A leptomycin B resistance gene of Schizosaccharomyces pombe encodes a protein similar to the mammalian P-glycoproteins

Screening for leptomycin B (LMB)-resistant transformants in a gene library constructed in Schizosaccharomyces pombe with the chromosomal DNA of an LMB-resistant mutant of S. pombe and with multicopy plasmid pDB248' as the vector led to the isolation of a gene, named pmd1+, encoding a 1362-amino-acid protein. This protein showed great similarity in amino acid sequence to the mammalian P-glycoprotein encoded by the multidrug resistance gene, mdr, and the Saccharomyces cerevisiae a-factor transporter encoded by STE6. In addition, computer analyses predicted that the protein encoded by pmd1+ formed an intramolecular duplicated structure and each of the halves contained six transmembrane regions as well as two ATP-binding domains, as observed with the P-glycoproteins and the STE6 product. Consistent with this was that S. pombe cells containing the pmd1+ gene on a multicopy plasmid showed resistance not only to LMB but also to several cytotoxic agents. The pmd1 null mutants derived by gene disruption were viable and hypersensitive to these agents. All these data suggest that the pmd1+ gene encodes a protein that is a structural and functional counterpart of mammalian mdr proteins.     

Bilateral giant myelolipoma in the adrenal of a cotton-top tamarin (Saguinus oedipus)

Abstract: A rare case of bilateral adrenal myelolipomas in a female cotton-top tamarin is reported. Large bilateral masses in the adrenal glands were composed of mature adipose cells containing varying amounts of hematopoietic cells of the myeloid, erythroid, and megakaryocyte series. The gross and histologic features of this case closely resemble human “giant” adrenal myelolipomas.     

Purification and properties of a novel enzyme,N-acylamino acid racemase,from Streptomyces atratus Y-53

A novel enzyme, N-acylamino acid racemase, was purified to homogeneity from Streptomyces atratus Y-53 and characterized. This enzyme catalyzes the interconversion of optically active N-acylamino acids. The relative molecular mass (M_r) of the enzyme was estimated to be about 41 000 and 244 000 on sodium dodecyl sulfate-polyacrylamide gel electrophoresis and gel filtration, respectively, indicating that the enzyme is composed of six subunits with an equal M_r. The enzyme showed a broad substrate specificity toward N-acylamino acids, such as N-acetylmethionine, N-chloroacetylphenylalanine and N-chloroacetylvaline. The apparent Michaelis constant (K_m) values for N-acetyl-l-methionine and N-acetyl-d-methionine were calculated to be 15.2 and 5.6 mm, respectively. Enzyme activity was markedly enhanced by divalent metal ions, such as Co²⁺, Mg²⁺ and Mn²⁺, and was inhibited by metal-chelating reagent, indicating that the enzyme is a metalloenzyme. We propose to name the enzyme N-acylamino acid racemase (acylamino acid racemase). Correspondence to: S. Tokuyama     

Transformation of fibroblasts into endothelial cells during angiogenesis

Light-and electron-microscopic autoradiography have been used to study fibroblast transformation into endothelial cells in the formation of new blood vessels during wound healing in rabbit ear chambers. When cultured fibroblasts labeled with tritium thymidine were transplanted autologously into the chambers, newly formed blood vessels contained endothelial cells labeled with tritium thymidine. This result suggests that fibroblasts play a pivotal role in angiogenesis, as progenitors of endothelial cells in newly formed blood vessels.     

Conditions for initiation of fertilization of eggs in the copulating elkhorn sculpin

The reason for failure to initiate fertilization internally was examined in a cottid fish, the elkhorn sculpin, Alcichthys alcicornis which has internal gametic association and external fertilization. While eggs could be activated in calcium free hypertonic media but not be fertilized, fertilization occurred in isotouic media rich in calcium ions. The rate of fertilization was dependent on calcium concentration, and eggs were not fertilized in solutions with a calcium ion concentration of less than 0·57 mmol kg⁻¹. Calcium ions could be replaced to some extent by magnesium ions, but the former were the more effective in fertilization. Since calcium ion concentration of ovarian fluid of A. alcicornis was 0·41 mmol kg⁻¹, it was inferred that low calcium concentration in the ovarian fluid was the cause of the failure of A. alcicornis eggs to fertilize internally.     

