Identification of three genetic loci controlling leaf senescence in Arabidopsis thaliana

Four mutants that show the delayed leaf senescence phenotype were isolated from Arabidopsis thaliana . Genetic analyses revealed that they are all monogenic recessive mutations and fall into three complementation groups, identifying three genetic loci controlling leaf senescence in Arabidopsis . Mutations in these loci cause delay in all senescence parameters examined, including chlorophyll content, photochemical efficiency of photosystem II, relative amount of the large subunit of Rubisco, and RNase and peroxidase activity. Delay of the senescence symptoms was observed during both age-dependent in planta senescence and dark-induced artificial senescence in all of the mutant plants. The results indicate that the three genes defined by the mutations are key genetic elements controlling functional leaf senescence and provide decisive genetic evidence that leaf senescence is a genetically programmed phenomenon controlled by several monogenic loci in Arabidopsis . The results further suggest that the three genes function at a common step of age-dependent and dark-induced senescence processes. It is further shown that one of the mutations is allelic to ein2-1 , an ethylene-insensitive mutation, confirming the role of ethylene signal transduction pathway in leaf senescence of Arabidopsis .     

Possible role of macrophage-derived soluble mediators in the pathogenesis of encephalomyocarditis virus-induced diabetes in mice.

Pancreatic islets from DBA/2 mice infected with the D variant of encephalomyocarditis (EMC-D) virus revealed lymphocytic infiltration with moderate to severe destruction of pancreatic beta cells. Our previous studies showed that the major population of infiltrating cells at the early stages of infection is macrophages. The inactivation of macrophages prior to viral infection resulted in the prevention of diabetes, whereas activation of macrophages prior to viral infection resulted in the enhancement of beta-cell destruction. This investigation was initiated to determine whether macrophage-produced soluble mediators play a role in the destruction of pancreatic beta cells in mice infected with a low dose of EMC-D virus. When we examined the expression of the soluble mediators interleukin-1 beta (IL-1beta), tumor necrosis factor alpha (TNF-alpha), and inducible nitric oxide synthase (iNOS) in the pancreatic islets, we found that these mediators were clearly expressed at an early stage of insulitis and that this expression was evident until the development of diabetes. We confirmed the expression of these mediators by in situ hybridization with digoxigenin-labelled RNA probes or immunohistochemistry in the pancreatic islets. Mice treated with antibody against IL-1beta or TNF-alpha or with the iNOS inhibitor aminoguanidine exhibited a significant decrease in the incidence of diabetes. Mice treated with a combination of anti-IL-1beta antibody, anti-TNF-alpha antibody, and aminoguanidine exhibited a greater decrease in the incidence of disease than did mice treated with one of the antibodies or aminoguanidine. On the basis of these observations, we conclude that macrophage-produced soluble mediators play an important role in the destruction of pancreatic beta cells, resulting in the development of diabetes in mice infected with a low dose of EMC-D virus.     

Involvement of cyclic GMP in the fusion of chick embryonic myoblasts in culture.

We found that a transient rise in cGMP levels, which was closely associated with the Ca2+ influx, occurred concomitant with the onset of myoblast fusion. The Ca2+ channel blocker D600 decreased both the cell fusion and the normal rise in cGMP levels. In contrast, the Ca2+ ionophore A23187 transiently increased cGMP levels and induced precocious fusion. In addition, the cGMP analog 8-Br-cGMP induced precocious fusion as A23187 did. The guanylate cyclase inhibitor, methylene blue delayed the fusion in a dose-dependent manner without significantly affecting cell alignment, proliferation, or muscle-specific protein expression. Furthermore, methylene blue delayed the normal rise in cGMP levels, and the fusion block imposed by methylene blue was significantly recovered by 8-Br-cGMP. On the basis of our present findings, we suggest that a Ca2+ influx-dependent rise in cGMP levels is an important step in myoblast fusion.     

Induction of an organ-specific autoimmune disease, lymphocytic hypophysitis, in hamsters by recombinant rubella virus glycoprotein and prevention of disease by neonatal thymectomy.

Glycosylated, membrane-associated E1 (58-kDa) and E2 (47- to 49-kDa) rubella virus proteins and unglycosylated nucleoprotein C (33 kDa), from separately expressed vaccinia virus recombinants, were injected into golden Syrian hamsters. Rubella virus E1 and E2 glycoproteins consistently induced an organ-specific autoimmune disease, autoimmune lymphocytic hypophysitis, which was evidenced by the induction of autoantibodies against pituitary cells and by lymphocytic infiltration of the pituitary. Neonatal thymectomy prevented the disease. In contrast, rubella virus nucleoprotein C did not induce either autoantibodies against pituitary cells or lymphocytic infiltration of the pituitary. This finding raises the possibility that virus-specific protein itself can induce an organ-specific autoimmune disease in certain circumstances.     

Platelet kinetics and other hematological profiles in experimental Plasmodium falciparum infection: a comparative study between Saimiri and Aotus monkeys.

