The c-SRC1 gene visualized by in situ hybridization on Xenopus laevis chromosomes

Fluorescent in situ hybridization followed by tyramide signal amplification was used to map the site of the c-SRC1 gene on XENOPUS LAEVIS chromosomes. Positive results were obtained with a cDNA probe of about 1 kb. The c-SRC1 gene is located in the subcentromeric region in the long arm of one of the acrocentric chromosomes of the G category (classified according to Graf and Kobel, 1991). The c-SRC1 gene and the XENOPUS major histocompatibility complex (MHC) 1b locus, which consists of 20 tandemly arranged gene copies, are situated on different chromosomes of the G category.     

Physiological functions of Pten in mouse tissues

PTEN is a tumor suppressor gene mutated in many human sporadic cancers and in hereditary cancer syndromes such as Cowden disease, Bannayan-Zonana syndrome and Lhermitte-Duclos disease. The major substrate of PTEN is PIP3, a second messenger molecule produced following PI3K activation induced by variety of stimuli. PIP3 activates the serine-threonine kinase PKB/Akt which is involved in anti-apoptosis, proliferation and oncogenesis. In mice, heterozygosity for a null mutation of Pten (Pten(+/-) mice) frequently leads to the development of a variety of cancers and autoimmune disease. Homozygosity for the null mutation (Pten (-/-) mice) results in early embryonic lethality, precluding the functional analysis of Pten in various organs. To investigate the physiological functions of Pten in viable mice, various tissue-specific Pten mutations have been generated using the Cre-loxP system. This review will summarize the phenotypes of conditional mutant mice lacking Pten function in specific tissues, and discuss how these phenotypes relate to the physiological roles of Pten in various organ systems.     

Salicylic acid inhibits indeterminate-type nodulation but not determinate-type nodulation

LCOs (lipochitin oligosaccharides, Nod factors) produced by the rhizobial symbiote of Vicia sativa subsp. nigra (vetch, an indeterminate-type nodulating plant) are mitogenic when carrying an 18:4 acyl chain but not when carrying an 18:1 acyl chain. This suggests that the 18:4 acyl chain specifically contributes to signaling in indeterminate-type nodulation. In a working hypothesis, we speculated that the 18:4 acyl chain is involved in oxylipin signaling comparable to, for example, signaling by derivatives of the 18:3 fatty acid linolenic acid (the octadecanoid pathway). Because salicylic acid (SA) is known to interfere with oxylipin signaling, we tested whether nodulation of vetch could be affected by addition of 10(-4) M SA. This concentration completely blocked nodulation of vetch by Rhizobium leguminosarum bv. viciae and inhibited the mitogenic effect of 18:4 LCOs but did not affect LCO-induced root-hair deformation. SA did not act systemically, and only biologically active SA derivatives were capable of inhibiting nodule formation. SA also inhibited R. leguminosarum bv. viciae association with vetch roots. In contrast, addition of SA to Lotus japonicus (a determinate-type nodulating plant responding to 18:1 LCOs) did not inhibit nodulation by Mesorhizobium loti. Other indeterminate-type nodulating plants showed the same inhibiting response toward SA, whereas SA did not inhibit the nodulation of other determinate-type nodulating plants. SA may be a useful tool for studying fundamental differences between signal transduction pathways of indeterminate- and determinate-type nodulating plants.     

Effect of methanol extract of Zanthoxylum piperitum leaves and of its compound,protocatechuic acid,on hepatic drug metabolizing enzymes and lipid peroxidation in rats

The effect of methanol extract and protocatechuic acid from the leaves of Zanthoxylum piperitum on lipid peroxidation and drug metabolizing enzymes were investigated in the liver of bromobenzene-treated rats. The methanol extract and protocatechuic acid reduced the level of lipid peroxide induced by bromobenzene. The methanol extract and protocatechuic acid reduced the activity of aniline hydroxylase that had been increased by bromobenzene, while did not affect the activities of aminopyrine N-demethylase and glutathione S-transferase. The methanol extract and compound effectively restored the activity of epoxide hydrolase which had been decreased by bromobenzene. These results may suggest that the methanol extract of Z. piperitum and protocatechuic acid prevented lipid peroxidation by reducing the activity of aniline hydroxylase, an epoxide-producing enzyme, and by enhancing the activity of epoxide hydrolase, an epoxide-removing enzyme, in rats that had been intoxicated with bromobenzene.     

