Excessive ingestion of long-chain polyunsaturated fatty acids during developmental stage causes strain- and sex-dependent eye abnormalities in mice

The eyes are riched in long-chain polyunsaturated fatty acids (LC-PUFAs) such as arachidonic acid [ARA; 20:4 (n−6)] and docosahexaenoic acid [DHA; 22:6 (n−3)]. Despite their abundance in the eyes, ARA and DHA cannot be sufficiently synthesized de novo in mammals. During gestation, eye development is exceptionally rapid, and substantial amounts of LC-PUFAs are needed to ensure proper eye development. Here, we studied the influences of dietary LC-PUFAs in dams (C57BL/6 and C3H/He) on the eye morphogenesis and organogenesis of their pups. Intriguingly, fetuses and newborn mice from C57BL/6 dams fed an LC-PUFA (particularly ARA)-enriched diet displayed a much higher incidence of eye abnormalities such as microphthalmia (small eye) and corneal opacity than those from dams fed an LC-PUFA-poor diet. The effects of LC-PUFAs on eye anomalies were evident only in the female pups of C57BL/6 inbred mice, not in those of C3H/He mice or male C57BL/6 mice. These results demonstrate a gene-by-environment (GxE) interaction in eye development in mice. Furthermore, our molecular analysis suggested the potential roles of Pitx3 and Pax6 in the above interaction involving ARA.     

Expression pattern of annelid <Emphasis Type="Italic">Zic</Emphasis> in embryonic development of the oligochaete <Emphasis Type="Italic">Tubifex tubifex</Emphasis>

Embryonic expression of a Zic homologue (Ttu-Zic) was examined in the oligochaete annelid Tubifex tubifex. The body plan of T. tubifex is characterized by obvious segmentation in the ectoderm and mesoderm. Ttu-Zic expression is detected in the mesodermal germ band and a subset of micromere descendants. Ttu-Zic is transiently expressed in primary m-blast cells (i.e., founder cells of mesodermal segments) as early as the time of their birth from M teloblasts. During its development, each mesodermal segment experiences two additional phases of Ttu-Zic expression. Ttu-Zic expression in micromere descendants is seen on the anterior surfaces of embryos undergoing teloblastogenesis; subsequently, these cells proliferate to form bilateral clusters, which then become internalized. Finally, clusters of Ttu-Zic-expressing cells are found in the center of the prostomium, corresponding to the cerebral ganglion. The Ttu-Zic expression profile in the early embryogenesis of T. tubifex may be homologous to those of evolutionarily distant animals.     

A proteolytic enzyme, trypsin, for immunofluorescence observation of microtubules in algal cells

The usefulness of a proteolytic enzyme, trypsin, in immunofluorescence microscopy was confirmed in algal ceils. Flagella of Umspora penicilliformis zoo-spores were visible using an anti-β tubulin antibody after trypsin treatment, and the cortical microtubules of vegetative cells could also be clearly detected. Interestingly, centrioles that were not detected in the control observation appeared in gametophyte cells of Acrosiphonia duriuscula and Monostroma angicava using the trypsin treatment.     

Rapid DNA extraction from conchocelis and ITS-1 rDNA sequences of seven strains of cultivated Porphyra yezoensis (Bangiales, Rhodophyta)

In order to extract DNA rapidly from cultivated Porphyra, we extracted total DNA from conchocelis using the ISOPLANT II kit (Nippon Gene) without liquid nitrogen treatment or CsCl-gradient ultracentrifugation. By confirming the reproducibility of RAPD patterns, it is concluded that the quality of the extracted DNA is sufficient to use as a template for molecular investigation. Using this rapid method, the nuclear ribosomal DNA of the internal transcribed spacer (ITS) regions was amplified from seven strains of cultivated Porphyra, which had been maintained as free-living conchocelis by subculturing in the laboratory. From the amplified DNAs, the ITS-1 sequences were determined in order to identify the species and genetic relationship of the strains. The sequences were identical in the seven strains, and all the strains were identified as P. yezoensis. Furthermore, the gametophytic blades of these strains showed long linear or oblanceolate shapes in the laboratory culture. It was concluded that these strains are P. yezoensis form. narawaensis. This rapid DNA extraction method from conchocelis will be a powerful tool for phylogenetic analysis and for genetic improvement of cultivated Porphyra.     

Cryptic species in the Pyropia yezoensis complex (Bangiales,Rhodophyta): Sympatric occurrence of two cryptic species even on same rocks

In a previous study on wild populations of Pyropia, the occurrence of two possible new species (Pyropia sp. 2 and Pyropia sp. 3) which are closely related to the two commercially important Pyropia species, P. yezoensis and P. tenera, was confirmed as the result of molecular phylogenetic analyses. To characterize the morphological features of the two wild Pyropia species, we collected Pyropia blades in a natural population in which Pyropia sp. 3 was known to occur, and carried out molecular identification before detailed morphological observations. Through the molecular identification we found, unexpectedly, that Pyropia sp. 2 blades grew sympatrically in the same site. Therefore, after molecular identification, we examined in detail the external morphology and anatomy of the two wild Pyropia species using more than 10 blades each. As a result, it is concluded that all of the blades of the two species are morphologically identical to P. yezoensis, but distinct from P. tenera. It is therefore considered that both of the two wild Pyropia species are cryptic species within the P. yezoensis complex. Furthermore, this study revealed that the two cryptic species grew sympatrically, even on the same rocks within the natural habitat.     

