Metabolites of mating pheromone,rhodotorucine A, by a cells of Rhodosporidiumtoruloides

Rhodotorucine $\text{A}$ which induces mating tube formation of $\text{a}$ cells in $\text{Rhodosporidium}\text{toruloides}$ is metabolized rapidly by $\text{a}$ cells. By use of labeled rhodotorucine $\text{A}$, the degradation was found to be proteolytic. Two peptide fragments Tyr-Pro-Glu-Ile-Ser-Trp-Thr-Arg and Asn-Gly-Cys(S-farnesyl) were identified as the metabolites. Proteolysis of the pheromone mainly occurred on the cell surface. Culture filtrate of $\text{a}$ cells at log phase did not metabolize rhodotorucine $\text{A}$.     

Heavy meromyosin and subfragment-1 from squid mantle myosin, and Ca-sensitivity of their Mg-ATPases

Heavy meromyosin (HMM) and subfragment-1 (S1) were obtained from squid mantle myosin by tryptic digestion and chymotryptic digestion, respectively. Squid HMM(T) and S1(CT) preparations contained stoichiometric amounts of the two types of light chain subunit; regulatory light chain, LC-2, and essential light chain, LC-1. No difference was detected in the chymotryptic digestibilities of squid mantle myosin in Ca-medium and in EDTA-medium. This is in contrast to the digestibility of scallop adductor myosin. The Mg-ATPase activity of HMM(T) alone and that of acto-HMM(T) were both sensitive to calcium ions. In contrast, the activity of S1(CT) alone and that of acto-S1(CT) were both insensitive to calcium ions. The affinity of HMM(T) for actin was not affected by calcium ions, but the amount of HMM(T) bound to actin was increased by calcium ions from 20% to 60% of the total amount of HMM(T). On the other hand, the actin affinity of S1(CT) and the amount of S1(CT) bound to actin were both unaffected by calcium ions. The role of calcium ions in the regulation of contraction in molluscan muscles is discussed.     

Increasing the Catalytic Performance of a Whole Cell Biocatalyst Harboring a Cytochrome P450cam System by Stabilization of an Electron Transfer Component

Catalytic activity of a recombinant Escherichia coli whole cell biocatalyst harboring a cytochrome P450cam monooxygenase system from Pseudomonas putida coupled with enzymatic co-factor regeneration was investigated. About 0.7 μmol camphor was hydroxylated per mg dry cells at 4°C in 50 mM Tris/HCl buffer (pH 7.4) when utilizing a stable putidaredoxin (Pdx) mutant, C73S/C85S-Pdx (Cys73Ser, Cys85Ser double mutant), instead of wild-type Pdx, which was about two-fold improvement in the substrate conversion. Ten-micromole camphor was completely hydroxylated at 20°C in 6 h by 15 mg dry cell weight of whole cell biocatalyst including C73S/C85S-Pdx. Thus, modulation of protein-protein interaction in multicomponent enzymatic catalysis in whole cells is important.     

A novel anti-plant viral protein from coelomic fluid of the earthworm Eisenia foetida: purification, characterization and its identification as a serine protease

A novel protein showing strong antiviral activities against cucumber mosaic virus (CMV) and tomato mosaic virus (TMV) was purified from the coelomic fluid of the earthworm Eisenia foetida. The protein was characterized as a cold-adapted serine protease. Its molecular weight was estimated to be 27,000 by SDS-PAGE. The enzyme was most active at pH 9.5 and 40–50 °C. The protease activity at 4 °C was 60% of that obtained at the optimal temperature. The activity was suppressed by various serine protease inhibitors. Partial N-terminal amino acid sequence of the enzyme showed homology with serine proteases of earthworms, E. foetida and Lumbricus rubellus previously studied. Our results suggest that the enzyme can be applicable as a potential antiviral factor against CMV, TMV, and other plant viruses.     

