WRN-targeted therapy using inhibitors NSC 19630 and NSC 617145 induce apoptosis in HTLV-1-transformed adult T-cell leukemia cells

Background

Human T-cell leukemia virus type 1 (HTLV-1) infection is associated with adult T-cell leukemia/lymphoma (ATLL), a lymphoproliferative malignancy with a dismal prognosis and limited therapeutic options. Recent evidence shows that HTLV-1-transformed cells present defects in both DNA replication and DNA repair, suggesting that these cells might be particularly sensitive to treatment with a small helicase inhibitor. Because the “Werner syndrome ATP-dependent helicase” encoded by the WRN gene plays important roles in both cellular proliferation and DNA repair, we hypothesized that inhibition of WRN activity could be used as a new strategy to target ATLL cells.

Methods

Our analysis demonstrates an apoptotic effect induced by the WRN helicase inhibitor in HTLV-1-transformed cells in vitro and ATL-derived cell lines. Inhibition of cellular proliferation and induction of apoptosis were demonstrated with cell cycle analysis, XTT proliferation assay, clonogenic assay, annexin V staining, and measurement of mitochondrial transmembrane potential.

Results

Targeted inhibition of the WRN helicase induced cell cycle arrest and apoptosis in HTLV-1-transformed leukemia cells. Treatment with NSC 19630 (WRN inhibitor) induces S-phase cell cycle arrest, disruption of the mitochondrial membrane potential, and decreased expression of anti-apoptotic factor Bcl-2. These events were associated with activation of caspase-3-dependent apoptosis in ATL cells. We identified some ATL cells, ATL-55T and LMY1, less sensitive to NSC 19630 but sensitive to another WRN inhibitor, NSC 617145.

Conclusions

WRN is essential for survival of ATL cells. Our studies suggest that targeting the WRN helicase with small inhibitors is a novel promising strategy to target HTLV-1-transformed ATL cells.

     

Heterologous expression of oxytetracycline biosynthetic gene cluster in Streptomyces venezuelae WVR2006 to improve production level and to alter fermentation process

Applied Microbiology and Biotechnology - Heterologous expression is an important strategy to activate biosynthetic gene clusters of secondary metabolites. Here, it is employed to activate and...     

microRNA-454-mediated NEDD4-2/TrkA/cAMP axis in heart failure: Mechanisms and cardioprotective implications

The current study aimed to investigate the mechanism by which miR-454 influences the progression of heart failure (HF) in relation to the neural precursor cell expressed, developmentally downregulated 4-2 (NEDD4-2)/tropomyosin receptor kinase A (TrkA)/cyclic adenosine 3',5'-monophosphate (cAMP) axis. Sprague-Dawley rats were used to establish a HF animal model via ligation of the left anterior descending branch of the coronary artery. The cardiomyocyte H9c2 cells were treated with H₂O₂ to stimulate oxidative stress injury in vitro. RT-qPCR and Western blot assay were subsequently performed to determine the expression patterns of miR-454, NEDD4-2, TrkA, apoptosis-related proteins and cAMP pathway markers. Dual-luciferase reporter gene assay coupled with co-immunoprecipitation was performed to elucidate the relationship between miR-454, NEDD4-2 and TrkA. Gain- or loss-of-function experiments as well as rescue experiments were conducted via transient transfection (in vitro) and adenovirus infection (in vivo) to examine their respective functions on H9c2 cell apoptosis and myocardial damage. Our results suggested that miR-454 was aberrantly downregulated in the context of HF, while evidence was obtained suggesting that it targeted NEDD4-2 to downregulate NEDD4-2 in cardiomyocytes. miR-454 exerted anti-apoptotic and protective effects on cardiomyocytes through inhibition of NEDD4-2, while NEDD4-2 stimulated ubiquitination and degradation of TrkA protein. Furthermore, miR-454 activated the cAMP pathway via the NEDD4-2/TrkA axis, which ultimately suppressed cardiomyocyte apoptosis and attenuated myocardial damage. Taken together, the key findings of the current study highlight the cardioprotective role of miR-454, which is achieved through activation of the cAMP pathway by impairing NEDD4-2-induced TrkA ubiquitination.     