Impaired cognitive function and altered hippocampal synapse morphology in mice lacking Lrrtm1, a gene associated with schizophrenia

Recent genetic linkage analysis has shown that LRRTM1 (Leucine rich repeat transmembrane neuronal 1) is associated with schizophrenia. Here, we characterized Lrrtm1 knockout mice behaviorally and morphologically. Systematic behavioral analysis revealed reduced locomotor activity in the early dark phase, altered behavioral responses to novel environments (open-field box, light-dark box, elevated plus maze, and hole board), avoidance of approach to large inanimate objects, social discrimination deficit, and spatial memory deficit. Upon administration of the NMDA receptor antagonist MK-801, Lrrtm1 knockout mice showed both locomotive activities in the open-field box and responses to the inanimate object that were distinct from those of wild-type mice, suggesting that altered glutamatergic transmission underlay the behavioral abnormalities. Furthermore, administration of a selective serotonin reuptake inhibitor (fluoxetine) rescued the abnormality in the elevated plus maze. Morphologically, the brains of Lrrtm1 knockout mice showed reduction in total hippocampus size and reduced synaptic density. The hippocampal synapses were characterized by elongated spines and diffusely distributed synaptic vesicles, indicating the role of Lrrtm1 in maintaining synaptic integrity. Although the pharmacobehavioral phenotype was not entirely characteristic of those of schizophrenia model animals, the impaired cognitive function may warrant the further study of LRRTM1 in relevance to schizophrenia.     

Induction and characterization of pigmentation mutants in Porphyra yezoensis (Bangiales, Rhodophyta)

Porphyra yezoensis Ueda conchospore germlings (1–4-cell stages) were treated with N-methyl-N′-nitro-N-nitrosoguanidine (MNNG) for inducing mutations. Three kinds of color-mutated gametophytic blades, which were composed of the mutated cells wholly, sectorially or spottedly, were obtained; and most of them were sectorially variegated blades. The highest frequency of these mutated blades was 1.3%. Four different pigmentation mutant strains were obtained by regenerating single cells and protoplasts that were enzymatically isolated from the mutated sectors of the sectorially variegated blades. The mutants were relatively stable in color in both gametophytic blade and conchocelis phases. In the two phases, each mutant strain showed characteristic differences in the in vivo absorption spectra, and had different pigment contents of major photosynthetic pigments (chlorophyll a, phycoerythrin and phycocyanin) as compared with the wild-type and with each other. The gametophytic blades from the four mutant lines showed significant differences in growth and photosynthetic rates, when they were cultured in the same conditions. By crossing the mutant with the wild-type, it was found that the color phenotypes of two mutants reported above, were resulted from two mutations in different genes, respectively. This revised version was published online in August 2006 with corrections to the Cover Date.     

Comparative photosynthetic studies of Ecklonia cava (Laminariales,Phaeophyta) bladelets with and without zoosporangial sori

Photosynthetic rates were compared between Ecklonia cava bladelets with and without zoosporangial sori sampled from the subtidal zone (about 5 m deep) in Nabeta Bay, Shimoda, Japan. Photosynthetic rates of bladelets were lower in the sorus portion than in the non-sorus portion on the basis of area, dry weight and chlorophyll a. Respiration rates were higher in the sorus portion than in the non-sorus portion on the basis of area and chlorophyll a, whereas they were almost the same on a dry weight basis. The differences were mainly due to a large difference in dry weight per unit bladelet area between the sorus and the non-sorus portion. Light compensation points were higher in the sorus portion than in the non-sorus portion.     

Immunofluorescence study of mitosis and cytokinesis in Acrosiphonia duriuscula (Acrosiphoniales,Chlorophyta)*

The behaviors of nuclei and microtubules (MT) in Acrosiphonia duriuscula (Ruprecht) Collins were observed in detail using fluorescence and electron microscopy. Numerous nuclei exist in cells of A. duriuscula (multinucleate cells). Cortical MT radiate from the apex of the tip cell and run parallel to its long axis. Between 30 and 40% of nuclei in the upper part of cytoplasm migrate downward to the region where cytokinesis will take place, and these numerous nuclei form a ‘nuclear ring’ before mitosis. The parallel array of the cortical MT changes to a transverse orientation at the region where cytokinesis will take place, and finally forms a characteristic circumferential band. Mitosis starts from the nuclei in the ring. Cortical MT disappear in the region of the nuclear ring and many mitotic spindles form. The band-shaped array of MT remains. Mitosis spreads in an apparent wave to the other nuclei. After mitosis, daughter nuclei that formed a nuclear ring migrate apically and repopulate the apical daughter cell. When the numerous daughter nuclei have relocated, a rearrangement of the cortical MT occurs. They are randomly arranged at first, but finally become parallel to the long axis of the cell. Cytokinesis occurs by furrowing of the cell, and the band-shaped array of MT could be detected at the leading edge of the furrow.     

A wide-range phylogenetic analysis of Zic proteins: implications for correlations between protein structure conservation and body plan complexity

We compared Zic homologues from a wide range of animals. Striking conservation was found in the zinc finger domains, in which an exon-intron boundary has been kept in all bilateralians but not cnidarians, suggesting that all of the bilateralian Zic genes are derived from a single gene in a bilateralian ancestor. There were additional conserved amino acid sequences, ZOC and ZF-NC. Combined analysis of the zinc finger, ZOC, and ZF-NC revealed the presence of two classes of Zic, based on the degree of protein structure conservation. The "conserved" class includes Zic proteins from the Arthropoda, Mollusca, Annelida, Echinodermata, and Chordata (vertebrates and cephalochordates), whereas the "diverged" class contains those from the Platyhelminthes, Cnidaria, Nematoda, and Chordata (urochordates). The result indicates that the ancestral bilateralian Zic protein had already acquired an entire set of conserved domains, but that this was lost and diverged in the platyhelminthes, nematodes, and urochordates.