Genotype/age interactions on aggressive behavior in gonadally intact estrogen receptor beta knockout (betaERKO) male mice

Estrogen, as an aromatized metabolite of testosterone, has a facilitatory effect on male aggressive behavior in mice. Two subtypes of estrogen receptors, alpha (ER-alpha) and beta (ER-beta), in the brain are known to bind estrogen. Previous studies revealed that the lack of ER-alpha gene severely reduced the induction of male aggressive behavior. In contrast, mice that lacked the ER-beta gene tended to be more aggressive than wild type (WT) control mice, although the behavioral effects of ER-beta gene disruption were dependent on their social experience. These findings lead us to hypothesize that estrogen may facilitate aggression via ER-alpha whereas it may inhibit aggression via ER-beta. In the present study, we further investigated the role of ER-beta in the regulation of aggressive behavior by examining developmental changes starting at the time of first onset, around the age of puberty. Aggressive behaviors of ER-beta gene knockout (betaERKO) mice were examined in three different age groups, puberty, young-adult, and adult. Each mouse was tested every other day for three times in a resident-intruder paradigm against olfactory bulbectomized intruder mice and their trunk blood was collected for measurements of serum testosterone after the completion of the study. Overall, betaERKO mice were significantly more aggressive than WT. These genotype differences were more pronounced in puberty and young adult age groups, but not apparent in the adult age group, in which betaERKO mice were less aggressive than those in two younger age groups. Serum testosterone levels of betaERKO mice were significantly higher than those of WT mice only in the pubertal age group, but not in young adult (when betaERKO mice were still significantly more aggressive than WT mice) and adult (when no genotype differences in aggression were found) age groups. These results suggest that ER-beta mediated actions of gonadal steroids may more profoundly be involved in the inhibitory regulation of aggressive behavior in pubertal and young adult mice.     

Characterization of a SINE species from vicuna and its distribution in animal species including the family Camelidae

Twenty-six sequences of a short interspersed repetitive element (SINE) with a size of approximately 150 base pairs (bp) were isolated from the genomic DNA of Vicugna vicugna (vicuna). RNA polymerase III split promoter sequence was observed in most of them, and many had direct repeats flanking to SINEs as well as a poly(A)-like structure. The SINE sequences were designated as ``vic-1' sequences. Comparison of the vic-1 consensus sequence with sequences registered in the DNA database (DDBJ/EMBL/GENBANK) revealed that the vic-1 sequence had a 79% homology with mouse ala-tRNA gene. In addition, the tRNA-related region of the consensus sequence was folded into a cloverleaf structure as with mouse ala-tRNA. These findings strongly indicated that vic-1 was a retroposon derived from ala-tRNA gene. The vic-1 sequences were used as a probe for dot-blot hybridization to examine the distribution of their homologous sequences in the genomes of various animal species spanning 14 orders, of which, homologous sequences were found only in the Camelidae family. In order to examine the phylogenetical relationship among vicuna, llama, and camel, vic-1 insertion analysis and homology analysis of vic-1 sequences were performed at each locus. The analyses indicated that vic-1 sequences were generated in a common ancestor of the animal species, and that camels first branched off from the clade Camelidae, followed by vicunas and llamas. Received: 5 July 2000 / Accepted: 14 November 2000     

Cultivation of a desulfurizing bacterium, Mycobacterium strain G3

Various carbon and sulfur sources on the growth and desulfurization activity of Mycobacterium strain G3, which is a dibenzothiophene (DBT)-degrading microorganism, were studied. Ethanol, glucose or glycerol as the sole carbon source and MgSO₄, taurine or dimethyl sulfoxide (DMSO) as the sole sulfur source were suitable for the growth. In addition, desulfurization activity was expressed in medium containing taurine, MgSO₄ or DMSO at 0.1 mM, when 217 mM ethanol was used as the sole carbon source. The highest desulfurization activity was in the stationary phase cells after 5 days' growth, rather than those harvested during active growth, when Mycobacterium G3 was cultivated in medium containing 217 mM ethanol and 0.1 mM MgSO₄. Thus alternative sulfur sources to DBT can be used for the cultivation of this desulfurizing microorganism.     