Molecular identification and functional analysis of Niemann-Pick type C2 protein in Macrocentrus cingulum Brischke (Hymenoptera: Braconidae)

Niemann-Pick type C2 (NPC2) proteins in arthropods have been extensively differentiated and possibly duplicated according to environmental conditions and are probable to have different functions. The participation of NPC2 proteins in chemical communication in arthropods brings new objectives in environmental-friendly strategies for pest population control. In this study, NPC2 gene in Macrocentrus cingulum (McinNPC2) was newly identified by rapid amplification cDNA ends (RACE) technology. McinNPC2 amino acid sequence alignment with other representative NPC2 annotates to evaluate the highly conserved consensus amino acids, but with odorant binding proteins in M. cingulum show that only one consensus amino acid. Primary six-cysteine structures that are same to odorant binding proteins in M. cingulum were observed in McinNPC2. Phylogenetic analysis of McinNPC2 indicated that the nearest monophyletic group forming one clade with high posterior probability values clusters as Cyphomyrmex costatus (CcosNPC2) whereas the nearest evolutionary relation group as some odorant binding proteins. Moreover, quantitative real-time PCR (qPCR) measurements show that the McinNPC2 gene expression level in various tissues of the female is significantly and ubiquitously higher than in male, whereas the highest expression level in female antennae. We further explore the binding characterization of recombinant McinNPC2 to candidate odor molecules and did the modeling and docking simulations. The results showed ligands binding specificity and docking tests results indicate that β-ionone, an aroma compound commonly found in essential oils, can strongly bind with McinNPC2. In conclusion, we proposed that McinNPC2 may be involved in chemical communication and play roles in perception of plant volatiles.     

Lnc-PFAR facilitates autophagy and exacerbates pancreatic fibrosis by reducing pre-miR-141 maturation in chronic pancreatitis

Chronic pancreatitis (CP) is described as progressive inflammatory fibrosis of pancreas, accompanied with irreversible impaired endocrine and exocrine insufficiency. Pancreatic stellate cells (PSCs) are widely distributed in the stroma of the pancreas and PSCs activation has been shown as one of the leading causes for pancreatic fibrosis. Our previous study has revealed that autophagy is dramatically activated in CP tissues, which facilitates PSCs activation and pancreatic fibrosis. Long non-coding RNAs (LncRNAs) have been recognized as crucial regulators for fibrosis-related diseases. LncRNAs interact with RNA binding protein or construct competitive endogenous RNA (ceRNA) hypothesis which elicited the fibrotic processes. Until now, the effects of lncRNAs on PSCs activation and pancreatic fibrosis have not been clearly explored. In this study, a novel lncRNA named Lnc-PFAR was found highly expressed in mouse and human CP tissues. Our data revealed that Lnc-PFAR facilitates PSCs activation and pancreatic fibrosis via RB1CC1-induced autophagy. Lnc-PFAR reduces miR-141 expression by suppressing pre-miR-141 maturation, which eventually upregulates the RB1CC1 and fibrosis-related indicators expression. Meanwhile, Lnc-PFAR enhanced PSCs activation and pancreatic fibrosis through trigging autophagy. Our study interrogates a novel lncRNA-induced mechanism in promoting the development of pancreatic fibrosis, and Lnc-PFAR is suggested to be a prospective therapeutic target in clinical scenarios.Subject terms: RNAi, Diagnostic markers, Chronic pancreatitis     

Scale specific determinants of tree diversity in an old growth temperate forest in China

The major processes generating pattern in plant community composition depend upon the spatial scale and resolution of observation, therefore understanding the role played by spatial scale on species patterns is of major concern. In this study, we investigate how well environmental (topography and soil variables) and spatial variables explain variation in species composition in a 25-ha temperate forest in northeastern China. We used new variation partitioning approaches to discover the spatial scale (using multi-scale spatial PCNM variables) at which environmental heterogeneity and other spatially structured processes influence tree community composition. We also test the effect of changing grain of the study (i.e. quadrat size) on the variation partitioning results. Our results indicate that (1) species composition in the Changbai mixed broadleaf-conifer forest was controlled mainly by spatially structured soil variation at broad scales, while at finer scales most of the explained variation was of a spatial nature, pointing to the importance of biotic processes. (2) These results held at all sampling grains. However, reducing quadrat size progressively reduced both spatially and environmentally explained variance. This probably partly reflects increasing stochasticity in species abundances, and the smaller proportion of quadrats occupied by each species, when quadrat size is reduced. The results suggest that environmental heterogeneity (i.e. niche processes) and other biotic processes such as dispersal work together, but at different spatial scales, to build up diversity patterns.     

