Comparative studies on the adhesiveness of granulocytes of guinea pig and man

Summary The high affinity of granulocytes of guinea pig and man to glass surfaces is modified by serum. Native serum contains both an adherence-promoting activity, which is related to complement, and components which reduce the adhesiveness of granulocytes. These components are stable at 56°C for 30 min and are tightly bound to the glass surface. -Lipoproteins are candidates for this adherence reducing ability of serum. Adherence promotion by native serum is mediated by coating the glass surface with C3b/C3bi. Human granulocytes from the peripheral blood adhered to glass surfaces coated by native human or guinea pig serum with C3b/C3bi to almost the same extent as in the presence of native serum, but on guinea pig granulocytes elicited in the peritoneal cavity, a cell surface metalloproteinase degraded the C3b/C3bi, thus reducing the adhesiveness of these cells. This proteinase was inhibited by MgEDTA, DTT, and 1,10-phenanthroline, whereby the high adhesiveness of granulocytes was restored to C3b/C3bi-coated glass.Abbreviations BA benzamidine hydrochloride - BTS Bacillus thuringiensis subtoxicus - DTT dithiothreitol - EAC -amino-caproic acid - gp guinea pig - LDL low density lipoproteins - SEM scanning electron microscopy     

The duplication in Charcot-Marie-Tooth disease type 1a spans at least 1100 kb on chromosome 17p11.2

Summary Recently, it has been shown that Charcot-Marie-Tooth disease type 1a (CMT1a) is linked with a duplication of a DNA segment that is detected by probe VAW409R3, and that is located on chromosome 17p11.2. Here, we show that this duplication also contains VAW412R3a, but not A10-41 and EW503. Accounting for the duplication in recombination analysis, we found recombinants between CMT1a and EW301 and EW502, but not with A10-41, VAW409R3, and VAW412R3. Using pulsed-field gel electrophoresis analysis, we estimated the minimal size of the duplicated region in CMT1a patients to be 1100 kb.     

Scavenging of Hydrogen Peroxide in the Endosperm of Ricinus communis by Ascorbate Peroxidase

The fat-storing endosperm of Ricinus communis L. was found tocontain an ascorbate peroxidase (EC 1.11.1.11 [EC] ), which is nearlyas active as catalase (EC 1.11.1.6 [EC] ) in degradation of hydrogenperoxide (H₂O₂) at its physiological concentrations. This ascorbateperoxidase probably functions together with monodehydroascorbatereductase (EC 1.6.5.4 [EC] ) or dehydroascorbate reductase (EC 1.8.5.1 [EC] )and glutathione reductase (EC 1.6.4.2 [EC] ) to remove the H₂O₂ producedduring the transformation of fat to carbohydrate in the glyoxysomes.The activities of these enzymes as well as the content of ascorbateand glutathione increase parallel to the activities of glyoxysomalmarker enzymes during the course of germination. Inhibitionof catalase by aminotriazole results in increases of the ascorbateperoxidase activity and of the glutathione content. All fourenzymes are predominantly localized in the cytosol of the Ricinusendosperm with low activities found in the plastids and themitochondria. The results suggest, that the ascorbate-dependentH₂O₂ scavenging pathway, which has been shown to be responsiblefor the reduction of photosynthetically derived H₂O₂ in thechloroplasts, operates also in the Ricinus endosperm. (Received June 5, 1990; Accepted July 31, 1990)     

New methods for estimating the numbers of synonymous and nonsynonymous substitutions

New methods for estimating the numbers of synonymous and nonsynonymous substitutions per site were developed. The methods are unweighted pathway methods based on Kimura's two-parameter model. Computer simulations were conducted to evaluate the accuracies of the new methods, Nei and Gojobori's (NG) method, Miyata and Yasunaga's (MY) method, Li, Wu, and Luo's (LWL) method, and Pamilo, Bianchi, and Li's (PBL) method. The following results were obtained: (1) The NG, MY, and LWL methods give overestimates of the number of synonymous substitutions and underestimates of the number of nonsynonymous substitutions. The major cause for the biased estimation is that these three methods underestimate the number of synonymous sites and overestimate the number of nonsynonymous sites. (2) The PBL method gives better estimates of the numbers of synonymous and nonsynonymous substitutions than those obtained by the NG, MY, and LWL methods. (3) The new methods also give better estimates of the numbers of synonymous and nonsynonymous substitutions than those obtained by the NG, MY, and LWL methods. In addition, estimates of the numbers of synonymous and nonsynonymous sites obtained by the new methods are reasonably accurate. (4) In some cases, the new methods and the PBL method give biased estimates of substitution numbers. However, from the number of nucleotide substitutions at the third position of codons, we can examine whether estimates obtained by the new methods are good or not, whereas we cannot make an examination of estimates obtained by the PBL method. (5) When there are strong transition/transversion and nucleotide-frequency biases like mitochondrial genes, all of the above methods give biased estimates of substitution numbers. In such cases, Kondo et al.'s method is recommended to be used for estimating the number of synonymous substitutions, although their method cannot estimate the number of nonsynonymous substitutions and is time-consuming. These results, particularly result (1), call for reexaminations of some genes. This is because evolutionary pictures of genes have often been discussed on the basis of results obtained by the NG, MY, and LWL methods, which are favorable for the neutral theory of molecular evolution.     

