Phylogeny of the insulin-like growth factors (IGFS) and receptors: A molecular approach

The IGFs (IGF-I and IGF-II) are essential for normal mammalian growth and development. Their actions are mediated primarily by their interactions with the type I IGF receptor (IGF-I receptor), a transmembrane tyrosine kinase. The ligands and the IGF-I receptor are structurally related to insulin and to the insulin receptor, respectively. Analysis of evolutionary conservation has often provided insights into essential regions of molecules such as hormones and their receptors. The genes for insulin and IGFs have been partially characterized in a number of vertebrate species extending evolutionarily from humans as far back as fish. The sequences of the exons encoding the mature insulin and IGF peptides are highly conserved among vertebrate species, and IGF-I-Iike molecules are found in species whose origins extend back as much as 550 million years. The insulin receptor is also highly conserved in vertebrate species, and an insulinreceptor-like molecule has been characterized in Drosophila. In contrast, IGF-I receptors have only been characterized in mammalian species and partially studied in Xenopus, in which the tyrosine kinase domain is highly conserved. Studies are presently being undertaken to analyze in more detail the regulation of the genes encoding this important family of growth factors and the structure/function relationships in the gene products themselves. © 1993 Wiley-Liss, Inc.     

Strong and regulated promoters in the cyanobacterium Anabaena PCC 7120

Abstract The strengths of several promoters were assessed in the cyanobacterium Anabaena PCC 7120 by fusing them to luxAB , encoding bacterial luciferase. Two promoters, P _tac and P _psbA , with sequences nearly identical to consensus Escherichia coli σ ⁷⁰ promoters, gave as high or higher expression than the strong Anabaena promoter, P _rbc . P _npt , the natural promoter driving expression of the kanamycin-resistance determinant from Tn5, was poorly expressed in Anabaena . The Lac repressor partially repressed expression from P _tac , permitting regulated expression in Anabaena after induction with isopropyl thiogalactoside to a level 4–5-fold higher than without inducer.     

The mosquito dihydrofolate reductase gene functions as a dominant selectable marker in transfected cells

An Aedes albopictus dihydrofolate reductase gene was used to construct two chimeric DNA vectors that functioned as dominant selectable markers in transfected, wild type mosquito cells. Stably transformed clones were recovered after 10–15 days in the presence of selective medium containing 1 μM methotrexate. The transformed clones contained an estimated 100–500 copies of transfected DNA per nucleus. Combined data from Southern blots and in situ hybridization to metaphase chromosomes indicated that transfected DNA was likely integrated into chromosomes both as repeated structures and as randomly integrated single copy molecules, with minimal rearrangement of coding sequences. Transfected DNA was stably maintained under selective conditions, but in some cases was lost when cells were maintained for prolonged periods in the absence of methotrexate. These observations provide a general framework for further development of stable gene transfer systems for mosquito cells in culture.     

Estimating tree-based dynamic treatment regimes using observational data with restricted treatment sequences

A dynamic treatment regime (DTR) is a sequence of decision rules that provide guidance on how to treat individuals based on their static and time-varying status. Existing observational data are often used to generate hypotheses about effective DTRs. A common challenge with observational data, however, is the need for analysts to consider “restrictions” on the treatment sequences. Such restrictions may be necessary for settings where (1) one or more treatment sequences that were offered to individuals when the data were collected are no longer considered viable in practice, (2) specific treatment sequences are no longer available, or (3) the scientific focus of the analysis concerns a specific type of treatment sequences (eg, “stepped-up” treatments). To address this challenge, we propose a restricted tree–based reinforcement learning (RT-RL) method that searches for an interpretable DTR with the maximum expected outcome, given a (set of) user-specified restriction(s), which specifies treatment options (at each stage) that ought not to be considered as part of the estimated tree-based DTR. In simulations, we evaluate the performance of RT-RL versus the standard approach of ignoring the partial data for individuals not following the (set of) restriction(s). The method is illustrated using an observational data set to estimate a two-stage stepped-up DTR for guiding the level of care placement for adolescents with substance use disorder.     

Murine FGF-4 gene expression is spatially restricted within embryonic skeletal muscle and other tissues

Fibroblast growth factors are believed to play many distinct roles in vertebrate development, owing to their ability to stimulate cell growth, prevent cell death, determine cell fate, and inhibit terminal differentiation in a variety of in vitro culture systems. We have used in situ hybridization to localize fibroblast growth factor-4 (FGF-4, also termed HST and K-FGF) gene expression in 7.5 to 16.5 day gestation mouse embryos. Seven discrete sites of gene expression were detected: (1) primitive streak (E7.5–8.5); (2) paraxial presomitic mesoderm in the trunk (E7.5–11.5); (3) primitive neuroectoderm (E8.0–8.5); (4) pharyngeal pouch endoderm (E8.5–9.5); (5) branchial arch ectoderm (E8.5–9.5); (6) limb apical ectoderm (E10.5–12.5), and (7) skeletal myoblast groups (E9.5–13.5). FGF-4 gene expression is spatially restricted within many of these sites. The profile of FGF-4 gene expression among skeletal muscle groups is overlapping, but distinct, from that of FGF-5, thereby revealing myoblast heterogeneity at the molecular level and suggesting distinct roles for multiple FGFs in muscle development.     