Quantitative analysis of the hemoglobin oxygenation state of rat brain in vivo by picosecond time-resolved spectrophotometry

Through the use of a picosecond laser pulse of near-infrared light at 1,064 nm, the temporal profile of the transmitted light through the anesthetized rat head has been investigated. The light intensity at a certain time after the input pulse was exponentially attenuated by the hemoglobin concentration with hematocrit values from 1.5 to 50%, although the transmitted pulse broadened markedly due to scattering by the cerebral tissue. The optical pathlength, which is required for quantitation of the absolute absorbance change, was directly determined, by the time of flight measurement of the light pulses, as the product of the velocity of light in tissue and time. The mean concentration of hemoglobin in the brain could be determined quantitatively by the use of this pathlength. The oxygen saturation of venous blood determined by our time of flight measurement was very close to that in the internal jugular vein determined directly with a gas analyzer. Thus, the picosecond laser technique is useful for quantifying the blood oxygenation in tissues.     

Study on placental transmission of Pneumocystis carinii in mice using immunodeficient SCID mice as a new animal model.

Placental transmission of Pneumocystis carinii in mice was examined in 39 animals obtained by caesarean section from 17 pregnant SCID females experimentally infected with P. carinii. When examined with toluidine blue O, DAPI and immunofluorescent antibody stains, P. carinii was detected in the lungs of infected mothers but not in the lungs of caesarean section-derived neonates even after the neonates were treated with dexamethasone for 8 weeks. However, 13 neonates born to five infected females developed P. carinii pneumonia. These results indicate that P. carinii cannot be transmitted transplacentally in mice.     

Three-Dimensional Labeling Program for Elucidation of the Geometric Properties of Biological Particles in Three-Dimensional Space

For all biological particles such as cells or cellular organelles, there are three-dimensional coordinates representing the centroid or center of gravity. These coordinates and other numerical parameters such as volume, fluorescence intensity, surface area, and shape are referred to in this paper as geometric properties, which may provide critical information for the clarification ofin situmechanisms of molecular and cellular functions in living organisms. We have established a method for the elucidation of these properties, designated the three-dimensional labeling program (3DLP). Algorithms of 3DLP are so simple that this method can be carried out through the use of software combinations in image analysis on a personal computer. To evaluate 3DLP, it was applied to a 32-cell-stage sea urchin embryo, double stained with FITC for cellular protein of blastomeres and propidium iodide for nuclear DNA. A stack of optical serial section images was obtained by confocal laser scanning microscopy. The method was found effective for determining geometric properties and should prove applicable to the study of many different kinds of biological particles in three-dimensional space.     

Nonsense and frameshift mutations in ZFHX1B, encoding Smad-interacting protein 1, cause a complex developmental disorder with a great variety of clinical features

Mutations in ZFHX1B, encoding Smad-interacting protein 1 (SIP1), have been recently reported to cause a form of Hirschsprung disease (HSCR). Patients with ZFHX1B deficiency typically show mental retardation, delayed motor development, epilepsy, microcephaly, distinct facial features, and/or congenital heart disease, in addition to the cardinal form of HSCR. To investigate the breadth of clinical variation, we studied DNA samples from six patients with clinical profiles quite similar to those described elsewhere for ZFHX1B deficiency, except that they did not have HSCR. The results showed the previously reported R695X mutation to be present in three cases, with three novel mutations-a 2-bp insertion (760insCA resulting in 254fs262X), a single-base deletion (270delG resulting in 91fs107X), and a 2-bp deletion (2178delTT resulting in 727fs754X)-newly identified in the other three. All mutations occurred in one allele and were de novo events. These results demonstrate that ZFHX1B deficiency is an autosomal dominant complex developmental disorder and that individuals with functional null mutations present with mental retardation, delayed motor development, epilepsy, and a wide spectrum of clinically heterogeneous features suggestive of neurocristopathies at the cephalic, cardiac, and vagal levels.     

