Histological Responses of Four Leguminous Crops Infected with Meloidogyne incognita

Histological responses to Meloidogyne incognita infection in Rhizobium nodules of clover, horsebean, lupine, and pea were investigated. The formation of giant cells in vascular bundles of nodules and roots, and the basal connection of the nodule, were usually associated with abnormal xylem and/or deformed xylem strands. However, giant cells did not disturb or prevent the development of nodular tissues. Areas in which galls formed, wall thickness of giant cells, and number of giant cells around the nematode head varied with plant species. Ranking by gall size and giant-cell wall thickness was horsebean > lupine and pea > clover. The multinucleate condition in giant cells resulted from repeated mitoses without subsequent cytokinesis. The resulting nuclei agglomerated in irregularly shaped masses in some giant cells.     

Pattern of binding of tritiated actinomycin D to Phaseolus coccineus polytene chromosomes. II. The entire chromosome complement

Abstract

The pattern of labelling by tritiated actinomycin D (³H-AMD) on polytene chromosomes in the embryo suspensor cells of Phaseolus coccineus is described in preparations exposed for different times to photographic emulsion. It is observed that: a) each chromosome pair displays a typical labelling pattern; b) heterochromatin close to the centromere does not label in the same way in different chromosome pairs; c) in some chromosome pairs, different bundles of strands in one an the same chromosome segment display differences in ³H-AMD labelling. These results allow an improved characterization of the Phaseolus polytene chromosomes and are discussed in relation to the possible factors determining the differential binding of ³H-AMD to the chromatin.     

Quadruplex formation by both G-rich and C-rich DNA strands of the C9orf72 (GGGGCC)8?(GGCCCC)8 repeat: effect of CpG methylation

Unusual DNA/RNA structures of the C9orf72 repeat may participate in repeat expansions or pathogenesis of amyotrophic lateral sclerosis and frontotemporal dementia. Expanded repeats are CpG methylated with unknown consequences. Typically, quadruplex structures form by G-rich but not complementary C-rich strands. Using CD, UV and electrophoresis, we characterized the structures formed by (GGGGCC)8 and (GGCCCC)8 strands with and without 5-methylcytosine (5mCpG) or 5-hydroxymethylcytosine (5hmCpG) methylation. All strands formed heterogenous mixtures of structures, with features of quadruplexes (at pH 7.5, in K⁺, Na⁺ or Li⁺), but no feature typical of i-motifs. C-rich strands formed quadruplexes, likely stabilized by G•C•G•C-tetrads and C•C•C•C-tetrads. Unlike G•G•G•G-tetrads, some G•C•G•C-tetrad conformations do not require the N7-Guanine position, hence C9orf72 quadruplexes still formed when N7-deazaGuanine replace all Guanines. 5mCpG and 5hmCpG increased and decreased the thermal stability of these structures. hnRNPK, through band-shift analysis, bound C-rich but not G-rich strands, with a binding preference of unmethylated > 5hmCpG > 5mCpG, where methylated DNA-protein complexes were retained in the wells, distinct from unmethylated complexes. Our findings suggest that for C-rich sequences interspersed with G-residues, one must consider quadruplex formation and that methylation of quadruplexes may affect epigenetic processes.     

Why Are Two Different Cross-linkers Necessary for Actin Bundle Formation In Vivo and What Does Each Cross-link Contribute?

In developing Drosophila bristles two species of cross-linker, the forked proteins and fascin, connect adjacent actin filaments into bundles. Bundles form in three phases: (a) tiny bundles appear; (b) these bundles aggregate into larger bundles; and (c) the filaments become maximally cross-linked by fascin. In mutants that completely lack forked, aggregation of the bundles does not occur so that the mature bundles consist of <50 filaments versus ∼700 for wild type. If the forked concentration is genetically reduced to half the wild type, aggregation of the tiny bundles occurs but the filaments are poorly ordered albeit with small patches of fascin cross-linked filaments. In mutants containing an excess of forked, all the bundles tend to aggregate and the filaments are maximally crossbridged by fascin. Alternatively, if fascin is absent, phases 1 and 2 occur normally but the resultant bundles are twisted and the filaments within them are poorly ordered. By extracting fully elongated bristles with potassium iodide which removes fascin but leaves forked, the bundles change from being straight to twisted and the filaments within them become poorly ordered. From these observations we conclude that (a) forked is used early in development to aggregate the tiny bundles into larger bundles; and (b) forked facilitates fascin entry into the bundles to maximally cross-link the actin filaments into straight, compact, rigid bundles. Thus, forked aligns the filaments and then directs fascin binding so that inappropriate cross-linking does not occur.     

