Rapid calcium exchange for protons and potassium in cell walls of Chara

Net fluxes of Ca²⁺, H⁺ and K⁺ were measured from intact Chara australis cells and from isolated cell walls, using ion-selective microelectrodes. In both systems, a stimulation in Ca²⁺ efflux (up to 100 nmol m^?2 s^?1, from an influx of ～40 nmol m^?2 s^?1) was detected as the H⁺ or K⁺ concentration was progressively increased in the bathing solution (pH 7.0 to 4.6 or K⁺ 0.2 to 10mol m^?3, respectively). A Ca²⁺ influx of similar size occurred following the reverse changes. These fluxes decayed exponentially with a time constant of about 10 min. The threshold pH for Ca²⁺ efflux (pH 5.2) is similar to a reported pH threshold for acid-induced wall extensibility in a closely related characean species. Application of NH₄⁺ to intact cells caused prolonged H⁺ efflux and also transient Ca²⁺ efflux. We attribute all these net Ca²⁺ fluxes to exchange in the wall with H⁺ or K⁺. A theoretical treatment of the cell wall ion exchanges, using the ‘weak acid Donnan Manning’ (WADM) model, is given and it agrees well with the data. The role of Ca²⁺ in the cell wall and the effect of Ca²⁺ exchanges on the measured fluxes of other ions, including bathing medium acidification by H⁺ efflux, are discussed.     

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.     

Cell wall turnover in phosphate and potassium limited chemostat cultures of Bacillus subtilis W23

Turnover in phosphate and potassium limited chemostat cultures of Bacillus subtilis W23 results in the release of over 80% of the wall material present at the time of chasing equilibrium-labelled cultures. The rate at which turnover proceeds is faster in potassium limited cultures than in phosphate limited cultures but in both cases a fraction of the wall material appears to be conserved, or to undergo turnover at a lower rate. Previously we have shown that the polar wall is less active metabolically than the cylindrical wall and it is possible that the apparently conserved wall is that present in the pole.     

Effects of ethylene on the orientation of microtubules and cellulose microfibrils of pea epicotyl cells with polylamellate cell walls

Summary Following a 5 hours ethylene treatment, cortical cells of Pea (Pisum sativum L. var Alaska) epicotyl third internode showed a change in the orientation of both microtubules near the plasma membrane and recently deposited cellulose microfibrils. Control cortical cells had mostly transverse microtubules. The ratio of the average frequency of transverse to longitudinal microtubules was 6.0. After 5 hours of ethylene treatment, cortical cells had mostly longitudinal microtubules, with the ratio of transverse to longitudinal microtubules equal to 0.1. Epidermal cells were more variable than cortical cells with regard to the frequency of longitudinal and transverse microtubules. Observation of cortical cell walls in conventionally stained thin sections revealed that recent deposition of microfibrils had been primarily transverse in almost all of the control cortical cells sampled. In contrast, more than half of the ethylene-treated cortical cells had recent deposition oriented primarily longitudinally. This change in microtubule and microfibril orientation may be early enough to constitute the primary effect of ethylene leading to radial cell expansion.Research supported by NSF grant PCM 78-03244, A1, 2 to PBG and by a Research Corporation grant to WRE.     

Auxin-induced changes in the cell wall structure: Changes in the sugar compositions, intrinsic viscosity and molecular weight distributions of matrix polysaccharides of the epicotyl cell wall of Vigna angularis

Rapid effects of indole-3-acetic acid (IAA) on the mechanical properties of cell wall, and sugar compositions, intrinsic viscosity and molecular weight distribution of cell wall polysaccharides were investigated with excised epicotyl segments of Vigna angularis Ohwi et Ohashi cv. Takara.

• 1 IAA caused cell wall loosening as studied by stress-relaxation analysis within 15 min after the IAA application.
• 2 IAA stimulated the decrease in the content of arabinose and galactose in the hemicellulose 1 h after its application. The amounts of other component sugars in the cell wall polysaccharides remained constant during the IAA-induced segment growth.
• 3 The intrinsic viscocity of the pectin increased as early as 30 min after the IAA application. This effect was not prevented when elongation growth of the segment was osmotically suppressed by 0.15 M mannitol.
• 4 Gel permeation chromatography of the pectin on a Sepharose 4 B column demonstrated that IAA caused increase in the mass-average molecular weight of the pectin. Analysis of the sugar compositions of the pectin eluted from the Sepharose 4 B column indicated that IAA increased the molecular weight of the polysaccharides composed of uronic acid, galactose, rhamnose and arabinose. This effect became apparent within 30 min after the IAA application. Furthermore, IAA increased the molecular weight of the pectin when elongation growth of the epicotyl segments was osmotically suppressed by 0.15 M mannitol.
• 5 Hemicellulose of the cell wall chromatographed on a Sepharose CL-4 B column. Analysis of the neutral sugar compositions and the iodine staining property (specific for xyloglucans) of the polysaccharide solution eluted from the column indicated that the hemicellulose consisted of xyloglucans, arabinogalactans and polysaccharides composed of xylose and/or mannose. IAA caused a decrease in the arabinogalactan content and depolymerization of xyloglucans. These IAA effects became apparent within 30 min after the IAA application. These changes occurred even when elongation growth of the epicotyl segments was osmotically suppressed by 0.15 M mannitol.

