Behavioral responses of males to estradiol-treated females in the budgerigar (<Emphasis Type="Italic">Melopsittacus undulatus</Emphasis>)

We investigated the effects of estradiol-treated females on the behavior of male budgerigars. In comparison to control females, females given implants of estradiol showed elevated nest behavior and darker cere color, which are characteristics of breeding females. After we confirmed the efficacy of estradiol treatment on behavior and morphology, each female was paired with a male mate. Males paired with estradiol-treated females showed more courtship behavior (auditory and visual display, and courtship feeding) to their mates than males paired with control females. These data indicate that budgerigar females with a high estrogen level enhance males' courtship behavior. Since males did not show response to estradiol-treated females soon after females were introduced, effects of estradiol-treated female budgerigars may be mediated by the endocrine system, rather than wholly by the nervous system, of males. Electronic Publication     

Analysis of Wnt8 for neural posteriorizing factor by identifying Frizzled 8c and Frizzled 9 as functional receptors for Wnt8

The dorsal ectoderm of vertebrate gastrula is first specified into anterior fate by an activation signal and posteriorized by a graded transforming signal, leading to the formation of forebrain, midbrain, hindbrain and spinal cord along the anteroposterior (A-P) axis. Transplanted non-axial mesoderm rather than axial mesoderm has an ability to transform prospective anterior neural tissue into more posterior fates in zebrafish. Wnt8 is a secreted factor that is expressed in non-axial mesoderm. To investigate whether Wnt8 is the neural posteriorizing factor that acts upon neuroectoderm, we first assigned Frizzled 8c and Frizzled 9 to be functional receptors for Wnt8. We then, transplanted non-axial mesoderm into the embryos in which Wnt8 signaling is cell-autonomously blocked by the dominant-negative form of Wnt8 receptors. Non-axial mesodermal transplants in embryos in which Wnt8 signaling is cell-autonomously blocked induced the posterior neural markers as efficiently as in wild-type embryos, suggesting that Wnt8 signaling is not required in neuroectoderm for posteriorization by non-axial mesoderm. Furthermore, Wnt8 signaling, detected by nuclear localization of beta-catenin, was not activated in the posterior neuroectoderm but confined in marginal non-axial mesoderm. Finally, ubiquitous over-expression of Wnt8 does not expand neural ectoderm of posterior character in the absence of mesoderm or Nodal-dependent co-factors. We thus conclude that other factors from non-axial mesoderm may be required for patterning neuroectoderm along the A-P axis.     

Studies on the Metabolic Activity of Muscle Proteins

The activities of glutamic oxaloacetic transaminase and Ca⁺⁺ ion-activated ATPase of muscle in the adult rats fed a protein-free diet for 8, 16 and 24 days were measured in order to clarify their metabolic responses with respect to reserve proteins. It was found that these enzyme activities, or presumably their enzyme proteins, decreased at the stage as early as the 8th day of protein depletion following the same pattern as seen in reserve proteins. Their responses, particularly those in unit activity, were somewhat different from each other. The metabolic significance of those responses was discussed in relation to protein nutrition.     

Structural analysis of an acidic, fatty acid ester-bonded extracellular polysaccharide produced by a pristane-assimilating marine bacterium, Rhodococcus erythropolis PR4

Rhodococcus erythropolis PR4 is a marine bacterium that can degrade various alkanes including pristane, a C(19) branched alkane. This strain produces a large quantity of extracellular polysaccharides, which are assumed to play an important role in the hydrocarbon tolerance of this bacterium. The strain produced two acidic extracellular polysaccharides, FR1 and FR2, and the latter showed emulsifying activity toward clove oil, whereas the former did not. FR2 was composed of D-galactose, D-glucose, D-mannose, D-glucuronic acid, and pyruvic acid at a molar ratio of 1:1:1:1:1, and contained 2.9% (w/w) stearic acid and 4.3% (w/w) palmitic acid attached via ester bonds. Therefore, we designated FR2 as a PR4 fatty acid-containing extracellular polysaccharide or FACEPS. The chemical structure of the PR4 FACEPS polysaccharide chain was determined by 1D (1)H and (13)C NMR spectroscopies as well as by 2D DQF-COSY, TOCSY, HMQC, HMBC, and NOESY experiments. The sugar chain of PR4 FACEPS was shown to consist of tetrasaccharide repeating units having the following structure: [structure: see text].     