Differential Fermentation of Cellulose Allomorphs by Ruminal Cellulolytic Bacteria

In addition to its usual native crystalline form (cellulose I), cellulose can exist in a variety of alternative crystalline forms (allomorphs) which differ in their unit cell dimensions, chain packing schemes, and hydrogen bonding relationships. We prepared, by various chemical treatments, four different alternative allomorphs, along with an amorphous (noncrystalline) cellulose which retained its original molecular weight. We then examined the kinetics of degradation of these materials by two species of ruminal bacteria and by inocula from two bovine rumens. Ruminococcus flavefaciens FD-1 and Fibrobacter succinogenes S85 were similar to one another in their relative rates of digestion of the different celluloses, which proceeded in the following order: amorphous > III_I > IV_I > III_II > I > II. Unlike F. succinogenes, R. flavefaciens did not degrade cellulose II, even after an incubation of 3 weeks. Comparisons of the structural features of these allomorphs with their digestion kinetics suggest that degradation is enhanced by skewing of adjacent sheets in the microfibril, but is inhibited by intersheet hydrogen bonding and by antiparallelism in adjacent sheets. Mixed microflora from the bovine rumens showed in vitro digestion rates quite different from one another and from those of both of the two pure bacterial cultures, suggesting that R. flavefaciens and F. succinogenes (purportedly among the most active of the cellulolytic bacteria in the rumen) either behave differently in the ruminal ecosystem from the way they do in pure culture or did not play a major role in cellulose digestion in these ruminal samples.     

The Minor Wall-Networks between Monolignols and Interlinked-Phenolics Predominantly Affect Biomass Enzymatic Digestibility in Miscanthus

Plant lignin is one of the major wall components that greatly contribute to biomass recalcitrance for biofuel production. In this study, total 79 representative Miscanthus germplasms were determined with wide biomass digestibility and diverse monolignol composition. Integrative analyses indicated that three major monolignols (S, G, H) and S/G ratio could account for lignin negative influence on biomass digestibility upon NaOH and H₂SO₄ pretreatments. Notably, the biomass enzymatic digestions were predominately affected by the non-KOH-extractable lignin and interlinked-phenolics, other than the KOH-extractable ones that cover 80% of total lignin. Furthermore, a positive correlation was found between the monolignols and phenolics at p<0.05 level in the non-KOH-extractable only, suggesting their tight association to form the minor wall-networks against cellulases accessibility. The results indicated that the non-KOH-extractable lignin-complex should be the target either for cost-effective biomass pretreatments or for relatively simply genetic modification of plant cell walls in Miscanthus.     

The guanine-rich fragile X chromosome repeats are reluctant to form tetraplexes

Using circular dichroism spectroscopy, UV absorption spectroscopy and polyacrylamide gel electrophoresis, we studied conformational properties of guanine-rich DNA strands of the fragile X chromosome repeats d(GGC)_n, d(GCG)_n and d(CGG)_n, with n = 2, 4, 8 and 16. These strands are generally considered in the literature to form guanine tetraplexes responsible for the repeat expansion. However, we show in this paper that the repeats are reluctant to form tetraplexes. At physiological concentrations of either Na⁺ or K⁺ ions, the hexamers and dodecamers associate to form homoduplexes and the longer repeats generate homoduplexes and hairpins. The tetraplexes are rarely observed being relatively most stable with d(GGC)_n and least stable with d(GCG)_n. The tetraplexes are exclusively formed in the presence of K⁺ ions, at salt concentrations higher than physiological, more easily at higher than physiological temperatures, and they arise with extremely long kinetics (even days). Moreover, the capability to form tetraplexes sharply diminishes with the oligonucleotide length. These facts make the concept of the tetraplex appearance in this motif in vivo very improbable. Rather, a hairpin of the fragile X repeats, whose stability increases with the repeat length, is the probable structure responsible for the repeat expansion in genomes.     