Marginal distribution and high heterozygosity of asexual Caloglossa vieillardii (Delesseriaceae,Rhodophyta) along the Australian coasts

In animals and land plants, many asexual species originate through inter‐ or intraspecific crosses, and such heterozygous asexuals frequently are more abundant than their sexual relatives in marginal habitats. Although asexual species have been reported in various macroalgal taxa, detailed information regarding their distribution, heterozygosity, and origin is limited. Because many asexual tetrasporophyte strains of Caloglossa vieillardii have been isolated from South Australia, far from their core tropical habitats, we re‐examined the distribution range of asexual C. vieillardii and genotyped these and other western Pacific strains using an actin gene marker. We confirmed the marginal distribution of the asexuals; however, a small patch of sexual thalli was newly discovered 450 km further west from asexual populations in South Australia. Three heterozygous genotypes and one homozygous genotypes were detected from nine asexual populations; 21 heterozygous strains were obligately asexual, but one homozygous strain suddenly produced sexual gametophytes after several years of culture. We hypothesized that the most abundant heterozygous genotype (defined as type 3/4) in asexual populations occurred by a cross between type 3 and type 4 allele gametophytes, both of which were isolated from the Australian coasts. In the crossing experiments, certain combinations between type 3 females and type 4 males produced tetrasporophytes, which recycled successive tetrasporophytes. In the culture experiments, whereas both sexual and asexual strains successfully produced tetraspores at 12°C, no sexual strains released carpospores below 14°C. However, it is uncertain whether this slight difference of maturation temperature was related to the marginal distribution of asexual C. vieillardii.     

Effect of repeated allogeneic bone marrow mononuclear cell transplantation on brain injury following transient focal cerebral ischemia in rats

Aims

Transplantation of bone marrow mononuclear cells (BMMCs) exerts neuroprotection against cerebral ischemia. We examined the therapeutic timepoint of allogeneic BMMC transplantation in a rat model of focal cerebral ischemia, and determined the effects of repeated transplantation outside the therapeutic window.

Main methods

Male Sprague–Dawley rats were subjected to 90 minute focal cerebral ischemia, followed by intravenous administration of 1 × 10⁷ allogeneic BMMCs or vehicle at 0, 3 or 6 h after reperfusion or 2 × 10⁷ BMMCs 6 h after reperfusion. Other rats administered 1 × 10⁷ BMMCs at 6 h after reperfusion received additional BMMC transplantation or vehicle 9 h after reperfusion. Infarct volumes, neurological deficit scores and immunohistochemistry were evaluated 24 or 72 h after reperfusion.

Key findings

Infarct volumes at 24 h were significantly decreased in transplantation rats at 0 and 3 h, but not at 6 h, after reperfusion, compared to vehicle-treatment. Even high dose BMMC transplantation at 6 h after reperfusion was ineffective. Repeated BMMC transplantation at 6 and 9 h after reperfusion reduced infarct volumes and significantly improved neurological deficit scores at 24 and 72 h. Immunohistochemistry showed repeated BMMC transplantation reduced ionized calcium-binding adapter molecule 1, 4-hydroxy-2-nonenal and 8-hydroxydeoxyguanosine expression at 24 and 72 h after reperfusion.

Significance

Intravenous allogeneic BMMCs were neuroprotective following transient focal cerebral ischemia, and the therapeutic time window of BMMC transplantation was > 3 h and < 6 h after reperfusion in this model. Repeated transplantation at 6 and 9 h after reperfusion suppressed inflammation and oxidative stress in ischemic brains, resulting in improved neuroprotection.     

Factors promoting maternal trophic egg provisioning in non-eusocial animals

The adaptive function of trophic egg-laying is generally regarded as extended parental investment to the offspring. However, the evolutionary factors promoting trophic egg-laying are still unclear, because the amount of maternal investment per offspring should be ideally equal between smaller offspring with trophic eggs and larger offspring without any additional investment. Several authors have suggested that trophic egg-laying should evolve only when egg size is constrained, but this hypothesis has not been evaluated. We investigated the evolutionary mechanisms of trophic egg-laying by two different approaches. First, we evaluated morphological constraints on egg size in two sibling ladybird species, Harmonia axyridis, which is known to produce trophic eggs, and H. yedoensis. Second, we theoretically predicted the optimal proportion of trophic eggs to total eggs and egg size in relation to environmental heterogeneity, predictability of environmental quality, and investment efficiency of trophic egg consumption. The intra- and interspecific morphological comparisons suggest that morphological constraints on the evolutionary determination of egg size are weak at best in the two ladybird species. Moreover, we theoretically showed that small egg size and trophic egg-laying are favoured in heterogeneous environments when mothers cannot adjust egg size plastically. We also showed that even a small reduction in investment efficiency makes a trophic egg strategy unlikely, despite relatively high environmental predictability. We conclude that trophic egg provisioning may be a flexible maternal adaptation to a highly heterogeneous environment rather than a response to a morphological constraint on egg size.     

