Remarkable retardation of the senecence of tobacco leaf disks by cordycepin, an inhibitor of RNA polyadenylation

The antibiotic cordycepin (3'-deoxyadenosine), a known specificinhibitor of nucleic acid synthesis and polyadenylation of RNA,remarkably retarded decrease in the chlorophyll and proteincontents of senescing tobacco leaf disks. The effectivenessof cordycepin, at its optimum concentration (ca. 4 x 10^–5M), was 53% as effective as a cytokinin, benzyladenine at 10^–6M. Adenosine and 2'-deoxyadenosine had no anti-senescence action. (Received May 29, 1975; )     

Alkali Protease and Alcohol Dehydrogenase Immobilized to Weakly Basic Anion Exchange Resins Using 2-Car-boxymethylamino-4,6-dichloro-s-triazine

Alkali protease partially purified from a strain of Bacillus subtilis and yeast alcohol dehydrogenase were immobilized to weakly basic anion exchange resins using a bifunctional reagent, 2-carboxymethyIamino-4,6-dichloro-s-triazine.

Properties of these immobilized enzymes were studied both in batchwise operation and in packed bed reactor systems.     

Nuclease C Polymorphism of Calcium-Dependent Nucleases in Chlamydomonas reinhardtii

DNases were extracted from cells of Chlamydomonas reinhardtii.Their polymorphism and metal ion requirements were investigated.Most of the nucleases from this organism required Ca²⁺ for fullactivation; therefore, the name nuclease C has been given tothem. Zn²⁺ and Mn²⁺ restored activation, but their respectivepotencies were 1/8 and 1/32 that of Ca²⁺. An in situ nucleaseassay revealed that there are at least 6 species of nucleaseC. The molecular weights of the components were 26,000 (C1),25,000 (C2), 22,000 (C5), 21,000 (C6), 18,000 (C3) and 17,000(C4) by SDS PAGE. Ca²⁺-dependent nuclease activity was slightlyhigher in the female gamete than in the male gamete. There wasno marked change in the apparent nuclease activity during thefirst 3 h after mating. An in situ assay, however, showed thatnuclease C6 and C4 are present in smaller amounts in the malegamete in comparison to the female gamete, and that the contentsor activities of nuclease C5 and C6 diminish during the first30–60 min after mating. This evidence is discussed interms of the possible participation of nuclease C in the maternalinheritance of chloroplast genes in this organism. (Received October 8, 1984; Accepted January 21, 1985)     

NKT cells are responsible for the clearance of murine norovirus through the virus-specific secretory IgA pathway

Norovirus infection cause epidemic nonbacterial gastroenteritis in patients. The immune mechanisms responsible for the clearance of virus are not completely understood. We examined whether NKT cells are effective against norovirus infection using CD1d KO mice. The body weights of 4-weeks-old CD1d KO mice that were infected with murine norovirus-S7 (MNV-S7) were significantly lower than those of non-infected CD1d KO mice. On the other hand, the body weights of infected WT mice were comparable to those of non-infected WT mice. Correspondingly, CD1d KO mice had an almost 1000-fold higher MNV-S7 burden in the intestine after infection in comparison to WT mice. The mechanism responsible for the insufficient MNV-S7 clearance in CD1d KO mice was attributed to reduced IFN-γ production early during MNV-S7 infection. In addition, the markedly impaired IL-4 production in CD1d KO mice resulted in an impaired MNV-S7-specific secretory IgA production after MNV-S7 infection which is associated with mucosal immunity. Thus, the present results provide evidence that NKT cells play an essential role in MNV-S7 clearance.     

Osmotic Stress Responses and Plant Growth Controlled by Potassium Transporters in Arabidopsis

Osmotic adjustment plays a fundamental role in water stress responses and growth in plants; however, the molecular mechanisms governing this process are not fully understood. Here, we demonstrated that the KUP potassium transporter family plays important roles in this process, under the control of abscisic acid (ABA) and auxin. We generated Arabidopsis thaliana multiple mutants for K⁺ uptake transporter 6 (KUP6), KUP8, KUP2/SHORT HYPOCOTYL3, and an ABA-responsive potassium efflux channel, guard cell outward rectifying K⁺ channel (GORK). The triple mutants, kup268 and kup68 gork, exhibited enhanced cell expansion, suggesting that these KUPs negatively regulate turgor-dependent growth. Potassium uptake experiments using ⁸⁶radioactive rubidium ion (⁸⁶Rb⁺) in the mutants indicated that these KUPs might be involved in potassium efflux in Arabidopsis roots. The mutants showed increased auxin responses and decreased sensitivity to an auxin inhibitor (1-N-naphthylphthalamic acid) and ABA in lateral root growth. During water deficit stress, kup68 gork impaired ABA-mediated stomatal closing, and kup268 and kup68 gork decreased survival of drought stress. The protein kinase SNF1-related protein kinases 2E (SRK2E), a key component of ABA signaling, interacted with and phosphorylated KUP6, suggesting that KUP functions are regulated directly via an ABA signaling complex. We propose that the KUP6 subfamily transporters act as key factors in osmotic adjustment by balancing potassium homeostasis in cell growth and drought stress responses.     

Hydroxyl Radical Generation Caused by the Reaction of Singlet Oxygen with a Spin Trap,DMPO, Increases Significantly in the Presence of Biological Reductants

Photosensitizers newly developed for photodynamic therapy of cancer need to be assessed using accurate methods of measuring reactive oxygen species (ROS). Little is known about the characteristics of the reaction of singlet oxygen (¹O²) with spin traps, although this knowledge is necessary in electron spin resonance (ESR)/spin trapping. In the present study, we examined the effect of various reductants usually present in biological samples on the reaction of ¹O² with 5,5-dimethyl-1-pyrroline-N-oxide (DMPO). The ESR signal of the hydroxyl radical (?OH) adduct of DMPO (DMPO-OH) resulting from ¹O²-dependent generation of ?OH strengthened remarkably in the presence of reduced glutathione (GSH), 6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid (Trolox), ascorbic acid, NADPH, etc. A similar increase was observed in the photosensitization of uroporphyrin (UP), rose bengal (RB) or methylene blue (MB). Use of 5-(diethoxyphosphoryl)-5-methyl-1-pyrroline-N-oxide (DEPMPO) as a spin trap significantly lessened the production of its ?OH adduct (DEPMPO-OH) in the presence of the reductants. The addition of DMPO to the DEPMPO-spin trapping system remarkably increased the signal intensity of DEPMPO-OH. DMPO-mediated generation of ?OH was also confirmed utilizing the hydroxylation of salicylic acid (SA). These results suggest that biological reductants enhance the ESR signal of DMPO-OH produced by DMPO-mediated generation of ?OH from ¹O², and that spin trap-mediated ?OH generation hardly occurs with DEPMPO.