Ultrastructural aspects of wall regeneration byPythium protoplasts

Electron microscope studies were made of wall regeneration byPythium protoplasts. Wall regeneration began with the formation of a loose network of fibrils on the surface of the protoplast followed by increase in density of the fibrillar mesh and deposition of granular matrix material. The majority of the protoplasts did not develop beyond the loose fibrillar network stage, however a small percentage were able to complete wall formation and to form hyphal tubes. A clear zone of demarcation was visible between the fibrillar surface of the protoplast and the smooth surface at the base of the developing hyphal tube.     

Ultrastructural specializations of the cell wall sleeve around plasmodesmata

Specialized structures were observed in the sleeve of wall surrounding plasmodesmata in the rhizome tips of Nephrolepis exaltata and root tips of Spirodela oligorrhiza, Azolla pinnata, and Hordeum vulgare. Material was prepared either by fixation in glutaraldehyde/paraformaldehyde/tannic acid followed by cell wall digestion, or by rapid freezing and freeze substitution, prior to resin embedding. Two structures were identified: rings of electron-opaque material encircling the neck region of plasmodesmata and electron-opaque helices spiraling around the length of plasmodesmata. In some instances, particle-like subunits were observed in the electron-opaque rings. The possible role of these structures as external sphincters involved in the control of permeability of plasmodesmata is discussed.     

Unique aspects of the grass cell wall

Grasses are amongst the most important crops worldwide, and the composition of their cell walls is critical for uses as food, feed, and energy crops. Grass cell walls differ dramatically from dicot cell walls in terms of the major structural polysaccharides present, how those polysaccharides are linked together, and the abundance and importance of pectins, proteins and phenolic compounds. Recent advances, spurred by the availability of genomic resources for several plant species, include the characterization of cellulose synthase like (Csl) gene families that are unique to the grasses and the demonstration that members of one of those gene families, CslF, are responsible for making the mixed linkage glucans that are unique to the order Poales.     

Ultrastructural study of the anomalous cell wall inrRhodotorula gracilis

Cells ofRhodotorula gracilis form under certain conditions equatorial rings, visible in the UV-fluorescence microscope after staining with a CFW brightener. The ultrastructural studies showed that the above rings are formed by a centripetal thickening of the cell wall. Mitochondria with circular cristae and anomalous, ring-like mitochondria were observed in cytoplasm of the cells with developed rings. The plates will be found at the end of the issue.     

Ultrastructural and biochemical aspects of cell wall reconstitution in recalcitrant (grapevine) and regenerating (tobacco) leaf protoplasts

Summary To identify possible reasons that may contribute to recalcitrance in plant protoplasts, the time course of new cell wall deposition was studied by scanning electron microscopy in protoplasts of a recalcitrant species, the grapevine. Results showed that microfibrils were developed after 2 days of culture, that complete cell wall formation occurred on Day 6 to 7 of protoplast culture, and its ultrastructural appearance was identical to that of grapevine leaf-derived callus cells. In addition, a comparative study was undertaken on [U-¹⁴C]glucose uptake and incorporation in ethanol-soluble, cellulosic, and noncellulosic polysaccharide fractions in protoplasts of grapevine and of a readily regenerating species, tobacco, during culture. There was a significantly higher [U-¹⁴C]glucose uptake by tobacco than by grapevine protoplasts. The label distribution in the ethanol-soluble, cellulosic, and noncellulosic fractions of newly synthesized cell walls differed quantitatively between the two species. In particular, the labeled glucose incorporated in the noncellulosic cell wall fraction was threefold greater in tobacco than in grapevine protoplasts. Differences were also revealed in the monosaccharide composition of this fraction between the two species. Addition of dimethyl sulfoxide to the culture medium resulted in a dramatic increase in [U-¹⁴C]glucose uptake by grapevine protoplasts, whereas it exhibited a limited effect in tobacco protoplasts. It showed no effect on the ultrastructural characteristics of new cell wall nor on the incorporation rate of labeled glucose in the cellulosic and noncellulosic cell wall fractions.     

Novel inhibitors of bacterial cell wall synthesis

Over the past forty years, efforts to discover antibacterials have yielded a wide variety of chemical structures, almost exclusively natural products, which inhibit many steps in cell wall synthesis. Although screening for new cell wall inhibitors has been continuous during that period, there have been few reports of new drugs. With the advent of genomics, high resolution X-ray crystallography and the recognition of the need for new antibiotics to combat resistant organisms, there has been a resurgence in interest in this validated target area.     

Regulation of cell wall synthesis and growth

The replication and growth of Mycobacterium tuberculosis are fundamentally linked to the synthesis and extension of its complex cell wall. Incorporation of new wall material must be tightly regulated so that its deposition does not compromise the extant structure. M. tuberculosis also produces an impressive array of complex bioactive lipids that are intimately involved in pathogenesis and protective immunity. The profiles of these lipids are regulated appropriately to allow the bacterium to respond to the prevailing conditions it faces in vivo. A number of regulatory strategies employed by M. tuberculosis to control cell wall biosynthesis and cell division have now been elucidated. The review highlights the role of alternative sigma factors with extracytoplasmic function in the activation of genes for biosynthesis of complex lipids involved in pathogenicity. Rel(Mtb) and CRP(Mt) play roles in cell wall responses to general nutrient deprivation by synthesis and sensing of starvation second messengers, respectively. Recently, the importance of protein phosphorylation networks in cell wall biosynthesis has attracted considerable interest. A plethora of two-component and eukaryotic-like serine/threonine protein kinases systems have been discovered and several are implicated in cell-division, morphogenesis and regulation of the profile of complex bioactive lipids elaborated by the pathogen.     