COMPARATIVE MORPHOLOGY, CROSSABILITY, AND TAXONOMY WITHIN THE CALOGLOSSA CONTINUA (DELESSERIACEAE, RHODOPHYTA) COMPLEX FROM THE WESTERN PACIFIC

Systematic studies were undertaken on the Caloglossa continua ( Okamura) King et Puttock complex from Japan, Singapore, and Australia, based on morphology and reproductive compatibility. Specimens from Japan had two to six cell rows derived from a nodal axial cell, at the margin opposite the branch, whereas those from Australia possessed only a single cell row. Specimens from Singapore formed one to four cell rows per nodal axial cell and always contained at least one single cell row on any one thallus. These differences were maintained in cultured materials over a range of temperatures or salinities. Type material of C. continua had the same morphology as the Japanese specimens in this study. Carpospores discharged from the Japanese isolate germinated at 10°C, whereas those from Singapore and Australia died at this temperature. In hybridization experiments, the Japanese entity was completely nonfertile with both the Singaporean and Australian isolates. Many pseudocystocarps were produced in the crossing between the male from Australia and the female from Singapore, although the reciprocal combination did not produce any such structures. On the basis of the discontinuous morphology coupled with the complete reproductive isolation, the entities from Singapore and Australia are described here as C. monosticha sp. nov. The entities with multiple cell rows likely expanded their geographic range from tropical regions, where the majority of Caloglossa species are now distributed, to high-latitude regions, and such an expansion would be associated with acquiring low-temperature tolerance .     

Analysis of the Epitopes Recognized by Mouse Monoclonal Antibodies Directed to Yersinia enterocolitica Heat-Shock Protein 60

To determine amino acid sequences of the epitopes recognized by monoclonal antibodies (mAbs) 3C8 and 5C3 directed against Yersinia enterocolitica heat-shock protein (HSP60), a dot blot analysis was perfomed using synthesized peptides of Y. enterocolitica HSP60 such as peptides p316-342, p327-359, p340-366, p316-326, p316-321, p319-323, and p321-326 which represent positions of amino acids in Y. enterocolitica HSP60. The dot blot analysis revealed that 5C3 mAb reacted with p316-342, p316-326 and p321-326, and 3C8 mAb p316-342 and p316-326. These results indicate that the epitopes recognized by the mAbs were associated with eleven amino acids, Asp Leu Gly Gln Ala Lys Arg Val Val Ile Asn, of p316-326. The sequence homology between p316-326 of Y. enterocolitica HSP60 and the rest of the HSP60 family suggests that the five amino acids of Lys, Arg, Val, Ile and Asn, which are highly conserved in the HSP60 family, might be related with the epitope recognized by 3C8. In contrast, it was also demonstrated that three amino acids of Leu, Gly and Val, which are not well conserved in the HSP60 family, might be related to the epitope recognized by 5C3.     

Reproduction,grazing, and development of the large subarctic calanoid <Emphasis Type="Italic">Eucalanus bungii</Emphasis>: is the spring diatom bloom the key to controlling their recruitment?

The response of large calanoid, Eucalanus bungii, to environmental fluctuation, particularly in relation to the spring diatom bloom in the Oyashio region, western subarctic Pacific Ocean, was examined by investigating egg production, grazing, development and starvation tolerance. Mean in situ egg production rate increased with ambient chlorophyll-a concentration, ranging from 0 to 47 eggs female⁻¹ d⁻¹, while no diurnal synchronous spawning behavior was observed. Under the spring bloom condition, E. bungii showed prey preference for less mobile and larger-sized prey (≥30 μm ESD) and bloom-forming diatom Thalassiosira spp. accounted for >80% of ingested carbon. In the laboratory, E. bungii was successfully reared from newly hatched nauplii to adult with the diatom, Thalassiosira nordenskioldi, as a food resource. Nauplii newly hatched from eggs reached the adult stage in ca. 150 days (5°C) with a sigmoidal developmental pattern and no sexual difference in development pattern. Starvation experiments indicated that the starved copepodids (C1–C4) became more vulnerable to high temperature with the progression of developmental stage, suggesting that the post-bloom condition with low food availability and increased temperature is harsh for their copepodids. The results of this study in conjunction with previous findings suggest that E. bungii is well adapted to utilize large-sized phytoplankton, such as a bloom-forming diatoms and, therefore, their recruitment processes, including egg production, development and mortality would be strongly affected by the duration and intensity of the spring bloom.