A point mutation in the IL-12R beta 2 gene underlies the IL-12 unresponsiveness of Lps-defective C57BL/10ScCr mice

Lps-defective C57BL/10ScCr (Cr) mice are homozygous for a deletion encompassing Toll-like receptor 4 that makes them refractory to the biological activity of LPS. In addition, these mice exhibit an inherited IL-12 unresponsiveness resulting in impaired IFN-gamma responses to different microorganisms. By positional cloning methods, we show here that this second defect of Cr mice is due to a mutation in a single gene located on mouse chromosome 6, in close proximity to the Igkappa locus. The gene is IL-12Rbeta2. Cr mice carry a point mutation creating a stop codon that is predicted to cause premature termination of the translated IL-12Rbeta2 after a lysine residue at position 777. The truncated beta2 chain can still form a heterodimeric IL-12R that allows phosphorylation of Janus kinase 2, but, unlike the wild-type IL-12R, can no longer mediate phosphorylation of STAT4. Because the phosphorylation of STAT4 is a prerequisite for the IL-12-mediated induction of IFN-gamma, its absence in Cr mice is responsible for their defective IFN-gamma response to microorganisms.     

Effects of citrinin on iron-redox cycle

The ability of the mycotoxin citrinin to act as an inhibitor of iron-induced lipoperoxidation of biological membranes prompted us to determine whether it could act as an iron chelating agent, interfering with iron redox reactions or acting as a free radical scavenger. The addition of Fe3+ to citrinin rapidly produced a chromogen, indicating the formation of citrinin-Fe3+ complexes. An EPR study confirms that citrinin acts as a ligand of Fe3+, the complexation depending on the [Fe3+]:[citrinin] ratios. Effects of citrinin on the iron redox cycle were evaluated by oxygen consumption or the o-phenanthroline test. No effect on EDTA-Fe2+-->EDTA-Fe3+ oxidation was observed in the presence of citrinin, but the mycotoxin inhibited, in a dose-dependent manner, the oxidation of Fe2+ to Fe3+ by hydrogen peroxide. Reducing agents such as ascorbic acid and DTT reduced the Fe3+-citrinin complex, but DTT did not cause reduction of Fe3+-EDTA, indicating that the redox potentials of Fe3+-citrinin and Fe3+-EDTA are not the same. The Fe2+ formed from the reduction of Fe3+-citrinin by reducing agents was not rapidly reoxidized to Fe3+ by atmospheric oxygen. Citrinin has no radical scavenger ability as demonstrated by the absence of DPPH reduction. However, a reaction between citrinin and hydrogen peroxide was observed by UV spectrum changes of citrinin after incubation with hydrogen peroxide. It was also observed that citrinin did not induce direct or reductive mobilization of iron from ferritin. These results indicate that the protective effect on iron-induced lipid peroxidation by citrinin occurs due to the formation of a redox inactive Fe3+-citrinin complex, as well as from the reaction of citrinin and hydrogen peroxide.     

The inhibitory activity of natural products on plant p-hydroxyphenylpyruvate dioxygenase

The inhibitory activity of 34 natural products of various structural classes on hydroxyphenylpyruvate dioxygenase (HPPD), the target site for triketone herbicides, and the mode of interaction of selected natural products were investigated. Recombinant HPPD from arabidopsis is sensitive to several classes of natural compounds including, in decreasing order of sensitivity, triketones, benzoquinones, naphthoquinones and anthraquinones. The triketone natural products acted as competitive tight-binding inhibitors, whereas the benzoquinones and naphthoquinones did not appear to bind tightly to HPPD. While these natural products may not have optimal structural features required for in vivo herbicidal activity, the differences in their kinetic behavior suggest that novel classes of HPPD inhibitors may be developed based on their structural backbones.     

Novel derivatives of 9,10-anthraquinone are selective algicides against the musty-odor cyanobacterium Oscillatoria perornata

Musty "off-flavor" in pond-cultured channel catfish (Ictalurus punctatus) costs the catfish production industry in the United States at least 30 million US dollars annually. The cyanobacterium Oscillatoria perornata (Skuja) is credited with being the major cause of musty off-flavor in farm-raised catfish in Mississippi. The herbicides diuron and copper sulfate, currently used by catfish producers as algicides to help mitigate musty off-flavor problems, have several drawbacks, including broad-spectrum toxicity towards the entire phytoplankton community that can lead to water quality deterioration and subsequent fish death. By use of microtiter plate bioassays, a novel group of compounds derived from the natural compound 9,10-anthraquinone have been found to be much more selectively toxic towards O. perornata than diuron and copper sulfate. In efficacy studies using limnocorrals placed in catfish production ponds, application rates of 0.3 micro M (125 micro g/liter) of the most promising anthraquinone derivative, 2-[methylamino-N-(1'-methylethyl)]-9,10-anthraquinone monophosphate (anthraquinone-59), dramatically reduced the abundance of O. perornata and levels of 2-methylisoborneol, the musty compound produced by O. perornata. The abundance of green algae and diatoms increased dramatically 2 days after application of a 0.3 micro M concentration of anthraquinone-59 to pond water within the limnocorrals. The half-life of anthraquinone-59 in pond water was determined to be 19 h, making it much less persistent than diuron. Anthraquinone-59 appears to be promising for use as a selective algicide in catfish aquaculture.     

Cryptdin-3 induces novel apical conductance(s) in Cl- secretory, including cystic fibrosis, epithelia

Opening ofanion-conductive pathways in apical membranes of secretory cells liningmucosal surfaces is a critical step in salt and water secretion and,thus, hydration of sites including airway and intestine. In intestine,Paneth cells are positioned at the base of the secretory gland (crypt)and release defensin peptide, in mice termed cryptdins, into the cryptlumen. Because at least some defensins have been shown to formanion-conductive channels in phospholipid bilayers, we tested whetherthese endogenous antimicrobial peptides could act as soluble inducersof channel-like activity when applied to apical membranes. To directlyevaluate the possibility of cryptdin-3-mediated apical anionconductance (G_ap), we have utilized amphotericinB to selectively permeabilize basolateral membranes of electricallytight monolayers of polarized human intestinal secretory epithelia (T84cells), thus isolating the apical membrane for study. Cryptdin-3induces G_ap that is voltage independent(G_ap = 1.90 ± 0.60 mS/cm²) and exhibits ion selectivity contrasting to thatelicited by forskolin or thapsigargin (for cryptdin-3,Cl = gluconate; for forskolin and thapsigargin,Cl gluconate). We cannot exclude the possibility thatthe macroscopic current induced by cryptdin could be the sum of cationand Cl currents. Cryptdin-3 induces a current inbasolaterally permeabilized epithelial monolayers derived from airwaycells harboring the F508 mutation of cystic fibrosis (CF;G_ap = 0.80 ± 0.06 mS/cm²), demonstrating that cryptdin-3 restores anionsecretion in CF cells; this occurs independently of the CFtransmembrane conductance regulator channel. These results support theidea that cryptdin-3 may associate with apical membranes ofCl-secreting epithelia and self-assemble into conductingchannels capable of mediating a physiological response.