A cis-acting element and a trans-acting factor involved in the wound-induced expression of a horseradish peroxidase gene

The mechanisms that control the wound-induced expression of the prxC2 gene for horseradish peroxidase (HRP) have been investigated. Analysis of the regulatory properties of 5′-deleted promoters showed that a positive element involved in the response to wounding was located between −307 and −99 bp from the site of initiation of translation. In in vitro binding assays of tobacco nuclear proteins and DNA fragments of prxC2 promoter, the binding site was the Box 1 from −296 to −283 containing the CACGTG motif. To identify the functional role of Box 1, the prxC2 promoter that has been digested from the 5′ end to −289 with a disrupted Box 1 was fused to a reporter gene for β-glucuronidase (GUS). No induction of GUS activity was observed in transgenic tobacco plants with the prxC2(−289)/GUS construct. These data indicated that the expression of prxC2 in response to wounding required the Box 1 sequence from −296 to −283. Furthermore, a tobacco cDNA expression library was screened and a cDNA clone for a protein, designated TFHP-1, that bound specifically to the Box 1 sequence was identified. The putative TFHP-1 protein contains a basic region and leucine zipper (bZip) motif and a helix—loop—helix (HLH) motif. The mRNA for TFHP-1 was abundant in roots and stems, and it was not induced by wounding in leaves. In tobacco protoplasts, antisense TFHP-1 suppressed the expression of prxC2 (−529)/GUS.     

Inhibition of the catalase reaction of Photosystem II by anions

A new binding site for anions which inhibit the water oxidizing complex (WOC) of Photosystem II in spinach has been identified. Anions which bind to this site inhibit the flash-induced S₂/S₀ catalase reaction (2H₂O₂2H₂O+O₂) of the WOC by displacing hydrogen peroxide. Using a mass spectrometer and gas permeable membrane to detect the ³²O₂ product, the yield and lifetime of the active state of the flash-induced catalase (to be referred to simply as flash-catalase) reaction were measured after forming the S₂ or S₀-states by a short flash. The increase in flash-catalase activity with H₂O₂ concentration exhibits a K_m=10–20 mM, and originates from an increase in the lifetime by 20-fold of the active state. The increased lifetime in the presence of peroxide is ascribed to formation of the long-lived S₀-state at the expense of the unstable S₂-state. The anion inhibition site differs from the chloride site involved in stimulating the photolytic water oxidation reaction (2H₂OO₂+4e^-+4H⁺). Whereas water oxidation requires Cl^- and is inhibited with increasing effectiveness by F^-CN^-N₃ ^-, the flash-catalase reaction is weakly inhibited by Cl^-, and with increasing effectiveness by F^-CN^-, N₃ ^-. Unlike water oxidation, chloride is unable to suppress or reverse inhibition of the flash-catalase reaction caused by these anions. The inhibitor effectiveness correlates with the pK_a of the conjugate acid, suggesting that the protonated species may be the active inhibitor. The reduced activity arises from a shortening of the lifetime of the flash-induced catalase active state by 3–10 fold owing to stronger anion binding in the flash-induced states, S₂ and S₀, than in the dark S-states, S₁ and S_-1. To account for the paradoxical result that higher anion concentrations are required to inhibit at lower H₂O₂ concentrations, where S₂ forms initially after the flash, than at higher H₂O₂ concentrations, where S₀ forms initially after the flash, stronger anion binding to the S₀-state than to the S₂-state is proposed. A kinetic model is given which accounts for these equilibria with anions and H₂O₂. The rate constant for the formation/release of O₂ by reduction of S₂ in the WOC is <0.4 s^-1.Abbreviations ADRY acceleration of the deactivation reactions of the water splitting enzyme system Y - BTP bis [tris(hydroxymethyl)methylamino]-propane - CCCP carbonylcyanide m-chlorophenylhyrazone - DCBQ 2,5-dichlorobenzoquinone - DMBQ 2,3-dimethylbenzoquinone - WOC water oxidizing complex     