An Inhibitor of Thrombin-Stimulated Blood Platelet Aggregation from the Salivary Glands of the Hard Tick Amblyomma variegatum (Acari: Ixodidae)

The tropical bont tick, Amblyomma variegatum can cause intense skin irritation and inflammation and bites that often develop into septic wounds or abscess in their host. Crude salivary gland extract (SGE) of partially engorged A. variegatum females as well as SGE protein fractions purified by three-step reverse phase HPLC procedure were tested for their anti-aggregatory effect on isolated human blood platelets stimulated with thrombin and compared with the effect of recombinant hirudin. At concentrations 10⁻³ and 5 × 10⁻³ μg protein/ml the following rank order of antiplatelet activity was detected: AV 16/3 (inhibitor purified from AV-III, third purification) > SGE > AV-II (fraction from first purification) > AV-III (fraction from first purification) > hirudin. The effect of all fractions tested was dose-dependent. For fraction AV 16/3, the inhibitory effect was 49 and 61% for 10⁻³ and 5 × 10⁻³ μg protein/ml, respectively. The results suggest that protein fractions from A. variegatum SGE possess an antithrombin effect on human blood platelets with hirudin-like activity. This revised version was published online in July 2006 with corrections to the Cover Date.     

A case of ovarian enterobiasis

A 36-year old Korean woman consulted a clinic for a regular gynecological examination, and a mass was noticed in her pelvis. She was referred to the Asan Medical Center, Seoul where transvaginal ultrasonography confirmed a pelvic mass exceeding 10 cm in diameter. She received total abdominal hysterectomy and bilateral salpingoophorectomy, and a borderline serous neoplasm with micropapillary features involving the left ovary and right ovarian serosa was histopathologically confirmed. In addition, a section of a nematode with numerous eggs was found in the parenchyma of the left ovary. The worm had degenerated but the eggs were well-preserved and were identified as those of Enterobius vermicularis. She is an incidentally recognized case of ovarian enterobiasis.     

Daunomycin binding to deoxypolynucleotides with alternating sequences: complete thermodynamic profiles of heterogeneous binding sites

Complete thermodynamic binding profiles for the interaction of the anticancer drug, daunomycin with natural DNA and synthetic deoxypolynucleotides were described. Fluorescence titration method was used to estimate the equilibrium binding constants. Binding isotherms were found to be surprisingly complex in some cases, presumably because there were heterogeneous sites even in simple deoxypolynucleotides of repeating sequence. Some polynucleotides consisting of alternating sequence contain at least two different binding sites for daunomycin. The binding affinity of the primary binding sites of alternating and non-alternating sequences was found to differ by two orders of magnitude. An isothermal microtitration calorimeter was used to directly measure the binding enthalpy at 25 degrees C with a high sensitivity. The binding enthalpy of poly[d(A-T)] was found to be -5.5 Kcal/mol, which was much lower than any other polynucleotides, while the binding constant of the high affinity sites, was similar. In this report, the complete thermodynamic profiles of daunomycin binding to deoxypolynucleotides were reliably shown for the first time.     

Enhancement of lysophosphatidic acid-induced ERK phosphorylation by phospholipase D1 via the formation of phosphatidic acid

We made stable cell lines overexpressing PLD1 (GP-PLD1) from GP+envAm12 cell, a derivative of NIH 3T3 cell. PLD1 activity and extracellular signal-regulated kinase (ERK) phosphorylation were enhanced in GP-PLD1 cells by the treatment of lysophosphatidic acid (LPA). In contrast, these LPA-induced effects were attenuated with the pretreatment of pertussis toxin (PTX) or protein kinase C (PKC) inhibitor. Moreover, accumulation of phosphatidic acid (PA), a product of PLD action, potentiated the LPA-induced ERK activation in GP-PLD1 cells while blocking of PA production with the treatment of 1-butanol attenuated LPA-induced ERK phosphorylation. From these results, we suggest that LPA activate PLD1 through pertussis toxin-sensitive G protein and PKC-dependent pathways, then PA produced from PLD1 activation facilitate ERK phosphorylation.     

Investigation of allosteric linkages in the regulation of tryptophan synthase: the roles of salt bridges and monovalent cations probed by site-directed mutation, optical spectroscopy, and kinetics

The tryptophan synthase bienzyme complex is the most extensively documented example of substrate channeling in which the oligomeric unit has been described at near atomic resolution. Transfer of the common metabolite, indole, between the alpha- and the beta-sites occurs by diffusion along a 25-A-long interconnecting tunnel within each alphabeta-dimeric unit of the alpha(2)beta(2) oligomer. The control of metabolite transfer involves allosteric interactions that trigger the switching of alphabeta-dimeric units between open and closed conformations and between catalytic states of low and high activity. This allosteric signaling is triggered by covalent transformations at the beta-site and ligand binding to the alpha-site. The signals are transmitted between sites via a scaffolding of structural elements that includes a monovalent cation (MVC) binding site and salt bridging interactions of betaLys 167 with betaAsp 305 or alphaAsp 56. Through the combined strategies of site-directed mutations of these amino acid residues and cation substitutions at the MVC site, this work examines the interrelationship of the MVC site and the alternative salt bridges formed between Lys beta167 with Asp beta305 or Asp alpha56 to the regulation of channeling. These experiments show that both the binding of a MVC and the formation of the Lys beta167-Asp alpha56 salt bridge are important to the transmission of allosteric signals between the sites, whereas, the salt bridge between betaK167 and betaD305 appears to be only of minor significance to catalysis and allosteric regulation. The mechanistic implications of these findings both for substrate channeling and for catalysis are discussed.