenger RNAs). Recent studies have shown that spraying dsRNAs or small RNAs (sRNAs) that target essential genes of pathogens on plant surfaces can confer protection against pests and pathogens. Also called spray-induced gene silencing (SIGS), this strategy can be used for disease control and for transient gene silencing to study the function of genes in plant-pathogen interactions. Furthermore, as sRNAs can move locally, systemically, and cross-kingdom during plant-microbe interactions, SIGS allows quick detection and characterization of gene functions in pathogens and plants.

Twigs of dove tree in high-latitude region tend to increase biomass accumulation in vegetative organs but decrease it in reproductive organs.

Adaptive traits are an important dimension for studying the interactions between rare plants and environment. Although the endangered mechanism of rare plants has been reported in many studies, how their twigs adapt to heterogeneous environments associated with latitude is still poorly known. Dove tree (Davidia involucrata Baill.), a monotypic rare species in China, was employed as a model species in our study, and the differences in functional traits, growth relationships and resource allocation among components of annual twig were investigated in three latitudinal regions (32°19' N, 30°08' and 27°55') in the Sichuan, Southwest China. Compared with low- and middle-latitude regions, the twig diameter in high-latitude region decreased by 36% and 26%, and dry mass decreased by 32% and 35%, respectively. Moreover, there existed an allometric growth between flower mass and stem mass or leaf mass in high-latitude region but an isometric growth in low- and middle-latitude regions. At the flower level, an isometric growth between bract area and flower stalk mass was detected among in three latitudinal regions, and the flower stalk mass in the low-latitude region was higher than in the middle- and high-latitude regions for a given bract area and flower mass. At the leaf level, the growth rate of petiole mass was significantly higher than those of leaf area, lamina mass and leaf mass among three latitudinal regions, and the petiole mass in the low-latitude region was higher than in the other two regions for a given leaf mass. Our research demonstrated that the twigs of dove tree in high-latitude region tend to become smaller, and resource input increase in stems and leaves but decrease in flowers, which reflects that dove tree can adapt to the environmental changes across different latitudes by adjusting phenotypic traits growth and biomass allocation of twigs.

A paralog of the MtN3/saliva family recessively confers race-specific resistance to Xanthomonas oryzae in rice.

Approximately one third of the identified 34 rice major disease resistance (R) genes conferring race-specific resistance to different strains of Xanthomonas oryzae pv. oryzae (Xoo), which causes rice bacterial blight disease, are recessive genes. However, only two of the recessive resistance genes have been characterized thus far. Here we report the characterization of another recessive resistance gene, xa25, for Xoo resistance. The xa25, localized in the centromeric region of chromosome 12, mediates race-specific resistance to Xoo strain PXO339 at both seedling and adult stages by inhibiting Xoo growth. It encodes a protein of the MtN3/saliva family, which is prevalent in eukaryotes, including mammals. Transformation of the dominant Xa25 into a resistant rice line carrying the recessive xa25 abolished its resistance to PXO339. The encoding proteins of recessive xa25 and its dominant allele Xa25 have eight amino acid differences. The expression of dominant Xa25 but not recessive xa25 was rapidly induced by PXO339 but not other Xoo strain infections. The nature of xa25-encoding protein and its expression pattern in comparison with its susceptible allele in rice-Xoo interaction indicate that the mechanism of xa25-mediated resistance appears to be different from that conferred by most of the characterized R proteins.

Vacuole Integrity Maintained by DUF300 Proteins Is Required for Brassinosteroid Signaling Regulation.

Brassinosteroid (BR) hormone signaling controls multiple processes during plant growth and development and is initiated at the plasma membrane through the receptor kinase BRASSINOSTEROID INSENSITIVE1 (BRI1) together with co-receptors such as BRI1-ASSOCIATED RECEPTOR KINASE1 (BAK1). BRI1 abundance is regulated by endosomal recycling and vacuolar targeting, but the role of vacuole-related proteins in BR receptor dynamics and BR responses remains elusive. Here, we show that the absence of two DUF300 domain-containing tonoplast proteins, LAZARUS1 (LAZ1) and LAZ1 HOMOLOG1 (LAZ1H1), causes vacuole morphology defects, growth inhibition, and constitutive activation of BR signaling. Intriguingly, tonoplast accumulation of BAK1 was substantially increased and appeared causally linked to enhanced BRI1 trafficking and degradation in laz1 laz1h1 plants. Since unrelated vacuole mutants exhibited normal BR responses, our findings indicate that DUF300 proteins play distinct roles in the regulation of BR signaling by maintaining vacuole integrity required to balance subcellular BAK1 pools and BR receptor distribution.

Characterization of a disease susceptibility locus for exploring an efficient way to improve rice resistance against bacterial blight.