Leaf and inflorescence peduncle anatomy: a contribution to the taxonomy of Rapateaceae

The anatomy of leaves and inflorescence peduncles was studied in species of Monotrema (4), Stegolepis (1) and Saxofridericia (1), aiming to contribute to the taxonomy of Rapateaceae. The form and structure of leaf blade midrib and the form of the inflorescence peduncle are diagnostic characteristics for the studied species. Monotrema is distinguished by: epidermal and vascular bundle outer sheath cells containing phenolic compounds in both organs; leaf blade with palisade and spongy chlorenchyma, arm-parenchyma, and air canals between the vascular bundles; leaf sheath with phenolic idioblasts in the mesophyll; inflorescence peduncle with tabular epidermal cells and air canals in the cortex and pith. Such characteristics support the recognition of Monotremoideae, which includes Monotrema. Stegolepis guianensis is distinguished by thick-walled epidermal cells and a plicate chlorenchyma in both organs; leaf blade with subepidermal fiber strands in abaxial surface and a heterogeneous mesophyll; inflorescence peduncle with rounded epidermal cells, a hypodermis with slightly thick-walled cells, and a pith with isodiametric cells and vascular bundles. Saxofridericia aculeata is distinguished by papillate epidermal cells in both organs; unifacial leaf blade with subepidermal fiber strands in both surfaces and a regular chlorenchyma; leaf sheath with a hypodermis in both surfaces and fiber bundles in the mesophyll; inflorescence peduncle with an undefined cortex and a hypodermis with thick-walled cells. S. guianensis shares few characteristics with S. aculeata, supporting their placement in different tribes.     

HIGH-RESOLUTION ELECTRON MICROSCOPIC ANALYSIS OF THE AMYLOID FIBRIL

The ultrastructural organization of the fibrous component of amyloid has been analyzed by means of high resolution electron microscopy of negatively stained isolated amyloid fibrils and of positively stained amyloid fibrils in thin tissue sections. It was found that a number of subunits could be resolved according to their dimensions. The following structural organization is proposed. The amyloid fibril, the fibrous component of amyloid as seen in electron microscopy of thin tissue sections, consists of a number of filaments aggregated side-by-side. These amyloid filaments are approximately 75–80 A in diameter and consist of five (or less likely six) subunits (amyloid protofibrils) which are arranged parallel to each other, longitudinal or slightly oblique to the long axis of the filament. The filament has often seemed to disperse into several longitudinal rows. The amyloid protofibril is about 25–35 A wide and appears to consist of two or three subunit strands helically arranged with a 35–50-A repeat (or, less likely, is composed of globular subunits aggregated end-to-end). These amyloid subprotofibrillar strands measure approximately 10–15 A in diameter.     

Biomass Enzymatic Saccharification Is Determined by the Non-KOH-Extractable Wall Polymer Features That Predominately Affect Cellulose Crystallinity in Corn

Corn is a major food crop with enormous biomass residues for biofuel production. Due to cell wall recalcitrance, it becomes essential to identify the key factors of lignocellulose on biomass saccharification. In this study, we examined total 40 corn accessions that displayed a diverse cell wall composition. Correlation analysis showed that cellulose and lignin levels negatively affected biomass digestibility after NaOH pretreatments at p<0.05 & 0.01, but hemicelluloses did not show any significant impact on hexoses yields. Comparative analysis of five standard pairs of corn samples indicated that cellulose and lignin should not be the major factors on biomass saccharification after pretreatments with NaOH and H₂SO₄ at three concentrations. Notably, despite that the non-KOH-extractable residues covered 12%–23% hemicelluloses and lignin of total biomass, their wall polymer features exhibited the predominant effects on biomass enzymatic hydrolysis including Ara substitution degree of xylan (reverse Xyl/Ara) and S/G ratio of lignin. Furthermore, the non-KOH-extractable polymer features could significantly affect lignocellulose crystallinity at p<0.05, leading to a high biomass digestibility. Hence, this study could suggest an optimal approach for genetic modification of plant cell walls in bioenergy corn.     

