Journal of ultrastructure research最新文献

Tubular smooth endoplasmic reticulum membranes have been found in the cytoplasm ofBlastocladiella emersonii, Blastocladiella britannica, andCatenaria anguillulae. These structures are formed and accumulate during the growth phase and are associated with cisternae of rough endoplasmic reticulum during sporangium formation. It is within the cisternae of rough of endoplasmic reticulum that the protein precursors of gamma particles are formed. It is probable that the tubular endomembranes represent a reservoir of membrane material that is formed during the growth phase and is converted into other endomembranes during sporogenesis.

Between birth and 10 days of age, the volume density (volume/unit cytoplasmic volume) of the matrix, and the surface density (area/unit cytoplasmic volume) of the inner membrane and cristae increased in both periportal and perihepatic hepatocytes, and did not differ significantly between the cells of the two zones. After 10 days of age, however, the volume density of the matrix decreased in perihepatic cells and remained unchanged in periportal cells, and, therefore, it became greater in periportal cells than in perihepatic cells in 20-day-old and adult animals. The surface density of the inner membrane and cristae decreased in the cells of both zones. Further, the hepatocyte volume increased markedly, especially in perihepatic zones between 20 days of age and the adult. The results show that, in postnatally differentiating hepatocytes, mitochondria are likely to develop during early postnatal period, then the structural heterogeneity of mitochondria arises, and hepatocyte volume increases markedly during late postnatal period after weaning. Thus, the process of postnatal hepatocyte differentiation includes such several phases of development.

Thin sectioning and freeze-fracture electron microscopy have been used to show that it is possible to obtain topologically closed vesicles by means of reconstitution of rat liver microsomal membrane “ghosts.” The reconstitution by 15 hr dialysis resulted in the formation of vesicles with intra-membrane particles (IMP) while after 40 hr dialysis no IMP were observed in the membranes. The protein/lipid ratio and functional activity of NADPH- and NADH-linked enzyme systems were similar in both cases. CytochromeP-450 (LM₂) was incorporated into liposomes of different composition (protein : lipid ratio — 1:200). IMP were observed only when the incorporation of cytochrome P-450 was performed in the presence of detergent Emulgen 913 as specific additive to the initial protein-lipid-sodium cholate mixture or in the course of incubation of proteoliposomal suspensions at 37°C. After the incorporation of cytochromeb₅ into azolectin liposomes vesicular membranes contain IMP if the incorporated membrane protein: lipid ratio is at least 1:50. Pronase-induced splitting off of a 11 kDa heme-containing fragment of cytochromeb₅ did not affect IMP content. The conditions of IMP formation in reconstituted membranes and in microsomal ghosts are discussed.

The validity of a method for localization of potential Ca²⁺ binding sites was analyzed in pollen tubes ofLilium longiflorum and calyptra cells ofZea mays. The Ca²⁺-enriched fixation produced electron opaque deposits mainly on the plasma membrane, endoplasmic reticulum, and mitochondria, with details of localization differing somewhat between pollen tubes and maize calyptra cells. Proton microprobe analysis indicated a relative enrichment of Ca in the sections. The newly introduced technique of electron spectroscopic imaging confirmed the presence of Ca (and possibly P) in the induced electron opaque structures.

The mechanical processes occurring around a cutting edge when a thin slice is formed are related to those effective when metal or plastic is lathed. Critically assigning to thin slicing of resins the force vectors proved or assumed to be true for shapping by chipping, the attempt is made to clarify qualitatively and quantitatively the complex interplay after having measured the effective weight force and the order of magnitude of the radius of cutting edges of glass and diamond knives. Practically, only five force vectors in a polygon determine shearing and shortening of chips or thin slices, shortening obeying a broken rational function. By computing the relative values of shortening, all other force vectors can be computed. The results obtained for the normal force on the edge surface and the tangential force in this surface shed light on the effect of systematical variation of setting angle, bevel angle, and chip angle for thin slicing; thus possibilities of improvement in conventional “ultra” microtomy and in unadulterated cryotomy are shown in concluding recommendations.

Treatment of purified preparations of Na,K-ATPase by phospholipase A₂ has led to the formation of two-dimensional crystals of the protein. Control tests with another phospholipase and two detergents have shown that crystallization occurs as the result of hydrolysis and/or solubilization of the phospholipids in the enzyme vesicles. Experimentation with various buffer systems has indicated that reduction in the amount of phospholipids alone is sufficient for inducing the formation of crystalline sheets. Inclusion of crystal inducing ions in the buffer facilitates the crystallization process, resulting in more extensive arrays. The new crystalline sheets are exclusively dimeric with average unit cell dimensions:a = 15.8 ± 0.4nm, b = 4.9 ± 0.2nm, and γ = 64 ± 3°. Examination of the micrographs shows that the initial intermolecular interaction leading to the formation of sheets is between the α subunits. Results from this study suggest that removal and/or modification of phospholipids by phospholipases could prove successful in crystallizing those membrane proteins in which excess lipid is the main barrier to the formation of two-dimensional arrays.

The effects of taxol on microtubule-associated proteins of high molecular weight (MAPs) were studiedin vitro. After negative staining, microtubules reconstituted in the presence of taxol from preparations of partially purified tubulin and MAPs, besides bundled, displayed prominent elongated or globular extensions without apparent regularity. These extensions, but not the tubulin polymer, were heavily decorated after immuno-gold-labeling using antibodies to MAP-1 and MAP-2. Microtubules reconstituted in the absence of taxol showed a much more regular, and apparently helical, arrangement of MAPs along their surfaces. The formation of polymeric structures was also observed when preparation of MAPs free of tubulin were incubated with taxol. In this case in addition to large network-type aggregates with little apparent substructure, more regular structures seemingly consisting of approximately 5-nm-thick filaments arrayed in parallel were observed. Taxol-induced MAP aggregation occurred rapidly and was directly proportional to the concentration of protein, as revealed by optical density measurements. It is concluded that taxol, aside from promoting the assembly of tubulin and stabilizing microtubules, promotes MAP/MAP interaction.

Suspension culture cells of carrot,Daucus carota L., and sycamore,Acer pseudoplatanus L., were freeze-fractured after ultrarapid freezing without fixation or cryoprotection in a propane-jet freezer. Infrequently, rosettes (ca. 24 nm diameter) of six (occasionally five) subunits (ca. 8 nm diameter) were observed in P-face views of the plasma membrane of both taxa. When present, rosette density was approximately 1/μm². Generally, rosettes were less frequently seen on plasma membranes exhibiting numerous vesicle fusion figures. Due to the high quality of the freezing, cellulose microfibril impressions were rarely seen on either PF or EF views of the plasma membrane, thus precluding correlations between microfibrils on the one hand and rosettes (and terminal globules) on the other. The presence of rosettes in suspension culture cells of these two species supports the putative role of rosettes in cellulose biosynthesis in higher plants.