摘要:

AbstractErythrocytes were bound to a lectin‐coated surface; the multivalent attachment to this surface resulted in a severe deformation of the cells and an alteration in the cellular phospholipid metabolism. Human erythrocytes were allowed to bind for 20 min at 20°C to polystyrene beads coated with wheat germ agglutinin (WGA beads). The bound erythrocytes were then lysed to produce stroma bound to WGA beads. Control stroma and stroma‐WGA beads were incubated at 37°C with γ‐32P‐ATP to examine the phospholipid labeling patterns. The control stroma incorporated32P‐label into phosphatidylinositol‐4‐phosphate and phosphatidylinositol‐4,5‐bisphosphate, in agreement with earlier studies. However, the stroma‐WGA beads showed incorporation of32P‐label into phosphatidic acid in addition to that in the phosphoinositides. The quantity of32P‐phosphatidic acid produced during the 20‐min assay was 3.23 ± 0.84 (n = 7) picomoles/μg stromal cholesterol; the amount synthesized, however, was dependent on the procedure used to prepare the stroma‐WGA beads. If the erythrocytes were bound to the WGA beads at 0°C instead of 20°C, the quantity of32P‐phosphatidic acid produced during the subsequent 37°C assay with γ‐32P‐ATP was decreased 4.2 fold; the phosphoinositide labeling pattern was unchanged. In addition, when the time for binding of intact erythrocytes to the WGA beads was varied from 1 to 20 minutes, there was a time‐dependent increase in the amount of32P‐phosphatidic acid produced. This induction of phosphatidic acid synthesis could not be duplicated with fluid phase WGA. Therefore, the multivalent binding of intact erythrocytes to WGA beads c

ISSN:0730-2312    

DOI:10.1002/jcb.240380102

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

年代:1988

数据来源: WILEY

摘要:

AbstractThe levels of a (2′‐5′) An‐dependent endonuclease (RNase L) were determined in extracts prepared from murine L cells and Ehrlich ascites tumor (EAT) cells by measuring specific binding of protein to a labeled derivative of (2′‐5′)An, (2′‐5′)A3[32P]pCp. RNase L levels were found to depend both on interferon (IFN) treatment and on cell growth conditions. Treatment of murine L cells and EAT cells with 100–2,000 IRU IFNβ or IFNγ resulted in a similar 2–4‐fold increase in the levels of RNase L when cells were present at low density. The levels of RNase L were also shown to increase 2–3‐fold as cells approached saturation density. Serum‐starved cells also displayed relatively high levels of RNase L. RNase L levels in cells maintained at high cell density did not change appreciably following treatment with IFNβ or IFNγ. Regulation of RNase L levels by cell growth conditions as well as by IFNβ or IFNγ treatment suggests that RNase L may play an important role in regulating the levels of cellular mRNAs as

ISSN:0730-2312    

DOI:10.1002/jcb.240380103

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

年代:1988

数据来源: WILEY

摘要:

AbstractThe cation‐independent mannose 6‐phosphate receptor (215,000 daltons) was isolated from embryonic bovine tracheal cells and embryonic human skin fibroblasts labelled with [3H]palmitic acid, the tritium label was detected in the protein upon fluorographic analysis of SDS‐polyacrylamide gels of the purified receptor. The label was not sensitive to hydroxylamine, methanolic KOH, or β‐mercaptoethanol, but labelled fatty acid was recovered from the protein by acidic methanolysis. Labelled receptor protein could not be isolated from cells grown in the presence of [3H]myristic acid. The results suggest the presence of amide‐linked palmitic acid in the structure of the cation‐independent mannose 6‐phos

ISSN:0730-2312    

DOI:10.1002/jcb.240380104

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

年代:1988

数据来源: WILEY

摘要:

