Bead rings at the endoplasmic reticulum-golgi complex boundary: morphological changes accompanying inihibition of intracellular transport of secretory proteins in arthropod fat body tissue

Golgi complex beads are 10-nm particles arranged in rings on the smooth surface of rough endoplasmic reticulum (ER) makind the forming face of the Golgi complex (GC). In arthropod cells they stain specifically with bismuth. Their morphology has been studied after treatment with reagents known to interfere with GC function. Inhibitors of oxidative phosphorylation (antimycin A, cyanide, and anoxia), but not an inhibitor of glycolysis (iodoacetate), both cause the bead rings to collapse and the GC saccules to round up, and inhibit transition vesicle (TV) formation. Cycloheximide blocks protein synthesis on ribosomes but does not stop TV formation or disrupt bead rings, even after prolonged treatment (6 h) to allow emptying of the rough ER cisternae. Thus the collapse of bead rings is not attributable to inhibition of protein synthesis, and the ring structure of beads does not require continued protein synthesis and secretion for its maintenance. Valinomycin has effects on the GC similar to those of antimycin A, but {"type":"entrez-nucleotide","attrs":{"text":"A23187","term_id":"833253","term_text":"A23187"}}A23187, monensin, and lasalocid do not affect bead ring structure or TV formation. These results are consistent with valinomycin’s secondarily uncoupling mitochondria, which collapses bead rings and prevents TV formation. Thus inhibitors of oxidative phosphorylation do not influence the beads through cation movement. Because mononsin and lasalocid block secretion at the level of the condensing vacuoles, bead rings are not influenced by blocks in secretion distal to them or by the backup of secretory material. These experiments are consistent with inhibitors of oxidative phosphorylation collapsing bead rings by decreasing intracellular ATP. The concomitant block to TV formation and the collapse of bead rings suggests that integrity of the bead rings is essential for the transport of secretory material from the rough ER to the GC.     

Individual variation of nucleoside triphosphate pyrophosphohydrolase activity in human erythrocytes,granulocytes, lymphocytes,and platelets

Analysis of the nucleoside triphosphate pyrophosphohydrolase specific activity of red cells obtained from a random Caucasian population indicated at least two subclasses. The specific activity of 18% of the population ranged from undetectable activity to 27.5 nmol ITP cleaved/20 min/mg hemoglobin. The remainder of the population had higher activity, 27.5–125 nmoles ITP cleaved/20 min/mg hemoglobin. The variation of NTPH activity evident in the red cells of an individual is reflected in granulocytes, lymphocytes, and platelets of that individual. Erythrocyte activity ranges from 0.7 to 21 units (nmol of ITP cleaved in 20 min)/10⁷ cells, granulocytes have 17–201 units/10⁷ cells, lymphocytes have 91–462 units/10⁷ cells, and platelets have 1.1–7.1 units/10⁷ platelets. These cell differences are discussed with respect to the hypothesis that NTPH prevents incorporation of ITP or dITP into nucleic acids.This work was supported by funds allocated by the Agricultural Experiment Station, Michigan State University. Michigan Agricultural Experiment Station Journal No. 8727.     

The proton-translocating adenosine triphosphatase of the obligately anaerobic bacterium Clostridium pasteurianum. 1. ATP phosphohydrolase activity.

1. The cell-membrane ATP phosphohydrolase of vegetatively grown Clostridium pasteurianum was specifically Mg2+-dependent, but demonstrated significant activity with GTP, CTP and UTP. It displayed approximate Michaelis-Menten kinetics only in the presence of certain effectors (e.g. phosphoenolpyruvate, fructose 1,6-bis-phosphate) which decreased the Km for ATP (to below 2 mM) but also V, whilst extending to pH 5.8 the effective pH range of activity of the enzyme. 2. ATP phosphohydrolase activity of the membrane ATPase (BF0F1) was inhibited by N,N'-dicyclohexylcarbodiimide, butyricin 7423, Dio-9, 4-chloro-7-nitrobenzofurazan, efrapeptin, leucinostatin and quercetin, and to a lesser degree by aurovertin and citreoviridin. The enzyme was not inhibited by oligomycin, spegazzinine, tributyl tin, triethyl tin or venturicidin. The soluble ATPase (BF1) component differed in not being inhibited by N,N'-dicyclohexylcarbodiimide, butyricin 7423 or leucinostatin. 3. The ATPase (BF0F1) complex and its soluble (BF1) component were separately purified. 4. Dodecylsulphate/polyacrylamide gel electrophoresis separated only four polypeptide components in the purified ATPase (BF0F1), with approximate molecular weights (+/- 10%) as follows: subunit a, 65 500; subunit c, 57 500; subunit da, 43 000; subunit fa, 15 000. The soluble (BF1 component contained only the three polypeptide subunits a, c and da. These were present in the BF0F1 preparation in the ratio 2 : 1 : 2; the contribution of subunit fa could not satisfactorily be quantified. 5. Subunit a was identified as the component binding 4-chloro-7-nitrobenzofurazan and subunit fa as the component binding N,N'-dicyclohexylcarbodiimide. The ATP phosphohydrolase activity of the membrane ATPase was not activated by trypsin treatment and the ATPase (BF0F1) contained no trypsin-sensitive inhibitor protein subunit. 6. Purified ATPase (BF0F1) was incorporated into artificial proteoliposomes which demonstrated ATP-dependent enhancement of 8-anilinonaphthalene-1-sulphonate fluorescence and ATP-dependent proton influx. These reactions were abolished by proton conductors (e.g. carbonylcyanide m-chlorophenylhydrazone) by valinomycin in the presence of a high external concentration of K+, or by N,N'-dicyclohexylcarbodiimide, butyricin 7423, Dio-9, 4-chloro-7-nitrobenzofurazan or leucinostatin. Oligomycin, tributyl tin, triethyl tin and venturicidin were not inhibitory. 7. When stripped of the soluble BF1 component, such ATPase-proteoliposomes demonstrated nil ATP phosphohydrolase activity and did not display ATP-dependent enhancement of 8-anilino-naphthalene-1-sulphonate fluorescence or ATP-dependent protein influx. All of these activities were restored by incubation of the BF1-depleted proteoliposomes with a purified preparation of the soluble BF1 component.     

