Nucleotide sequence of the Syrian hamster intracisternal A-particle gene: close evolutionary relationship of type A particle gene to types B and D oncovirus genes.

We determined the complete nucleotide sequence of the intracisternal A-particle gene, IAP-H18, cloned from the normal Syrian hamster liver DNA. IAP-H18 was 7,951 base pairs in length with two identical long terminal repeats of 376 base pairs at both ends. On the coding strand, imperfect open reading frames corresponding to gag and pol of the retrovirus genome were observed, whereas many stop codons were present in the region corresponding to env. The putative H18 gag gene (809 amino acids) had a sequence homologous to the N-terminal half of the mouse mammary tumor virus gag gene and locally to the Rous sarcoma virus gag gene. The putative H18 pol gene (900 residues) was homologous to the Rous sarcoma virus pol gene almost throughout the entire region. Two conserved regions among the retrovirus pol genes have been reported. One presumably corresponds to the DNA polymerase and the RNase H domain, and the other corresponds to the DNA endonuclease domain of the multifunctional protein pol. By the comparison of the deduced amino acid sequences of the putative endonuclease domain of six representative oncovirus genomes, a phylogenetic tree of the oncovirus genomes was constructed, and the intracisternal A-particle (type A) genome was found to be more closely related to the mouse mammary tumor virus (type B) and squirrel monkey retrovirus (type D) genomes.     

Fine-granular cationic iron colloid. Its preparation, physicochemical characteristics and histochemical use for the detection of ionized anionic groups

In order to obtain distinct and reliable information concerning the localization of ionized anionic groups in tissues, fine-granular cationic ferric hydroxide colloid solution (Fe-Cac-f) was newly devised. This can be obtained by boiling a mixture of ferric chloride and ammonium cacodylate solutions. The colloid particles of Fe-Cac-f are about 1.0 nm in size, i.e., one-fifth of the size of ferric cacodylate colloid (Fe-Cac; Seno et al. 1983a). As with Fe-Cac, Fe-Cac-f particles in the pH range of 1.6-7.6 carry a positive electric charge, but the latter show a better permeation of tissues. Using the Prussian blue reaction, Fe-Cac-f gives a distinct deep-blue color and can be used for the detection of anionic groups of acid mucopolysaccharides and proteins by light microscopy. It is also useful for detecting the exact sites of ionized anionic groups in deep tissue areas using electron microscopy.     

IgE heavy chain constant region genes in atopic dermatitis and senile erythroderma patients

By Southern hybridization using a genomic DNA fragment carrying a human IgE heavy chain constant region gene (C ) as a probe, we analyzed the organization of human C genes and their flanking regions in 23 atopic dermatitis and 6 senile erythroderma patients with elevated serum IgE levels, and 6 atopic dermatitis patients with normal IgE levels. On Barn HI, Hind III, and Eco RI digestions, we detected three hybridizable fragments containing three human C genes, C 1, C 2, and C 3, respectively, in all leukocyte DNAs. These fragments were almost identical in size among patients and healthy donors. Pst I digestion generated a genetic polymorphism. We, however, could find no correlation between this polymorphism and the disorders. It was concluded that among the patients and healthy donors, there was no marked difference in the organization of the functional C gene and its flanking region containing a class switch region. Our conclusion cannot rule out the presence of genetic abnormalities of this region in some atopic dermatitis patients which are not resolvable by our method. In the course of this study, we found a novel C -like gene in placenta DNA which differs from the three C genes commonly present in normal human DNA.     

Epitope-specific regulation. IV. In vitro studies with suppressor T cells induced by carrier/hapten-carrier immunization

Sequential immunization with a carrier molecule and a new epitope (hapten) conjugated to the carrier (carrier/hapten-carrier immunization) induces specific suppression for IgG antibody production to the new epitope (hapten) on the carrier. Once induced, this "epitope-specific" suppression persists and specifically suppresses subsequent in vivo IgG antibody responses to the hapten presented on the same or on an unrelated carrier molecule. In vitro studies presented here characterize the surface markers and specificity of suppressor T cells generated in carrier/hapten-carrier-immunized animals. Thus we show (1) that spleen cells from these donors suppress in vitro IgG anti-hapten antibody production by cocultured hapten-primed spleen cells; (2) that some but not all of the suppressor cells carry surface Lyt-2; (3) that at least some of the suppressor cells have receptors for the inducing hapten (DNP); and (4) that, unlike the suppression obtained in vivo, the in vitro suppression extends to IgG responses to unrelated carrier protein epitopes presented in association with the inducing hapten.     

Spatial relationship among individuals of the Japanese lacertidTakydromus tachydromoides (Sauria,Lacertidae)

The home range ofTakydromus tachydromoides was studied in a grassland area from April 1977 to November 1978. The mean size of home range did not differ markedly between sexes; 136.5 m² for males and 130.8 m² for females. Home ranges of adults overlapped greatly in each sex, and the lizard was considered to be non-territorial. Individuals showed return movement to a definite area (sleeping site) within the home range, and the home range did not shift within a year or between years. Characteristics of the home range of this grassland-inhabiting lizard were discussed in relation to resource abundance and predation pressure.     

