Effects of salinity on cell growth and docosahexaenoic acid content of the heterotrophic marine microalga Crypthecodinium cohnii

The effects of salinity on cell growth and docosahexaenoic acid (DHA) content of three marine microalgal strains, Crythecodinium cohnii ATCC 30556, C. cohnii ATCC 50051 and C. cohnii RJH were investigated. The lag phases of the three strains increased with increasing salinity in Porphyridium medium. The specific growth rate of C. cohnii ATCC 30556 was the highest at 9 g L⁻¹ NaCl while the other two strains had their highest specific growth rates at 5 g L⁻¹ NaCl. The highest cell dry weight concentrations of 2.51 g L⁻¹ and 1.56 g L⁻¹ were achieved at 9 g L⁻¹ NaCl for C. cohnii ATCC 30556 and ATCC 50051, respectively, while the highest dry weight concentration of 2.49 g L⁻¹ was achieved at 5 g L⁻¹ NaCl for C. cohnii RJH. The highest cell growth yield coefficient on glucose was 0.5 g g⁻¹ for both C. cohnii ATCC 30556 and C. cohnii RJH and 0.45 g g⁻¹ for C. cohnii ATCC 50051. All three strains responded to the change of salinity by modifying their cellular fatty acid compositions. At 9 g L⁻¹ NaCl, C. cohnii ATCC 30556 had the highest total fatty acid content and DHA (C22:6) proportion. In contrast, C. cohnii ATCC 50051 and C. cohnii RJH had the highest DHA content at 5 g L⁻¹ NaCl. C. cohnii ATCC 30556 and ATCC 50051 had the highest DHA yield (131.55 and 68.24 mg L⁻¹ respectively) at 9 g L⁻¹ NaCl while C. cohnii RJH had the highest DHA yield (128.83 mg L⁻¹) at 5 g L⁻¹ NaCl. Received 27 May 1999/ Accepted in revised form 27 August 1999     

Metabolic engineering of Corynebacterium glutamicum for increasing the production of l-ornithine by increasing NADPH availability

The experiments presented here were based on the conclusions of our previous proteomic analysis. Increasing the availability of glutamate by overexpression of the genes encoding enzymes in the l-ornithine biosynthesis pathway upstream of glutamate and disruption of speE, which encodes spermidine synthase, improved l-ornithine production by Corynebacterium glutamicum. Production of l-ornithine requires 2 moles of NADPH per mole of l-ornithine. Thus, the effect of NADPH availability on l-ornithine production was also investigated. Expression of Clostridium acetobutylicum gapC, which encodes NADP-dependent glyceraldehyde-3-phosphate dehydrogenase, and Bacillus subtilis rocG, which encodes NAD-dependent glutamate dehydrogenase, led to an increase of l-ornithine concentration caused by greater availability of NADPH. Quantitative real-time PCR analysis demonstrates that the increased levels of NADPH resulted from the expression of the gapC or rocG gene rather than that of genes (gnd, icd, and ppnK) involved in NADPH biosynthesis. The resulting strain, C. glutamicum ΔAPRE::rocG, produced 14.84 g l^?1 of l-ornithine. This strategy of overexpression of gapC and rocG will be useful for improving production of target compounds using NADPH as reducing equivalent within their synthetic pathways.     

Nitidine chloride inhibited the expression of S phase kinase-associated protein 2 in ovarian cancer cells

Nitidine chloride (NC) has been reported to exert its anti-tumor activity in various types of human cancers. However, the molecular mechanism of NC-mediated tumor suppressive function is largely unclear. In the current study, we used several approaches such as MTT, FACS, RT-PCR, Western blotting analysis, invasion assay, transfection, to explore the molecular basis of NC-triggered anti-cancer activity. We found that NC inhibited cell growth, induced cell apoptosis, caused cell cycle arrest in ovarian cancer cells. Emerging evidence has demonstrated that Skp2 plays an important oncogenic role in ovarian cancer. Therefore, we also explored whether NC exerts its biologic function via downregulation of Skp2 in ovarian cancer cells. We observed that NC significantly inhibited the expression of Skp2 in ovarian cancer cells. Notably, overexpression of Skp2 abrogated the anti-cancer activity induced by NC in ovarian cancer cells. Consistently, downregulation of Skp2 expression enhanced the sensitivity of ovarian cancer cells to NC treatment. Thus, inactivation of Skp2 by NC could be a novel strategy for the treatment of human ovarian cancer.     

Short exposure to paclitaxel induces multipolar spindle formation and aneuploidy through promotion of acentrosomal pole assembly

Paclitaxel is a widely used microtubule drug and cancer medicine. Here we report that by short exposure to paclitaxel at a low dose, multipolar spindles were induced in mitotic cells without centrosome amplification. Both TPX2 depletion and Aurora-A overexpression antagonized the multipolarity. Live cell imaging showed that some paclitaxel-treated cells accomplished multipolar cell division and a portion of the daughter cells went on to the next round of mitosis. The surviving cells grew into clones with varied genome content. The results indicated that an aneuploidy population could be induced by short exposure to paclitaxel at a low dose, implicating potential side effects of paclitaxel.     