Green synthesis of graphene oxide sheets decorated by silver nanoprisms and their anti-bacterial properties

A widely soluble graphene oxide sheets decorated by silver nanoprisms were prepared through green synthesis at the room temperature using gelatin as reducing and stabilizing agent. The samples were characterized by X-ray diffraction (XRD), scanning electron microscopy (SEM), transmission electron microscopy (TEM), UV-visible spectroscopy and fluorescence spectra. The results demonstrate that these silver-nanoprisms assembled on graphene oxide sheets are flexible and can form stable suspensions in aqueous solutions. Furthermore, the formation mechanism of soluble graphene oxide sheets decorated by silver nanoprisms was successfully explained. The anti-bacterial properties of graphene oxide sheets decorated by silver nanoprisms were tested against Escherichia coli. This work provides a simple and “green” method for the synthesis of graphene oxide sheets decorated by silver nanoprisms in aqueous solution with promising antibacterial property.     

Use of Aqueous Silver To Enhance Inactivation of Coliphage MS-2 by UV Disinfection

A synergistic effect between silver and UV radiation has been observed that can appreciably enhance the effectiveness of UV radiation for inactivation of viruses. At a fluence of ca. 40 mJ/cm², the synergistic effect between silver and UV was observed at silver concentrations as low as 10 μg/liter (P < 0.0615). At the same fluence, an MS-2 inactivation of ca. 3.5 logs (99.97%) was achieved at a silver concentration of 0.1 mg/liter, a significant improvement (P < 0.0001) over the ca. 1.8-log (98.42%) inactivation of MS-2 at ca. 40 mJ/cm² in the absence of silver. Modified Chick-Watson kinetics were used to model the synergistic effect of silver and UV radiation. For an MS-2 inactivation of 4 logs (99.99%), the coefficient of dilution (n) was determined to be 0.31, which suggests that changes in fluence have a greater influence on inactivation than does a proportionate change in silver concentration.     

Kinetic and thermodynamic characterization of single-mismatch discrimination using single-molecule imaging

A single-molecule detection setup based on total internal reflection fluorescence (TIRF) microscopy has been used to investigate association and dissociation kinetics of unlabeled 30mer DNA strands. Single-molecule sensitivity was accomplished by letting unlabeled DNA target strands mediate the binding of DNA-modified and fluorescently labeled liposomes to a DNA-modified surface. The liposomes, acting as signal enhancer elements, enabled the number of binding events as well as the residence time for high affinity binders (K_d < 1 nM, k_off < 0.01 s⁻¹) to be collected under equilibrium conditions at low pM concentrations. The mismatch discrimination obtained from the residence time data was shown to be concentration and temperature independent in intervals of 1–100 pM and 23–46°C, respectively. This suggests the method as a robust means for detection of point mutations at low target concentrations in, for example, single nucleotide polymorphism (SNP) analysis.     

Structural characterizations of some adducts of silver(I) nitrate and perchlorate with some cyclic di- (or tri-) ene organic ligands

Single crystal X-ray structural characterizations of some adducts of silver(I) nitrate and perchlorate with assorted organic poly-ene ligands (nbd = norbornadiene, bicyclo[2.2.1]hepta-2,5-diene; cod = 1,5-cyclooctadiene; cdt ≡ trans,trans,cis-cyclododeca-1,5,9-triene) are reported, all being polymeric in form (with the exception of mononuclear ionic AgClO₄:cod (1:2)), with chains comprised of alternating silver and nitrate/perchlorate components substituted or linked by unsaturated donors which complete the coordination spheres of the silver atoms. In AgNO₃:nbd (2:1) (a redetermination), pairs of silver/nitrate strands are linked in a one-dimensional polymer by the nbd ligands. In AgNO₃:nbd (1:1)_∞, meandering silver/nitrate strands containing pairs of independent silver and nitrate units in a crystallographic mirror plane are linked to either side with parallel planes by nbd ligands. In AgNO₃:cod (1:1)_∞, the cod ligands ‘chelate’ to the silver atoms in a silver/nitrate chain. In AgNO₃:cdt (1:2)_∞, pairs of ‘unidentate’ cdt ligands are pendant from a silver/nitrate chain, while in the (1:1)_∞ adduct, the cdt ligands bridge pairs of silver atoms from an adjacent chain forming a two-dimensional web. A common form of the bridging nitrate group in the above is as an O,O′-NO₃-O′,O″ bis-chelate, the pair of the bis-oxygen chelates having a common oxygen atom.     

