Nutritive effects of d-amino acids on the silkworm, Bombyx mori

Nutritive effects of d-amino acids on the silkworm, Bombyx mori, were investigated by growth experiments using defined diets and also by analysis of free amino acids in the larval haemolymph. None of the d-forms of the usual ten essential amino acids could be utilized effectively, although d-methionine was utilized in lieu of the l-form only to a limited extent and d-histidine gave a positive but smaller effect than d-methionine. d-Proline, its l-form being semi-essential for the silkworm, was not utilized. d-Leucine, and to a lesser extent d-alanine and d-serine, were found to be somewhat toxic. Comparison of free amino acid patterns in the haemolymph of the fifth-instar larvae, which fed on diets either lacking l-forms of histidine, methionine and leucine singly or including the d-forms singly in place of these l-forms, supported the results of the growth experiments.     

Involvement of Ca2+-dependent proteasome in the degradation of both cyclin B1 and Mos during spontaneous activation of matured rat oocytes

In matured rat oocytes, spontaneous activation from the metaphase-II (MII) stage occurred after collection from the oviducts. It is well known that the mitogen-activated protein kinase (MAPK) pathway and p34(cdc2) kinase play an important role in the arrest at MII in other species. However, there is no information about the difference in these factors among strains of rats. In the present study, in spontaneously activated oocytes from the Wistar rat, the Mos protein level and the activity of MAPK kinase (MEK)/MAPK were decreased at 120 min (13.8, 25.7, and 19.3, respectively, P<0.05), whereas Sprague-Dawley (SD) oocytes, which were not spontaneously activated, had a high level of Mos protein and MEK/MAPK activity (75.9, 76.2, and 87.9, respectively, P<0.05). Phosphorylation of MAPK in the SD oocytes was significantly suppressed by MEK inhibitor, U0126 at 60 min; this treatment decreased p34(cdc2) kinase activity via cyclin B1 degradation in a time-dependent manner. The treatment with proteasome inhibitor, MG132 or Ca2+-chelator, BAPTA-AM, overcame the spontaneous degradation of both Mos and cyclin B1 in a dose-dependent manner in Wistar oocytes. More than 90% of Wistar oocytes treated with BAPTA-AM were arrested at MII until 120 min. In conclusion, SD oocytes carrying Mos/MEK/MAPK, maintained a high activity of p34(cdc2) kinase by stabilizing cyclin B1, thus involved in their meiotic arrest. In contrast, Wistar oocytes had a relatively low cytostatic factor activity; rapid decrease of Mos/MEK/MAPK failed to stabilize both cyclin B1 and Mos, and these oocytes were likely to spontaneously activate.     

A test of the effect of floral color change on pollination effectiveness using artificial inflorescences visited by bumblebees

Floral color change has been recognized as a pollination strategy, but its relative effectiveness has been evaluated insufficiently with respect to other floral traits. In this study, effects of floral color change on the visitation pattern of bumblebees were empirically assessed using artificial flowers. Four inflorescence types were postulated as strategies of flowering behavior: type 1 has no retention of old flowers, resulting in a small display size; type 2 retains old flowers without nectar production; type 3 retains old flowers with nectar; and type 4 retains color-changed old flowers without nectar. Effects of these treatments varied depending on both the total display size (single versus multiple inflorescences) and the pattern of flower-opening. In the single inflorescence experiment, a large floral display due to the retention of old flowers (types 2–4) enhanced pollinator attraction, and the number of flower visits per stay decreased with color change (type 4), suggesting a decrease in geitonogamous pollination. Type-4 plants also reduced the foraging time of bees in comparison with type-2 plants. In the multiple inflorescence experiment, the retention of old flowers did not contribute to pollinator attraction. When flowering occurred sequentially within inflorescences, type-4 plants successfully decreased the number of visits and the foraging time in comparison with type-2 plants. In contrast, floral color change did not influence the number of visits, and it extended the foraging time when flowering occurred simultaneously within inflorescences but the opening of inflorescences progressed sequentially within a plant. Therefore, the effectiveness of floral color change is highly susceptible to the display size and flowering pattern within plants, and this may limit the versatility of the color change strategy in nature.     

Sensitive detection of FGFR3 mutations in bladder cancer and urine sediments by peptide nucleic acid-mediated real-time PCR clamping

Somatic mutations of the fibroblast growth factor receptor 3 (FGFR3) gene were detected by peptide nucleic acid (PNA)-mediated real-time PCR clamping. Mutation was detected in negative control containing only wild-type DNA due to a misincorporation of dNTPs to PNA binding sites when the amount of template DNA was decreased to 1 ng. Thus, the amount of template DNA was critical determinant of the assay sensitivity in PNA-mediated PCR clamping. Assay conditions were optimized to detect FGFR3 mutations in exons 7, 10, and 15, at a concentration of more than 1% mutated DNA using 50 ng of genomic DNA as the template. Mutations were detected in 12 of 13 (92.3%) tumor tissues and 11 of 13 (84.6%) urine samples from patients with superficial bladder cancer, while no mutations were detected in tissues and/or urine samples from patients with muscle-invasive bladder cancer or chronic cystitis.     