Extradural haematoma: effect of delayed treatment.

The case records of patients with extradural haematomas treated in the Lothian region during 1951-60 and 1968-77 were analysed to assess the effect of delay in treatment on morbidity and mortality. Delay was defined as the time from deterioration in level of consciousness to surgical evacuation of clot. There were 83 supratentorial extradural haematomas unassociated with intradural clot or contusion. The mean delay in patients who died was 15.7 hours, while in good-quality survivors the mean delay was 1.9 hours. Mortality decreased from 33.3% during 1951-60 to 8.9% during 1968-77. In addition, good recovery without morbidity occurred in 40.7% of patients in the earlier period and 67.9% in the later period. Mean delays from deterioration in level of consciousness to operation were 9.8 and 2.4 hours in the earlier and later periods respectively. The results emphasise the need for immediate operation in patients deteriorating with extradural haematomas. Direct admission of all head-injured patients to a head and spinal injuries unit staffed by neurosurgeons resulted in minimal delay times as well as a reduction in morbidity and mortality.     

Prolyl hydroxylase activity in normal and diseased human liver.

Prolyl hydroxylase activity was determined in liver biopsy samples obtained from 10 patients. The liver prolyl hydroxylase values in patients with active hepatitis distribute into two numerical populations based on the extent of elevation over control. The first of these groups includes those with enzyme levels elevated approximately 2.5-fold over normal. Included in this group are patients with active (but nonagrressive) hepatitis and patients where advanced portal fibrosis is already established. The second group where prolyl hydroxylase is elevated approximately nine-fold is comprised of two patients with advanced clinical symptoms of active alcoholic hepatitis with evidence of aggressive cirrhosis but with only early minimal evidence of existing fibrosis.     

Conformational studies of peptides. The crystal structures of N-acetyl-L-prolinamide and N-acetyl-(S)-thiazolidine-4-carboxamide

The crystal structure of N′-acetyl-L -prolinamide and its isomorphous alalog N′-acetyl-(S)-thiazolidine-4-carboxamide was determined using highly accurate parameters obtained by room- and low-temperature data-collecting systems. Both crystals are orthorhombic, space group P2₁2₁2₁, with four molecules per unit cell held together by a hydrogen-bond system that extends in all directions. Both molecules exhibit the following conformational features: the acetyl group is in the trans configuration in respect to the tertiary amide. The primary amide is almost at right angles with respect to the mean plane of the ring and the –NH₂ group is over the ring. The pyrrolidine and thiazolidine rings were found to be rather flexible with C^β puckering in the former and sulfur in the latter.     

Incidence of Aflatoxin in California Almonds

In a survey of California almonds, aflatoxin was found in 14% of 74 samples of unsorted, in-shell almonds as received by the processor in 1972, but it occurred at very low levels (below 20 parts per billion [ppb]) in 90% of the contaminated samples. The overall proportion of individual nuts contaminated was especially low and is estimated with 95% probability to have been in the range of 1 nut/55,300 nuts to 1 nut/14,700 nuts. Aflatoxin contamination is not restricted to any particular section of the almond-growing region of California. Commercial sorting procedures are effective in removing most aflatoxin-contaminated nutmeats, since none of 26 samples of processed, whole nutmeats contained aflatoxin. In contrast, 13 of 27 samples of diced almonds were contaminated, but nine of these 13 samples contained less than 20 ppb. Only one of 25 samples of sliced nutmeats contained aflatoxin (4 ppb). Thus, aflatoxin incidence in almonds varies greatly with the category of finished product. The apparent high incidence in diced nutmeats is probably due mostly to the more uniform distribution of aflatoxin occurring in this product (because of its small particle size) than that occurring in the other products. Sample size requirements for monitoring aflatoxin in almonds are discussed.