Geldanamycin induces CHOP expression through a 4-（2-aminoethyl）-benzenesulfonyl fluoride-responsive serine protease

Dear Editor： Geldanamycin is a benzoquinone ansamycin, which was originally described as a tyrosine kinase inhibitor. However, subsequent studies have revealed that geldanamycin binds to and inhibits heat-shock protein 90 （Hsp90） activity [1]. Hsp90 is a molecular chaperone involved in the conformational maturation of proteins such as mutated p53, Raf- 1, Akt, Bcr-Abl, and ErbB2. It is suggested that agents inhibiting Hsp90 have anti-cancer properties, although the precise molecular mechanisms underlying the anti-cancer effects of geldanamycin are not well understood.     

Hinokiresinol is not a precursor of agatharesinol in the norlignan biosynthetic pathway in Japanese cedar

The biosynthetic relationship between the two norlignans agatharesinol and trans-hinokiresinol was investigated. Fresh sapwood sticks of Cryptomeria japonica were fed with stable isotope-labeled compounds, namely p-coumaryl alcohol-[9,9-²H], p-coumaryl alcohol-[9-¹⁸O] and trans-hinokiresinol-[1-²H], and then incubated under high-humidity for approximately 20 days, during which the two norlignans were produced simultaneously. While trans-hinokiresinol was strongly deuterium-labeled after feeding with p-coumaryl alcohol-[9,9-²H], agatharesinol was only lightly labeled after feeding with either p-coumaryl alcohol-[9,9-²H] or -[9-¹⁸O]. These results suggest that p-coumaryl alcohol, which is a precursor of hinokiresinol, is not involved in the biosynthesis of agatharesinol. Therefore, the norlignan carbon skeleton of agatharesinol must be framed from different types of phenylpropanoid monomers compared to those utilized by the trans-hinokiresinol pathway. The biosynthesis of these two norlignans seems to branch at an early stage, i.e., before the framing of the norlignan carbon skeleton. Furthermore, agatharesinol was not labeled with deuterium after feeding with ²H-labeled trans-hinokiresinol, which has the simplest norlignan structure. This result strongly supports the suggestion that the conversion of trans-hinokiresinol to agatharesinol is not part of the biosynthesis of norlignans and that early branching occurs instead.     

Fbxw7 acts as an E3 ubiquitin ligase that targets c-Myb for nemo-like kinase (NLK)-induced degradation

The c-myb proto-oncogene product (c-Myb) is degraded in response to Wnt-1 signaling via a pathway involving TAK1 (transforming growth factor-beta-activated kinase 1), HIPK2 (homeodomain-interacting protein kinase 2), and NLK (Nemo-like kinase). NLK directly binds to c-Myb, which results in the phosphorylation of c-Myb at multiple sites, and induces its ubiquitination and proteasome-dependent degradation. Here, we report that Fbxw7, the F-box protein of an SCF complex, targets c-Myb for degradation in a Wnt-1- and NLK-dependent manner. Fbxw7alpha directly binds to c-Myb via its C-terminal WD40 domain and induces the ubiquitination of c-Myb in the presence of NLK in vivo and in vitro. The c-Myb phosphorylation site mutant failed to interact with Fbxw7alpha, suggesting that the c-Myb/Fbxw7alpha interaction is enhanced by NLK phosphorylation of c-Myb. Treatment of M1 cells with Fbxw7 small interfering RNA (siRNA) rescued the Wnt-induced c-Myb degradation and also the Wnt-induced inhibition of cell proliferation. NLK bound to Cul1, a component of the SCF complex, while HIPK2 interacted with both Fbxw7alpha and Cul1, suggesting that both kinases enhance the c-Myb/SCF interaction. In contrast to c-Myb, the v-myb gene product (v-Myb) encoded by the avian myeloblastosis virus was resistant to NLK/Fbxw7alpha-induced degradation. Thus, Fbxw7 is an E3 ubiquitin ligase of c-Myb, and the increased c-Myb levels may contribute, at least partly, to transformation induced by mutation of Fbxw7.