The ability of whale haploid spermatogenic cells to induce calcium oscillations and its relevance to oocyte activation

Interspecies microinsemination assay was applied to examine the ability of minke whale haploid spermatogenic cells to induce Ca2+ oscillations and oocyte activation. Populations of round spermatids (RS), early-stage elongating spermatids (e-ES), late-stage elongating spermatids (1-ES) and testicular spermatozoa (TS) were cryopreserved in the presence of 7.5% glycerol on board ship in the Antarctic Ocean. Repetitive increases of intracellular Ca2+ concentration occurred in 0, 65, 81 and 96% of BDF1 mouse oocytes injected with the postthaw RS, e-ES, 1-ES and TS, respectively. A normal pattern of the Ca2+ oscillations was observed in 26-47% of the responding oocytes. Most oocytes that exhibited Ca2+ oscillations, regardless of the oscillation pattern, resumed meiosis (83-94%). These results indicate that whale spermatogenic cells acquire SOAF activity, which is closely related to their Ca2+ oscillation-inducing ability at the relatively early stage of spermiogenesis.     

Carbohydrate-recognition domains of galectin-9 are involved in intermolecular interaction with galectin-9 itself and other members of the galectin family

Galectin-9 (Gal-9) is a tandem-repeat-type member of the galectin family associated with diverse biological processes, such as apoptosis, cell aggregation, and eosinophil chemoattraction. Although the detailed sugar-binding specificity of Gal-9 has been elucidated, molecular mechanisms that underlie these functions remain to be investigated. During the course of our binding study by affinity chromatography and surface plasmon resonance (SPR) analysis, we found that human Gal-9 interacts with immobilized Gal-9 in the protein-protein interaction mode. Interestingly, this intermolecular interaction strongly depended on the activity of the carbohydrate recognition domain (CRD), because the addition of potent saccharide inhibitors abolished the binding. The presence of multimers was also confirmed by Ferguson plot analysis of result of polyacrylamide gel electrophoresis and matrix-assisted laser-desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry (MS). Moreover, this intermolecular interaction was observed between Gal-9 and other galectin members, such as Gal-3 and Gal-8, but not Gal-1. Because such properties have not been reported yet, they may explain an unidentified mechanism underlying the diverse functions of Gal-9.     

Genetic and epigenetic alteration among three homoeologous genes of a class E MADS box gene in hexaploid wheat

Bread wheat (Triticum aestivum) is a hexaploid species with A, B, and D ancestral genomes. Most bread wheat genes are present in the genome as triplicated homoeologous genes (homoeologs) derived from the ancestral species. Here, we report that both genetic and epigenetic alterations have occurred in the homoeologs of a wheat class E MADS box gene. Two class E genes are identified in wheat, wheat SEPALLATA (WSEP) and wheat LEAFY HULL STERILE1 (WLHS1), which are homologs of Os MADS45 and Os MADS1 in rice (Oryza sativa), respectively. The three wheat homoeologs of WSEP showed similar genomic structures and expression profiles. By contrast, the three homoeologs of WLHS1 showed genetic and epigenetic alterations. The A genome WLHS1 homoeolog (WLHS1-A) had a structural alteration that contained a large novel sequence in place of the K domain sequence. A yeast two-hybrid analysis and a transgenic experiment indicated that the WLHS1-A protein had no apparent function. The B and D genome homoeologs, WLHS1-B and WLHS1-D, respectively, had an intact MADS box gene structure, but WLHS1-B was predominantly silenced by cytosine methylation. Consequently, of the three WLHS1 homoeologs, only WLHS1-D functions in hexaploid wheat. This is a situation where three homoeologs are differentially regulated by genetic and epigenetic mechanisms.     

Natural dental caries in molars of osteogenic disorder Shionogi rats

Osteogenic disorder Shionogi (ODS) rats are genetically defective in ascorbic acid biosynthesis. They exhibit a gait abnormality due to dysfunctional bone formation and display various dental abnormalities. Conditions of the oral cavity and tooth quality both influence the development of dental caries. This study was designed to determine the characteristics of dental caries in ODS/ ShiJclod/od rats. Caries were scored and compared among ODS/ShiJclod/od, ODS/ShiJcl+/+, and Jcl:Wistar retired breeders. Among male rats, the caries scores of the ODS/ShiJclod/od and ODS/ShiJcl+/+ groups were similar to each other but greater than those in Jcl:Wistar rats, whereas among female rats, caries scores in ODS/ShiJclod/od animals were equivalent to or somewhat greater than those in ODS/ShiJcl+/+ rats, whose scores were markedly greater than those of Jcl:Wistar rats. The results suggest that ODS/ShiJcl rats were more susceptible to dental caries than were Jcl:Wistar rats. Under the conditions of the study, caries scores between ODS/ ShiJclod/od and ODS/ShiJcl+/+ rats differed only among parous females.