摘要:

AbstractThe effects of 4,6‐dioxoheptanoic acid (succinylacetone, SA), an inhibitor of δ‐aminolevulinic acid dehydratase, on total iron uptake, heme synthesis, and globin synthesis were studied in rat marrow cells in culture in order to examine the coordination of heme and globin synthesis. SA inhibited heme synthesis in both control and erythropoietin‐stimulated cells in a dose‐dependent fashion; at 10−3M, inhibition was complete, whereas at 10−7M, there was no significant effect. Inhibition of total iron uptake was also dose‐dependent although, at 10−3M, it was not complete. The inhibition of heme synthesis by SA was partially overcome by addition of 10−4M porphobilinogen or protoporphyrin IX. SA caused an almost complete suppression of globin formation in both erythropoietin‐stimulated and unstimulated cells as early as five hours after the addition of the inhibitor. When inhibition of heme synthesis was incomplete, globin synthesis was partially inhibited. These results indicate that heme synthesis is required for erythropoietin‐mediated induction of globin synthesis in cult

ISSN:0730-2312    

DOI:10.1002/jcb.240210201

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1983

数据来源: WILEY

摘要:

AbstractSwainsonine, an indolizidine alkaloid, found in plants of the genus Swainsona, has been shown to be a strong inhibitor in vitro of the α‐D‐mannosidase activity in normal human fibroblasts. Therefore, inhibition of α‐D‐mannosidase activity in extracts of harvested cells grown with swainsonine in the medium has been used to follow the association of the alkaloid with normal human fibroblasts in culture. Swainsonine that could not be removed by extensive washing became associated with the cells within 1 min, and it is concluded that the alkaloid is internalized rapidly by the cells. The amount of swainsonine taken up into the cells depended on the length of time in contact and the concentration of swainsonine in the medium, but at 37°C a plateau of internalized swainsonine occurred after 2 hr with extracellular concentrations of swainsonine of 100 μM or greater. At lower concentrations of swainsonine the rate of uptake was found to be temperature‐dependent, increasing greatly at 20°C. The rapidity and temperature sensitivity of the uptake, together with the observation that mannose or mannose‐6‐phosphate did not prevent the association, suggest that swainsonine enters the cells by permeation rather than by endocytosis. When swainsonine is withdrawn from the culture medium, there is a decrease with time of cell‐associated swainsonine. The kinetics of uptake and release of swainsonine and its slightly basic nature make it likely that swainsonine is concentrated initially in the lysosomes. This rapid, but reversible, concentration of swainsonine in lysosomes would be consistent with the observed effects

ISSN:0730-2312    

DOI:10.1002/jcb.240210202

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1983

数据来源: WILEY

摘要:

AbstractA β‐galactoside‐binding endogenous lectin extracted from bovine heart binds to the surface of baby hamster kidney (BHK) cells. The binding to and agglutination of cells is reduced in certain ricin‐resistant mutants (Ric cells) in parallel with the decreased number of binding sites for the selective agent, ricin, a galactose‐specific plant lectin. However, clear differences in the binding specificities of bovine lectin and ricin are shown by the effect of neuraminidase. BHK cells and Ric mutant cells treated with neuraminidase bind similar amounts of the bovine lectin compared with untreated cells, and ricin binding is greatly increased.The mammalian lectin immobilised on inert glass mediates the attachment and spreading of normal BHK cells and agglutinates these cells in solution. Ricin‐resistant mutant cells respond poorly. These results are consistent with a role of endogenous lectins in cellular adhesiveness and show that cell adhesion may be regulated by the density of specific surface receptors f

ISSN:0730-2312    

DOI:10.1002/jcb.240210203

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1983

数据来源: WILEY

摘要:

AbstractThe biosynthesis and secretion of cellular fibronectin from human melanoma cells have been investigated by pluse‐chase/immunoprecipitation analysis. Melanoma cells synthesize endolglycosidase H (Endo H)‐sensitive glycoprotein precursors of fibronectin glycoproteins which chase to an Endo H‐resistant monomer with an apparent Mrof 240,000 (240 K). This molecule, which has a significantly higher molecular weight than normal plasma or cellular fibronectin, is rapidly secreted by melanoma cells, resulting in the secretion of 80% of newly synthesized fibronectin in 120 min, following a 10‐min biosynthetic pulse. This active secretory process can be inhibited by brief exposure of melanoma cells to sodium monensin (10−7M), which also results in a modified fibronectin of lower apparent Mr. Monosaccharide‐incorporation studies of melanoma fibronectin reveal that monensin significantly inhibits galactose and fucose incorporation into this glycoprotein, correlating with reported effects of monensin on Golgi apparatus functions. These studies indicate that this tumor‐associated and biosynthetically altered cellular fibronectin is a rapidly secreted major N‐linked glycoprotein of metastatic human

