Developmental expression of HpNanos, the Hemicentrotus pulcherrimus homologue of nanos

The Hemicentrotus pulcherrimus homologue of nanos (HpNanos), that encodes a protein containing two CCHC zinc finger motifs, was isolated from a gastrula cDNA library. The accumulation of HpNanos mRNA during embryonic development and the spatial expression pattern are reported. Developmental northern blot analysis revealed that HpNanos mRNA markedly accumulated during the blastula stages, and then decreased in abundance at the mesenchyme blastula stage. The second phase of HpNanos mRNA expression occurred during gastrulation, after which the expression returned to a low level. Whole-mount in situ hybridization showed that the HpNanos was exclusively expressed in four to six small micromere-descendant cells at the blastula stage. The expression of HpNanos was restricted to the coelomic pouch, which gives rise to the mesoderm of the ventral surface of the adult rudiment, at the prism stage. These results suggest that HpNanos expression will be instrumental for future analyses of the function of small micromere-descendant cells and of the origin of germ cells during sea urchin development.     

Occurrence of the Fimbria Gene hifA in clinical isolates of nonencapsulated Haemophilus influenzae

The adherence of Haemophilus influenzae to epithelial cells plays a crucial role in infections. However, little is known about the occurrence of fimbriae. In this study, we examined the distribution of the fimbria gene (hifA) by PCR among 167 H. influenzae strains isolated from patients with respiratory infections. Almost all (163; 98%) of the isolates were nonencapsulated strains. The carriage rate of hifA by the nonencapsulated strains was 18.4%. Electron microscopy showed that fimbriae were abundantly present on the cell surface of hifA-positive strains tested. Only four (2.4%) isolates were encapsulated, all of which were type b and did not possess hifA. The present work suggests that fimbriae may play a considerable role as adhesins in nonencapsulated H. influenzae strains.     

Purification and properties of soluble and bound gamma-glutamyltransferases from radish cotyledon

Soluble and cell wall bound gamma-glutamyltransferases (GGTs) were purified from radish (Raphanus sativus L.) cotyledons. Soluble GGTs (GGT I and II) had the same M(r) of 63,000, and were composed of a heavy subunit (M(r), 42,000) and a light one (M(r), 21,000). The properties of GGT I and II were similar. Bound GGTs (GGT A and B) were purified to homogeneity from the pellet after the extraction of soluble GGTs. GGT A and B were monomeric proteins with an M(r) of 61,000. The properties of GGT A and B were similar. Thus, bound GGTs were distinguished from soluble GGTs. The optimal pHs of soluble and bound GGTs were about 7.5. Both soluble and bound GGTs utilized glutathione, gamma-L-glutamyl-p-nitroanilide, oxidized glutathione and the conjugate of glutathione with monobromobimane as substrates, and were inhibited by acivicin, but soluble GGTs were also distinguished from bound GGTs with regard to these properties.     

Growth strategy of an emergent macrophyte, Typha orientalis Presl, in comparison with Typha latifolia L. and Typha angustifolia L.

The growth strategy of an emergent plant, Typha orientalis Presl, was examined in experimental ponds in comparison with two other Typha species distributed in Japan, Typha latifolia L. and Typha angustifolia L. T. orientalis showed the greatest ability of vegetative reproduction at the expense of growth in height. T. orientalis started to produce new ramets earlier than T. latifolia and T. angustifolia. These results suggest that T. orientalis should be a rather pioneer-like species and would be restricted to disturbed habitats.     

The family 42 carbohydrate-binding module of family 54 alpha-L-arabinofuranosidase specifically binds the arabinofuranose side chain of hemicellulose

Alpha-L-arabinofuranosidase catalyses the hydrolysis of the alpha-1,2-, alpha-1,3-, and alpha-1,5-L-arabinofuranosidic bonds in L-arabinose-containing hemicelluloses such as arabinoxylan. AkAbf54 (the glycoside hydrolase family 54 alpha-L-arabinofuranosidase from Aspergillus kawachii) consists of two domains, a catalytic and an arabinose-binding domain. The latter has been named AkCBM42 [family 42 CBM (carbohydrate-binding module) of AkAbf54] because homologous domains are classified into CBM family 42. In the complex between AkAbf54 and arabinofuranosyl-alpha-1,2-xylobiose, the arabinose moiety occupies the binding pocket of AkCBM42, whereas the xylobiose moiety is exposed to the solvent. AkCBM42 was found to facilitate the hydrolysis of insoluble arabinoxylan, because mutants at the arabinose binding site exhibited markedly decreased activity. The results of binding assays and affinity gel electrophoresis showed that AkCBM42 interacts with arabinose-substituted, but not with unsubstituted, hemicelluloses. Isothermal titration calorimetry and frontal affinity chromatography analyses showed that the association constant of AkCBM42 with the arabinose moiety is approximately 10(3) M(-1). These results indicate that AkCBM42 binds the non-reducing-end arabinofuranosidic moiety of hemicellulose. To our knowledge, this is the first example of a CBM that can specifically recognize the side-chain monosaccharides of branched hemicelluloses.     

Chara myosin and the energy of cytoplasmic streaming

Recently, it was found that myosin generating very fast cytoplasmic streaming in Chara corallina has very high ATPase activity. To estimate the energy consumed by this myosin, its concentration in the internodal cells of C. corallina was determined by quantitative immunoblot. It was found that the concentration of Chara myosin was considerably high (200 nM) and the amount of ATP consumed by this myosin would exceed that supplied by dark respiration if all myosin molecules were fully activated by the interaction with actin. These results and model calculations suggested that the energy required to generate cytoplasmic streaming is very small and only one-hundredth of the existing myosin is enough to maintain the force for the streaming in the Chara cell.     