一种新型T细胞免疫原的原核表达及对口蹄疫疫苗的免疫增强作用

主要探讨了T细胞免疫原TI对口蹄疫疫苗的免疫增强作用。设计并原核表达产生了一种包含口蹄疫病毒VP1,VP4,3A和3D蛋白上多个T细胞表位与两个通用T细胞表位的T细胞免疫原,命名为TI;同时表达了O和Asia 1两个型口蹄疫病毒 VP1 蛋白的串联编码基因,表达产物命名为OA-VP1。将上述T细胞免疫原分别与OA-VP1和口蹄疫灭活疫苗按不同剂量组合免疫小鼠,于免疫后不同时间测定各组小鼠的体液与细胞免疫应答情况。采用微量中和试验检测小鼠O型和Asia1型中和抗体,采用流式细胞检测技术和测定γ-干扰素的水平来分析不同免疫组小鼠细胞免疫的水平。结果显示,与灭活疫苗或OA-VP1单独免疫组相比,添加TI抗原后灭活疫苗 (P＜0.01) 和OA-VP1免疫组(P＜0.05)小鼠均能产生高水平的特异性中和抗体;且CD4+ T细胞数量显著增多,IFN-γ产生水平显著升高 (P＜0.01)。说明TI抗原具有很好的诱导特异性体液与细胞免疫应答的作用,是一种很好的免疫增效剂,可作为口蹄疫蛋白亚单位疫苗和灭活疫苗中的一种有效成分,以提高疫苗的免疫效果。     

A novel xylanase,XynA4-2, from thermoacidophilic <Emphasis Type="Italic">Alicyclobacillus</Emphasis> sp. A4 with potential applications in the brewing industry

A xylanase gene, xynA4-2, was obtained from the genome sequence of thermoacidophilic Alicyclobacillus sp. A4 and expressed in Escherichia coli BL21 (DE3). xynA4-2 encodes a mature protein of 411 residues with a calculated molecular weight of 46.8 kDa. Based on the amino acid sequence similarities (highest identity of 61%), the enzyme was confined into glycoside hydrolase family 10. The purified recombinant XynA4-2 exhibited maximum activity at pH 6.2 and 55°C. The enzyme was stable over a broad pH range, retaining more than 90% of the original activity at pH 5.8–12.0, 37°C for 1 h. The substrate specificity of XynA4-2 was relatively narrow, exhibiting 100, 93, and 35% of the relative activity towards birchwood xylan, oat spelt xylan, and wheat arabinoxylan, respectively. Supplementation of XynA4-2 to mash caused the reduction of mash filtration rate (5.6%) and viscosity (4.0%). When combined with the commercial glucanase from Sunson, higher reduction was detected in the filtration rate (12.0%) and viscosity (17.2%). These favorable properties make XynA4-2 a good candidate in the brewing industry.     

Can the Study of Natural Vegetation Succession Assist in the Control of Soil Erosion on Abandoned Croplands on the Loess Plateau,China?

In the Loess Plateau, China, arable cultivation of slope lands is common and associated with serious soil erosion. Planting trees or grass may control erosion, but planted species may consume more soil water and can threaten long‐term ecosystem sustainability. Natural vegetation succession is an alternative ecological solution to restore degraded land, but there is a time cost, given that the establishment of natural vegetation, adequate to prevent soil erosion, is a longer process than planting. The aims of this study were to identify the environmental factors controlling the type of vegetation established on abandoned cropland and to identify candidate species that might be sown soon after abandonment to accelerate vegetation succession and establishment of natural vegetation to prevent soil erosion. A field survey of thirty‐three 2 × 2–m plots was carried out in July 2003, recording age since abandonment, vegetation cover, and frequency of species together with major environmental and soil variables. Data were analyzed using correspondence analysis, classification tree analysis, and species response curves. Four vegetation types were identified and the data analysis confirmed the importance of time since abandonment, total P, and soil water in controlling the type of vegetation established. Among the dominant species in the three late‐successional vegetation types, the most appropriate candidates for accelerating and directing vegetation succession were King Ranch bluestem (Bothriochloa ischaemum) and Lespedeza davurica (Leguminosae). These species possess combinations of the following characteristics: tolerance of low water and nutrient availability, fibrous root system and strong lateral vegetative spread, and a persistent seed bank.