Diacylglycerol kinase from pig brain. Purification and phospholipid dependencies

Diacylglycerol kinase (EC 2.7.1.-) was purified 1,650-fold from pig brain cytosol. The purified enzyme showed a single protein band on polyacrylamide gel electrophoresis in the presence and absence of sodium dodecyl sulfate. The molecular weight of the kinase was estimated to be 78,000 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. A similar value (76,000) was obtained by Sephadex G-150 gel filtration. The activity of the purified enzyme was markedly enhanced by either deoxycholate or phospholipids. The extent of activation by phospholipids was in the order of phosphatidylcholine greater than lysophosphatidylcholine greater than phosphatidylethanolamine approximately equal to phosphatidylserine greater than sphingomyelin. Other phospholipids and unsaturated fatty acids were ineffective. Phosphatidylcholines from egg yolk and pig brain, and dioleoyl phosphatidylcholine were similarly effective. Saturated phosphatidylcholines with acyl chain lengths shorter than palmitate also gave a considerable activation. The activity with phosphatidylcholine was from 1.5- to 2.5-fold higher than that measured with deoxycholate. A very small amount of phosphatidylinositol or phosphatidylglycerol potently inhibited the phosphatidylcholine-dependent (but not deoxycholate-dependent) kinase activity. The inhibition by phosphatidylinositol was varied according to its molar ratio to phosphatidylcholine. As little as about 2.5 mol per cent of phosphatidylinositol resulted in 50% inhibition of the phosphatidylcholine-dependent kinase activity. The deoxycholate- and phosphatidylcholine-dependent kinase activities showed almost the same Km values for the substrates. In both cases, the apparent Km values for ATP and diacylglycerol were 300 microM and about 60 microM, respectively. The kinase required Mg2+ for its activity. When compared to deoxycholate, phosphatidylcholine was more effective at higher Mg2+ concentrations. The deoxycholate-dependent activity showed a broad pH optimum at around 8.0, whereas the phosphatidylcholine-dependent activity formed a clear peak at pH 7.4.     

Histochemical application of mild alkaline hydrolysis for selective elimination of O-glycosidically linked glycoproteins

A new technique to eliminate O-glycosidically linked glycoprotein (mucin-type glycoprotein) selectively has been developed. Composite paraffin sections were collodionized before and after alkaline treatment with 0.5 M NaOH in 70% ethanol; the effect of this procedure on mucosubstances was examined using the periodic acid-Schiff reaction. Exposure to alkaline hydrolysis for 72 to 144 hours at 4 C led to a complete loss of periodic acid-Schiff reactivity of epithelial mucins in rat sublingual gland, stomach and small intestine, but that of fuzzy coat, thyroid colloid, collagen fibers and tracheal cartilage was well preserved. These results agreed fairly well with biochemical findings. The present study also revealed that materials prepared by freeze-substitution provided the most satisfactory results.     

Fluorescence polarization as a tool to study lectin-sugar interaction

The binding ofRicinus communis agglutinin andAbrus agglutinin to 4-methylumbelliferyl β-D-galactopyranoside was studied by equilibrium dialysis, fluo-rescence quenching and fluorescence polarization. The number of binding sites and the association constant value obtained by fluorescence polarization for bothRicinus communis agglutinin andAbrus agglutinin are in close agreement with those obtained by the other methods. This indicates the potential of ligand-fluorescence polarization measurements in the investigation of lectin-sugar interactions.     

Lectins from<Emphasis Type="Italic">Griffonia simplicifolia</Emphasis> seeds

The physical-chemical and carbohydrate binding specificity ofGriffonia simplicifolia I (GS I) isolectins, one of the 4 lectins isolated fromGriffonia simplicifolia seeds, are described.Association constants for the binding of methyl α- and β-D-galactopyranoside and methyl 2-acetamido-2-deoxy-α-D-galactopyranoside to the A₄, A₂ B₂ and B₄ isolectins are reported.Precipitation reactions of theGriffonia simplicifolia isolectins with guaran and type B blood group substance are described.The hypothesis that subunit B is a precursor of subunit A, a process involving proteolytic cleavage of the B subunit, was tested by conducting structural studies on the 2 subunits. The results indicated that the A and B subunits are probably products of 2 separate but closely related, possibly contiguous genes.     

Haemolysis of rabbit erythrocytes by a lectin from the seeds of<Emphasis Type="Italic">Croton tiglium</Emphasis>

The kinetics of haemolysis of rabbit erythrocytes byCroton tiglium lectin was studied as a function of concentration of the lectin and erythrocytes. The length of the prelytic period decreased with increasing lectin concentrations, indicating that the secondary events at the membrane which follow the binding of the lectin to cell surface carbohydrate receptors are accelerated at higher surface concentrations of the lectin. The rate or extent of haemolysis was not affected by the inclusion of ions like K⁺, Ca²⁺ and Mg²⁺ in the medium or by the substitution of ionic medium by a non-ionic medium. The inhibition of haemagglutination and haemolysis of rabbit red cells byCroton tiglium lectin by antilectin rabbit serum was observed. A possible mechanism of haemolysis by the lectin is discussed.