Regular Substructure Research Articles

Phosphorylated paxillin and phosphorylated FAK constitute subregions within focal adhesions.

Integrin-mediated adhesions are convergence points for multiple signaling pathways. Their inner structure and diverse functions can be studied with super-resolution microscopy. Here, we examined the spatial organization within focal adhesions by analyzing several adhesion proteins with structured illumination microscopy (SIM). Paxillin (Pax) serves as a scaffold protein and signaling hub in focal adhesions, and focal adhesion kinase (FAK, also known as PTK2) regulates the dynamics of adhesions. We found that their phosphorylated forms, pPax and pFAK, form spot-like, spatially defined clusters within adhesions in several cell lines and confirmed these findings with additional super-resolution techniques. These clusters showed a more regular separation from each other compared with more randomly distributed signals for FAK or paxillin. Mutational analysis indicated that the active (open) FAK conformation is a prerequisite for the pattern formation of pFAK. Live-cell super-resolution imaging revealed that organization in clusters is preserved over time for FAK constructs; however, distance between clusters is dynamic for FAK, while paxillin is more stable. Combined, these data introduce spatial clusters of pPax and pFAK as substructures in adhesions and highlight the relevance of paxillin-FAK binding for establishing a regular substructure in focal adhesions.

Stick-slip substructure in rapid tape peeling

The peeling of adhesive tape is known to proceed with a stick-slip mechanism and produces a characteristic ripping sound. The peeling also produces light and when peeled in a vacuum, even X-rays have been observed, whose emissions are correlated with the slip events. Here we present direct imaging of the detachment zone when Scotch tape is peeled off at high speed from a solid surface, revealing a highly regular substructure, during the slip phase. The typical 4-mm-long slip region has a regular substructure of transverse 220 μm wide slip bands, which fracture sideways at speeds over 300 m/s. The fracture tip emits waves into the detached section of the tape at ∼ 100 m/s, which promotes the sound, so characteristic of this phenomenon.

Structural characterisation of protein-caged iron minerals in biological systems

In this paper we have collected multiple high angle annular dark field (HAADF) images of purified horse spleen ferritin mineral cores with an aberration corrected STEM (SuperSTEM). The images of individual cores have been separated into groups to determine characteristic views, and averaged to reduce the noise. We have found that these isolated cores do not have the same clear substructure as ferritin cores in plastic-embedded human liver sections (from a patient with haemochromatosis). There is a clear discrepancy between the in situ human liver and purified horse spleen ferritin, but also between Massover's observation of regular substructure in purified horse spleen ferritin by cryo-electron microscopy [1] and these results. A possible basis for this discrepancy is the difference in purification methods used between the two horse spleen ferritin preparations, with the one here purified using CdSO4 precipitation, and the one used by Massover purified without the use of CdSO4.

Some Known Results and an Open Problem of Tree - Wheel Graph Ramsey Numbers

There are many famous problems on finding a regular substructure in a sufficiently large combinatorial structure, one of them i.e. Ramsey numbers. In this paper we list some known results and an open problem on graph Ramsey numbers. In the special cases, we list to determine graph Ramsey numbers for trees versus wheels

Regular Substructure Inhomogeneity of Textured Materials by the Example of Rolled Ti-Ni Single Crystals

The substructure inhomogeneity of rolled Ti-48%Ni-2%Fe single crystals was studied by the X-ray method of Generalized Pole Figures. Under rolling of single crystals, along with texture formation, the nonuniform substructure develops. It is conditioned by inhomogeneous reorientation of the crystalline lattice and local differences in predominant plastic deformation mechanisms. The distribution of residual deformation effects is controlled by the orientation of separate regions relative to texture maxima. Rolled single crystals prove to be divided into subcomponents with close orientations and opposite signs of elastic microstrains.