AbstractBacteroids having a high level of respiration‐supported nitrogenase activity were isolated from nitrogen‐fixing alfalfa root nodules. Gentle maceration under anaerobic conditions in the presence of sodium succinate and a fatty acid scavenging agent were employed in this method.A large proportion of isolated bacteroids retained a triple membrane structure as shown by transmission electron microscopy. Dicarboxylic acids of the TCA cycle (malate, fumarate, succinate), but not glutamate or aspartate, supported sufficient respiratory activity to supply the nitrogenase system with ATP and reducing equivalents and to protect the nitrogenase system from inactivation by 4% oxygen over a period of 20–30 min. Sugars did not support nitrogenase activity in intact bacteroids. The properties of the isolated bacteroids were ascribed to minimal damage to the cytoplasmic membrane and peribacteroidal membrane during isolation.With succinate as substrate and oxygen as terminal electron acceptor, initial nitrogenase activity was determined at 4% oxygen in the gas phase of the assay system employed. At this oxygen concentration, the sustained rate of acetylene reduction by respiring bacteroids was linear up to 30 min. Bacteroid activity declined rapidly with time of exposure to oxygen above 4% in the gas phase. The optimum temperature range for this activity was 10–20°C. Nitrogenase activity was measurable at incubation tempertures below 10°C under 4% oxygen. Functionally intact bacteroids had little nitrogenase activity under anaerobic conditions in the presence of an external source of ATP and reductant. Treatment of the bacteroids with chlorpromazine eliminated respirtation‐supported activity and rendered the bacteroid cell membrane permeable to external ATP. Bacteroids treated with chlorpromazine had high acetylene reducing activity with external ATP and dithionite in the absenc

ISSN:0730-2312    

DOI:10.1002/jcb.240380105

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

年代:1988

数据来源: WILEY

摘要:

AbstractEpidermal growth factor (EGF) induces the degradation of EGF receptors in both human foreskin fibroblasts and A‐431 cells. Similar degradation products of125I‐EGF covalently linked to its receptor appeared at the same times in both A‐431 cells and fibroblasts when the cells were exposed to a concentration of 10 ng/ml EGF. Although the products between the two cell types differed in molecular weight, this was at least partly caused by an actual difference in the receptor proteins from the two cell types (as shown by partial proteolysis) rather than from different pathways of receptor degradation. However, when EGF receptors were biosynthetically labeled, no receptor degradation products could be observed, even when the receptor was labeled with radioactive mannose or phosphate, molecules which would predominantly label the outside or inside face of the receptor, respectively. At 20°C, degradation of the receptor slowed and a 150,000‐dalton degradation product was observed. This degradation product has previously been observed in cell homogenates produced in the presence of calcium, mediated by calpain. Thus, calpain may play a role in the intracellular degradation of the EGF

ISSN:0730-2312    

DOI:10.1002/jcb.240380106

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

年代:1988

数据来源: WILEY

摘要:

AbstractWe have previously purified a Mr75,000 protein, cytovillin, from cultured human choriocarcinoma cells (JEG‐3) and shown that this protein was specifically confined to the microvillus membrane of these cells. I have now studied the expression and the subcellular distribution of cytovillin in eighteen normal and transformed human cell lines and strains by using immunoblotting and indirect immunofluorescence microscopy. In all cell types, cytovillin was highly enriched in cell surface protrusions. When cell types were ranked according to their staining intensity, choriocarcinoma was highest, then amniotic epithelial cells, other choriocarcinoma cells and tumor cells, and finally fibroblastoid cells. The latter only gave faint diffuse fluorescence on the plasma membrane and, occasionally, on the microvilli. However, detergent extracts of all cell types could be shown to contain cytovillin by the use of immunoblotting techniques. Metabolic pulse‐chase labelling experiments with JEG‐3 cells demonstrated synthesis of cytovillin as a single‐chain polypeptide. No precursor forms or specific proteolytic cleavage products could be seen either by immunoblotting or immunoprecipitation. The protein was found to be very stable with a biologic half‐life of about 25 hours. The pI determined by isoelectric focusing was 6.1. These results were consistent with cytovillin being an integral component of the microvilli and other surface extensions of all human cell types

ISSN:0730-2312    

DOI:10.1002/jcb.240380107

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

年代:1988

数据来源: WILEY

ISSN:0730-2312    

DOI:10.1002/jcb.240380101

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

年代:1988

数据来源: WILEY