QCM detection of DNA targets with single-base mutation based on DNA ligase reaction and biocatalyzed deposition amplification

A novel biosensing technique for highly specific identification of gene with single-base mutation is proposed based on the implementation of the DNA ligase reaction and the biocatalyzed deposition of an insoluble product. The target gene mediated deposition of an insoluble precipitate is then transduced by quartz crystal microbalance (QCM) measurements. In this method, the DNA target hybridizes with a capture DNA probe tethered onto the gold electrode and then with a biotinylated allele-specific detection DNA. A ligase reaction is performed to generate the ligation between the capture and the detection probes, provided there is perfect match between the DNA target and the detection probe. Otherwise even when there is an allele mismatch between them, no ligation would take place. After thermal treatment at an elevated temperature, the formed duplex melts apart that merely allows the detection probe perfectly matched with the target to remain on the electrode surface. The presence of the biotinylated allele-matched probe is then detected by the QCM via the binding to streptavidin-peroxide horseradish (SA-HRP), which catalyzes the oxidative precipitation of 3,3-diaminobenzidine (DAB) by H2O2 on the electrode and provides an amplified frequency response. The proposed approach has been successfully implemented for the identification of single-base mutation in -28 site of the beta-thalassemia gene with a detection limit of 0.1 nM, demonstrating that this method provides a highly specific and cost-efficient approach for point mutation detection.     

Cardiac mitochondrial ATP-sensitive potassium channel is activated by nitric oxide in vitro

Previous observations on the activation of the mitochondrial ATP-sensitive potassium channel (mitoK(ATP)) by nitric oxide (NO) in myocardial preconditioning were based on indirect evidence. In this study, we have investigated the direct effect of NO on the rat cardiac mitoK(ATP) after reconstitution of the inner mitochondrial membranes into lipid bilayers. We found that the mitoK(ATP) was activated by exogenous NO donor S-nitroso-N-acetyl penicillamine or PAPA NONOate. This activation was inhibited by mitoK(ATP) blockers 5-hydroxydecanoate or glibenclamide. Our observations confirm that NO can directly activate the cardiac mitoK(ATP), which may underlie its contribution to myocardial preconditioning.     

Novel mechanisms of tube-size regulation revealed by the Drosophila trachea

The size of various tubes within tubular organs such as the lung, vascular system and kidney must be finely tuned for the optimal delivery of gases, nutrients, waste and cells within the entire organism. Aberrant tube sizes lead to devastating human illnesses, such as polycystic kidney disease, fibrocystic breast disease, pancreatic cystic neoplasm and thyroid nodules. However, the underlying mechanisms that are responsible for tube-size regulation have yet to be fully understood. Therefore, no effective treatments are available for disorders caused by tube-size defects. Recently, the Drosophila tracheal system has emerged as an excellent in vivo model to explore the fundamental mechanisms of tube-size regulation. Here, we discuss the role of the apical luminal matrix, cell polarity and signaling pathways in regulating tube size in Drosophila trachea. Previous studies of the Drosophila tracheal system have provided general insights into epithelial tube morphogenesis. Mechanisms that regulate tube size in Drosophila trachea could be well conserved in mammalian tubular organs. This knowledge should greatly aid our understanding of tubular organogenesis in vertebrates and potentially lead to new avenues for the treatment of human disease caused by tube-size defects.     

The transposable element landscape of the model legume Lotus japonicus

The largest component of plant and animal genomes characterized to date is transposable elements (TEs). The availability of a significant amount of Lotus japonicus genome sequence has permitted for the first time a comprehensive study of the TE landscape in a legume species. Here we report the results of a combined computer-assisted and experimental analysis of the TEs in the 32.4 Mb of finished TAC clones. While computer-assisted analysis facilitated a determination of TE abundance and diversity, the availability of complete TAC sequences permitted identification of full-length TEs, which facilitated the design of tools for genomewide experimental analysis. In addition to containing all TE types found in previously characterized plant genomes, the TE component of L. japonicus contained several surprises. First, it is the second species (after Oryza sativa) found to be rich in Pack-MULEs, with >1000 elements that have captured and amplified gene fragments. In addition, we have identified what appears to be a legume-specific MULE family that was previously identified only in fungal species. Finally, the L. japonicus genome contains many hundreds, perhaps thousands of Sireviruses: Ty1/copia-like elements with an extra ORF. Significantly, several of the L. japonicus Sireviruses have recently amplified and may still be actively transposing.     

Exogenous maltose enhances Zebrafish immunity to levofloxacin-resistant Vibrio alginolyticus

Understanding the interplay between bacterial fitness, antibiotic resistance, host immunity and host metabolism could guide treatment and improve immunity against antibiotic-resistant pathogens. The acquisition of levofloxacin (Lev) resistance affects the fitness of Vibrio alginolyticus in vitro and in vivo. Lev-resistant (Lev-R) V. alginolyticus exhibits slow growth, reduced pathogenicity and greater resistance to killing by the host, Danio rerio (zebrafish), than Lev-sensitive (Lev-S) V. alginolyticus, suggesting that Lev-R V. alginolyticus triggers a weaker innate immune response in D. rerio than Lev-S V. alginolyticus. Differences were detected in the metabolome of D. rerio infected with Lev-S or Lev-R V. alginolyticus. Maltose, a crucial metabolite, is significantly downregulated in D. rerio infected with Lev-R V. alginolyticus, and exogenous maltose enhances the immune response of D. rerio to Lev-R V. alginolyticus, leading to better clearance of the infection. Furthermore, we demonstrate that exogenous maltose stimulates the host production of lysozyme and its binding to Lev-R V. alginolyticus, which depends on bacterial membrane potential. We suggest that exogenous exposure to crucial metabolites could be an effective strategy for treating and/or managing infections with antibiotic-resistant bacteria.