Expression of human lactotransferrin receptors in phytohemagglutinin-stimulated human peripheral blood lymphocytes. Isolation of the receptors by antiligand-affinity chromatography

In the resting rate, the human peripheral blood lymphocytes did not show detectable surface and intracellular receptors for human lactotransferrin. However, both types of lactotransferrin receptors were expressed during stimulation of lymphocytes with phytohemagglutinin. The appearance of receptors was time-dependent and the number of receptors reached a plateau after at least two days of mitogen stimulation. These results suggest that the presence of surface receptors on mitogen-stimulated lymphocytes is not consecutive to a modification of subcellular distribution but to an induction of biosynthesis of the receptors. As measured by incorporation of [3H]thymidine into DNA, addition of human lactotransferrin in a serum-free medium increased the proliferative activity of phytohemagglutinin-stimulated lymphocytes. Optimal enhancement of [3H]thymidine incorporation was obtained by adding 30% iron-saturated lactotransferrin at a concentration of 0.17 microM. Therefore, the role of lactotransferrin in the response of lymphocytes to mitogen stimulation appears to be similar to that previously described for serotransferrin. The lactotransferrin receptor was visualized using 125I-labeled lactotransferrin on nitrocellulose paper after electroblotting of the Triton X-100 extract of the phytohemagglutinin-stimulated lymphocytes as two protein bands of 100 and 110 kDa molecular mass. Purification of the lactotransferrin receptor from the Triton-X-100-soluble extract of stimulated lymphocytes was performed by antiligand-affinity chromatography. The binding of lactotransferrin to the purified receptors was reversible and dependent on concentration and pH.     

A differential scanning calorimetric study of the binding of sulfate ion and of Cibacron blue F3GA to yeast phosphoglycerate kinase

In continuation of earlier work [Hu, C. Q., & Sturtevant, J.M. (1987) Biochemistry 26, 178-182], differential scanning calorimetry has been employed in a study of the effects on the thermal denaturation of yeast phosphoglycerate kinase of two inhibitors of the enzyme, sulfate ion and the dye Cibracron blue F3GA. Sulfate ion, as is usual with ligands that dissociate during unfolding of the host protein, raises t1/2, the temperature of half-completion of the denaturation, has only a modest effect, stemming from the enthalpy of dissociation of the ligand, on the enthalpy of denaturation, and has little or no effect on the heat capacity change resulting from denaturation. In sharp contrast, Cibacron blue F3GA lowers t1/2 and drastically decreases both the enthalpy and heat capacity changes due to denaturation. The DSC results with sulfate ion are consistent with previous kinetic data [Scopes, R. K. (1978) Eur. J. Biochem. 91, 119-129; Khamis, M. H., & Larsson-Raznikiewicz, M. (1981) Biochim. Biophys. Acta 657, 190-194], which indicate two binding sites for sulfate ion at one of which the ligand acts as a competitive inhibitor. The results with Cibacron blue F3GA indicate that the dye induces a major destabilizing structural change in the enzyme in addition to rendering it enzymically inactive.     

Cooperative binding of Agrobacterium tumefaciens VirE2 protein to single-stranded DNA.

The VirE2 protein of Agrobacterium tumefaciens Ti plasmid pTiA6 is a single-stranded-DNA-binding protein. Density gradient centrifugation studies showed that it exists as a tetramer in solution. Monomeric VirE2 active in DNA binding could also be obtained by using a different protein isolation procedure. VirE2 was found to be thermolabile; brief incubation at 37 degrees C abolished its DNA-binding activity. It was insensitive to the sulfhydryl-specific reagent N-ethylmaleimide. Removal of the carboxy-terminal 37 residues of the 533-residue VirE2 polypeptide led to complete loss of DNA-binding activity; however, chimeric fusion proteins containing up to 125 residues of the VirE2 C terminus were inactive in DNA binding. In nuclease protection studies, VirE2 protected single-stranded DNA against degradation by DNase I. Analysis of the DNA-VirE2 complex by electron microscopy demonstrated that VirE2 coats a single-stranded DNA molecule and that the binding of VirE2 to its substrate is cooperative.     

A toxic substance produced by Nocardia otitidiscaviarum isolated from cutaneous nocardiosis

During our studies on toxic substances from clinically isolated Nocarida, a new isolate identified as Nocardia otitidiscaviarum from cutaneous nocardiosis was found to produce a toxic substance called HS-6 that had strong in vitro as well as in vivo toxicity. The mouse intraperitoneal LD₅₀ value was 1.25 mg/kg and the ED₅₀ value for L1210 cultured cells was 0.3 ng/ml. The structure of HS-6 was determined and found to belong to the 16-membered macrocyclic group with a molecular formula of C₄₃H₆₈O₁₂. HS-6 also showed activity against pathogenic fungi such as Cryptococcus neoformans.     

抗阿特拉津转基因大豆植株后代的遗传分析

本试验用阿特拉津溶液涂抹、荧光诱导动力学检测、分子杂交等方法对抗阿特拉津转基因大豆植株的后代进行了鉴定,在第二代及第三代中检测到了抗性基因的存在,表明从龙葵中得到的此抗阿特拉津 psbA 基因不仅能导人大豆叶绿体基因组中获得表达,而且可以遗传到后代。     

Plant regeneration from mesophyll protoplasts of Diplotaxis muralis,a wild crucifer

Mesophyll protoplasts from leaves of aseptically grown shoot tips of Diplotaxis muralis were isolated (6.2–7.1×10⁵ protoplasts/g fresh weight of tissue) using one step enzyme digestion. The protoplasts (71% viability) underwent divisions (4.2+0.1%) on plating in M8PS₂ medium and ultimately formed calli with 0.45+0.03% plating efficiency. Plant regeneration could be achieved both through embryogenesis and organogenesis. The efficiency of plant regeneration through organogenesis was 9 times higher than embryogenesis. Forty eight out of 52 plants regenerated so far from 3 independent experiments were normal with respect to fertility and meiotic chromosomal behavior.Abbreviations BAP 6-benzylaminopurine - GA₃ Gibberellic acid - A Kao and Michayluk, 1981 - KM Kao and Michayluk, 1975 - MK₃ Modified K₃ - M8P Modified 8P - MS Murashige and Skoog, 1962 - NAA 1-naphthalene acetic acid - PE Plating efficiency     

The effect of carbamylcholine on CFU-s differentiation

The proportion of spleen colony-forming units (CFU-s) killed by hydroxyurea was greatly increased after bone marrow cells (BMCs) from LACA mice were exposed to carbamylcholine (Cach; 1 X 10(-13) to 1 X 10(-9) in vitro and there was a marked change in the proportion of spleen colony types. Following treatment with Cach, granulocytic and mixed erythroid-type colonies increased from 20 to 26.3% and 16.1 to 29.6% in 9-day colonies and from 8.3 to 28.2% and 21.7 to 39.4% in 13-day colonies, respectively. Single cell suspensions of spleen colonies were made for granulocyte-macrophage progenitor (CFU-gm) and late erythroid progenitor (CFU-e) assays. The number of CFU-gm from Cach-treated BMC was about twice that from control BMC for both day 9 and day 13 groups; the number of CFU-e decreased relatively. The results suggest that cholinergic receptors on CFU-s may increase the tendency to differentiate into the granulocytic/monocytic line.