Computer modelling of Tetrahymena axonemes at macromolecular resolution. Interpretation of electron micrographs

A computer-generated model of the structural arrangement of the complete 9+2 ciliary axoneme of Tetrahymena at macromolecular resolution (4 nm) is presented. The model reconciles detailed information about subcomponents from negative-stained, thin-section and freeze-fracture electron micrographs, integrating the images into a consistent three-dimensional picture. This illuminates problems such as the requirement for compaction of dynein to form the arm, difficulties in visualization of the circumferential links, construction of the central sheath, and the comparative periodicities of the inner and outer arms. The model is pragmatic in that it is flexible and easily changed, as new information becomes available. It is also useful in the development of dynamic concepts, such as a spatial description of the dynein cross-bridge cycle, which is illustrated, or relationships between adjacent doublets during sliding and bending.

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Structural cross-bridges between microtubules and mitochondria in central axons of an insect (Periplaneta americana)

Journal of Cell Science ◽

10.1242/jcs.27.1.255 ◽

1977 ◽

Vol 27 (1) ◽

pp. 255-272

Author(s):

D.S. Smith ◽

U. Jarlfors ◽

M.L. Cayer

Keyword(s):

Electron Micrographs ◽

Mitochondrial Membrane ◽

Periplaneta Americana ◽

Freeze Fracture ◽

Outer Mitochondrial Membrane ◽

Thin Sections ◽

Cross Bridge ◽

Multiple Association ◽

Random Models ◽

Cross Bridges

The distribution of microtubules and mitochondria in central axons of an insect (Periplaneta americana) is assessed by comparison between counts on micrographs and computed axon random ‘models’. These studies show that the observed multiple association of microtubules with individual mitochondria is statistically highly significant. Electron micrographs of thin sections show that linkage is effected by physical cross-bridge, possibly comprising components from the microtubule and mitochondrion. Linear particle arrays are described on the outer mitochondrial membrane in freeze-fracture replicas, and tentatively related to the bridges seen in thin sections. The results are discussed in terms of proposed roles of microtubules in neurons and other cells.

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A method of estimating radial distribution function of protein particles in membranes from freeze-fracture electron micrographs

Computers in Biology and Medicine ◽

10.1016/0010-4825(82)90035-x ◽

1982 ◽

Vol 12 (4) ◽

pp. 319-322 ◽

Cited By ~ 2

Author(s):

Jacek T. Duniec ◽

David J. Goodchild ◽

Sidney W. Thorne

Keyword(s):

Distribution Function ◽

Radial Distribution Function ◽

Electron Micrographs ◽

Radial Distribution ◽

Freeze Fracture ◽

Protein Particles ◽

Fracture Electron

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Morphology of isolated triads.

The Journal of Cell Biology ◽

10.1083/jcb.96.4.1017 ◽

1983 ◽

Vol 96 (4) ◽

pp. 1017-1029 ◽

Cited By ~ 37

Author(s):

R D Mitchell ◽

A Saito ◽

P Palade ◽

S Fleischer

Keyword(s):

Sarcoplasmic Reticulum ◽

Thin Section ◽

Freeze Fracture ◽

Structural Features ◽

Negative Staining ◽

Transverse Tubule ◽

Freeze Fracture Electron Microscopy ◽

Fracture Electron ◽

Terminal Cisternae

The triad is the junctional association of transverse tubule with sarcoplasmic reticulum terminal cisternae. A procedure for the isolation of highly enriched triads from skeletal muscle has been described in the previous paper. In the present study, the structural features of isolated triads have been examined by thin-section, negative-staining, and freeze-fracture electron microscopy. In isolated triads, key features of the structure observed in situ have been retained, including the osmiophilic "feet," junctional structures between the transverse tubule and terminal cisternae. New insight into triad structure is obtained by negative staining, which also enables visualization of feet at the junctional face of the terminal cisternae, whereas smaller surface particles, characteristic of calcium pump protein, are not visualized there. Therefore, the junctional face is different from the remainder of the sarcoplasmic reticulum membrane. Junctional feet as viewed by thin section or negative staining have similar periodicity and extend approximately 100 A from the surface of the membrane. Freeze-fracture of isolated triads reveals blocklike structures associated with the membrane of the terminal cisternae at the junctional face, interjunctional connections between the terminal cisternae and t-tubule, and intragap particles. The intragap particles can be observed to be closely associated with the t-tubule. The structure of isolated triads is susceptible to osmotic and salt perturbation, and examples are given regarding differential effects on transverse tubules and terminal cisternae. Conditions that adversely affect morphology must be considered in experimentation with triads as well as in their preparation and handling.