Newly made enzymes determine ongoing cell wall synthesis and the antibacterial effects of cell wall synthesis inhibitors.

Cell wall synthesis can continue with less than the total complement of cell wall synthetic enzymes present in normal growing cells. A method was developed to investigate whether there exists an excess of cell wall-synthesizing enzymes (penicillin-binding proteins [PBPs]) which all remain functional or whether a mixed population of functional and nonfunctional enzymes characterize normal cells. Surprisingly, cells in which less than 10% of the PBPs were functional could grow at a normal rate, as evidenced by increases in viable counts, culture turbidity, and rates of peptidoglycan, protein, and RNA synthesis. This subset of functional enzymes was biosynthetically new. Penicillin-induced lysis occurred contingent on the acylation of this same small fraction of PBPs, the copy number and affinities of which were below the level of detection by current fluorographic assay techniques. We propose that PBPs have a short functional half-life and that cell wall synthesis and bacterial lysis reflect the activity of newly synthesized PBPs.     

The cell wall of Chlamydomonas eugametos. Immunological aspects

An antiserum was raised against the major cell wall glycoprotein of Chlamydomonas eugametos which after purification reacted specifically with all individual wall components but not with intact cell walls. The antigenic sites in intact walls appear to be cryptic but become exposed on partial enzymatic degradation or in situ during daughter-cell release when the insoluble component is digested. Using the antiserum as a specific label for cell walls in various stages of disintegration, cell wall digestion during asexual and sexual reproduction was studied. It is also shown that while cell wall material is associated with isolated flagella, it is not normally associated with the flagella of intact cells.     

Galactosyl transferases in mycobacterial cell wall synthesis

Two galactosyl transferases can apparently account for the full biosynthesis of the cell wall galactan of mycobacteria. Evidence is presented based on enzymatic incubations with purified natural and synthetic galactofuranose (Galf) acceptors that the recombinant galactofuranosyl transferase, GlfT1, from Mycobacterium smegmatis, the Mycobacterium tuberculosis Rv3782 ortholog known to be involved in the initial steps of galactan formation, harbors dual β-(1→4) and β-(1→5) Galf transferase activities and that the product of the enzyme, decaprenyl-P-P-GlcNAc-Rha-Galf-Galf, serves as a direct substrate for full polymerization catalyzed by another bifunctional Galf transferase, GlfT2, the Rv3808c enzyme.     

Ultrastructural aspects of cryofixed nerves

Summary The ultrastructure of the optic and trigeminal nerves of the rat, cryofixed by use of a liquid nitrogenpropane jet, was examined, paying special attention to the myelin sheath and the cytoskeleton of the axoplasm. The cytoskeleton of the axoplasm is formed by a meshwork of neurofilaments and microtubules connected both to each other and also to the cell organelles and axolemma. These cross-linkers are fixed to the longitudinal neurofilaments in a helical arrangement, which could be a morphological substrate for the diverse axonal transport phenomena. The myelin sheath is formed by concentrically apposed membrane pairs, which are not fused together. The corresponding major and intraperiod lines seen using classical electron microscopy are in fact fissures that are obscured by the pattern of the selective deposition of osmium at certain sites and cannot be interpreted as specific structures. The cryofixed myelin membranes have the appearance of predominantly globular subunits arranged in an asymmetrical bilayer. The globular particles are of diverse diameter and occupy varying positions within the membrane. The tight junctions or zonulae occludentes of the myelin are formed by arrays of isolated particles, and consequently the fibril formation seems to be a result of the chemical fixation.     

Ultrastructural aspects of infertility.

Organ culture studies have demonstrated that 17beta oestradiol can cause the rapid deterioration and dissolution of pre-existing nuclear channel systems in the human endometrial glandular cell. The importance of the nuclear channel system in implantation is emphasized and the implications of secretory activity in the endometrium of an infertile woman are discussed.     

Ultrastructural aspects of preglioma]

Investigation of submicroscopic changes in the astrocytes after the intracerebral injection of the carcinogen 9,10-dimethyl-1-2-benzanthracene suggested that the so-called substantial period of preglioma began from the 45th day of the experiment, when structurally atypical astrocytes appeared against the background of distrophic changes. The changes in the submicroscopic organization of the astrocytes during chemical carcinogenesis can be divided into three stages: 1) intracellular hyperplasia due to traumatic injury of the brain; 2) dystrophic changes connected with the vascular tissue disturbances and immediate action of the carcinogen on the tissue; 3) atypical ultrastructure reconstruction of the astrocytes.