Conversion of the bifunctional 8-oxoguanine/beta-delta apurinic/apyrimidinic DNA repair activities of Drosophila ribosomal protein S3 into the human S3 monofunctional beta-elimination catalyst through a single amino acid change

The Drosophila S3 ribosomal protein has important roles in both protein translation and DNA repair. In regards to the latter activity, it has been shown that S3 contains vigorous N-glycosylase activity for the removal of 8-oxoguanine residues in DNA that leaves baseless sites in their places. Drosophila S3 also possesses an apurinic/apyrimidinic (AP) lyase activity in which the enzyme catalyzes a beta-elimination reaction that cleaves phosphodiester bonds 3' and adjacent to an AP lesion in DNA. In certain situations, this is followed by a delta-elimination reaction that ultimately leads to the formation of a single nucleotide gap in DNA bordered by 5'- and 3'-phosphate groups. The human S3 protein, although 80% identical to its Drosophila homolog and shorter by only two amino acids, has only marginal N-glycosylase activity. Its lyase activity only cleaves AP DNA by a beta-elimination reaction, thus further distinguishing itself from the Drosophila S3 protein in lacking a delta-elimination activity. Using a hidden Markov model analysis based on the crystal structures of several DNA repair proteins, the enzymatic differences between Drosophila and human S3 were suggested by the absence of a conserved glutamine residue in human S3 that usually resides at the cleft of the deduced active site pocket of DNA glycosylases. Here we show that the replacement of the Drosophila glutamine by an alanine residue leads to the complete loss of glycosylase activity. Unexpectedly, the delta-elimination reaction at AP sites was also abrogated by a change in the Drosophila glutamine residue. Thus, a single amino acid change converted the Drosophila activity into one that is similar to that possessed by the human S3 protein. In support of this were experiments executed in vivo that showed that human S3 and the Drosophila site-directed glutamine-changed S3 performed poorly when compared with Drosophila wild-type S3 and its ability to protect a bacterial mutant from the harmful effects of DNA-damaging agents.     

Fate of nitrogen applied as Azolla and blue-green algae (Cyanobacteria) in waterlogged rice soils—a15N tracer study

Summary ¹⁵N tracer was used to detect the extent to which nitrogen of appliedAzolla caroliniana, Anabaena variabilis andNostoc muscorum was available for assimilation by the growing rice plants in pots under 4 cm flood water for 60 days. The rate of release of nitrogen from the above biofertilizers, the amount of nitrogen remaining in the soils and the amount that was lost from the soils during this period were also examined. Previously¹⁵N-labelled biomass of Azolla, Anabaena and Nostoc to provide 40 mg N was mixed thoroughly with 0.5 kg silt loam Bangladesh soil (Sonatola series) in each of three pots used for a single treatment. Each pot received four 16 days old IR8 rice seedlings. A parallet set of experiments was conducted without rice plants.It was found that nitrogen uptake in the rice plants was increased by 91, 176 and 215% on using Azolla, Anabaena and Nostoc which resulted in increased total dry matter yields (shoot plus root) of 74, 105 and 125%, respectively. Of the total¹⁵N applied at the start, 26, 49 and 53% was released from Azolla, Anabaena and Nostoc; about 7, 14 and 13% was lost by denitrification and 74, 51 and 47% remained in the soils as the undecomposed part of the biofertilizers, respeciively, after 60 days. Of 15.76, 22.72 and 25.92 mg N assimilated by the rice plants, 48, 61 and 62% was supplied by Azolla, Anabaena and Nostoc, respectively. The rest was obtained from the soil used.In the absence of the rice plants 30, 43 and 45% of applied¹⁵N of Azolla, Anabaena and Nostoc was released, respectively, in 60 days of which 93–96% was lost as N₂ through denitrification.     