Biochemical and Structural Analysis of Isolated Mature Cores of Human Immunodeficiency Virus Type 1

Mature human immunodeficiency virus type 1 (HIV-1) particles contain a cone-shaped core structure consisting of the internal ribonucleoprotein complex encased in a proteinaceous shell derived from the viral capsid protein. Because of their very low stability after membrane removal, HIV-1 cores have not been purified in quantities sufficient for structural and biochemical analysis. Based on our in vitro assembly experiments, we have developed a novel method for isolation of intact mature HIV-1 cores. Concentrated virus suspensions were briefly treated with nonionic detergent and immediately centrifuged in a microcentrifuge for short periods of time. The resuspended pellet was subsequently analyzed by negative-stain and thin-section electron microscopy and by immunoelectron microscopy. Abundant cone-shaped cores as well as tubular and aberrant structures were observed. Stereo images showed that core structures preserved their three-dimensional architecture and exhibited a regular substructure. Detailed analysis of 155 cores revealed an average length of ca. 103 nm, an average diameter at the base of ca. 52 nm, and an average angle of 21.3 degrees. There was significant variability in all parameters, indicating that HIV cores are not homogeneous. Immunoblot analysis of core preparations allowed semiquantitative estimation of the relative amounts of viral and cellular proteins inside the HIV-1 core, yielding a model for the topology of various proteins inside the virion.

Separation of Scrapie Prion Infectivity from PrP Amyloid Polymers

The prion protein (PrP) undergoes a profound conformational change when the cellular isoform (PrP C) is converted into the scrapie form (PrP Sc). Limited proteolysis of PrP Scproduces PrP 27 – 30 which readily polymerizes into amyloid. To study the structure of PrP amyloid, we employed organic solvents that perturb protein conformation. Hexafluoro-2-propanol (HFIP), which promotes α-helix formation, modified the ultrastructure of rod-shaped PrP amyloids; flattened ribbons with a more regular substructure were found. As the concentration of HFIP was increased, the β-sheet content and proteinase K resistance of PrP 27-30 as well as prion infectivity diminished. HFIP reversibly decreased the binding of Congo red dye to the rods while inactivation of prion infectivity was irreversible. In contrast to 10% HFIP, 1,1,1-trifluoro-2-propanol (TFIP) did not inactivate prion infectivity but like HFIP, TFIP did alter the morphology of the rods and abolish Congo red binding. This study separates prion infectivity from the amyloid properties of PrP 27-30 and underscores the dependence of prion infectivity on PrP Scconformation. The results also demonstrate that the specific β-sheet-rich structures required for prion infectivity can be differentiated from those needed for amyloid formation as determined by Congo red binding.

On combinatorial structures of line drawings of polyhedra

Combinatorial structures of line drawings of polyhedra were recently clarified by Sugihara in terms of the regularity of line drawings (references [6,8] in the paper). The definition of the regularity of drawings is quite similar to that mf matroids, and we here clarify both matroidal aspects and non-matroidal ones of the regularity of line drawings, especially concerning the problem of finding a maximum regular substructure of line drawing. A new class of regular line drawings and an algorithm for finding a maximal regular substructure are also discussed.

Ferritin in plastids of Nicotiana alata pollen grains

In the present paper the presence of ferritin (phytoferritin) aggregates inside the plastids of mature Nicotiana alata pollen grains is reported. Phytoferritin forms a dark mass which sometimes possesses a regular substructure with a crystalline appearance. The presence of phytoferritin aggregates could be related to the hydration and activation of pollen grains and subsequent pollen tube emission.

The polar ring of coccidian sporozoites: a unique microtubule-organizing centre.

The infective stages, or 'zoites, of coccidian parasites possess an organized network of spirally arranged microtubules that closely follow the helical body shape of these vermiform cells. These subpellicular microtubules are anchored anteriorly by insertion into a highly structured circular microtubule-organizing centre (MTOC) known as the polar ring. This MTOC has been examined both in situ and in isolated, critical-point-dried whole cytoskeletons. The 24 microtubules attach laterally to the MTOC through shallow depressions on the inner face of the ring: the ends do not appear to be physically capped. The polar ring has no obvious or regular substructure, although it has a faintly fibrous appearance. The polarity of the microtubules, determined by 'hook decoration', is such that the plus or fast-growing end is distal to the MTOC. The coccidian 'zoite MTOC is unique both in its highly defined structure and in the degree of organization it confers upon the developing cell in terms of the number, spacing, orientation and polarity of the subpellicular microtubules.