Distribution of various nickel compounds in rat organs after oral administration

In this study, eight kinds of nickel (Ni) compounds were orally administered to Wistar male rats and the distribution of each compound was investigated 24 h after the administration. The Ni compounds used in this experiment were nickel metal [Ni−M], nickel oxide (green) [NiO(G)], nickel oxide (black) [NiO(B)], nickel subsulfide [Ni₃S₂], nickel sulfide [NiS], nickel sulfate [NiSO₄], nickel chloride [NiCl₂], and nickel nitrate [Ni(NO₃)₂]. The solubilities of the nickel compounds in saline solution were in the following order; [Ni(NO₃)₂>NiCl₂>NiSO₄]≫[NiS>Ni₃S₂]>[NiO(B)>Ni−M>NiO(G)]. The Ni level in the visceral organs was higher in the rats given soluble Ni compounds; Ni(NO₃)₂, NiCl₂, NiSO₄, than that in the rats receiving other compounds. In the rats to which soluble Ni compounds were administered, 80–90% of the recovered Ni amounts in the examined organs was detected in the kidneys. On the other hand, the Ni concentration in organs administered scarcely soluble Ni compounds; NiO(B), NiO(G), and Ni−M were very low. The estimated absorbed fraction of each Ni compounds was increased with the increase of the solubility. These results suggest that the kinetic behavior of Ni compounds administered orally is closely related with the solubility of Ni compounds, and that the solubility of Ni compounds is one of the important factors for determining the health effect of Ni compounds.     

外消旋薄荷醇对映选择性酶促酯化中酸酐与羧酸的比较研究

利用脂肪酶在有机溶剂中催化对映选择性酯化反应对外消旋薄荷醇进行了有效的光学拆分。对分别使用酸酐和相应的游离羧酸作酰基给体时的反应性能进行了比较。发现酸酐的反应性远高于对应的游离羧酸,但在酶的催化作用下酸酐易水解成为游离羧酸;在微水系统中使用过高浓度的酸酐会导致酶缺水而失活,同时会促进手性醇的非选择性酯化,从而降低产物的光学纯度。然而,在连续流加丙酸酐的半批式反应系统中,所有这些缺点均可有效地克服。与使用游离丙酸的批式反应系统相比,ｄｌ-薄荷醇的反应时间缩短了一半,酶的稳定性大幅度提高,而产物ｌ薄荷醇酯的光学纯度不相上下（＞９８％ｅ）。     

Genetic variations within and between troops of the crab-eating macaque (Macaca fascicularis) on Sumatra,Java, Bali,Lombok and Sumbawa,Indonesia

Genetic variations within and between troops of the Indonesian crab-eating macaque (Macaca fascicularis) were studied. A total of 456 blood samples were collected from 29 free-ranging troops in 19 different localities on Sumatra, Java, Bali, Lombok and Sumbawa. Blood protein polymorphisms were examined electrophoretically. Mean genetic variability within troops was estimated to be P_poly=12.22% and =3.84%. Troops inhabiting small islands showed lower variability. Genetic differences were more marked between troops on different islands than between troops on the same island. Additionally, clinal variations of allelic frequencies at some loci were detected. The genetic features and socio-ecological and evolutionary implications are discussed.     

A case of pituitary adenoma producing both growth hormone (GH) and adrenocorticotropic hormone (ACTH).

The authors report a very rare case of pituitary adenoma producing both GH and ACTH. A 29-year-old female was admitted with obesity, amenorrhea, acromegaly, hirsutism, excessive pigmentation, acne, and diabetes mellitus. Computed tomography revealed an intrasellar tumor 16 mm in height, with a destroyed sellar floor. The blood concentrations of GH, ACTH and cortisol were increased (GH: 92 ng/ml, ACTH: 94 pg/ml, cortisol: 18.3 micrograms/dl). No diurnal variation in the amount of cortisol was observed. The urinary 17-OHCS was suppressed by 8 mg but not by 2 mg of dexamethasone. A subtotal adenomectomy was then performed through the transsphenoidal approach, which led to a sufficient reduction of both blood GH and ACTH (cortisol). Histologically the tumor was an acidophilic pituitary adenoma. Immunoperoxidase staining showed diffuse GH and sporadic ACTH producing cells, but failed to show any cells producing both hormones. The electron micrograms of neoplastic cells showed the ultrastructural characteristics of respective GH and ACTH cells. Another increase in both GH and cortisol, which occurred 19 months after the operation, has been controlled by bromocriptine administration. This case may be the first reported case of a pituitary adenoma producing both GH and ACTH, not accompanied by prolactin (PRL) hypersecretion, which has been fully confirmed endocrinologically and histopathologically.     

