Histone chaperone activity of Fanconi anemia proteins, FANCD2 and FANCI, is required for DNA crosslink repair

Fanconi anaemia (FA) is a rare hereditary disorder characterized by genomic instability and cancer susceptibility. A key FA protein, FANCD2, is targeted to chromatin with its partner, FANCI, and plays a critical role in DNA crosslink repair. However, the molecular function of chromatin-bound FANCD2-FANCI is still poorly understood. In the present study, we found that FANCD2 possesses nucleosome-assembly activity in vitro. The mobility of histone H3 was reduced in FANCD2-knockdown cells following treatment with an interstrand DNA crosslinker, mitomycin C. Furthermore, cells harbouring FANCD2 mutations that were defective in nucleosome assembly displayed impaired survival upon cisplatin treatment. Although FANCI by itself lacked nucleosome-assembly activity, it significantly stimulated FANCD2-mediated nucleosome assembly. These observations suggest that FANCD2-FANCI may regulate chromatin dynamics during DNA repair.     

Chemiluminescent immunoassay (CLIA) for salmon growth hormone (GH)

A highly sensitive and specific chemiluminescent immunoassay (CLIA) was developed for quantification of growth hormone (GH) in salmonid species. The CLIA for salmon GH was performed using the sandwich method with anti-GH IgG as the first antibody and chemiluminescent acridinium ester-labelled specific anti-GH F(ab′)₂ as the second antibody. The measurable range of salmon GH in the CLIA was 39–1250 pg/mL using a short assay (1 day) protocol and 3.9–125 pg/mL in a longer (2-day) assay. The dilution curve in the CLIA of serum from masu salmon (Oncorhynchus masou) was parallel to the standard curve of recombinant chum salmon (Oncorhynchus keta) GH. Seasonal changes of serum GH levels were measured in 1 year-old masu salmon cultivated in a pond from March to November. Their serum GH levels increased during smoltification from March to April, achieved a maximum level of 21 ng/mL in August, and then declined gradually to 11 ng/mL in October. © 1997 John Wiley & Sons, Ltd.     

A Similar Expression Pattern of Adhesion Molecules between Intermediate TCR Cells in the Liver and Intraepithelial Lymphocytes in the Intestine

Two major populations of extrathymically differentiated T cells exist in the liver and intestine. Such T cells in the liver have TCR of intermediate intensity (i.e., intermediate TCR cells) and constitutively express IL-2 receptor β-chain (IL-2Rβ), whereas those in the intestine, especially intraepithelial lymphocytes, have TCR of bright intensity, consisting of a mixture of IL-2Rβ⁺ and IL-2Rβ^–. All mature thymocytes and thymus-derived T cells seen in the peripheral immune organs are TCR-bright⁺IL-2Rβ^– under resting conditions. When the expression pattern of adhesion molecules, including CD44, L-selectin, LFA-1 and ICAM-1, was compared among these T-cell populations, they displayed quite unique patterns of expression. All extrathymic T cells in the liver, intestine, and even other organs were CD44⁺L-selectin^– LFA-1⁺⁺ICAM-1⁺, whereas thymocytes and thymus-derived T cells were CD44^– L-selectin⁺LFA-1⁺ICAM-1^–. This inverted expression of adhesion molecules between extrathymic T cells and thymus-derived T cells might be associated with their unique tissue-localization.     

Neurotransmitter-Mediated Regulation of Brain Aromatase: Protein Kinase C- and G-Dependent Induction

Abstract: Aromatase in the diencephalic neurons, the level of which increases transiently during the prenatal to neonatal period, has been suggested to be involved in control of sexual behavior and differentiation of the CNS. Effects of neurotransmitters on levels of aromatase mRNA in cultured neurons were investigated to determine factors regulating the developmental increase that occurs in level of fetal brain aromatase. The expression of aromatase in diencephalic neurons of fetal mice at embryonic day 13, cultured in vitro, was significantly affected by α₁-adrenergic receptor ligands. Aromatase mRNA levels were higher in neurons treated with the α₁-agonist phenylephrine than in control neurons, whereas prazosin, an α₁-antagonist, suppressed this increase, and ligands for α₂- or β-adrenergic receptors did not exert any influence. The profile of α₁-adrenergic receptor subtypes during actual development in vivo suggested that the α_1B subtype is in fact responsible for the signal transduction. Substance P, cholecystokinin, neurotensin, and brain natriuretic peptide also increased the level of expression along with phorbol 12-myristate 13-acetate and dibutyryl-cyclic GMP, whereas forskolin and dibutyryl-cyclic AMP caused a decrease. These data indicate that stimulation via α₁ (possibly α_1B)-adrenergic receptors, as well as receptors of specific neuropeptides, controls the expression of aromatase in embryonic day 13 diencephalic neurons through activation of protein kinase C or G. β-Adrenergic receptors would not appear to participate in the regulation, judging from their developmental profile, although cyclic AMP might be a suppressive second messenger.     