Effects of cadmium,copper, and zinc on growth and thiol content of aquatic hyphomycetes

The effects of cadmium, copper, and zinc on the growth of ten strains ofaquatic hyphomycetes were investigated. On a solid medium, Cd and Cu reducedradial growth of most strains by 50% at concentrations between150–400 µM; in a liquid medium, the strains were more sensitive.The inhibitory effects of zinc were much less severe. Two isolates(Articulospora tetracladia and Tetracladium marchalianum) from a copper-minestream were more resistant against copper than conspecific strains from anon-polluted stream. Heliscus lugdunensis and Varicosporium elodeaeresponded to Cd exposure, but not to Cu or Zn exposure, by increasedsynthesis of SH-containing compounds. Glutathione levels showed a unimodalresponse to increasing Cd and Zn exposure. With copper, glutathionedecreased at intermediate levels of contamination. In the presence of Cd, H. lugdunensis synthesized several unknown sulfur-rich substances that wereabsent or produced at reduced rates in control cultures.     

Short-term progressive early diagenesis in density bands of recent corals:Porites Colonies,Mauritius Island,Indian Ocean

Summary The growth history of some recentPorites colonies of Mauritius Island (Indian Ocean) was dated by sclerochronological methods. Couples of high-density and low-density bands represent the annual growth rate of the corals and allow the growth pattern of every year in the corallum to be counted. The growth and structure of the skeletons ofPorites solida andPorites lutea were investigated. Older parts of the aragonitic skeleton in these 10 to 20 year old corals show various secondary microstructures resulting from alterations and thickenings of the elements of the skeleton. The primary needle-like aragonite crystals are absent in older parts of the corallum and blocky aragonitic cements can occur. Pores and primary skeletal elements are overgrown by new microstructures. These microstructures are caused by secondary cementation and exhibit frontal zones (Stirnzonen), zigzag-like and pseudolamellar-structures. The lamellar structures can be compared with similar structures in the exoskeleton of some Rugosa. A very short early diagenesis within the recent corals is responsible for the thickening and alteration of skeletal elements. It occurs only 4 to 5 years after the formation of the skeleton and tends to increase in importance in older parts of the corallum. Nevertheless, there is no proof for any alteration of aragonite to calcite.     

FURTHER STUDIES ON MOBILIZATION OF CFUs

Mobilization of CFUs from haemopoietic tissues into circulation was studied after injection of different bacterial lipopolysaccharides (LPS), zymosan, phytohaemagglutinin (PHA), concanavalin A (Con A), trypsin and di-isopropyl-fluorophosphate-inhibited trypsin. All bacterial LPS used gave an increase of CFUs in the peripheral blood at 1 h after i.v. injection. Some variation in activity could not be excluded. As with Salmonella typhosa LPS, zymosan gave an increase in circulating CFUs during the first few hr and a second peak a few days later. After injection of zymosan as well as S. typhosa LPS the second peak in the blood was accompanied by a large increase in CFUs numbers in the spleen. PHA gave an immediate mobilization of CFUs, but the mobilization after injection of Con A during the first few hr occurred more slowly. After injection of S. typhosa LPS, zymosan and PHA the blood C3 level was found to be depressed considerably. This might indicate that the complement system is involved in the early mobilization of CFUs. Dexamethasone, a synthetic hormone which has been reported to give sequestration of several cell types in the bone marrow, did not inhibit the early and late mobilization of CFUs which normally occurs after injection of S. typhosa LPS.     

Tandem affinity purification of functional TAP-tagged proteins from human cells

Tandem affinity purification (TAP) is a generic two-step affinity purification protocol for isolation of TAP-tagged proteins together with associated proteins. We used bacterial artificial chromosome to heterologously express TAP-tagged murine Sgo1 protein in human HeLa cells. This allowed us to test the functionality of the Sgo1-TAP protein by RNA interference-mediated depletion of the endogenous human Sgo1. Here, we present an optimized protocol for purification of TAP-tagged Sgo1 protein as well as KIAA1387 from HeLa cells with detailed instructions. The purification protocol can be completed in 1 day and it should be applicable to other proteins.