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New Evidence for Membrane Specialization in the Diatom Thalassiosira Antarctica Comber

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s1431927600002737 ◽

1980 ◽

Vol 38 ◽

pp. 624-625

Author(s):

Gregory J. Doucette

Keyword(s):

Electron Microscopy ◽

Thin Section ◽

Freeze Fracture ◽

Marine Diatom ◽

Supplementary Data ◽

Ultra Structure ◽

Membrane Specialization ◽

Freeze Fracture Electron Microscopy ◽

New Evidence ◽

Fracture Electron

The present investigation was undertaken to examine the nonsiliceous ultra-structure of diatoms (BacilIariophyta) by means of freeze fracture electron microscopy. Freeze fracture procedures are complicated by the difficulties encountered in fracturing silica-based components and the removal of these materials subsequent to replica casting. Supplementary data was obtained through conventional thin section methodologies. This report is a prelim¬inary account of observations made on selected nonsiliceous, cellular con¬stituents of Thalassiosira antarctica Comber, a planktonic, marine diatom.

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Analysis of freeze-fracture electron micrographs by a computer-based technique

Biophysical Journal ◽

10.1016/s0006-3495(76)85716-5 ◽

1976 ◽

Vol 16 (6) ◽

pp. 613-625 ◽

Cited By ~ 13

Author(s):

R.J. Mehlhorn ◽

L. Packer

Keyword(s):

Electron Micrographs ◽

Freeze Fracture ◽

Computer Based ◽

Fracture Electron

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Coalescence of microsomal vesicles from rat liver: a phenomenon occurring in parallel with enhancement of the glycosylation activity during incubation of stripped rough microsomes with GTP.

The Journal of Cell Biology ◽

10.1083/jcb.86.1.29 ◽

1980 ◽

Vol 86 (1) ◽

pp. 29-37 ◽

Cited By ~ 32

Author(s):

J Paiement ◽

H Beaufay ◽

D Godelaine

Keyword(s):

Electron Microscopy ◽

Thin Section ◽

Rat Liver ◽

Freeze Fracture ◽

Endogenous Protein ◽

Previously Treated ◽

Nucleotide Sugars ◽

Freeze Fracture Electron Microscopy ◽

Fracture Electron

Rough microsomes from rat liver have been subjected to various treatments and incubated afterwards with UDP-N-acetyl-[14C]glucosamine and GDP-mannose in the presence of GTP (0.5 mM), or of other nucleotides. In agreement with earlier results from this laboratory, the preparations previously treated to strip off the ribosomes and incubated in the presence of GTP assembled dolichol-linked oligosaccharides and transferred these oligosaccharides to endogenous protein acceptors much more actively than untreated preparations, or stripped preparations incubated in the absence of GTP. Thin-section and freeze-fracture electron microscopy have revealed that pyrophosphate-treated preparations incubated with GTP are aggregated and contain numerous vesicles as large as 1-4 micrometer, or more. Such large vesicles were not present before incubation and thus were considered to have been formed through coalescence of regular-sized ones. Like glycosylation, the coalescence phenomenon depends upon the removal of ribosomes, because it occurred whether ribosomes had been stripped, at least partly, with pyrophosphate, KCl, or puromycin, but not when rough microsomes had been washed with 0.25 M sucrose or with KCl and MgCl2. Like glycosylation, it also depends on the addition of GTP and was not induced by ATP, UTP, CTP, and nonhydrolysable analogues of GTP. Rough microsomes coalesced, however, when pyrophosphate-treated preparations were incubated with GTP in the absence of nucleotide sugars, or in the presence f tunicamycin, indicating that the coalescence phenomenon does not result from the glycosylation of some membrane constituents.

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