Evidence for inserted sequences in the head region of nonmuscle myosin specific to the nervous system. Cloning of the cDNA encoding the myosin heavy chain-B isoform of vertebrate nonmuscle myosin.

The complete amino acid sequence of a vertebrate nonmuscle myosin heavy chain-B isoform (MHC-B, 1976 amino acids, 229 kDa) has been deduced by using cDNA clones from chicken brain libraries. The chicken nonmuscle MHC-B shows overall similarity in primary structure to other MHCs in the areas contributing to the ATP-binding site and actin-binding site. Similar to other nonsarcomeric MHC IIs, there is a short uncoiled tail sequence at the carboxyl terminus of the molecule. It is in the uncoiled tail sequence that the greatest number of differences in amino acids sequence between MHC-A and B were found, which allowed generation of isoform-specific antibodies. These antibodies were used to determine the relative content of MHC-A and MHC-B in various tissues. During the cloning of the cDNA encoding chicken brain MHC-B, we found a 63-nucleotide insertion encoding 21 amino acids located in the head region of the MHC near to the actin-binding site and a 30 nucleotide insertion encoding 10 amino acids near to the ATP-binding site. Analysis using S-1 nuclease showed that both inserts are expressed in a tissue-dependent manner; mRNA containing the inserts is present in tissues of the nervous system, but is absent from other non-muscle cells, which contain the noninserted isoform of MHC-B. Similar inserts were found in corresponding positions in human cerebellar mRNA. Antibodies raised against a peptide synthesized based on the 21 amino acid insert found in chickens recognize a MHC isoform in the same tissues that are enriched for the mRNA. These insertions appear to be a mechanism for generating additional MHC-B isoforms specific to the nervous system.     

Pseudomeningocele: an unusual complication of craniofacial surgery.

A previously unreported complication of pseudomeningocele following monoblock frontofacial advancement is described. The defect was repaired by means of an extracranial approach with a satisfactory outcome.     

Isolation of a full-length cDNA encoding mouse aromatase P450

A full-length cDNA clone for aromatase P450 has been isolated from a pregnant mouse ovarian cDNA library. The insert of this clone (2394 bp) contains a 1509-bp open reading frame encoding 503 amino acid residues together with a 46-bp 5'-untranslated stretch and an 839-bp 3'-untranslated region to which a poly(A) tract is attached. Northern blot analysis of ovarian RNA from pregnant mice reveals a major mRNA band of 2.5 kb with a minor band of 2.1 kb. Comparison of mouse aromatase P450 with that of rat, human, and chicken shows 91, 81, and 69% identity in the nucleotide sequence and 92, 79, and 69% identity in the deduced amino acid sequence, respectively. The membrane-spanning domain of mouse aromatase P450 is estimated to be an extremely hydrophobic segment located within the N-terminal region of the molecule. Furthermore, a highly conserved heme-binding domain is noticed.     

Adenosine deaminase deficiency due to heterozygous abnormality consisting of a deletion of exon 7 and the absence of enzyme mRNA

An adenosine deaminase (ADA;EC 3.5.4.4)-deficient B lymphoblastoid cell line BADO5 derived from a Japanese patient with severe combined immunodeficiency disease and two B lymphoblastoid cell lines, BAMO5 from his mother and BAFO5 from his father, were characterized. To identify mutations affecting ADA activity, we prepared cDNAs to ADA mRNAs of the BADO5 cell line for nucleotide sequencing. Sequence analysis of one of the BADO5 ADA cDNA clones revealed deletion of exon 7, and one point mutation of base 629 from G to A that did not affect the amino acid sequence. All clones of the BADO5 cell line so far examined showed the absence of exon 7 by Southern blotting analysis. Ribonuclease protection assay with an RNA probe spanning from exon 5 to exon 11 showed that the BADO5 ADA mRNA had a deletion of exon 7, the BAMO5 mRNA had normal length, and the BAFO5 mRNA had two species with a deletion of exon 7 and with normal length. Consequently, the patient's ADA genes resulted from one allele of the BAMO5 ADA gene that did not produce a detectable mRNA, and the other allele of the BAFO5 ADA gene producing an aberrant mRNA without exon 7.