Slug,Twist, and E-Cadherin as Immunohistochemical Biomarkers in Meningeal Tumors

The overexpression of Twist and Slug and subsequent down-regulation of E-cadherin facilitate the acquirement of invasive growth properties in cancer cells. It is unclear which of these molecules are expressed in mesenchymal tumors in the central nervous system. Here, we investigated 10 cases each of hemangiopericytoma, solitary fibrous tumor, meningothelial, fibrous, angiomatous, and atypical meningiomas, and 5 cases of anaplastic meningioma for Slug, Twist, E-cadherin, and N-cadherin immunoexpression. Nuclear Slug expression was observed in 9/10 (90%) hemangiopericytomas and 5/10 (50%) solitary fibrous tumors, but not in any meningiomas, except for 1 case. Similarly, nuclear Twist expression was more extensive in hemangiopericytomas and solitary fibrous tumors than meningiomas. In contrast to Slug and Twist, the positive expression of E-cadherin was observed in 39/45 (87%) meningiomas, but not in any hemangiopericytomas or solitary fibrous tumors (P<0.0001). The fraction of tumor cells expressing E-cadherin in meningeal tumors was negatively correlated to those of Twist (P = 0.004) and Slug (P<0.0001). The overexpression of Slug and Twist with down-regulation of E-cadherin was characteristic findings in hemangiopericytomas and solitary fibrous tumors, but not in meningiomas. The immunohistochemical profiles of the two tumor groups may be useful as diagnostic markers in cases that present a differential diagnosis challenge.     

Comparison of Fungal Laccases and Redox Mediators in Oxidation of a Nonphenolic Lignin Model Compound

Several fungal laccases have been compared for the oxidation of a nonphenolic lignin dimer, 1-(3,4-dimethoxyphenyl)-2-(2-methoxyphenoxy)propan-1,3-diol (I), and a phenolic lignin model compound, phenol red, in the presence of the redox mediators 1-hydroxybenzotriazole (1-HBT) or violuric acid. The oxidation rates of dimer I by the laccases were in the following order: Trametes villosa laccase (TvL) > Pycnoporus cinnabarinus laccase (PcL) > Botrytis cinerea laccase (BcL) > Myceliophthora thermophila laccase (MtL) in the presence of either 1-HBT or violuric acid. The order is the same if the laccases are used at the same molar concentration or added to the same activity (with ABTS [2,2′-azinobis (3-ethylbenzothiazoline-6-sulfonic acid)] as a substrate). During the oxidation of dimer I, both 1-HBT and violuric acid were to some extent consumed. Their consumption rates also follow the above order of laccases, i.e., TvL > PcL > BcL > MtL. Violuric acid allowed TvL and PcL to oxidize dimer I much faster than 1-HBT, while BcL and violuric acid oxidized dimer I more slowly than BcL and 1-HBT. The oxidation rate of dimer I is dependent upon both k_cat and the stability of the laccase. Both 1-HBT and violuric acid inactivated the laccases, violuric acid to a greater extent than 1-HBT. The presence of dimer I or phenol red in the reaction mixture slowed down this inactivation. The inactivation is mainly due to the reaction of the redox mediator free radical with the laccases. We did not find any relationship between the carbohydrate content of the laccases and their inactivation. When the redox potential of the laccases is in the range of 750 to 800 mV, i.e., above that of the redox mediator, it does not affect k_cat and the oxidation rate of dimer I.     

Lignin Degradation by Streptomyces viridosporus: Isolation and Characterization of a New Polymeric Lignin Degradation Intermediate