Bacterial blight caused by Xanthomonas oryzae pv. oryzae (Xoo) is the most harmful bacterial disease of rice worldwide. Previously, we characterized major disease resistance (MR) gene xa25, which confers race-specific resistance to Xoo strain PXO339. The xa25 is a recessive allele of the SWEET13 locus, but SWEET13's interaction with PXO339 and how efficiently using this locus for rice breeding still need to be defined. Here we show that the SWEET13 allele from rice Zhenshan 97 is a susceptibility gene to PXO339. Using this allele's promoter to regulate xa25 resulted in disease, suggesting that the promoter is a key determinant in SWEET13 caused disease in Zhanshan 97 after PXO339 infection. PXO339 transcriptionally induces SWEET13 to cause disease. Partial suppressing SWEET13 expression leads to a high level of resistance to PXO339. Thus, the transcriptionally suppressed SWEET13 functions as xa25 in resistance to PXO339. Hybrid rice is widely grown in many countries. However, recessive MR genes have not been efficiently used for disease resistance breeding in hybrid rice production for both parents of the hybrid have to carry the same recessive gene. However, the suppressed SWEET13 functions dominantly, which will have advantage to improve the resistance of hybrid rice to xa25-incomptible Xoo.

Chromatin assembly factor CAF-1 represses priming of plant defence response genes.

Plants have evolved efficient defence systems against pathogens that often rely on specific transcriptional responses. Priming is part of the defence syndrome, by establishing a hypersensitive state of defence genes such as after a first encounter with a pathogen. Because activation of defence responses has a fitness cost, priming must be tightly controlled to prevent spurious activation of defence. However, mechanisms that repress defence gene priming are poorly understood. Here, we show that the histone chaperone CAF-1 is required to establish a repressed chromatin state at defence genes. Absence of CAF-1 results in spurious activation of a salicylic acid-dependent pathogen defence response in plants grown under non-sterile conditions. Chromatin at defence response genes in CAF-1 mutants under non-inductive (sterile) conditions is marked by low nucleosome occupancy and high H3K4me3 at transcription start sites, resembling chromatin in primed wild-type plants. We conclude that CAF-1-mediated chromatin assembly prevents the establishment of a primed state that may under standard non-sterile growth conditions result in spurious activation of SA-dependent defence responses and consequential reduction of plant vigour.

Molecular rationale for the use of PI3K/AKT/mTOR pathway inhibitors in combination with crizotinib in ALK-mutated neuroblastoma.

Mutations in the ALK tyrosine kinase receptor gene represent important therapeutic targets in neuroblastoma, yet their clinical translation has been challenging. The ALK(F1174L) mutation is sensitive to the ALK inhibitor crizotinib only at high doses and mediates acquired resistance to crizotinib in ALK-translocated cancers. We have shown that the combination of crizotinib and an inhibitor of downstream signaling induces a favorable response in transgenic mice bearing ALK(F1174L)/MYCN-positive neuroblastoma. Here, we investigated the molecular basis of this effect and assessed whether a similar strategy would be effective in ALK-mutated tumors lacking MYCN overexpression. We show that in ALK-mutated, MYCN-amplified neuroblastoma cells, crizotinib alone does not affect mTORC1 activity as indicated by persistent RPS6 phosphorylation. Combined treatment with crizotinib and an ATP-competitive mTOR inhibitor abrogated RPS6 phosphorylation, leading to reduced tumor growth and prolonged survival in ALK(F1174L)/MYCN-positive models compared to single agent treatment. By contrast, this combination, while inducing mTORC1 downregulation, caused reciprocal upregulation of PI3K activity in ALK-mutated cells expressing wild-type MYCN. Here, an inhibitor with potency against both mTOR and PI3K was more effective in promoting cytotoxicity when combined with crizotinib. Our findings should enable a more precise selection of molecularly targeted agents for patients with ALK-mutated tumors.

Effects of budesonide on the expression of the glucocorticoid receptor-α in nasal polyp epithelial cells.

BACKGROUND: This study explores effects of budesonide on the proliferation of nasal polyp epithelial cells and expression of the glucocorticoid receptor (GR) alpha in nasal polyp epithelial cells. METHODS: Primary cultured, purified, and identified the epithelial cells collected from nasal polyps. The proliferation of nasal polyp epithelial cells was examined by a cell counting kit, and expression of GR-alpha mRNA in nasal polyp epithelial cells was examined by reverse transcription polymerase chain reaction, after training nasal polyp epithelial cells in budesonide solution. RESULTS: The average survival rate of nasal polyp epithelial cells was the lowest in 1 × 10(-6) M budesonide solution (29.284 ± 0.311%), compared with other concentrations. Budesonide at 1 × 10(-8) M caused down-regulation of GR-alpha mRNA expression levels at 6 and 12 ours, compared with the 0-hour group (p < 0.001); compared with the 0-hour group, there were significantly lower expression levels of GR-alpha mRNA at both 24 and 48 hours (p < 0.001); Expression of GR-alpha mRNA at either 48 or 12 hours was not significantly different from that at 24 hours. CONCLUSION: Budesonide can significantly inhibit the proliferation of nasal polyp epithelial cells, down-regulate the expression of GR-alpha mRNA in nasal polyp epithelial cells with time dependence.