Fine Structure of Lipid-Depleted Mitochondria

Fleischer, Sidney; Fleischer, Becca; Stoeckenius, Walther

doi:10.1083/jcb.32.1.193

Cited by 301 publications

(70 citation statements)

References 10 publications

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“…Some of the samples were fixed in 2.5% glutaraldehyde for 2 h at 4~ and then postfixed in 1% OsO4 in 0.1 M cacodylate buffer pH 7.2 (glutaraldehydeOsO4). All samples were block-stained with 0.5% uranyl acetate in veronal-acetate buffer (pH 6.0) for 2 h at room temperature, dehydrated in a series of increasing ethanol concentrations, the ethanol was replaced with propylene oxide, and the samples were embedded in Epon 812 (7). Thin sections were cut on an LKB Ultratome (LKB Instruments, Inc., Rockville, Md.…”

Section: Embed-dingmentioning

confidence: 99%

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Membrane asymmetry and enhanced ultrastructural detail of sarcoplasmic reticulum revealed with use of tannic acid.

Saito¹,

Wang²,

Fleischer³

1978

The Journal of Cell Biology

111

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Fixation of purified sarcoplasmic reticulum (SR) membrane vesicles, using glutaraldehyde supplemented with 1% tannic acid, reveals newly visualized ultrastructure in thin sections. The trilaminar appearance of the membrane is highly asymmetric; the outer electron-opaque layer is appreciably wider (70 ~) than the inner layer (20 ~). The asymmetry is not referable to lack of penetration of the tannic acid since: (a) SR vesicles made permeable with 1 mM EDTA, pH 8.5, show similar asymmetry; (b) treatment of SR with trypsin results in progressive loss in protein content and decrease in the thickness of the outer layer, until in the limit the trilayer has a symmetric appearance; (c) within the same muscle section, the SR membrane appears highly asymmetric whereas the sarcolemma has a more symmetric appearance; (d) reconstituted SR vesicles have a symmetric appearance with equally broad inner and outer layers (-70 •); the symmetric structure is confirmed by freeze-fracture and negative staining electron microscopy. Heavy and light SR vesicles obtained by isopycnic density sedimentation of purified SR have the same asymmetric appearance of the membrane and seem to differ mainly in that the heavy vesicles contain internal contents consisting largely of Ca++-binding protein. The asymmetry of the SR membrane is referable mainly to the unidirectional alignment of the Ca ++ pump protein, the major component (90% of the protein) of the membrane. The asymmetry of the SR membrane can be visualized now for the first time in situ in thin sections of muscle.

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Section: Embed-dingmentioning

confidence: 99%

“…NEGATIVE STAINING: Negative staining with 2% phosphotungstic acid (pH 7.2) or 1% uranyl acetate was carded out as described previously (7). FREEZE-FRACTURE: Freeze-fracture and preparation of the replicas was carried out in a Balzers BAF 300 apparatus (Balzers High Vacuum Corp., Santa Ana, Calif.) in a manner previously described (3).…”

Section: Embed-dingmentioning

confidence: 99%

Membrane asymmetry and enhanced ultrastructural detail of sarcoplasmic reticulum revealed with use of tannic acid.

Saito¹,

Wang²,

Fleischer³

1978

The Journal of Cell Biology

111

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“…(19) : 85-90% of the PLP were extracted by the treatment . Since, however, no distinct changes in the electrophoretograms appeared, delipidation was routinely omitted .…”

Section: Gel Electrophoresismentioning

confidence: 99%

“…In the acid gel most of the fast-moving bands (Nos . [13][14][15][16][17][18][19] appear to be contributed predominantly by ribosomes . Hence, they are present in pure ribosome preparations (Figs .…”

Section: Gel Electrophoresismentioning

confidence: 99%

Composition of Cellular Membranes in the Pancreas of the Guinea Pig

Meldolesi

Cova

1972

The Journal of Cell Biology

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Two methods of polyacrylamide gel electrophoresis (the acid method of Eytan and Ohad and the Na dodecylsulfate (SDS) disc method of Maizel) have been used for analyzing the proteins of gel fractions isolated from the guinea pig pancreatic exocrine cells and in particular the proteins bound to the membranes involved in the synthesis, intracellular transport, and discharge of secretory enzymes : rough (RM) and smooth microsome (SM) membranes, zymogen granule (ZG) membranes, and plasma membranes (PM) . Since in the two systems the electrophoretic mobility of proteins depends on different factors (size, shape, and net charge of molecules in the acid system ; size only in the SDS system) a deeper insight into the protein composition of the fractions could be obtained . The gel patterns of RM, SM, and ZG membranes turned out to be accounted for mainly by segregated secretory enzymes (in rough microsomes also by ribosome proteins) and thus were found to share most of the bands . In contrast, with highly purified membrane fractions different patterns were obtained : RM and SM membrane proteins turn out to contain a large number of different proteins with molecular weights varying between 150,000 and 15,000 daltons . The pattern of ZG membranes was greatly different in the two systems : only two bands were separated by the acid method and as many as 23 by the SDS m ethod. PM gave a rather complex pattern in either system . Both ZG membranes and PM were found to contain a large proportion of low molecular weight proteins . Nothing appears in common between the proteins of SM membranes (primarily of Golgi origin) and those of ZG membranes, while the latter and PM exhibit a certain degree of similarity . By amino acid analysis we found only slight differences : relative to the other fractions : RM membranes were higher in basic amino acids and ZG membranes contained a larger amount of methionine . Taken together with recent data on lipid composition and enzyme activities of the same fractions, these results indicate that the membranes of the pancreatic exocrine cells are chemically and functionally distinct, and hence do not mix randomly with one another during the transport of secretory products .It is well known that in pancreatic acinar cells functionally interconnected membrane-bound secretory proteins, following their synthesis in the compartments : in sequence, the RER, the Golgi attached ribosomes, are segregated within the complex (GC), and the zymogen granules (ZG) . cisternae of the rough endoplasmic reticulum (RER)' and then transported through a series of ' Abbreviations used : RER, rough-surfaced endo-

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“…The aqueous acetone treatment extracted about 70% of the mitochondrial phospholipid although the degree of extraction was not as much as reported with rat liver mitochondria (about 80%) (4). With rat liver mitochondria, an aqueous acetone treatment containing ammonium extracted larger amounts (95%) of the phospholipid than the aqueous acetone treatment (4); however, in sweet potato mitochondria, the treatment extracted only about 50% of the phospholipid.…”

Section: Resultsmentioning

confidence: 74%