A new, quantitatively significant intermediate formed during lignin degradation by Streptomyces viridosporus T7A was isolated and characterized. In Streptomyces-inoculated cultures, the intermediate, an acid-precipitable, polyphenolic, polymeric lignin (APPL), accumulated in the growth medium. The APPL was a water-soluble polymer probably consisting of a heterogeneous mixture of molecular weight components of ≥20,000. APPLs were precipitable from culture filtrates after they had been acidified to pH <3 to 5. Noninoculated controls yielded little APPL, but supernatant solutions from inoculated cultures produced quantities of APPL that correlated with the biodegradability of the lignocellulose type. Maximal recovery of APPL was obtained from corn lignocellulose, reaching 30% of the initial lignin present in the substrate. APPLs contained small amounts of carbohydrate, organic nitrogen, and inorganic materials. The lignin origin of APPLs was confirmed by chemical analyses, which included acidolysis, permanganate oxidation, elemental analyses, functional group analyses, nuclear magnetic resonance spectroscopy, and ¹⁴C isotopic techniques. Analyses of APPLs from corn lignocelluloses showed that S. viridosporus-degraded APPLs were lignin derived but significantly different in structure from APPLs derived from uninoculated controls or from a standard corn milled-wood lignin. Degraded APPLs were enriched in phenolic hydroxyl groups and, to a small extent, in carboxyl groups. Degradative changes appeared to be largely oxidative and were thought to involve substantial cleavage of p-hydroxy ether linkages and methoxyl groups in the lignin.     

Myosin heavy chains: Detection by immuno-blotting in higher plants and localization by immunofluorescence in the alga chara

A monoclonal antibody to the heavy chain of myosin from mouse 3T3 cells was used to identify myosin heavy chains in four flowering plants and to identify and localize them in the green alga Chara. The M_r of the immunoreactive bands varied from ca 200 000 in Chara and Arabidopsis to 170 000 in mung beans, peas and wheat. An additional band of 158 000-M_r was resolved in roots and shoots of mung beans. Chara contained a second, immunoreactive band of 110 000-M_r whose possible relationship to the tail-less myosin I enzymes is discussed.Immunofluorescence of giant internodal cells of Chara showed that myosin was almost entirely confined to the streaming endoplasm. Individual organelles and beaded endoplasmic strands were heavily labelled as were the sub-cortical filament bundles. Actin, in contrast, was confined to the sub-cortical bundles. It is proposed that force is generated by interaction of the actin in the subcortical bundles with myosin on individual organelles and on the beaded endoplasmic strands. By ramifying through the endoplasm, the strands may ensure the cohesive movement of the whole mass of endoplasm.     

Crystal structure of d(GCGAAAGCT) containing a parallel-stranded duplex with homo base pairs and an anti-parallel duplex with Watson-Crick base pairs

A DNA fragment d(GCGAAAGCT), known to adopt a stable mini-hairpin structure in solution, has been crystallized in the space group I4₁22 with the unit-cell dimensions a = b = 53.4 Å and c = 54.0 Å, and the crystal structure has been determined at 2.5 Å resolution. The four nucleotide residues CGAA of the first half of the oligomer form a parallel duplex with another half through the homo base pairs, C₂:C₂⁺ (singly-protonated between the Watson– Crick sites), G₃:G₃ (between the minor groove sites), A₄:A₄ (between the major groove sites) and A₅:A₅ (between the Watson–Crick sites). The two strands remaining in the half of the parallel duplex are split away in different directions, and they pair in an anti-parallel B-form duplex with the second half extending from a neighboring parallel duplex, so that an infinite column is formed in a head-to-tail fashion along the c-axis. It seems that a hexa-ammine cobalt cation supports such a branched and bent conformation of the oligomer. One end of the parallel duplex is stacked on the corresponding end of the adjacent parallel duplex; between them, the guanine base of the first residue is stacked on the fourth ribose of another duplex.     

VASCULAR PATTERNS IN STEMS OF THE CYCLANTHACEAE

A survey was made of the distribution of stem vascular bundles in representatives of ten genera of the tropical monocotyledonous family Cyclanthaceae. Films of series of serial transverse sections were used to reconstruct the stem vasculature. Each leaf trace, followed in a basipetal direction from its level of insertion at the stem periphery, describes an obliquely downward course, initially contacting from 1 to 4 (or more) existing axial bundles. The associated bundles form a compound vascular bundle in which the original bundles initially remain discrete, most commonly in a tetrapolar arrangement, with four separate strands. Followed further in the basipetal direction, the strands eventually fuse partly or completely, usually to form a collateral or amphivasal axial bundle which participates in a new structural cycle. Quantitative variation between different taxa includes a simple pattern in Ludovia, in which only bipolar bundles are developed. More elaborate forms have multipolar bundles with more than four separate strands. A systematically useful observation is that stem vasculature in Cyclanthus, representing the subfamily Cyclanthoideae, does not differ significantly from that in subfamily Carludovicoideae although there are some distinctive structural features.