A petal‐specific InMYB1 promoter from Japanese morning glory: a useful tool for molecular breeding of floricultural crops

Production of novel transgenic floricultural crops with altered petal properties requires transgenes that confer a useful trait and petal‐specific promoters. Several promoters have been shown to control transgenes in petals. However, all suffer from inherent drawbacks such as low petal specificity and restricted activity during the flowering stage. In addition, the promoters were not examined for their ability to confer petal‐specific expression in a wide range of plant species. Here, we report the promoter of InMYB1 from Japanese morning glory as a novel petal‐specific promoter for molecular breeding of floricultural crops. First, we produced stable InMYB1_1kb::GUS transgenic Arabidopsis and Eustoma plants and characterized spatial and temporal expression patterns under the control of the InMYB1 promoter by histochemical β‐glucuronidase (GUS) staining. GUS staining patterns were observed only in petals. This result showed that the InMYB1 promoter functions as a petal‐specific promoter. Second, we transiently introduced the InMYB1_1 kb::GUS construct into Eustoma, chrysanthemum, carnation, Japanese gentian, stock, rose, dendrobium and lily petals by particle bombardment. GUS staining spots were observed in Eustoma, chrysanthemum, carnation, Japanese gentian and stock. These results showed that the InMYB1 promoter functions in most dicots. Third, to show the InMYB1 promoter utility in molecular breeding, a MIXTA‐like gene function was suppressed or enhanced under the control of InMYB1 promoter in Arabidopsis. The transgenic plant showed a conspicuous morphological change only in the form of wrinkled petals. Based on these results, the InMYB1 promoter can be used as a petal‐specific promoter in molecular breeding of floricultural crops.     

Synthesis of Very-Long-Chain Fatty Acids in the Epidermis Controls Plant Organ Growth by Restricting Cell Proliferation

Plant organ growth is controlled by inter-cell-layer communication, which thus determines the overall size of the organism. The epidermal layer interfaces with the environment and participates in both driving and restricting growth via inter-cell-layer communication. However, it remains unknown whether the epidermis can send signals to internal tissue to limit cell proliferation in determinate growth. Very-long-chain fatty acids (VLCFAs) are synthesized in the epidermis and used in the formation of cuticular wax. Here we found that VLCFA synthesis in the epidermis is essential for proper development of Arabidopsis thaliana. Wild-type plants treated with a VLCFA synthesis inhibitor and pasticcino mutants with defects in VLCFA synthesis exhibited overproliferation of cells in the vasculature or in the rib zone of shoot apices. The decrease of VLCFA content increased the expression of IPT3, a key determinant of cytokinin biosynthesis in the vasculature, and, indeed, elevated cytokinin levels. These phenotypes were suppressed in ipt3;5;7 triple mutants, and also by vasculature-specific expression of cytokinin oxidase, which degrades active forms of cytokinin. Our results imply that VLCFA synthesis in the epidermis is required to suppress cytokinin biosynthesis in the vasculature, thus fine-tuning cell division activity in internal tissue, and therefore that shoot growth is controlled by the interaction between the surface (epidermis) and the axis (vasculature) of the plant body.     

In vivo and in vitro studies on the appearance of LHRH neurons in the hypothalamus of perinatal rats

Summary Ontogenetic development of LHRH-containing neurons was studied by fluorescence and enzyme immunohistochemistry in rats. In in vitro studies, the tissues of the septal-chiasmatic and mediobasal hypothalamic areas of fetal rats on day 16.5 or 18.5 of gestation were trypsinized separately for dissociation of the neural cells, and cultured for several days. Immunopositive reaction against LHRH was first detected in nerve cells derived from both areas of the hypothalamus of the fetuses on days 16.5 and 18.5 of gestation, after 8 and 6 days culture, respectively. The cells were small, and seemed to be bipolar in morphology indicating an axon and arborized dendrites. Immunopositive material occurred in the cell soma as well as in the cellular processes. In in vivo studies, immunopositive material, possibly deposited in nerve fibers, appeared first in OVLT and simultaneously in the external layer of the median eminence of fetuses on day 20.5 of gestation. The immunoreactive fibers increased in number in both parts with development, especially after birth in the median eminence. No immunopositive material was detected within any neural cell bodies nor in the cytoplasm of any ependymal cells.This work was financed by the Ministry of Education, Japan. No. 257008. We would like to thank Dr. Katsuhiko Saito (Department of Surgery, Tokushima University) for his kind advice on the preparation of the antibody used for the immunofluorescence study.