Induction of erythroid differentiation by cytoplast fusion in mouse erythroleukemia (Friend) cells

An intracellular activity, which is induced by dimethyl sulfoxide (DMSO) or hexamethylenebisacetamide (HMBA) and leads to erythroid differentiation in mouse Friend cells, was characterized by cell fusion between genetically marked intact cells and cytoplasts. For this, a procedure for rapid selection of cybrids was devised by sensitizing non-fused cells with oligomycin. We were able to demonstrate that cytoplasts derived from DMSO- (or HMBA)-treated cells trigger erythroid differentiation upon fusion with UV-irradiated cells. The activity in the cytoplasts remained only transiently and its induction was inhibited by biologically active phorbol esters or cycloheximide. The activity, however, was not induced in cytoplasts by directly treating them with DMSO (or HMBA). These results indicate that (1) the intracellular erythroid-inducing activity is located in cytoplasts, (2) it acts in trans and induces erythroid differentiation as a dominant factor and (3) its production requires de novo nuclear protein synthesis. The mechanisms of the induction of the intracellular activity and of how it triggers erythroid differentiation are discussed.     

Pharmacological and Physicochemical Properties of Pre-Versus Postsynaptic 5-Hydroxytryptamine1A Receptor Binding Sites in the Rat Brain: A Quantitative Autoradiographic Study

Numerous data suggested that the pharmacological and biochemical properties of 5-hydroxytryptamine1A (5-HT1A) receptors exhibit some regional differences in the CNS, notably within the raphe nuclei compared with various forebrain areas (such as the hippocampus). This possibility has been further investigated in the dorsal raphe nucleus and two areas within the hippocampus, the dentate gyrus and the CA1 area, using the quantitative autoradiographic technique. The potencies of 5'-guanylylimidodiphosphate to inhibit the specific binding of 125I-Bolton-Hunter-8-methoxy-2-(N-propyl-N-propylamino)tetralin (125I-BH-8-MeO-N-PAT) to 5-HT1A sites and of N-ethylmaleimide to block these sites irreversibly were identical in the dorsal raphe nucleus and the hippocampal areas in rat brain sections. In contrast, slight but significant differences were noted in the pH dependence and pharmacological properties of 5-HT1A sites labeled by 125I-BH-8-MeO-N-PAT in these three regions. Similarly, heat denaturation experiments and tissue exposure to either phospholipase A2 or the alkylating agent 8-methoxy-2-(N-2'-chloropropyl,N-propyl)aminotetraline revealed regional differences in the properties of 5-HT1A sites. However, in most cases, the observed variations were of greater amplitude between the CA1 area and the dentate gyrus, where 5-HT1A sites are located postsynaptically, than between any one of these areas and the dorsal raphe nucleus where they act as (presynaptic) somatodendritic autoreceptors. These data further support that subtypes of 5-HT1A receptors probably exist in the rat brain, but this heterogeneity seems unrelated to the pre- or post-synaptic location of these receptors.     

Effect of tween 80 on exploratory behavior and locomotor activity in rats

Exploratory behavior and locomotor activity is enhanced in male rat pups (aged 10 to 20 days) whose dams received a chronic dose (1.25 ml/l) of Tween 80 via their drinking water. This enhancement manifests itself during the diurnal period of the day. These data suggest that Tween 80 has an effect on the CNS which could lead to misinterpretation of results in toxicology studies that use this compound as a dosage vehicle.     

Growth-related expression of a double-stranded RNA-dependent protein kinase in 3T3 cells

Cultured mouse 3T3-F442A and 3T3-C2 fibroblasts exhibit a transient double-stranded RNA (dsRNA)-dependent phosphorylation of a 67,000-dalton protein (67K) without prior treatment with interferon (IFN). This phosphoprotein is similar but not identical to the dsRNA-dependent eukaryotic initiation factor-2 (eIF-2) alpha protein kinase (dsI), which regulates protein synthesis in rabbit reticulocytes. We have studied the relationship between cell growth and phosphorylation of the 67K protein (designated 3T3-dsRNA-dependent eIF-2 alpha kinase). A low level of dsRNA-dependent phosphorylation of 3T3-dsI was detectable in extracts prepared from cells not treated with IFN and grown at a low cell density. The phosphorylation of dsI and the phosphorylation of a 38K protein identified as the alpha-subunit (38K) of 3T3-eIF-2 (eIF-2 alpha) occurred concomitantly; the levels of these phosphorylations confluent and thereafter decreased markedly. Treatment of cells with IFN at all stages of growth resulted in an increase in phosphorylation of dsI. 3T3-F442A and 3T3-C2 fibroblasts were found to produce and secrete IFN at levels sufficient to induce an elevated dsI activity.     