Trichothecene nonproducer Gibberella species have both functional and nonfunctional 3-O-acetyltransferase genes

The trichothecene 3-O-acetyltransferase gene (FgTri101) required for trichothecene production by Fusarium graminearum is located between the phosphate permease gene (pho5) and the UTP-ammonia ligase gene (ura7). We have cloned and sequenced the pho5-to-ura7 regions from three trichothecene nonproducing Fusarium (i.e., F. oxysporum, F. moniliforme, and Fusarium species IFO 7772) that belong to the teleomorph genus Gibberella. BLASTX analysis of these sequences revealed portions of predicted polypeptides with high similarities to the TRI101 polypeptide. While FspTri101 (Fusarium species Tri101) coded for a functional 3-O-acetyltransferase, FoTri101 (F. oxysporum Tri101) and FmTri101 (F. moniliforme Tri101) were pseudogenes. Nevertheless, F. oxysporum and F. moniliforme were able to acetylate C-3 of trichothecenes, indicating that these nonproducers possess another as yet unidentified 3-O-acetyltransferase gene. By means of cDNA expression cloning using fission yeast, we isolated the responsible FoTri201 gene from F. oxysporum; on the basis of this sequence, FmTri201 has been cloned from F. moniliforme by PCR techniques. Both Tri201 showed only a limited level of nucleotide sequence similarity to FgTri101 and FspTri101. The existence of Tri101 in a trichothecene nonproducer suggests that this gene existed in the fungal genome before the divergence of producers from nonproducers in the evolution of Fusarium species.     

Synthesis of the cyclic and acyclic acetal derivatives of 1-(3-C-ethynyl-beta-D-ribo-pentofuranosyl)cytosine,a potent antitumor nucleoside. Design of prodrugs to be selectively activated in tumor tissues via the bio-reduction-hydrolysis mechanism

We have designed and synthesized the acetal derivatives of 1-(3-C-ethynyl-beta-D-ribo-pentofuranosyl)cytosine (ECyd, 1), the 2',3'-O-nitrobenzylidene derivatives 2 and 3 and the 5'-O-(alkoxy)(nitrophenyl)methyl derivatives 6-10 as potential prodrugs of ECyd. These prodrugs can be selectively activated in tumor tissues via a bio-reduction-hydrolysis mechanism owing to the characteristic properties of tumor tissues, such as hypoxia and lower pH. Although the 2',3'-O-(4-nitrobenzylidene) derivatives 2 and 3 were converted bio-reductively into the corresponding 4-aminobenzylidene derivatives by rat S-9 mix, the reduction products, that is, the corresponding amino congeners 4 and 5, proved to be rather stable in an aqueous solution at pH 6.5 used as a pH model for acidic tumor tissues. In contrast, the 5'-O-(alkoxy)(4-nitropheny)methyl derivatives 6-8 were also reduced by rat S-9 mix to the corresponding amino congeners 11-13, which were hydrolyzed to release ECyd more effectively at pH 6.5 than at pH 7.4. Accordingly, the acyclic acetals 6-8 may be efficient prodrugs of ECyd, that are effectively reduced under physiological conditions releasing ECyd in acidic tumor tissues.     

Insights into tRNA-Dependent Amidotransferase Evolution and Catalysis from the Structure of the Aquifex aeolicus Enzyme

Many bacteria form Gln-tRNA^Gln and Asn-tRNA^Asn by conversion of the misacylated Glu-tRNA^Gln and Asp-tRNA^Asn species catalyzed by the GatCAB amidotransferase in the presence of ATP and an amide donor (glutamine or asparagine). Here, we report the crystal structures of GatCAB from the hyperthermophilic bacterium Aquifex aeolicus, complexed with glutamine, asparagine, aspartate, ADP, or ATP. In contrast to the Staphylococcus aureus GatCAB, the A. aeolicus enzyme formed acyl-enzyme intermediates with either glutamine or asparagine, in line with the equally facile use by the amidotransferase of these amino acids as amide donors in the transamidation reaction.A water-filled ammonia channel is open throughout the length of the A. aeolicus GatCAB from the GatA active site to the synthetase catalytic pocket in the B-subunit. A non-catalytic Zn²⁺ site in the A. aeolicus GatB stabilizes subunit contacts and the ammonia channel. Judged from sequence conservation in the known GatCAB sequences, the Zn²⁺ binding motif was likely present in the primordial GatB/E, but became lost in certain lineages (e.g., S. aureus GatB). Two divalent metal binding sites, one permanent and the other transient, are present in the catalytic pocket of the A. aeolicus GatB. The two sites enable GatCAB to first phosphorylate the misacylated tRNA substrate and then amidate the activated intermediate to form the cognate products, Gln-tRNA^Gln or Asn-tRNA^Asn.