The myosin filament: VII. Changes in internal structure along the length of the filament

Improved fixation procedures have enabled substructure to be observed by electron microscopy in transverse sections of vertebrate skeletal muscle thick filaments as thin as 140 nm. Optical diffraction combined with digital autocorrelation analysis, focal series and tilting experiments have confirmed the presence of a regular substructure having a repeat near 4 nm and shown that it is highly unlikely to be an artifact associated with the electron microscope imaging system. The results obtained strongly suggest that the thick filament is constructed from a bundle of rod-like subfilaments arranged parallel to the thick filament axis to within less than a degree. This cannot easily be reconciled with the general theory of thick filament structure proposed by Squire (1973), but it is consistent with the model proposed by Pepe (1966,1967). Optical diffraction of 140 nm thick serial transverse sections has also suggested a structural change along the length of the filament that is manifest by a variation in the proportion of filaments showing strong diffraction maxima in one, two or three directions.

A comparison of the periodic substructure of the trichocysts of the cryptophyceae and prasinophyceae

A study of the trichocysts of the Cryptophyceae and Prasinophyceae has confirmed the general concept of their structures as advanced by Hovasse, Mignot, and Joyon (1967) and Manton (1968) respectively. In Cryptomonas 58 a staining method using ruthenium red reveals a uniform and regular substructure within the fabric of the trichocyst ribbon. A finer and less regular substructure was observed in the trichocyst of Pyramimonas parkeae.

THE ULTRASTRUCTURE OF CULTIVABLE TREPONEMES.

Treponema phagedenis, Treponema vincentii, and Treponema refringens were studied in the electron microscope by means of negative staining and sectioning techniques. The length and width of the cells were within the same limits for all three strains examined. Statistical analysis, however, demonstrated a difference in the wavelengths of the cells. Organisms of T. phagedenis and T. vincentii had blunt ends and possessed cell wall surface layers in which a regular substructure was only occasionally revealed, while cells of T. refringens had tapered ends and a distinctly structured cell wall surface layer. Generally, cells of the strains investigated had 4–6 flagella inserted at each end. Two bundles of flagella, one from each end of the cell, wound around the organism and overlapped in the middle of the cell. The flagella with their insertion organelles were identical for the three strains studied. Two bundles of cytoplasmic tubules were detected in the interior of the treponemes after treatment with either sodium deoxycholate or Myxobacter AL‐1 protease 1. The two bundles of cytoplasmic tubules overlapped in the middle of the cell. The results of the present morphological study are compared with results obtained by other investigators studying other characteristics of these strains and it is concluded that the three strains studied belong to three different species.

Absence of an electron microscopic substructure in intact flagella of Proteus mirabilis and Bacillus subtilis

Flagella of Proteus mirabilis D 52 appeared without any regular substructure in close to focus, high resolution electron micrographs of specimens negatively contrasted with potassium phosphotungstate or uranyl acetate. The same was true for many flagella of Bacillus subtilis ATCC 6051, but frequently these flagella exhibited a longitudinal linear pattern, interpretable as a bundle of about 3 nm-thick subfibrils. However, this pattern was probably caused by a slight degradation of the flagella, since the lined specimens were about 30% wider than the structureless ones. Moreover the Bacillus flagella were easily lost when washed with water or buffer. As controls, close to focus micrographs of mixtures of flagella and T6 bacteriophages were taken. The cross striations of the phage tails were clearly seen, spaced at 4 nm.

Chromatin sub-structure. The digestion of chromatin DNA at regularly spaced sites by a nuclear deoxyribonuclease

Evidence is presented that indicates a regular sub-structure in nuclear nucleoprotein with regularly distributed sites that are specifically susceptible to the cellular Ca-Mg endonuclease.

Correlation between the sub-structure of the luminal plasma membrane of the bladder in man and other mammals

