Impact of Nuclear Envelope Stress on Physiological and Pathological Processes in Central Nervous System

Neurochemical Research - The nuclear envelope (NE) separates genomic DNA from the cytoplasm and provides the molecular platforms for nucleocytoplasmic transport, higher-order chromatin...     

Photochemical intermediates of Arabidopsis phototropin 2 LOV domains associated with conformational changes

The photochemical reactions of Arabidopsis phototropin 2 light- oxygen-voltage domain 2 (LOV2) with the linker region (LOV2-linker), without the linker (LOV2), and LOV1 were studied using the time-resolved transient grating (TG) and transient lens (TrL) methods. Although the absorption spectra did not change after the formation of the adduct species, a small volume expansion process with a time constant of 9 ms was observed for LOV2. For the LOV2-linker, at 293 K, a volume contraction process with a time constant of 140 mus was observed in addition to a volume expansion process with 9 ms and the diffusion coefficient change with 2 ms. The reaction intermediate species were characterized on the basis of their thermodynamic properties, such as changes in enthalpy, thermal expansion, and heat capacity. For the first intermediate (S(390)), the values of these properties were similar to those of the ground state for both LOV2 and LOV2-linker. A relatively large thermal expansion volume (0.09 cm(3)mol(-1)K(-1)) and a positive heat capacity change (4.7 kJ mol(-1)K(-1)) were detected for the intermediates of LOV2-linker. These characteristic features were interpreted in terms of structural fluctuation and exposure of hydrophobic residues in the linker domain, respectively. The enthalpy change of S(390) of the LOV1 domain was significantly greater than changes for the LOV2 or LOV2-linker samples. Data from this study support a major conformational change of the linker region in the photochemical reaction of phototropin.     

Subtype Specification of Cerebral Cortical Neurons in Their Immature Stages

Neurochemical Research - The diversification of neuronal subtypes during corticogenesis is fundamental to the establishment of the complex cortical structure. Although subtype specification has...     

Genes outside the S supergene suppress S functions in buckwheat (Fagopyrum esculentum)

BACKGROUND AND AIMS: Common buckwheat (Fagopyrum esculentum) is a dimorphic self-incompatible plant with either pin or thrum flowers. The S supergene is thought to govern self-incompatibility, flower morphology and pollen size in buckwheat. Two major types of self-fertile lines have been reported. One is a type with long-homostyle flowers, Kyukei SC2 (KSC2), and the other is a type with short-homostyle flowers, Pennline 10. To clarify whether the locus controlling flower morphology and self-fertility of Pennline 10 is the same as that of KSC2, pollen tube tests and genetic analysis have been performed. METHODS: Pollen tube growth was assessed in the styles and flower morphology of KSC2, Pennline 10, F1 and F2 plants that were produced by the crosses between plants with pin or thrum and Pennline 10. KEY RESULTS: Pollen tubes of Pennline 10 reached ovules of all flower types. The flower morphology of F1 plants produced by the cross between thrum and Pennline 10 were thrum or pin, and when pin plants were used as maternal plants, all the F1 plants were pin. Both plants with pin or short-pin flowers, whose ratio of style length to anther height was smaller than that of pin, appeared in F2 populations of thrum x Pennline 10 as well as in those of pin x Pennline 10. CONCLUSION: The results suggest that Pennline 10 possesses the s allele as pin does, not an allele produced by the recombination in the S supergene, and that the short style length of Pennline 10 is controlled by multiple genes outside the S supergene.     

Production of cloned pigs from adult somatic cells by chemically assisted removal of maternal chromosomes

The present study demonstrated that brief treatment of in vitro-matured porcine oocytes with demecolcine results in a membrane protrusion that contains a condensed chromosome mass, which can be easily removed by aspiration. This simple, chemically assisted method for removing maternal chromosomes enabled the production of a large number of nuclear-transferred porcine eggs. The development of eggs whose chromosomes were removed by this procedure following transfer of somatic cell nuclei to the blastocyst stage was not significantly different among groups activated using different procedures (6% to 11%) and was also not different among donor cells of different origins (3% to 9%), except for cumulus cells (0.4%). After transfer of 180 to 341 nuclear-transferred eggs that received somatic cells to 6 recipients, 2 of the recipients produced 8 healthy cloned piglets from the heart cells of a female pig. The chemically assisted method for removing maternal chromosomes was also effective for bovine and rabbit eggs.     

Hydroxylation-induced stabilization of the collagen triple helix. Acetyl-(glycyl-4(R)-hydroxyprolyl-4(R)-hydroxyprolyl)(10)-NH(2) forms a highly stable triple helix

