The ultrastructure of the erythrocyte cytoskeleton at neutral and reduced pH

Timme, A. H.

doi:10.1016/s0022-5320(81)80041-x

Cited by 19 publications

(12 citation statements)

References 16 publications

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“…This array exhibited a strong resemblance to that seen by scanning electron microscopy of RBC ghosts. In one study (Hainfeld & Steck, 1977), the cells were lysed osmotically, fixed and freeze dried prior to examination; in another (Timme, 1981), the ghosts were exposed to low pH, which induced aggregation of the spectrin. AFM images of ghosts on polylysine treated substrates were particularly likely to have such very coarse features.…”

Section: Resultssupporting

confidence: 92%

Atomic force microscopy of the erythrocyte membrane skeleton

Swihart

Mikrut

Ketterson

et al. 2001

Journal of Microscopy

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SummaryThe atomic force microscope was used to examine the cytoplasmic surface of untreated as well as fixed human erythrocyte membranes that had been continuously maintained under aqueous solutions. To assess the effects of drying, some membranes were examined in air. Erythrocytes attached to mica or glass were sheared open with a stream of isotonic buffer, which allowed access to the cytoplasmic membrane face without exposing cells to non-physiological ionic strength solutions. Under these conditions of examination, the unfixed cytoplasmic membrane face revealed an irregular meshwork that appeared to be a mixture largely of triangular and rectilinear openings with mesh sizes that varied from 35 to 100 nm, although few were at the upper limit. Fixed ghosts were similar, but slightly more contracted. These features represent the membrane skeleton, as when the ghosts were treated to extract spectrin and actin, these meshworks were largely removed. Direct measurements of the thickness of the membrane skeleton and of the lateral dimensions of features in the images suggested that, especially when air dried, spectrin can cluster into large, quite regularly distributed aggregates. Aggregation of cytoskeletal components was also favoured when the cells were attached to a polylysine-treated substrate. In contrast, the membrane skeletons of cells attached to substrates rendered positively charged by chemical derivatization with a cationic silane were much more resistant to aggregation. As steps were taken to reduce the possibility of change of the skeleton after opening the cells, the aggregates and voids were eliminated, and the observed structures became shorter and thinner. Ghosts treated with Triton X-100 solutions to remove the bilayer revealed a meshwork having aggregated components resembling those seen in air. These findings support the proposition that the end-to-end distance of spectrin tetramers in the cell in the equilibrium state is much shorter than the contour length of the molecule and that substantial rearrangements of the spectrin-actin network occur when it is expanded by low ionic strength extraction from the cell. This study demonstrates the applicability of AFM for imaging the erythrocyte membrane skeleton at a resolution that appears adequate to identify major components of the membrane skeleton under near-physiological conditions.

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Section: Resultssupporting

confidence: 92%

Atomic force microscopy of the erythrocyte membrane skeleton

Swihart

Mikrut

Ketterson

et al. 2001

Journal of Microscopy

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“…In this Letter, we report that after crosslinking of the VI polypeptide a filamentous network appears after Triton X-100 extraction of ACh-receptor rich membranes isolated from Torpedo marmorata electric organ. This structure strikingly resembles the skeleton recently described in the erythrocyte membrane [29,30].…”

Section: Introductionsupporting

confidence: 81%

“…Finally, it is worth mentioning that in the red blood cells a cytoskeleton membrane complex which includes a limited number of interconnected proteins, actin [45] in particular, displays structural features comparable to the network described in the present paper [29,30]. their helpful advice, Mrs. M.A.…”

Section: Discussionmentioning

confidence: 61%

Evidence for a skeleton in acetylcholine receptor‐rich membranes from Torpedo marmorata electric organ

Cartaud¹,

Oswald²,

Clément³

et al. 1982

FEBS Letters

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“…Earlier electron microscopic studies of the membrane skeleton were complicated by the high density of the native skeleton. Using various techniques of specimen preparation, such as negative staining 03, 39,47), thin sectioning (40,41), shadowing (30), rapid freezing (12, 27), and scanning electron microscopy (17), these studies revealed only a filamentous weblike structure containing a variety of poorly defined elements of various sizes. Recently, however, the resolution was drastically improved when the membrane skeleton was either fragmented or artificially spread and then examined by negative staining electron microscopy.…”

mentioning

confidence: 99%

Visualization of the hexagonal lattice in the erythrocyte membrane skeleton.

Liu¹,

Derick²,

Palek³

1987

The Journal of Cell Biology

375

276

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Abstract. The isolated membrane skeleton of human erythrocytes was studied by high resolution negative staining electron microscopy. When the skeletal meshwork is spread onto a thin carbon film, clear images of a primarily hexagonal lattice of junctional F-actin complexes crosslinked by spectrin filaments are obtained. The regularly ordered network extends over the entire membrane skeleton. Some of the junctional complexes are arranged in the form of pentagons and septagons, •3 and 8 %, respectively. At least five forms of spectrin crosslinks are detected in the spread skeleton including (a) a single spectrin tetramer linking two junctional complexes, (b) three-armed Y-shaped spectrin molecules linking three junctional complexes, (c) three-armed spectrin molecules connecting two junctional complexes with two arms bound to one complex and the third arm bound to the adjacent complex, (d) double spectrin filaments linking two junctional complexes, and (e) four-armed spectrin molecules linking two junctional complexes. Of these, the crosslinks of single spectrin tetramers and three-armed molecules are the most abundant and represent 84 and 11% of the total crosslinks, respectively. These observations are compatible with the presence of spectrin tetramers and oligomers in the erythrocyte membrane skeleton. Globular structures (9-12 nm in diameter) are attached to the majority of the spectrin tetramers or higher order oligomer-like molecules, "080 nm from the distal ends of the spectrin tetramers. These globular structures are ankyrinor ankyrin/band 3-containing complexes, since they are absent when ankyrin and residual band 3 are extracted from the skeleton under hypertonic conditions.

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The ultrastructure of the erythrocyte cytoskeleton at neutral and reduced pH

Cited by 19 publications

References 16 publications

Atomic force microscopy of the erythrocyte membrane skeleton

Atomic force microscopy of the erythrocyte membrane skeleton

Evidence for a skeleton in acetylcholine receptor‐rich membranes from Torpedo marmorata electric organ

Visualization of the hexagonal lattice in the erythrocyte membrane skeleton.

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