ISSN:0730-2312    

DOI:10.1002/jcb.240210204

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1983

数据来源: WILEY

摘要:

AbstractDigitonin was used as a tool to investigate the organization and function of cholesterol in gastric microsomes. Microsomal vesicles were treated with digitonin for different time at 0–4°C under isotonic conditions. The effects of digitonin treatment of the vesicles on removal of cholesterol, ultrastructural changes, (H++ K+)‐ATPase activity, and gastric ATPase‐dependent H+uptake ability were investigated. Microsomal cholesterol was extracted in an exponential manner with a t1/2of 32 min. There was no release of microsomal phospholipids by digitonin treatment during the same period. Digitonin treatment (30 min) produced visible “holes” in the vesicles; at the same time (H++ K+)‐ATPase‐dependent H+uptake was abolished. Under the same conditions the K+‐stimulated ATPase activity, however, was moderately (about 35%) reduced, although the response of K+stimulation to valinomycin was obliterated. Longer digitonin treatment resulted in gradual diffusion and eventual disappearance of the “holes” with the generation of distorted cup‐shaped microsomes. The data strongly suggest that membrane lipids are freely mobile and that there is a certain degree of specialization in the organization of gastric microsomal cholesterol for the proper maintenance of the membrane

ISSN:0730-2312    

DOI:10.1002/jcb.240210205

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1983

数据来源: WILEY

ISSN:0730-2312    

DOI:10.1002/jcb.240210206

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1983

数据来源: WILEY

摘要:

AbstractSeveral investigators have reported that there are both large and small insulinbinding proteins in plasma membranes; the larger protein demonstrates nonlinear Scatchard binding, and the smaller protein has linear binding. We now present evidence that the larger insulin‐binding species consists of four proteins of different sizes. Rat epididymal adipocyte plasma membranes were prebound with125I‐insulin and then exposed to 1 mM disuccinimidyl suberate for 15 min at 2°C. The membranes were solubilized in 0.1% Triton X‐100 and applied to a Sepharose 6B column. Peaks of radioactivity from the column were dialyzed, lyophilized, and analyzed by dodecyl‐sulphate gel electrophoresis (5%, 100/1; mono/bisacrylamide). Autoradiograms of the gels were scanned with a densitometer. The Sepharose chromatogram revealed four radioactive peaks: peak 1 at column void volume; peak 2, Kav= 0.27; peak 3, Kav= 0.77; and peak 4, Kav= 1.09. Dodecyl sulphate electrophoresis of fractions in peak 2 demonstrated four bands on autoradiography; peak 1 did not enter the gel and peaks 3 and 4 ran with the dye front. Molecular weight estimates of the four insulin‐binding species in peak 2 were 600, 500, 420, and 350 K. On dithiothreitol reduction each insulin‐binding species yielded subunits of Mr≅ 135 and 18 K. The three largest binding species demonstrated an additional 45‐K dalton protein on dithiothreitol reduction, and the 500‐K and 420‐K binding species also yielded a 49‐K dalton protein. These results suggest that the large insulin‐binding protein in rat epididymal adipocytes contains several insulin‐binding species, and that these insulin‐binding species differ in the number of and the type of subunits they contain. In addition, it may be postulated that the nonlinear Scatchard binding associated with the larger binding protein is a consequence of the heterogeneity of the insulin‐binding

ISSN:0730-2312    

DOI:10.1002/jcb.240210207

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1983

数据来源: WILEY

摘要:

AbstractWe immunized mice with insulin and found that those strains that develop insulin antibodies subsequently produce insulin‐like activity in amount equivalent to 300–400 ng insulin per ml serum. The activity was due exclusively to IgG2 antibodies. Bioactivity could be blocked efficiently by insulin antibodies from guinea pigs and from mice. The active IgG2 also displaced labeled insulin from fat cells. Preliminary in vivo studies have indicated that the appearance of insulin‐like antibodies in the mouse resulted in abnormal glucose homeostasis and “down regulation” of insulin receptors. These results indicate that immunization to insulin can initiate an idiotype‐anti‐idiotype network resulting in antibodies to the hor

ISSN:0730-2312    

DOI:10.1002/jcb.240210208

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1983

数据来源: WILEY