Effect of lysosomotropic amines on the secretory pathway and on the recycling of the asialoglycoprotein receptor in human hepatoma cells

We studied the intracellular transport of secretory and membrane proteins in the human hepatoma cell line HepG-2 infected with vesicular stomatitis virus. Cells were pulse-labeled in the presence of [35S]methionine and chased in the presence of the lysosomotropic agent primaquine. At a concentration of 0.3 mM primaquine effectively inhibited the secretion of albumin and, to a lesser extent, that of orosomucoid and transferrin. The drug also prevented the budding of virus particles at the cell surface. The intracellular transport to the Golgi complex of the membrane protein VSV-G was not affected by primaquine as it acquires resistance to endo-beta-N-acetylglucosaminidase H at the same rate as in control cells. Addition of primaquine at various times after the initiation of the chase period indicates that the effect of primaquine occurs just before secretion. In confirmation of the biochemical data, immunocytochemical localization of albumin in cells treated with NH4Cl demonstrated that albumin accumulated in vesicles at the trans side of the Golgi complex. The effect of primaquine on secretion was also compared with its effect on receptor recycling. The dose-response characteristics of the effect of primaquine on receptor recycling are identical to those of the effects on protein secretion and virus budding. These results indicate that both processes involve the same transport mechanism, and/or that they occur via at least one identical intracellular compartment.     

The role of nerve lysosomal enzymes in the pathogenesis of denervation atrophy. Electromyographic and histochemical study in rats

Section of sciatic nerves of rats produced fibrillations within 3 days. Foci of hyalination leading to necrosis corresponded to segments of muscles containing end plates. The electrolyte content, mainly Ca, was increased, NADH2-TR activity was decreased and membrane ATP-ase was increased. The known increase in hydrolytic enzyme activities in denervated muscles was due to spilling of lysosomal enzymes from degenerating axons at the myoneural junction. This explains the discrepancy between morphological studies indicating paucity of lysosomes in normal muscles and the high hydrolytic enzyme activities in denervation. We propose that denervation changes are at least partly due to the effect of lysosomal spillage from degenerating axons.     

Genetic mapping of pheU, an Escherichia coli gene for phenylalanine tRNA

We report the genetic mapping of pheU , an Escherichia coli gene for phenylalanine tRNA. This gene was located near 94.5 min on the E. coli map. There are no other known tRNA or ribosomal genes in its immediate vicinity.     

Cat pancreatic eicosapeptide and its biosynthetic intermediate. Conservation of a monobasic processing site.

Pancreatic eicosapeptide is synthesized together with the hormone pancreatic polypeptide in a common precursor in the major endocrine cell type of the duodenal pancreas. This processing has been previously demonstrated in man and in the dog. In the present study the cat pancreatic eicosapeptide and a C-terminally extended form of this were isolated and characterized from acid/ethanol extracts of pancreas by gel filtration and reverse-phase h.p.l.c. The sequence homology in the C-terminal part of the eicosapeptides from different species was shown to continue to the other side of the monobasic cleavage site in the extended intermediate form, whereas the end of the extension differed both in chain length and amino acid sequence. It is concluded that the processing site in the intermediate form of the pancreatic eicosapeptide is an example of a proline-directed monobasic cleavage site that has been conserved during evolution.     

A new genus and species of adapid primate from the middle Eocene of Alsace, and a new genus for 'Adapis' ruetimeyeri Stehlin, 1912

A new primate genus and species, represented by isolated teeth, is described from the middle Eocene Alsatian site of Bouxwiller. This new form is closely related to Egerkingen 'Adapis' ruetimeyeri, for which a new genus is proposed because of the important distinctions from type Adapis present in its dentition. Nonetheless, both new genera are closely related to Adapis and Leptadapis.     

Catalase and superoxide dismutase in Escherichia coli

We assessed the roles of intrabacterial catalase and superoxide dismutase in the resistance of Escherichia coli to killing by neutrophils. E. coli in which the synthesis of superoxide dismutase and catalase were induced by paraquat 10-fold and 5-fold, respectively, did not resist killing by neutrophils. When bacteria were allowed to recover from the toxicity of paraquat for 1 h on ice and for 30 min at 37 degrees C, they still failed to resist killing by neutrophils. Induction of the synthesis of catalase 9-fold by growth in the presence of phenazine methosulfate did not render E. coli resistant to killing by either neutrophils or by H2O2 itself. The lack of protection by intrabacterial catalase from killing by neutrophils could not be attributed to an impermeable bacterial membrane; the evolution of O2 from H2O2 was no less rapid in suspensions of E. coli than in lysates. The failure of intrabacterial catalase or superoxide dismutase to protect bacteria from killing by neutrophils might indicate either that the flux of O-2 and H2O2 in the phagosome is too great for the intrabacterial enzymes to alter or that the site of injury is at the bacterial surface.     

Ecdysteroids regulate the release and action of eclosion hormone in the tobacco hornworm, Manduca sexta (L.)

The relationship between the ecdysteroid titre and eclosion hormone was explored for the pupal and adult ecdyses of Manduca sexta. Ecdysteroid treatment late during either moult caused a dosedependant delay in the time of ecdysis. Sensitivity to exogenous steroid treatment dropped off as the respective moults neared completion and in both cases coincided with the time of the low point in the endogenous ecdysteroid titre. It was concluded that an ecdysteroid decline is a normal prerequisite for the ecdyses of both stages. The steroid drop is important for two aspects of the eclosion hormone system: it causes target tissues to become sensitive to the peptide and it is a prerequisite for the subsequent release of eclosion hormone itself. Thus, the dual action of the declining ecdysteroid titre insures that when eclosion hormone is released, the tissues will be competent to respond to it.     

Immunological evidence for two distinct chondroitin sulfate proteoglycan core proteins: Differential expression in cartilage matrix deficient mice

The expression and core protein structure of two proteoglycans, the major cartilage proteoglycan isolated from a rat chondrosarcoma and a small molecular weight chondroitin sulfate proteoglycan isolated from a rat yolk sac tumor, have been compared. The cartilage proteoglycan was not detectable in the cartilage tissue of cartilage matrix deficient ($\text{cmd}\text{cmd}$) neonatal mice by immunofluorescence, but the cmd cartilage did react with antibodies against the core protein of the yolk sac tumor proteoglycan. Radioimmunoassays showed that the core proteins of these proteoglycans are not cross-reactive with each other. Analysis of the core proteins by sodium dodecyl sulfate/polyacrylamide gel electrophoresis after chondroitinase ABC treatment of the proteoglycan revealed a large difference in their sizes. The cartilage proteoglycan core protein had a molecular weight of about 200,000 while the yolk sac tumor proteoglycan core protein migrated with an apparent molecular weight of about 20,000. In addition, the cultured yolk sac tumor cells that make the small proteoglycan did not react with antiserum against the cartilage proteoglycan. These results indicate that the proteoglycan isolated from the yolk sac tumor is similar to the small chondroitin sulfate proteoglycan species found in cartilage and support the existence of at least two dissimilar and genetically independent chondroitin sulfate proteoglycan core proteins.     

Reinitiation of chromosome replication in a thermosensitive DNA mutant of Escherichia coli. II. Synchronization of chromosome replication after temperature shifts

A short incubation at the non-permissive temperature, 10 to 15 minutes at 40 °C, suffices to induce chromosome reinitiation in CRT 266, a thermosensitive DNA mutant of Escherichia coli. In order to acquire the potentiality to reinitiate chromosome replication, protein synthesis is necessary, both during the 40 °C incubation and also during the first 15 minutes after returning to 30 °C.