The sites of synthesis and transport of extracellular polysaccharides in the root tissues of maize

The secreted slime from root cap cells of corn (Zea mays, cv. SX-17) was studied. Production of slime by excised root tips is stimulated by the addition of 40 mM sucrose or fucose and half-strength Hoagland's solution to the incubation medium.Secreted slime was recovered from aqueous solution by precipitation with ethanol. The polymer has a molecular weight greater than 2 X 10-8 daltons and a density of 1.63 g cm-'. Protein is not present in material purified by density gradient centrifugation with cesium chloride. Fucose (39 %) and galactose (30%) are the principle neutral sugars found in the purified polymer. Galacturonic and glucuronic acids, arabinose, xylose, mannose, and glucose are also present.Several investigators have studied the mucilage or slime secreted by corn roots, but there is limited agreement on the chemical composition and properties of this secretion (9,10,13,14). This could result from differences between the varieties of corn studied or from variations in culture methods or purification procedures. Before analyzing the secreted polysaccharide produced by roots of cultivar SX-17 of Zea mays, we undertook a detailed study of the culture conditions required for production of the material. We report results of our investigation into the production of the secreted polysaccharide and some of the chemical and physical characteristics of the purified polymer. MgSOJ, 25 jam boric acid, 40 mm sucrose or fucose, 20 jug/ml of chloramphenicol, and 5 jug/ml of streptomycin. In those experiments using entire seedlings rather than excised root tips, the primary root was passed through a hole in a plastic support into the standard incubation medium in a 50-ml beaker. Root tips were excised at the end of the incubation and retained for analysis.The incubation was terminated by making the medium to 80% (v/v) ethanol by the addition of absolute alcohol and keeping it at 4 C for a minimum of 12 hr. The precipitated material and root tips were collected together on glass fiber discs (Whatman GF/C) previously heated at 500 C for 1 hr to remove any carbohydrate (10). The material was washed five times with 95% (v/v) alcohol to remove soluble sugars. The samples were then dried and transferred to 20-X 150-mm culture tubes with Teflon-lined screw caps. The internal standard, normally 2 or 4 mg of myoinositol, was added. Then the sample was hydrolyzed and alditol acetates were prepared as described below.The secretory product from root tips of whole seedlings was collected in two ways. The material designated as crude material was collected by wiping the root tip onto a 2.5-cm diameter glass fiber filter disc (Whatman GF/C, heated for 1 hr at 500 C). Discs carrying the adhering material were dried over anhydrous magnesium perchlorate in a vacuum desiccator. Material to be purified was prepared by aspirating the secreted material from the root tip and collecting it in a test tube. There it was diluted with glass-distilled H20 to reduce its viscosity, and it was centrifuged at 20,000g for 30 min to remove sl...

Section: Methodssupporting

confidence: 73%

Studies on the Secretion of Maize Root Cap Slime

Paull¹,

Johnson²,

Jones³

1975

“…Golgi-and ER-derived vesicles are capable of transporting membrane components and secretory products, including enzymic protein, to the cell surface in plants (3,13,18). Radioautographic (3,24) and biochemical (3, 21) studies of higher plant tissues indicate that neither of these organelles forms cellulose or low mol wt cellodextrins in vivo, despite the /8-1,4-glucan synthetase associated with them.…”

Section: Resultsmentioning

confidence: 99%

“…Radioautographic (3,24) and biochemical (3, 21) studies of higher plant tissues indicate that neither of these organelles forms cellulose or low mol wt cellodextrins in vivo, despite the /8-1,4-glucan synthetase associated with them. Since relatively much higher levels of cellulose synthetase activity could be detected at undisturbed cell surfaces and these were well correlated with actual rates of cellulose deposition in vivo (Table I), the suggestion (21) is reiterated that the intracellular B-1,4-glucan synthetase represents normally inactive enzyme which is in the process of being transported to the surface.…”

Section: Resultsmentioning

confidence: 99%

The Site of Cellulose Synthesis

Shore¹,

Raymond²,

Maclachlan³

1975

“…Recent evidence suggests that the Golgi apparatus is involved in the biosynthesis and the secretion of the hemicellulosic component of the cell wall (1,18) and of other extracellular polysaccharides (9). In animal cells, the Golgi apparatus functions in the secretion and terminal glycosylation of extracellular proteins and mucopolysaccharides (10,16,21,24).…”

Section: Discussionmentioning

confidence: 99%

Involvement of the Golgi Apparatus in the Synthesis and Secretion of Hydroxyproline-rich Cell Wall Glycoproteins

Gardiner¹,

Chrispeels²

1975

115

Pulse labeling of carrot root phloem parenchyma (Daucus carota L. cv. Nantes) tissue with '4C-proline followed by fractionation of the cytoplasmic organelles on sucrose gradients was used to determine the identity of the membranous organelies involved in the secretion of the hydroxyproline-rich gly. coproteins of the cell wall. Identification of the organelles was done through electron-microscopical observations and through the localization of marker enzymes on the sucrose gradients. Enrichment of the organelles involved in secretion was determined by measuring the percentage of the incorporated radioactivity present as '4C-hydroxyproline. The Golgi apparatus (dictyosome) was found to be a major site of glycoprotein transport. This identification was based on the observed enrichment of dictyosomes paralleling the purification of newly synthesized cell-wall glycoproteins. A marker enzyme for the Golgi apparatus, inosinediphosphatase, banded with the newly synthesized cell wall glycoproteins on sequential isopycnic and rate zonal sucrose gradients. Marker enzymes for the endoplasmic reticulum and the plasma membrane were clearly separated from the dictyosome-rich fraction. UDParabinose arabinosyl transferase, an enzyme involved in the glycosylation of the peptide moiety of this glycoprotein, also banded with the dictyosomes on both kinds of gradients. The results suggest an important role of the Golgi apparatus in the biosynthesis and the secretion of the cell wall glycoproteins of higher plants.The primary walls of plant cells contain a structural hydroxyproline-rich glycoprotein called "extensin" (for review see Lamport [12]). This protein is synthesized in the cytoplasm as a proline-rich polypeptide and the proline residues are hydroxylated by a soluble peptidyl proline hydroxylase (20). The 'This investigation was supported by the Atomic Energy Commission under Contract AT(04-3)-34 PA 159.