Characterization of the Factors that Influence Sinapine Concentration in Rapeseed Meal during Fermentation

We analyzed and compared the difference in sinapine concentration in rapeseed meal between the filamentous fungus, Trametes sp 48424, and the yeast, Saccharomyces cerevisiae, in both liquid and solid-state fermentation. During liquid and solid-state fermentation by Trametes sp 48424, the sinapine concentration decreased significantly. In contrast, the liquid and solid-state fermentation process by Saccharomyces cerevisiae just slightly decreased the sinapine concentration (P ≤ 0.05). After the solid-state fermented samples were dried, the concentration of sinapine in rapeseed meal decreased significantly in Saccharomyces cerevisiae. Based on the measurement of laccase activity, we observed that laccase induced the decrease in the concentration of sinapine during fermentation with Trametes sp 48424. In order to eliminate the influence of microorganisms and the metabolites produced during fermentation, high moisture rapeseed meal and the original rapeseed meal were dried at 90°C and 105°C, respectively. During drying, the concentration of sinapine in high moisture rapeseed meal decreased rapidly and we obtained a high correlation coefficient between the concentration of sinapine and loss of moisture. Our results suggest that drying and enzymes, especially laccase that is produced during the solid-state fermentation process, may be the main factors that affect the concentration of sinapine in rapeseed meal.     

Mechanochemical modeling of neutrophil migration based on four signaling layers,integrin dynamics,and substrate stiffness

Directional neutrophil migration during human immune responses is a highly coordinated process regulated by both biochemical and biomechanical environments. In this paper, we developed an integrative mathematical model of neutrophil migration using a lattice Boltzmann-particle method built in-house to solve the moving boundary problem with spatiotemporal regulation of biochemical components. The mechanical features of the cell cortex are modeled by a series of spring-connected nodes representing discrete cell–substrate adhesive sites. The intracellular signaling cascades responsible for cytoskeletal remodeling [e.g., small GTPases, phosphoinositide-3-kinase (PI3K), and phosphatase and tensin homolog] are built based on our previous four-layered signaling model centered on the bidirectional molecular transport mechanism and implemented as reaction–diffusion equations. Focal adhesion dynamics are determined by force-dependent integrin–ligand binding kinetics and integrin recycling and are thus integrated with cell motion. Using numerical simulations, the model reproduces the major features of cell migration in response to uniform and gradient biochemical stimuli based on the quantitative spatiotemporal regulation of signaling molecules, which agree with experimental observations. The existence of multiple types of integrins with different binding kinetics could act as an adaptation mechanism for substrate stiffness. Moreover, cells can perform reversal, U-turn, or lock-on behaviors depending on the steepness of the reversal biochemical signals received. Finally, this model is also applied to predict the responses of mutants in which PTEN is overexpressed or disrupted.     

Deregulated Expression of Aurora Kinases Is Not a Prognostic Biomarker in Papillary Thyroid Cancer Patients

A number of reports indicated that Aurora-A or Aurora-B overexpression represented a negative prognostic factor in several human malignancies. In thyroid cancer tissues a deregulated expression of Aurora kinases has been also demonstrated, butno information regarding its possible prognostic role in differentiated thyroid cancer is available. Here, weevaluated Aurora-A and Aurora-B mRNA expression and its prognostic relevance in a series of 87 papillary thyroid cancers (PTC), with a median follow-up of 63 months. The analysis of Aurora-A and Aurora-B mRNA levels in PTC tissues, compared to normal matched tissues, revealed that their expression was either up- or down-regulatedin the majority of cancer tissues. In particular, Aurora-A and Aurora-B mRNA levels were altered, respectively, in 55 (63.2%) and 79 (90.8%) out of the 87 PTC analyzed.A significant positive correlation between Aurora-A and Aurora-B mRNAswas observed (p=0.001). The expression of both Aurora genes was not affected by the BRAF^V600E mutation. Univariate, multivariate and Kaplan-Mayer analyses documented the lack of association between Aurora-A or Aurora-B expression and clinicopathological parameterssuch as gender, age, tumor size, histology, TNM stage, lymph node metastasis and BRAF status as well asdisease recurrences or disease-free interval. Only Aurora-B mRNA was significantly higher in T(3-4) tissues, with respect to T(1-2) PTC tissues. The data reported here demonstrate that the expression of Aurora kinases is deregulated in the majority of PTC tissues, likely contributing to PTC progression. However, differently from other human solid cancers, detection of Aurora-A or Aurora-B mRNAs is not a prognostic biomarker inPTC patients.     