Polymerization of the pectin, degradation of arabinogalactans and depolymerization of xyloglucans appear to be involved in the mechanism by which IAA induces cell wall loosening and therefore extension growth of cells.     

A thermodynamic study of Cu++−Zn++ ion exchange in theNitella flexilis cell wall

Summary Exchange isotherms of Cu²⁺ vs Zn²⁺-ions were performed on the cell wall of a fresh water alga,Nitella flexilis. The relevant thermodynamic parameters are calculated. The cell wall absorbs copper selectively. The selectivity is explained by a stronger chelation of the cupric ion, due to the Jahn-Teller effect. The wall acts like a two-site model, based on the nature of the ligands: a first group of aminesites reacts exothermically and a second of hydroxylic-carboxylic sites of lower affinity, reacts endothermically.     

Carotenoids in the outer cell-wall layer of Scenedesmus (Chlorophyceae)

Scenedesmus obliquus, strain 633, which synthesizes ketocarotenoids and sporopollenin, also forms pink-red-colored cell walls. Both the cell walls left over after autospore liberation and those from homogenates of disrupted green cells have similar carotenoid pigmentation. Canthaxanthin, astaxanthin, an unidentified ketocarotenoid, and lutein were found as integral cell wall components. They are bound to the outer (trilaminar) layer of the complete cell wall which also contains sporopollenin.Abbreviations CWH complete cell walls isolated from the homogenates - CWM maternal cell walls accumulated in the medium - KC ketocarotenoid - SC secondary carotenoids - SP sporopollenin     

Isolation and analysis of cell wall material from beeswing wheat bran (Triticum aestivum)

Cell wall material (CWM) isolated from beeswing wheat bran contains 66% carbohydrate, 12% Klason lignin, 6% protein and 4% ash. The relative proportions of sugars in the CWM are arabinose 34%, xylose 26%, galactose 2%, glucose 32% and uronic acid 6%. The uronic acid was shown to consist of glucuronic acid and its 4-O-Me analogue in the ratio 1.8:1. Partial acid hydrolysis of the CWM yielded neutral sugars and a uronic acid fraction. The latter was shown to contain Glc p A-(1→2)-Xyl p and Glc p A-(1→2)-O-Xyl p-(1→4)-Xyl p and their 4-O-Me substituted uronic acid analogues. Methylation analysis of the whole CWM and partially degraded methylated CWM revealed the nature of the constituent glycosidic linkages. From the combined evidence we infer that the major structural features of the non-cellulosic polysaccharides are a linear chain of xylopyranose units joined by (1→4)-linkages, and arabinofuranose, xylose, galactose (and uronic acid) end groups, which in at least some of the polysaccharides, are attached directly by (1→2)- and/or (1→3)-linkages to the xylan chain. The CWM has been fractionated by successive extractions with water at 80°, 0.2 M (NH₄)₂C₂O₄ at 80°, Na chlorite/HOAc at 70°, 0.2 M (NH₄)₂C₂O₄ at 80°, 1 M and 4 M KOH, and the neutral sugar composition of the fractions determined. It is concluded from these and other experiments that the CWM contains two main types of polysaccharides, the arabinoxylans and cellulosic polymers, and that phenolic ester linkages play a role in holding them together.     

Lysis of growing fissin-yeast cells induced by aculeacin A,a new antifungal antibiotic

Cells of Schizosaccharomyces pombe grown in the presence of aculeacin A, a peptide antibiotic, were lysed resulting the death of cells. Under high osmolarity, the cellular lysis induced by aculeacin A was considerably reduced. The use of synchronous-culture systems distinguished cell elongation from cell division revealed that the sites of aculeacin A-induced lysis on the fission yeast were the end(s) and the cell plate region, corresponded to the regions of the cell wall synthesis. Aculeacin A-resistant survivors exhibited morphological alterations which were swollen at one or both ends of the cell and appeared drumstick or dumbbel like; the wall of the bulge region was observed to be stained with a fluorescent brightener, as well as that of the cell plate region. These effects of aculeacin A are discussed as compared with effects of 2-deoxy-D-glucose.