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J Heuser

Publications and source records attributed to J Heuser.

65 records · Page 4Linked to original sources
PubMed1980

Three-dimensional visualization of coated vesicle formation in fibroblasts.

Fibroblasts apparently ingest low density lipoproteins (LDL) by a selective mechanism of receptor-mediated endocytosis involving the formation of coated vesicles from the plasma membrane. However, it is not known exactly how coated vesicles collect LDL receptors and pinch off from the plasma membrane. In this report, the quick-freeze, deep-etch, rotary-replication method has been applied to fibroblasts; it displays with unusual clarity the coats that appear under the plasma membrane at the start of receptor-mediated endocytosis. These coats appear to be polygonal networks of 7-nm strands or struts arranged into 30-nm polygons, most of which are hexagons but some of which are 5- and 7-sided rings. The proportion of pentagons in each network increases as the coated area of the plasma membrane puckers up from its planar configuration (where the network is mostly hexagons) to its most sharply curved condition as a pinched-off coated vesicle. Coats around the smallest vesicles (which are icosahedrons of hexagons and pentagons) appear only slightly different from "empty coats" purified from homogenized brain, which are less symmetrical baskets containing more pentagons than hexagons. A search for structural intermediates in this coat transformation allows a test of T. Kanaseki and K. Kadota's (1969. J. Cell Biol. 42:202--220.) original idea that an internal rearrangement in this basketwork from hexagons to pentagons could "power" coated vesicle formation. The most noteworthy variations in the typical hexagonal honeycomb are focal juxtapositions of 5- and 7-sided polygons at points of partial contraction and curvature in the basketwork. These appear to precede complete contraction into individual pentagons completely surrounded by hexagons, which is the pattern that characterizes the final spherical baskets around coated vesicles.

Animals↗
PubMed1980

The vitelline layer of the sea urchin egg and its modification during fertilization. A freeze-fracture study using quick-freezing and deep-etching.

Eggs of the sea urchin Strongylocentrotus purpuratus were quick-frozen, freeze fractured, and deep-etched to reveal the detailed structure of the vitelline layer (VL), an extracellular coat. The VL consisted of a network of fibers lying in sheet raised 20 nm off the plasma membrane and connected to it by a series of short processes. Sperm attached to the fibers of this sheet and upon fertilization the VL rose off the egg surface to form the fertilization envelope (FE). By 1 min postinsemination (p.i.), the FE had become augmented by a new set of smaller fibrils, and the original fibers of the VL appeared to be undergoing degradation. The FE exhibited casts of microvilli the VL had once covered. These were rounded at 1 min p.i., but by 2 min they had become angular and coated with an orderly array of repeating macromolecular units. In areas between casts, the coating process was slower; incomplete rows of units were seen at 5 min p.i. and complete rows at 10 min. Deep-etching of FE isolated from eggs by homogenization and differential centrifugation showed that both top and bottom surfaces were coated. The coat pattern was made up of 17.5-nm wide rows of parallelogram-like units that repeated every 12.2 nm along the row axis. Units in adjacent rows were in register to produce a secondary axis 76 degrees from the row axis. The results of this and previous studies suggest that the coating process plays a major role in "hardening" the FE to produce a tough barrier that protects the early embryo from chemical and mechanical injury.

Animals↗
PubMed1979

Membrane fusion during secretion: cortical granule exocytosis in sex urchin eggs as studied by quick-freezing and freeze-fracture.

Exocytosis of cortical granules was observed in sea urchin eggs, either quick-frozen or chemically fixed after exposure to sperm. Fertilization produced a wave of exocytosis that began within 20 s and swept across the egg surface in the following 30 s. The front of this wave was marked by fusion of single granules at well-separated sites. Toward the rear of the wave, granule fusion became so abundant that the egg surface left with confluent patches of granule membrane. The resulting redundancy of the egg surface was accommodated by elaboration of characteristic branching microvilli, and by an intense burst of coated vesicle formation at approximately 2 min after insemination. Freeze-fracture replicas of eggs fixed with glutaraldehyde and soaked in glycerol before freezing displayed forms of granule membrane interaction with the plasma membrane which looked like what other investigators have considered to be intermediates in exocytosis. These were small disks of membrane contact or membrane fusion, which often occurred in multiple sites on one granule and also between adjacent granules. However, such membrane interactions were never found in eggs that were quick-frozen fixation, or in eggs fixed and frozen without exposure to glycerol. Glycerination of fixed material appeared to be the important variable; more concentrated glycerol produced a greater abundance of such "intermediates." Thus, these structures may be artifacts produced by dehydrating chemically fixed membranes, and may not be directly relevant to the mechanism by which membranes naturally fuse.

Animals↗
PubMedDate not supplied

Deep-etch views of clathrin assemblies.

Clathrin assemblies were adsorbed to mica and freeze-dried by a new procedure that yields 3-D images with much topological detail. These permitted renewed inquiry into how clathrin trimers (i.e. "triskelions") assemble into polygonal coats or baskets. Freeze-drying revealed unsuspected differences in the relative shapes and dimensions of individual trimer building blocks, as compared with the completed polygonal networks, which indicate that the assembly scheme first proposed by Crowther and Peare (1) requires modification. Specifically, the freeze-etch images display the following new features: (1) Trimer arms possess terminal scroll-shaped hooks that can open or close and thus determine their lengths. (2) When extended, trimer arms are sufficiently long to pass around three sides of the final polygonal facet. Since current views hold that the arms pass around only two sides, the remaining length, including the terminal hook, must point into the basket interior. (3) Freeze-dried trimers display bends in their arms at specific loci that determine their final distribution in the completed baskets. (4) The completed struts of the final assemblies are uniformed in the calibre, cylindrical in profile, and travel directly between the vertices of each polygon, without any sign of the slew or width-variation that is predicted by the Crowther and Pearse model. Based on this direct comparison of promoter vs product, by a single technique that can image both, we offer a modified scheme for clathrin coat assembly, in which we predict that the individual arms in each clathrin triskelion emanate from its center in a slewed manner, but the final assembled struts of the basket need not be slewed. Attempts were made to capture assembly intermediates on mica to obtain support for the scheme, but these unfortunately yielded ambiguous images of incomplete polygons with blunt projections, rather than the expected "halo" of uncommitted trimer arms. These we interpret to be "dead ends" that failed to polymerize further because they included proteolyzed components. Further assembly experiments, avoiding such hazards, are indicated.

Aluminum Silicates↗