依达拉奉对硝普钠诱导的PC12细胞凋亡的保护作用

目的：探讨依达拉奉对硝普钠诱导的PCI2细胞凋亡的影响。方法：体外培养PCI2细胞,并分为依达拉奉对硝普钠保护组（含500μmol／L硝普钠和75μmol／L依达拉奉）、硝普钠诱导组（含500μmol／L硝普钠）和对照组。采用MTT法检测细胞的增殖率：流式细胞术检测细胞的凋亡情况;Western-blot检i受4凋亡抑制蛋白Bcl-2和凋亡促进蛋白Bad的表达。结果：与对照组相比,硝普钠处理的PCI2细胞增殖率显著降低,而细胞凋亡率显著升高,细胞内Bcl-2的表达显著减少,而Bad的表显著增加,差异均具有统计学意义（P〈0．05）;与单纯硝普钠诱导组相比,依达拉奉处理组细的胞增殖率显著增加而细胞凋亡率显著减少,同时Bcl－2的表达显著增加,而Bad的表达明显减少,差异均具有统计学意义（P〈0．05）。结论：依达拉奉对硝普钠诱导的PCI2细胞凋亡具有抑制作用,可能通过增加Bcl-2的表达并降低Bad的表达发挥抗凋亡作用。     

Enhancement of chondrocyte autophagy is an early response in the degenerative cartilage of the temporomandibular joint to biomechanical dental stimulation

Autophagy is a cell protective mechanism for maintaining cellular homeostasis. The present study aimed to investigate whether autophagy is enhanced in the biomechanically induced degenerative cartilage of the temporomandibular joint (TMJ) and the potential role of mitogen-activated protein kinase kinase kinase kinase 3 (MAP4K3) and mammalian Target of rapamycin (mTOR) in this observation. To induce degenerative changes in the TMJs, rats were subjected to biomechanical dental stimulation by moving 4 molars away from their original position as we previously reported. The ultrastructure of autophagosome was observed by transmission electron microscopy. The number of lysosomes was analyzed by flow cytometry. The expression levels of Beclin1 and LC3 and the involvement of MAP4K3 activity were detected by immunohistochemistry, real-time PCR and western blot. The activity of the mTOR pathway indicated by p-mTOR and p-p70S6 K was assayed by western blot. TMJ degeneration, characterized by irregular cell arrangement and cell-free area, was induced in the experimental groups. Under transmission electron microscopy, we observed the presence of autophagosomes, small patches of condensed chromatin, abundant rough endoplasmic reticulum and Golgi apparatus. The number of lysosomes and the expression levels of Beclin1 and LC3 increased, while the activity of mTOR and the expression level of MAP4K3 decreased in the experimental groups. Cartilage in TMJ which was induced to be degenerative biomechanically exhibited autophagy accompanied by reduced mTOR and MAP4K3 activity.     

TULIP study: Trail of Lurasidone in bipolar disorder in Pakistan

BackgroundThis study examined usefulness and efficiency of Lurasidone in appraisal with the placebo as for the treatment of Bipolar Disorders.MethodsSeven treatment centers in Pakistan were selected for the purpose of starting a six week-long control trial (randomized and double-blind placebo). 76 subjects, already diagnosed with Bipolar I or II based on DSM 5 diagnosis, were selected after randomization. Patients were allocated in one of the two groups. Primary efficacy of the drug was measured using Young Mania Rating Scale. Positive response of the drug was defined as 50% reduction in symptoms from the baseline/13 point less than the baseline score on Young Mania Rating Scale. Efficacy and safety of the drug was assessed using variety of markers such as administering extra-pyramidal symptoms rating scale, adverse side effects reported, electrocardiograms, body weight, vital signs changes, and laboratory investigations.ResultsPatients treated with Lurasidone showed enhanced improvement in their overall health and symptoms manifestation in comparison to patients who were given placebo. Lurasidone treated patients showed a better response to the drug (66%), in comparison with the placebo treated patients (42%).LimitationsStudy was conducted on small scale due to complexity.ConclusionPatients treated with Lurasidone showed reduction in bipolar symptoms and tolerate the drug well.