Effects of phenoxy acid herbicides and glyphosate on nitrogenase activity (acetylene reduction) in root-associated Azospirillum, Enterobacter and Klebsiella

Abstract Nitrogenase activity (C₂H₂ reduction) in root-associated Azospirillum lipoferum, Klebsiella pneumoniae, Enterobacter agglomerans and Pseudomonas sp. isolated from roots of Finnish grasses was assayed in the presence of glyphosate, the phenoxy acid herbicides 2-methyl-4-chlorophenoxy acetic acid (MCPA), 2,4-dichlorophenoxy acetic acid (2,4-D), (±)-2-(2-methyl-4-chlorophenoxy)propionic acid (mecoprop) and (±)-2-(2,4-dichlorophenoxy)propionic acid (dichlorprop), and the commercial products Roundup, Nurmikko-Hedonal, Mepro, and Dipro. In the presence of the phenoxy acid herbicides the nitrogenase activity of K. pneumoniae was significantly inhibited, but that of E. agglomerans was stimulated. With the exception of Mepro and mecoprop no phenoxy acid herbicides inhibited the nitrogenase activity of A. lipoferum and none that of Pseudomonas sp. Nurmikko-Hedonal considerably stimulated the nitrogenase activity of E. agglomerans , and Pseudomanas sp. On the other hand, the nitrogenase activity of both K. pneumoniae and E. agglomerans was considerably repressed by glyphosate and Roundup, which also inhibited the growth of the bacteria. These chemicals had no effect on the growth of A. lipoferum and Pseudomonas sp., but stimulated their nitrogenase activity.     

Identification of tumor antigens as potential target antigens for immunotherapy by serological expression cloning

The presence of tumor infiltrating T cells has been shown to be associated with a favorable prognosis in different tumor types. Several strategies have been developed to identify relevant tumor antigens which can be used for active immunotherapy strategies. The SEREX technique (serological analysis of cDNA expression libraries) identifies tumor antigens based on a spontaneous humoral immune response in cancer patients. This technique is not limited to tumor types that can be grown in cell culture or depends on established T cell clones recognizing the autologous tumor. Several steps of analysis are mandatory to evaluate SEREX-defined antigens before they become new target antigens for active immunotherapy: expression analysis; serological analysis with sera from tumor patients and normal individuals; identification of potential peptide epitopes for CD8 T cells and evaluation in T cell assays. This article summarizes our approach of antigen identification and evaluation giving the example of the recently cloned breast cancer antigen NY-BR-1.This work was presented at the first Cancer Immunology and Immunotherapy Summer School, 8–13 September 2003, Ionian Village, Bartholomeio, Peloponnese, Greece.     

Potent 2-[(pyrimidin-4-yl)amine}-1,3-thiazole-5-carbonitrile-based inhibitors of VEGFR-2 (KDR) kinase

Pyrimidino-thiazolyl carbonitriles were prepared that are potent VEGFR-2 (KDR) kinase inhibitors. The modification of lead structures resulted in 3m which exhibited the best overall profile in KDR inhibitory activity, iv/po pharmacokinetics, and reduced hERG affinity.     

Human tankyrases are aberrantly expressed in colon tumors and contain multiple epitopes that induce humoral and cellular immune responses in cancer patients

Purpose Tankyrases 1 and 2 are telomere-associated poly(ADP-ribose) polymerases (PARP) that can positively regulate telomere elongation and interact with multiple cellular proteins. Recent reports implicated tankyrases as tumor antigens and potential targets of anticancer treatment. We examined expression of tankyrases in colon tumors and immune response to these enzymes in patients with different types of cancer. Methods mRNA and protein expression was evaluated by quantitative real-time RT-PCR and Western blotting, respectively. Humoral immune response to recombinant tankyrases was investigated by modified enzyme-linked immunoassays. Cellular immune response was analysed by ELISPOT and ⁵¹Cr release assays. Results We found that both mRNA and protein levels of tankyrase 2 (TNKL) are upregulated in colon tumors. In contrast, protein level of tankyrase 1 (TNKS) is downregulated, while mRNA level shows variable changes. More than a quarter of colon cancer patients develop humoral immune response to at least one of the two tankyrases. In this study we mapped common and unique B-cell epitopes located in different domains of the two proteins. Additionally, we present evidence for T-cell responses both to epitopes that are unique for TNKL and to those shared between TNKL and TNKS. Conclusion Our study favors a biomarker usage of antibody response to tankyrases. Spontaneous CD8⁺ T-cell responses to these enzymes are rare and further investigation is needed to evaluate tankyrases as potential targets for cancer immunotherapy.     

Coxsackievirus B3-induced cellular protrusions: structural characteristics and functional competence

Virus-induced alterations in cell morphology play important roles in the viral life cycle. To examine the intracellular events of coxsackievirus B3 (CVB3) infection, green monkey kidney (GMK) cells were either inoculated with the virus or transfected with the viral RNA. Various microscopic and flow cytometric approaches demonstrated the emergence of CVB3 capsid proteins at 8 h posttransfection, followed by morphological transformation of the cells. The morphological changes included formation of membranous protrusions containing viral capsids, together with microtubules and actin. Translocation of viral capsids into these protrusions was sensitive to cytochalasin D, suggesting the importance of actin in the process. Three-dimensional (3D) live-cell imaging demonstrated frequent contacts between cellular protrusions and adjacent cells. Markedly, in spite of an increase in the cellular viral protein content starting 8 h postinfection, no significant decrease in cell viability or increase in the amount of early apoptotic markers was observed by flow cytometry by 28 h postinfection. Comicroinjection of viral RNA and fluorescent dextran in the presence of neutralizing virus antibody suggested that these protrusions mediated the spread of infection from one cell to another prior to virus-induced cell lysis. Altogether, the CVB3-induced cellular protrusions could function as a hitherto-unknown nonlytic mechanism of cell-to-cell transmission exploited by enteroviruses.     

Crystal structure of a member of a novel family of dioxygenases (PF10014) reveals a conserved cupin fold and active site

PF10014 is a novel family of 2‐oxyglutarate‐Fe²⁺‐dependent dioxygenases that are involved in biosynthesis of antibiotics and regulation of biofilm formation, likely by catalyzing hydroxylation of free amino acids or other related ligands. The crystal structure of a PF10014 member from Methylibium petroleiphilum at 1.9 Å resolution shows strong structural similarity to cupin dioxygenases in overall fold and active site, despite very remote homology. However, one of the β‐strands of the cupin catalytic core is replaced by a loop that displays conformational isomerism that likely regulates the active site. Proteins 2014; 82:164–170. © 2013 Wiley Periodicals, Inc.