The luminal plasma membrane of the mammalian urinary bladder possesses a sub-structure consisting of a hexagonal lattice of sub-units which has previously been observed in rat and mouse by means of negative-staining and in rabbit both by negative-staining and freeze-etching. In the present work, the same regular sub-structure has been found in various other mammals including man. By the use of two independent optical techniques (linear integration and optical filtering) it has also been demonstrated that the similarity in structure even extends to that of the individual sub-units which at high resolution can be shown to exhibit a dodecameric pattern. The correlation between the structure of the membrane lining the bladder in man and other mammals indicates that, contrary to some previous reports, the bladders of the other mammals may be satisfactory models for studying the morphology and physiology of this region in man. Le plasma de la membrane luminaire de la vessie des mammifères possède une sous-structure consistant d'un réseau hexagonal de sous-unités observées au préalable chez le rat et la souris par coloration négative et attaque par congélation localisée. La même sous-structure a été observée chez différents autres ammifères, y copris l'homme, dns les travaux présentés cidessous. La similarité de structure s'éntendant même jusqu'au niveau de la sous-unité individuelle qui possède un motif dodécamérique lorsque la résolution est suffisament élevée, a été observée par deux techniques optiques indépendantes (intégration linéaire et filtrage optique). La corrélation entre la structure de la membrane recouvrant la vessie chez l'homme et d'autres animaux indique que contrairement à certaines publications antérieures, la vessie chez d'autres animaux peut être considérée comme un modèle satisfaisant pour des études morphologiques et physiologiques de cette partie du corps humain. Plasmamembrane der Innenseite von Säugetierharnblasen weisen eine sechsseitige Mikrogitterstruktur auf. Diese Struktur ist schon in der Ratte und in der Maus mittels Negativfärbung und im Kaninchen mittels Negativfärbung und Gefrierätzung dargestellt worden. Im Laufe dieser Untersuchungsreihe fand man die gleiche regelmäßige Struktur in verschiedenen anderen Säugetieren und auch beim Menschen. Mit zwei voneinander unabhängigen optischen Verfahren (Linearintegration und optischer Filterung) konnte gezeigt werden, daß sich die Ähnlichkeit auch auf die einzelnen Untereinheiten erstreckt, welche bei hoher Auflösung ein zwölfgliedriges Muster aufweisen. Die Ähnlichkeit in den Membranstrukturen des Blaseninneren des Menschen und anderer Säugetiere deutet darauf hin, daß Säugerharnblasen befriedigende Vorlagen für da Studium der Morphologie und Physiologie dieser Zone im Menschen sein können, obwohl dies in früheren Arbeiten bezweifelt wurde.

Extremembranous deposits and "membranous" glomerulonephritis. Their fine structure, development, and correlation.

In 15 renal puncture biopsies from 14 patients, electron microscopy revealed extramembranous deposits (EMD) in various developmental stages (or their residues) in glomerular capillaries. Florid EMD exhibited a regular granular substructure as well as a typical localization within the subpodocytal space. Ultrastructurally, a gradual regression of EMD and their incorporation into the basement membrane could be observed. Early florid EMD were found in acute glomerulonephritis or in the nephrotic syndrome persisting for several months, with only discrete to absent “membranous” changes with light microscopy. Late stages of EMD with predominantly degenerative changes of glomerular basement membranes and apparent “membranous” thickening in light microscopy, on the other hand, were characteristic of the nephrotic syndrome lasting for several years. At early stages of EMD, long-lasting remission up to clinical healing were observed in a half of the cases studied, whereas at a late “membranous” stage long-lasting remission took place only once in six cases.

Structure and formation of pores in fenestrated capillaries

The pores of the fenestrated capillary endothelium of rat kidney are spanned by a diaphragm which has a regular substructure. This was established by the comparison of conventionally fixed and sectioned material with replicas obtained by the freeze-etching method. The substructure of the diaphragm consists of two centrally located fibrous rings which are connected to the pore rim by radiating fibers. Additional evidence for such a structural arrangement was found in forming pores where the membrane differentiated into structures comparable to the ones in completed pores. There is some indication that the endothelial pores are octagonal in outline. A dynamic concept of pore formation is presented. The observed structural arrangement of the diaphragm or central density of the pores may account for the “small pores” as required by the “pore theory” of capillary permeability and for rapid passage of peroxidase molecules, but a purely structural function cannot be excluded.

The fine structure of the testicular interstitial cells in men of normal androgenic status.

The ultrastructural features of the testicular interstitial cells were examined in a group of men of normal androgenic status. In osmium-fixed material three cell types were identified, namely, an immature cell, a light cell, and a dark interstitial cell not previously described. The dark cells possessed increased amounts of tubular agranular endoplasmic reticulum, variable numbers of membrane-bounded bodies, and accumulations of glycogen granules. These differed from the light interstitial cells which possessed both tubular and vesicular forms of agranular endoplasmic reticulum, no demonstrable accumulation of glycogen, and in general the membranous components of the light cells were less electron dense than in the dark cell. Intermediate forms between the cell types were observed. Tubular crystalline inclusions with a regular substructure were described and a hypothesis linking these structures with the formation of the crystals of Reinke is discussed. A new cytoplasmic inclusion consisting of aggregations of short dense rod-shaped structures seen in these cells, was also described.