Influence of a Small Number of Mature T Cells in Donor Bone Marrow Inocula on Reconstitution of Lymphoid Tissues and Negative Selection of a T Cell Repertoire in the Recipient

Allo-chimerism and clonal elimination of self antigen (Ag) (Ia + Mls-1^a) reactive Vβ6+ T cells were analyzed and compared between allogeneic bone marrow (BM) chimeras reconstituted with BM cells which had been treated with anti-Thy-1 monoclonal antibody (mAb) plus complement (C) (T^– chimeras) and BM chimeras which had been reconstituted with BM cells pretreated with anti-Thy-1 mAb alone (T⁺ chimeras). When lethally irradiated AKR (Mls-1^a) mice were reconstituted with BM cells from B10 or B10 H-2 congenic mice, both T⁺ and T^– chimeras were entirely free of signs of graft-versus-host reaction (GVHR). However, complete replacement of the AKR lymphoid tissues by donor BM cells was accomplished at an early stage in T⁺ chimeras but not in T^– chimeras. On the other hand, clonal elimination of Vβ6⁺ T cells reactive to the recipient Ag (Mls-1^a) was abolished in T⁺ chimeras but successfully induced in T^– chimeras. The Vβ6⁺ T cells not eliminated in T⁺ chimeras showed depressed responses against Mls-1^a antigens. The findings herein demonstrate that T cells which contaminate a BM inoculum survive in recipient mice after treatment with anti-Thy-1 mAb without C in vitro followed by BMT. The surviving T cells have been estimated to represent fewer than 0.5% of the BM cells inoculated. These cells appear to accelerate the full replacement of recipient lymphoid tissues by donor cells. Furthermore, the T cells which survive in the marrow inoculum influence eventually the development of a tolerant state in the T cell repertoire of the donor.     

Immunocytochemical localization of 5α-reductase type 1 in the prostate of normal and castrated rats

We immunohistochemically studied the localization of 5-reductase type 1 in combination with androgen receptor (AR) expression in individual lobes of the prostates of intact and castrated rats. In the normal rat prostate, 5-reductase was localized in the cytoplasm of most epithelial cells in the ventral, dorsal, and lateral type 1 (L1) lobes. Epithelial cells of lateral type 2 (L2) lobes were negative for 5-reductase. AR was present in the nuclei of all epithelial and stromal cells throughout the prostate. The number of 5-reductase-immunoreactive cells rapidly decreased in the ventral and L1 lobes after castration, whereas many positive cells remained in the dorsal lobe even at 4 weeks after castration. AR immunostaining was lost in the ventral, dorsal, and L1 lobes at 1 week after castration, but remained in the L2 lobe of 4-week-castrated rats. Electron microscopic immunocytochemistry showed that 5-reductase was exclusively localized in the rough endoplasmic reticulum membranes and that there were no distinct structural differences between the positively and negatively stained epithelial cells. These findings suggested that the expression of 5-reductase type 1 in the epithelial cell is heterogeneous within and among the individual lobes of the rat prostate, and does not correspond to AR expression.     

Photo-induced activation of cytochrome P450/reductase fusion enzyme coupled with spinach chloroplasts

Summary Photoactivation of cytochrome P450 monooxygenase was studied using a combination of spinach chloroplasts and yeast microsomes containing rat P4501A1/yeast reductase fusion enzyme. Under illumination, in the reaction mixture, NADP was reduced, transferring electrons to the P450/reductase fusion enzyme to convert 7-ethoxycoumarin to 7-hydroxycoumarin.     

The role of virulence antigens (VW) in the protection of mice againstYersinia pestis infection

Subcutaneous infection withYersinia enterocolitica harboring plasmid responsible for Ca²⁺ dependence at 37°C induced cell-mediated protective immunity against a lethal challenge withYersinia pestis; the isogenic derivative strain cured from this plasmid subverted the immunity in mice. This is the first identification of the antigen(s) responsible for the induction of cell-mediated protective immunity against the facultatively intracellular bacteria.     

Ejaculations induced by p-chloroamphetamine (PCA) in rats, hamsters and mice.

The ejaculatory response induced by p-chloroamphetamine (PCA) in male rats, hamsters and mice was observed during 2 hours after the injection. The animals were treated intraperitoneally with PCA at doses ranging from 0.78125 to 160 mg/kg. The ED50 (effective dose in 50% of animals) values of PCA for the initiation of ejaculation in rats and hamsters were 1.3397 (1.0732-1.6725) and 0.1105 (0.0802-0.1522) mg/kg, respectively. On the other hand, no ejaculation was observed in any mice at any doses examined. So we concluded that there are species differences in the ejaculatory response, induced by PCA, among rats, hamsters and mice.