Light-induced phase-shifting of the peripheral circadian oscillator in the hearts of food-deprived mice.

In the present study, we investigated the effect of fasting on photoentrainment of the peripheral circadian oscillator in the mammalian heart. Northern blotting showed that a single light pulse applied at an appropriate time in constant darkness, caused obvious phase-shifting in the circadian expression rhythm of the mammalian clock gene Period2 (mPer2) even in the hearts of food-deprived mice. Fasting did not significantly affect either the phase or the light-induced phase-shifts of the mPer2 rhythm. Although several studies of temporal feeding restriction have indicated that feeding is the dominant timing cue for mammalian peripheral oscillators, our findings suggest that feeding is not essential for mammals to induce phase resetting of the circadian oscillator in the heart.     

Analyses of beta-1 syntrophin, syndecan 2 and gem GTPase as candidates for chicken muscular dystrophy.

Despite intensive studies of muscular dystrophy of chicken, the responsible gene has not yet been identified. Our recent studies mapped the genetic locus for abnormal muscle (AM) of chicken with muscular dystrophy to chromosome 2q using the Kobe University (KU) resource family, and revealed the chromosome region where the AM gene is located has conserved synteny to human chromosome 8q11-24.3, where the beta-1 syntrophin (SNTB1), syndecan 2 (SDC2) and Gem GTPase (GEM) genes are located. It is reasonable to assume those genes might be candidates for the AM gene. In this study, we cloned and sequenced the chicken SNTB1, SDC2 and GEM genes, and identified sequence polymorphisms between parents of the resource family. The polymorphisms were genotyped to place these genes on the chicken linkage map. The AM gene of chromosome 2q was mapped 130 cM from the distal end, and closely linked to calbindin 1 (CALB1). SNTB1 and SDC2 genes were mapped 88.5 cM distal and 27.6 cM distal from the AM gene, while the GEM gene was mapped 18.5 cM distal from the AM gene and 9.1 cM proximal from SDC2. Orthologues of SNTB1, SDC2 and GEM were syntenic to human chromosome 8q. SNTB1, SDC2 and GEM did not correspond to the AM gene locus, suggesting it is unlikely they are related to chicken muscular dystrophy. However, this result also suggests that the genes located in the proximal region of the CALB1 gene on human chromosome 8q are possible candidates for this disease.     

Nuclear localization activity of phytochrome B

Phytochromes are soluble red/far-red-light photoreceptor proteins which mediate various photomorphogenic responses of plants. Despite much effort, the signal transduction mechanism of phytochrome has remained obscure. Phytochromes are encoded by a small multigene family in Arabidopsis . Among the members of the family, phytochrome A (phyA) and B (phyB) are the best characterized. PhyB contains putative nuclear localization signals within its C-terminal region. Transgenic Arabidopsis plants were produced which expressed a fusion protein consisting of GUS and C-terminal fragments of phyB. GUS staining from the fusion protein in these transgenic plants was observed in the nucleus, which suggests that the nuclear localization signal of the fragment is functional. Next, it was examined whether the endogenous phyB was detected in the nucleus. Nuclei were isolated from the light-grown wild-type Arabidopsis leaves and subjected to the immunoblot analysis. The result indicated that a substantial fraction of total phyB was recovered in the isolated nuclei. This result was further confirmed by the immunocytochemical analysis of the protoplasts. Finally, the effects of light treatments on the levels of phyB in the isolated nuclei were examined. Dark adaptation of the plants before the nuclear isolation reduced the levels of phyB. The reduction was accelerated by irradiation of plants with far-red light before the transfer to darkness. Thus, nuclear localization of phyB was suggested to be light-dependent.     

Reproductive success in male Japanese minnows,Pseudorasbora parva: observations under experimental conditions

To evaluate the spawning success of male Japanese minnows,Pseudorasbora parva, and female mate choice, spawning behaviour was observed under both artificial and experimental conditions. Larger males had larger territories and greater reproductive success. The body weight of territorial males decreased during the maintenance of territories, while that of non-territorial males increased significantly. When the weight of non-territorial males exceeded that of territorial males, the former began to establish new territories on the substrate, suggesting a conditional strategy by non-territorial males to trade off immediate reproductive success with growth and hence improve future reproductive success. Females chose males with larger body size, probably based on dominance rank rather than the quality (or size) of territory. It was concluded that females choose males of higher dominance rank and that males compete for large territories, both of which play an important part in male reproductive success.     