The collagen triple helix is one of the most abundant protein motifs in animals. The structural motif of collagen is the triple helix formed by the repeated sequence of -Gly-Xaa-Yaa-. Previous reports showed that H-(Pro-4(R)Hyp-Gly)(10)-OH (where '4(R)Hyp' is (2S,4R)-4-hydroxyproline) forms a trimeric structure, whereas H-(4(R)Hyp-Pro-Gly)(10)-OH does not form a triple helix. Compared with H-(Pro-Pro-Gly)(10)-OH, the melting temperature of H-(Pro-4(R)Hyp-Gly)(10)-OH is higher, suggesting that 4(R)Hyp in the Yaa position has a stabilizing effect. The inability of triple helix formation of H-(4(R)Hyp-Pro-Gly)(10)-OH has been explained by a stereoelectronic effect, but the details are unknown. In this study, we synthesized a peptide that contains 4(R)Hyp in both the Xaa and the Yaa positions, that is, Ac-(Gly-4(R)Hyp-4(R)Hyp)(10)-NH(2) and compared it to Ac-(Gly-Pro-4(R)Hyp)(10)-NH(2), and Ac-(Gly-4(R)Hyp-Pro)(10)-NH(2). Ac-(Gly-4(R)Hyp-4(R)Hyp)(10)-NH(2) showed a polyproline II-like circular dichroic spectrum in water. The thermal transition temperatures measured by circular dichroism and differential scanning calorimetry were slightly higher than the values measured for Ac-(Gly-Pro-4(R)Hyp)(10)-NH(2) under the same conditions. For Ac-(Gly-4(R)Hyp-4(R)Hyp)(10)-NH(2), the calorimetric and the van't Hoff transition enthalpy DeltaH were significantly smaller than that of Ac-(Gly-Pro-4(R)Hyp)(10)-NH(2). We postulate that the denatured states of the two peptides are significantly different, with Ac-(Gly-4(R)Hyp-4(R)Hyp)(10)-NH(2) forming a more polyproline II-like structure instead of a random coil. Two-dimensional nuclear Overhauser effect spectroscopy suggests that the triple helical structure of Ac-(Gly-4(R)Hyp-4(R)Hyp)(10)-NH(2) is more flexible than that of Ac-(Gly-Pro-4(R)Hyp)(10)-NH(2). This is confirmed by the kinetics of amide (1)H exchange with solvent deuterium of Ac-(Gly-4(R)Hyp-4(R)Hyp)(10)-NH(2), which is faster than that of Ac-(Gly-Pro-4(R)Hyp)(10)-NH(2). The higher transition temperature of Ac-(Gly-4(R)Hyp-4(R)Hyp)(10)-NH(2), can be explained by the higher trans/cis ratio of the Gly-4(R)Hyp peptide bonds than that of the Gly-Pro bonds, and this ratio compensates for the weaker interchain hydrogen bonds.     

Pyridinium cationic-dimer antimalarials, unlike chloroquine, act selectively between the schizont stage and the ring stage of Plasmodium falciparum

Malaria is a leading cause of death in developing countries, and the emergence of strains resistant to the main therapeutic agent, chloroquine, has become a serious problem. We have developed cationic-dimer type antimalarials, MAP-610 and PMAP-H10, which are structurally different from chloroquine. In this study, we introduced several substituents on the terminal phenyl rings of PMAP-H10. The electronic and hydrophobic properties of the substituents were correlated with the antimalarial activity and cytotoxicity of the compounds, respectively. Studies with synchronized cultures of malarial plasmodia showed that our cationic-dimers act selectively between the schizont stage and the ring stage of the parasitic cycle, unlike chloroquine, which has a stage-independent action. Thus, the mechanism of action of our antimalarials appears to be different from that of chloroquine, and our compounds may be effective against chloroquine-resistant strains.     

Immunolocalization of calcineurin and FKBP12, the FK506-binding protein, in Hassall's corpuscles of human thymus and epidermis

Calcineurin, a Ca2+/calmodulin-dependent protein phosphatase, plays an important role in various physiological functions including T cell activation. This enzyme is a target molecule for the immunosuppressants, cyclosporin A and FK506. In the present study, we investigated immunohistochemical localization of calcineurin and FKBP12, an FK506-binding protein, in human thymus and epidermis. The catalytic subunit (calcineurin A) of calcineurin was abundantly expressed in Hassall's corpuscles which were localized in the thymic medulla and represented the terminal stages of thymic medullary epithelium. The regulatory subunit (calcineurin B) of calcineurin was also expressed in high amounts in Hassall's corpuscles. In the epidermis, which shows similarities to Hassall's corpuscles, both subunits were also abundantly expressed, and their expression increased with the differentiation of keratinocytes. FKBP12 was observed to be expressed abundantly, both in Hassall's corpuscles and the entire epidermis. These findings suggest that the differentiated forms of the two cell types, which are the thymic medullary epithelial cell and the keratinocyte, are the target for pharmacological actions of FK506.     

A possible role for calmodulin in Ca2+-induced swelling of mitochondria

Ca2+-induced mitochondrial swelling was inhibited by a low concentration of calmodulin antagonists. Two affinities of Ca2+ to mitochondrial swelling were observed: high (2-5 microM) and low (more than 100 microM) systems. The high-affinity change was inhibited by micromolar level of trifluoperazine and W-7, but not by W-5. The possible mechanism of this inhibition and the role of CaM in mitochondria are discussed.     

Plasma membrane-associated sialidase as a crucial regulator of transmembrane signalling

Mammalian sialidases, glycosidases responsible for the removal of sialic acids from glycoproteins and glycolipids, has been implicated to participate in many biological processes as well as in lysosomal catabolism. Among those forms identified to date, plasma membrane-associated sialidase, Neu3, is a key enzyme in degradation of gangliosides, for which it exhibits a special substrate preference. This sialidase has been shown to control transmembrane signalling for many cellular processes, including cell differentiation, cell growth and apoptosis, and human orthologue NEU3 is markedly up-regulated in various cancers. It is known to suppress apoptosis in cancer cells. Furthermore, its overexpression causes impaired glucose tolerance and hyper-insulinaemia together with overproduction of insulin in enlarged islets in the transgenic mice. The present review primarily summarizes our recent results, focusing on Neu3 as a regulator of transmembrane signalling.