Raman and infrared spectra of toxin gamma from the venom of the scorpion Tityus serrulatus

Toxin gamma is a basic, low-molecular-weight, neurotoxic protein, isolated from the venom of the Brazilian scorpion, Tityus serrulatus. Raman spectra (400-1800 cm-1 region) of this toxin in both the lyophilized state and in 0.1 M acetate buffer (pH 4.5) and the infrared spectrum (700-4000 cm-1 region) of a solid film were investigated. From the vibrational spectra, it can be concluded that the polypeptide backbone of toxin gamma consists of a mixture of the different secondary structures, with predominance of beta-sheet, followed by unordered structure and alpha-helix, with some evidence of beta-turn structures. The four disulfide bridges assume the gauche-gauche-gauche conformation of the CCSSCC fragments. The intensity ratio of the doublet at 853 and 828 cm-1 suggests that four out of the five tyrosine residues are exposed. The three tryptophan residues are exposed on the surface, and the single methionine residue assume the gauche-gauche conformation. Toxin gamma retains full activity in the pH 4.5-7.5 range, but is almost completely inactivated at pH 11.5.     

Mutual recognition between polymerized liposomes. IV. Polysaccharide-lectin system.

As a model of cell-cell recognition processes, the association processes of a polysaccharide (mannan)-carrying liposome with a lectin (Concanavalin A, Con A)-carrying polymerized liposome were followed by turbidimetry. The association process was strongly inhibited by the addition of a low molecular weight sugar, methyl-alpha-D-mannopyranoside, which shows that the association between the liposomes is due to the specific interaction between Con A and mannan. The association rate constant obtained was much smaller than the theoretical value for a diffusion-controlled binary association process. This implies that the association rate of liposomes is limited by the recognition between complementary ligands bound on the liposome surfaces. Another reason for the smaller association rate constant in the liposome-liposome system is the repulsive hydration effect. The effect of the surface density of the lectin immobilized on the liposome on the recognition was also examined.     

Pentazocine-induced agranulocytosis.

A 46-year-old man with pentazocine-induced agranulocytosis is described. In previously reported cases of a complete absence of mature neutrophils in the peripheral blood and bone marrow the patients had undergone marrow-depressing treatment with radiation and antineoplastic drugs. This case is unique in that the patient had complete agranulocytosis without predisposing factors.     

Subunit composition and ATP site labeling of the coated vesicle proton-translocating adenosinetriphosphatase

The partially purified proton-translocating adenosinetriphosphatase [(H+)-ATPase] from clathrin-coated vesicles has been reported to contain eight polypeptides of molecular weights 15,000-116,000 [Xie, X.S., & Stone, D.K. (1986) J. Biol. Chem. 261, 2492-2495]. To determine whether these polypeptides form a single macromolecular complex, we have isolated three monoclonal antibodies which recognize the reconstitutively active (H+)-ATPase in the native, detergent-solubilized state. All three monoclonal antibodies precipitate the same set of polypeptides from either the partially purified enzyme or the detergent-solubilized coated vesicle membrane proteins. The immunoprecipitated polypeptides have molecular weights of 100,000, 73,000, 58,000, 40,000, 38,000, 34,000, 33,000, 19,000, and 17,000. These results thus indicate that this set of polypeptides forms a single macromolecular complex and suggest that they correspond to subunits of the coated vesicle (H+)-ATPase. To identify the ATP-hydrolytic subunit of the coated vesicle (H+)-ATPase, the purified enzyme was reacted with N-ethylmaleimide (NEM) and 7-chloro-4-nitro-2,1,3-benzoxadiazole (NBD-Cl), both of which inhibit activity in an ATP-protectable manner. Labeling was carried out by using [3H]NEM or [14C]NBD-Cl, and the specificity of the reaction was increased by prelabeling of the protein with the nonradioactive reagents in the presence of ATP and by taking advantage of the nucleotide specificity of protection. The principal polypeptide labeled by both [3H]NEM and [14C]NBD-Cl had a molecular weight of 73,000. In addition, this protein was the only polypeptide whose labeling was significantly reduced in the presence of ATP.(ABSTRACT TRUNCATED AT 250 WORDS)