Distribution of various nickel compounds in rat organs after oral administration

In this study, eight kinds of nickel (Ni) compounds were orally administered to Wistar male rats and the distribution of each compound was investigated 24 h after the administration. The Ni compounds used in this experiment were nickel metal [Ni−M], nickel oxide (green) [NiO(G)], nickel oxide (black) [NiO(B)], nickel subsulfide [Ni₃S₂], nickel sulfide [NiS], nickel sulfate [NiSO₄], nickel chloride [NiCl₂], and nickel nitrate [Ni(NO₃)₂]. The solubilities of the nickel compounds in saline solution were in the following order; [Ni(NO₃)₂>NiCl₂>NiSO₄]≫[NiS>Ni₃S₂]>[NiO(B)>Ni−M>NiO(G)]. The Ni level in the visceral organs was higher in the rats given soluble Ni compounds; Ni(NO₃)₂, NiCl₂, NiSO₄, than that in the rats receiving other compounds. In the rats to which soluble Ni compounds were administered, 80–90% of the recovered Ni amounts in the examined organs was detected in the kidneys. On the other hand, the Ni concentration in organs administered scarcely soluble Ni compounds; NiO(B), NiO(G), and Ni−M were very low. The estimated absorbed fraction of each Ni compounds was increased with the increase of the solubility. These results suggest that the kinetic behavior of Ni compounds administered orally is closely related with the solubility of Ni compounds, and that the solubility of Ni compounds is one of the important factors for determining the health effect of Ni compounds.     

Purification and Characterization of a Ca2+-Dependent Protein Kinase from the Halotolerant Green Alga Dunaliella tertiolecta

A Ca²⁺-dependent protein kinase (CDPK) that has been partiallypurified and characterized previously [Yuasa and Muto (1992)Arch. Biochem. Biophys. 296: 175] was further purified to about20,000-fold from the soluble fraction of Dunaliella tertiolecta.The enzyme preparation contained 60- and 52-kDa polypeptidesboth of which phosphorylated casein as a substrate. Both polypeptidesshowed a Ca²⁺-dependent increase in mobility during SDS-PAGEand ⁴⁵Ca²⁺-binding activity after SDS-PAGE and electroblottingonto a nitrocellulose membrane, suggesting that both the 60-and 52-kDa CDPKs directly bind Ca²⁺. The protein kinase inhibitors,K-252a and staurosporine, inhibited the CDPK competitively withrespect to ATP. An antibody raised against the 60-kDa CDPK crossreactedwith both the 60- and 52-kDa polypeptides. Both molecular specieswere autophosphorylated in the presence of Ca²⁺, and a highlyphosphorylated 80-kDa band appeared in addition to these phosphorylatedbands at 60 and 52 kDa in SDS-PAGE. However, the specific activityof CDPK was not changed by prior autophosphorylation when theautophosphorylated enzyme was assayed as a mixture of thesephosphorylated molecular species. Only the 60-kDa polypeptidewas immunodetected in subcellular fractions of Dunaliella cells.The 52-kDa polypeptide increased during storage of the enzyme.These results suggest that the 52-kDa polypeptide is a proteolyticartifact produced during purification. Immunoreactive bandsof 60-kDa were detected in extracts of several green algae butnot in extracts of higher plants or a brown alga. ¹This research was partly supported by Grants-in-Aid from theMinistry of Education, Science and Culture, Japan (No. 06454013and 06304023) and Research Fellowship of the Japan Society forthe Promotion of Science for Young Sciencists. ²Research Fellow (PD) of the Japan Society for the Promotionof Science.     

Formation of a metallocycle by dimerization of acrylonitrile. Crystal structure of the hexakis{(1,4-dicyanobutane-1,4-diyl)-nickel(II)} complex

A novel hexanickel(II) complex [Ni₆(NCCHCH₂CH₂CHCN)₆] (2) with 1,4-dicyanobutane-1,4-diyl (L) which was produced by the metal-induced dimerization of acrylonitrile (AN) has been isolated and the structure has been determined crystallographically. Complex 2 is triclinic, space group

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. Each nickel atom is coordinated by two carbon atoms of L and two nitrogen atoms of the cyano group of two other L, providing a square-plenar geometry. The six nickel atoms are bridged by the cyano group and carbon atom to form the slightly distorted octahedral Ni₆ core.     

In vitro neurotoxicity of amyloid β-peptide cross-linked by transglutaminase

Transglutaminase catalyzes the intermolecular cross-linking of peptides between Gln and Lys residues, forming an -(-glutamyl) lysine bond. Amyloid -peptide, a major constituent of the deposits in Alzheimer disease, contains Lys16, Lys28, and Gln15 which may act as substrates of transglutaminase. Transglutaminase treatment of amyloid -peptide (1–28) and amyloid -peptide (1–40) yielded cross-linked oligomers. Transglutaminase-treated A retarded neurite extension of PC12 cells, and rat cultured neurons of hippocampus and septum, brain areas severely affected by Alzheimer disease, and subsequently caused cell death, whereas the transglutaminase-untreated counterparts did not show harmful effects. The transglutaminase-catalyzed oligomers of amyloid -peptide and their neurotoxicity may be involved in two characteristics in Alzheimer disease, neuronal degeneration and formation of the insoluble deposits.Abbreviations: AD – Alzheimer disease, A – amyloid -peptide, DMEM – Dulbecco's modified Eagle's medium, DMEM/F–12–1:1 mixture of DMEM and Ham's F–12 medium, FCS – fetal calf serum, HS – horse serum, PAGE – polyacrylamide gel electrophoresis, MTT – 3-(4,5-dimethylthiazol–2-yl)–2,5-diphenyltetrazolium bromide, NGF – nerve growth factor, TGase – transglutaminase.