Structural Optimization,Fungicidal Activities Evaluation,DFT Study and Structure-Activity Relationship of Dopamine Derivatives with Benzothiazole Fragment from Polyrhachis dives

In our previous work, two dopamine derivatives with benzothiazole fragment were isolated and identified from Polyrhachis dives (P. dives). Based on their characteristic structure, we used them as lead compound to carry out structural optimization and subsequent fungicidal evaluation. Here 20 dopamine derivatives with benzothiazole fragment were designed and synthesized by a facile method, and their structures were characterized by ¹H-NMR, ¹³CNMR and HMRS. In bioassays, most of the title compounds possess potential fungicidal activities against Altenaia alternala (A. alternala) and Botrytis cinerea (B. cinerea). Especially, (E)-N-(2-(benzo[d]thiazol-6-yl)ethyl)-3-(p-tolyl)acrylamide and (E)-N-(2-(benzo[d]thiazol-6-yl)ethyl)-3-(4-(trifluoromethyl)phenyl)acrylamide displayed 29.3 mg/L and 10.7 mg/L EC₅₀ value against A. alternala, respectively, which possessed equivalent fungicidal activities level to hymexazol.     

Chemical and Electron Microscopic Studies of Cattle (Bos taurus) With Four Types of Phenotypic Pigmentation

The biological behavior of the pigmentary phenotypes of four breeds of cattle has been analysed: the black pigmentation of Holstein Friesian; the red pigmentation of Limousin; the dilution in Charolais; and the postnatal disappearance of red pigmentation in Chianina. The analytic techniques included the characterization of melanins by high-performance liquid chromatography, the examination of follicular melanocytes by light microscopy, and the examination of melanosomes by electron microscopy. The black phenotype was very strongly eumelanogenic. The red phenotype in Limousin is polymorphic: individual follicular melanocytes contain both mature eumelanosomes and pheomelanosomes. Charolais and Chianina cattle exhibited a dramatic reduction in melanogenic activity, which was characterized by the almost exclusive presence of prephaoemelanosomes in Charolais and of immature premelanosomes in Chianina. In the dilute Charolais phenotype, the density of distribution of follicular melanocytes also seemed to be reduced. The genes that are responsible for these four phenotypes seem to act on the maturation, differentiation, and density of distribution of the melanosomes.     

Characterization of a 46 kda insect chitinase from transgenic tobacco

A 46 kDa Manduca sexta (tobacco hornworm) chitinase was isolated from leaves of transgenic tobacco plants containing a recombinant insect chitinase cDNA, characterized, and tested for insecticidal activity. The enzyme was purified by ammonium sulfate fractionation, Q-Sepharose anion-exchange chromatography and mono-S cation-exchange chromatography. Although the gene for the chitinase encoded the 85 kDa full-length chitinase as previously reported by Kramer et al. [Insect Biochem. Molec. Biol. 23, 691–701 (1993)], the enzyme is produced in tobacco as a 46 kDa protein that is approximately four-fold less active than the 85 kDa chitinase. The N-terminal amino acid sequence of the 46 kDa chitinase is identical to that of the 85 kDa chitinase. The former enzyme is not glycosylated, whereas the latter contains approximately 25% carbohydrate. The pH and temperature optima of the 46 kDa chitinaseare similar to those of the 85 kDa chitinase. The former enzyme is more basic than the latter. The 46 kDa chitinase likely consists of the N-terminal catalytic domain of the 85 kDa chitinase and lacks the C-terminal domain that contains several potential sites for glycosylation. The 46 kDa chitinase is expressed in a number of plant organs, including leaves, flowers, stems and roots. Enzyme levels are higher in leaves and flowers than in stems and roots, and leaves from the middle portion of the plant have more chitinase than leaves from the top and bottom portions. Little or no enzyme is secreted outside of the plant cells because it remains in the intracellular space, even though its transit sequence is processed. When fed at a 2% dietary level, the 46 kDa chitinase caused 100% larval mortality of the merchant grain beetle, Oryzaephilis mercator. The results of this study support the hypothesis that insect chitinase is a biopesticidal protein for insect pests feeding on insect chitinase gene-containing transgenic plants.     

RT－cDNA克隆－PCR法获取犬瘟热病毒融合蛋白基因（F）

纯化鸡胚成纤维细胞培养的犬瘟热病毒（ＣａｎｉｎｅＤｉｓｔｅｍｐｅｒＶｉｒｕｓ,ＣＤＶ）,获得病毒基因组ＲＮＡ后,反转录合成双链病毒Ｆ基因ｃＤＮＡ。将此双链ｃＤＮＡ平端插入ＰＵＣ１９质粒ＳａｍⅠ位点构建重组质粒,进行ｃＤＮＡ克隆。以重组克隆质粒为模板ＰＣＲ扩增,获得ＣＤＶ全长Ｆ基因。将此Ｆ基因插入表达载体ＰＢＶ２２０,在大肠杆菌中表达,通过对表达产物的最终鉴定,可确认所获片段为ＣＤＶ全长Ｆ基因．