Levels of platelets and other hematological values were monitored in 21 Saimiri and 12 Aotus monkeys over a period of three weeks post-infection with monkey-adapted Indochina CDC-1 strain of Plasmodium falciparum. In both Saimiri sciureus boliviensis and Aotus nancymai karyotype-1 monkeys the severest thrombocytopenia was observed at 14 days post-infection coinciding with peak parasitemia, neutropenia, lymphocytosis, and anemia associated with severe hemoglobinemia and elevated fibrinogen degeneration products(FDP's). MCH and MCV profiles in Aotus monkeys decreased with ascending parasitemia. In contrast, these parameters in Saimiri were characterized by a significant compensatory increase correlating with parasitemia. In general, thrombocytopenia was one of the earliest clinical manifestations of the infection with the platelets returning to normal levels shortly after peak parasitemia at 14 days. Platelet kinetics had a strong correlation with hematologic and parasitologic values in the Aotus model. No consistent associations were observed between platelet kinetics and other parameters in the Saimiri model. These data indicate that the Aotus model for malaria is more predictable than the Saimiri. Further, platelet turnover rates and recovery provide a useful prognostic parameter during malaria infection. The results are discussed in relation to the value of the two species of monkeys as models for the pathogenesis of human malaria.     

Substrate Binding Mechanism of a Type I Extradiol Dioxygenase

A meta-cleavage pathway for the aerobic degradation of aromatic hydrocarbons is catalyzed by extradiol dioxygenases via a two-step mechanism: catechol substrate binding and dioxygen incorporation. The binding of substrate triggers the release of water, thereby opening a coordination site for molecular oxygen. The crystal structures of AkbC, a type I extradiol dioxygenase, and the enzyme substrate (3-methylcatechol) complex revealed the substrate binding process of extradiol dioxygenase. AkbC is composed of an N-domain and an active C-domain, which contains iron coordinated by a 2-His-1-carboxylate facial triad motif. The C-domain includes a β-hairpin structure and a C-terminal tail. In substrate-bound AkbC, 3-methylcatechol interacts with the iron via a single hydroxyl group, which represents an intermediate stage in the substrate binding process. Structure-based mutagenesis revealed that the C-terminal tail and β-hairpin form part of the substrate binding pocket that is responsible for substrate specificity by blocking substrate entry. Once a substrate enters the active site, these structural elements also play a role in the correct positioning of the substrate. Based on the results presented here, a putative substrate binding mechanism is proposed.     

Control of the pericentrosomal H2O2 level by peroxiredoxin I is critical for mitotic progression

Proteins associated with the centrosome play key roles in mitotic progression in mammalian cells. The activity of Cdk1-opposing phosphatases at the centrosome must be inhibited during early mitosis to prevent premature dephosphorylation of Cdh1—an activator of the ubiquitin ligase anaphase-promoting complex/cyclosome—and the consequent premature degradation of mitotic activators. In this paper, we show that reversible oxidative inactivation of centrosome-bound protein phosphatases such as Cdc14B by H₂O₂ is likely responsible for this inhibition. The intracellular concentration of H₂O₂ increases as the cell cycle progresses. Whereas the centrosome is shielded from H₂O₂ through its association with the H₂O₂-eliminating enzyme peroxiredoxin I (PrxI) during interphase, the centrosome-associated PrxI is selectively inactivated through phosphorylation by Cdk1 during early mitosis, thereby exposing the centrosome to H₂O₂ and facilitating inactivation of centrosome-bound phosphatases. Dephosphorylation of PrxI by okadaic acid–sensitive phosphatases during late mitosis again shields the centrosome from H₂O₂ and thereby allows the reactivation of Cdk1-opposing phosphatases at the organelle.     

Lab-scale study of an anaerobic membrane bioreactor (AnMBR) for dilute municipal wastewater treatment

Anaerobic bioreactors supplemented with membrane technology have become quite popular, owing to their favorable energy recovery characteristics. In this study, a lab-scale anaerobic Membrane Bioreactor (AnMBR) was assessed in experimental treatments of pre-settled dilute municipal wastewater obtained from a full-scaled wastewater treatment plant. The MBR system was operated in continuous flow mode for 440 days. To evaluate the performance of the AnMBR under various loading rates, the hydraulic retention time (HRT) was reduced in a stepwise manner (from 2 to 0.5 days). Afterward, the mixed liquor suspended solids (MLSS) were reduced from 7,000 to 3,000 mg/L in increments of 1,000 mg/L, resulting in a decrease in solids retention time (SRT) at a constant HRT of 1.0 day. The soluble chemical oxygen demand (SCOD) concentration in the feed varied between 38 and 131 mg/L, whereas the average permeate SCOD ranged between 18 and 37 mg/L, reflecting excellent effluent quality. The AnMBR performance in terms of COD removal proved stable, despite variations in influent characteristics and HRT and SRT changes. The concentration of extracellular polymeric substance (EPS) was reduced with decreases in HRT from 42 to 22 mg VS/mg of MLSS, thereby indicating that the increased biomass concentration biodegraded the EPS at lower HRTs. AnMBR is, therefore, demonstrably a feasible option for the treatment of dilute wastewater with separate stage nitrogen and phosphorus removal processes.