Contribution of ethylene biosynthesis for resistance to blast fungus infection in young rice plants

The role of ethylene (ET) in resistance to infection with blast fungus (Magnaporthe grisea) in rice (Oryza sativa) is poorly understood. To study it, we quantified ET levels after inoculation, using young rice plants at the four-leaf stage of rice cv Nipponbare (wild type) and its isogenic plant (IL7), which contains the Pi-i resistance gene to blast fungus race 003. Small necrotic lesions by hypersensitive reaction (HR) were formed at 42 to 72 h postinoculation (hpi) in resistant IL7 leaves, and whitish expanding lesions at 96 hpi in susceptible wild-type leaves. Notable was the enhanced ET emission at 48 hpi accompanied by increased 1-aminocyclopropane-1-carboxylic acid (ACC) levels and highly elevated ACC oxidase (ACO) activity in IL7 leaves, whereas only an enhanced ACC increase at 96 hpi in wild-type leaves. Among six ACC synthase (ACS) and seven ACO genes found in the rice genome, OsACS2 was transiently expressed at 48 hpi in IL7 and at 96 hpi in wild type, and OsACO7 was expressed at 48 hpi in IL7. Treatment with an inhibitor for ACS, aminooxyacetic acid, suppressed enhanced ET emission at 48 hpi in IL7, resulting in expanding lesions instead of HR lesions. Exogenously supplied ACC compromised the aminooxyacetic acid-induced breakdown of resistance in IL7, and treatment with 1-methylcyclopropene and silver thiosulfate, inhibitors of ET action, did not suppress resistance. These findings suggest the importance of ET biosynthesis and, consequently, the coproduct, cyanide, for HR-accompanied resistance to blast fungus in young rice plants and the contribution of induced OsACS2 and OsACO7 gene expression to it.     

Monodentate, didentate chelating, and bridging 1,8-naphthyridine complexes of pentamethylcyclopentadienyliridium(III): Syntheses and structures in the solid states and in solution

A series of new iridium(III) complexes containing pentamethylcyclopentadienyl (Cp^∗ = η⁵-C₅Me₅) and 1,8-naphthyridine (napy) have been prepared. X-ray crystallography revealed that napy acted as a monodentate, a didentate chelating, and a bridging ligand in complexes of [Cp^∗IrCl₂(napy)] (1), [Cp^∗IrCl(napy)]PF₆ (2), and [(Cp^∗IrCl)₂(H)(napy)]PF₆ (4), respectively. The crystal structure of [Cp^∗Ir(napy)₂](PF₆)₂ (3) has also been determined; the dicationic complex bore both monodentate and chelating napy ligands. Dinuclear Cp^∗Ir^III complex bridged by napy was only isolable if two Ir^III centers were supported by a hydride (H⁻) bridge. In complexes 2 and 3, the four-membered chelate rings formed by napy exhibited a large steric strain; in the rings the NIrN bond angles were only 60.5(2)-61.0(4)° and the IrNC angles were 94.7(8)-96.7(8)°. The bridging coordination of napy in complex 4 also afforded a large strain, i.e., the Ir^III centers were displaced by 0.84(3) Å from the napy plane, due to the steric interaction between two Cp^∗IrCl moieties. The monodentate napy complex 1 in CDCl₃ or CD₂Cl₂ at ambient temperature showed a rapid coordination-site exchange reaction, which gave two N sites of napy equivalent; at temperatures below −40 °C, the ¹H NMR spectra corresponded to the molecular structure of [Cp^∗IrCl₂(napy-κN)]. The analogous diazido complex of [Cp^∗Ir(N₃)₂(napy)] (5) has also been prepared, and the crystal structure has been determined. In contrast to the dichloro complex 1, the diazido complex 5 exhibited a dissociation equilibrium of coordinated napy in solution.     

Rho2 is a target of the farnesyltransferase Cpp1 and acts upstream of Pmk1 mitogen-activated protein kinase signaling in fission yeast

We have previously demonstrated that knockout of the calcineurin gene or inhibition of calcineurin activity by immunosuppressants resulted in hypersensitivity to Cl- in fission yeast. We also demonstrated that knockout of the components of the Pmk1 mitogen-activated protein kinase (MAPK) pathway, such as Pmk1 or Pek1 complemented the hypersensitivity to Cl-. Using this interaction between calcineurin and Pmk1 MAPK, here we developed a genetic screen that aims to identify new regulators of the Pmk1 signaling and isolated vic (viable in the presence of immunosuppressant and chloride ion) mutants. One of the mutants, vic1-1, carried a missense mutation in the cpp1+ gene encoding a beta subunit of the protein farnesyltransferase, which caused an amino acid substitution of aspartate 155 of Cpp1 to asparagine (Cpp1(D155N)). Analysis of the mutant strain revealed that Rho2 is a novel target of Cpp1. Moreover, Cpp1 and Rho2 act upstream of Pck2-Pmk1 MAPK signaling pathway, thereby resulting in the vic phenotype upon their mutations. Interestingly, compared with other substrates of Cpp1, defects of Rho2 function were more phenotypically manifested by the Cpp1(D155N) mutation. Together, our results demonstrate that Cpp1 is a key component of the Pck2-Pmk1 signaling through the spatial control of the small GTPase Rho2.