Extensins

The ability of PO to interact with the acetyl residues of chitin allows us to compare them with monovalent lectins (i.e. extensins) which when binding with hemicellulose are only affected in a medium with a high ionic strength (Brownleader et al., 2006). As a rule, POs are bound with the plant cell wall and act as its modifiers. Some POs can form complexes with an extensin of cell walls (Brownleader et al., 2006). Consequently, chitin-specific sites that are capable of interacting with polysaccharides exist in the molecules of PO, and these sites can resemble the membrane receptor binding sites or else be similar to the domains of heparinbinding proteins (Kim et al., 2001). 

Some divalent cations such as Cu and Pb form very stable complexes with pectate, but are unlikely to be present at sufiScient concentration in the apoplast of plants to form a major fraction of the counterions associated with the pectic fraction in vivo. The Al ion may deserve closer examination, as it is certainly able to displace Ca from cell walls and reaches substantial concentrations in plant roots under some conditions [60,61]. aluminium is not usually considered to be freely translocated, however. Basic peptides with their negative charges spaced at a similar interval to galacturonans (0.43 nm or a small multiple thereof) can in principle have a very high afiBnity for pectate [62,63], but the extensins that are associated with the most insoluble pectic fractions [M] do not appear to have this type of structure. The possibility that the non-extractable pectic polymers in most cell walls are very strongly complexed with some cation other than Ca " cannot be ruled out, but there is little evidence to support it at present. 

Bryodin 1 Suspension Nicotiana tabacum (tobacco) Microparticle bombardment of suspension CaMV 35S Extensin 30 mg r1 (e) 74... 

Bryodin 1 CaMV 35S promoter/ nos terminator Extensin LP N. tabacum NT-1 30 mg L-1 82... 

A breakthrough for tissue printing, at least for the botanical sciences, came when Cassab and Varner (3) combined the use of nitrocellulose and antibody technology. They placed sections of a freshly cut soybean seed on nitrocellulose membranes and probed the resulting imprints using specific antibodies. They were able to show that soluble extensin protein is primarily localized in the seed coat and vascular tissues. 

Cassab GI, Varner JE. Immunocytolocalization of extensin in developing soybean seed coats by immunogold-silver staining and by tissue printing on nitrocellulose paper. J Cell Biol 1987 105 2581-2588. 

Cellulose microfibrils make up the basic framework of the primary wall of young plant cells (3), where they form a complex network with other polysaccharides. The linking polysaccharides include hemicellulose, which is a mixture of predominantly neutral heterogly-cans (xylans, xyloglucans, arabinogalactans, etc.). Hemicellulose associates with the cellulose fibrils via noncovalent interactions. These complexes are connected by neutral and acidic pectins, which typically contain galac-turonic acid. Finally, a collagen-related protein, extensin, is also involved in the formation of primary walls. 

In order to think about the nature and consequences of cell wall polymer phenolic cross-linking, we need a working model of the mode of assembly and the final structure of the primary cell wall. Unfortunately, there is no universally acceptable model that proposed by Albersheim and co-workers (3) is not now widely accepted because the postulated interpolysaccharide glycosidic bonds have not been demonstrated (4) and the warp-weft model of Lamport (5) rests on the assumptions that extensin (i) forms a defined-porosity network (not proven) (ii) is orientated anti-clinally to the cell surface [some evidence against (6)] and (iii) is a major component of all primary cell walls (not true). 

The basic extensins. These are hydroxyproline-, lysine- and tyrosine-rich glycoproteins consisting of rigid molecular rods about 80 nm long (14,15), bearing short mono- to tetrasaccharide side-chains (2,14). When newly secreted they bind ionically to the acidic polysaccharides of the cell wall and can be extracted with cold salt solutions later they become much more resistant to salt-extraction and are said to be covalently bound, probably via dimerization of their tyrosine residues to form isodityrosine (15). 

Feruloylated pectins have been found in the parenchymatous cell walls of many Dicotyledons (mainly in the Centrospermae and Solanaceae), but UV-fluorescence microscopy suggests that at least the epidermal cell walls of all Dicotyledons contain phenolic residues it remains to be seen whether these phenolic residues are attached to polysaccharides or to cutin, but location of even a small quantity of, say, feruloyl-pectin in the epidermal wall would be particularly significant in the control of growth because the extensibility of the epidermis controls the expansion of whole stems (23) and leaves (Fry, unpublished observations). The extensins, as already mentioned, are rich in the phenolic amino acid tyrosine (2). 

Background. In order to maximize the efficiency of cross-linking based on a small number of phenolic groups, it is important that these groups should be sited on the wall polymers at appropriate loci rather than randomly. In the case of the phenolic side-chains of extensins this criterion is met since the siting of the tyrosines is genetically encoded (27). 

The previous section has presented evidence that phenolic units are carefully positioned within the wall polymers. When these units undergo oxidative phenolic coupling reactions in the cell wall, the coupling reactions themselves are also remarkably specific. This can be illustrated by reference to the tyrosine residues of extensin. 

In animal structural proteins in vivo, the only known dimer of tyrosine is dityrosine (40,41) in the extensin of plant cell walls, in contrast, the only dimer formed in vivo is isodityrosine (16). How is the coupling of tyrosine in plants confined to the formation of isodityrosine There is nothing unique about the local environment of the tyrosine residues in (pure) extensin, since... 

Extensin in vivo Dicot primary cell wall < 7 Idt (16)... 

Extensin in vitro Bovine serum peroxidase + H2O2 9 DiT (38)... 

Role of Neighboring Polysaccharide Molecules in Determining the Orientation of Tyrosine Residues During Coupling. These considerations suggest a third possible explanation for the exclusive formation of isodityrosine in the plant cell wall in vivo that the neighboring structural molecules of the wall constrain extensin to prevent dityrosine formation. This would mean that the biologically relevant substrate for peroxidase in the plant cell wall is not naked extensin but extensin complexed with another wall component, possibly an acidic polysaccharide to which the extensin would bind ionically. 

In conclusion, it seems fair to say that specificity exists in both the biosynthesis and in the oxidative coupling of polymer-bound phenols in the growing cell wall, (a) Tyrosine residues are placed at specific sites along the extensin molecule by genetically-encoded information, (b) Tyrosine cross-linking in vivo is a very specific, carefully steered process in that it occurs... 

In plants, ascorbate is required as a substrate for the enzyme ascorbate peroxidase, which converts H202 to water. The peroxide is generated from the 02 produced in photosynthesis, an unavoidable consequence of generating 02 in a compartment laden with powerful oxidation-reduction systems (Chapter 19). Ascorbate is a also a precursor of oxalate and tartrate in plants, and is involved in the hydroxylation of Pro residues in cell wall proteins called extensins. Ascorbate is found in all subcellular compartments of plants, at concentrations of 2 to 25 mM—which is why plants are such good sources of vitamin C. 

Lamport40 considered the primary cell-wall to be a single, bagshaped macromolecule having a coherent, cross-linked structure, with bonds both between the hydroxy-L-proline-rich, wall protein extensin and wall polysaccharides, and between individual polysaccharides. 

The hydroxy-L-proline of the crude, mung-bean-wall preparation is not associated with the lectin259 this indicates a difference between the mung-bean-wall lectin and the lectin isolated from potato tubers. The latter is rich in hydroxy-L-proline, arabinose, and galactose260 in this respect, it shows features similar to those of the hydroxy-L-proline-rich glycoprotein ( extensin ) of the primary cell-wall, first reported by Lamport261 (see Section VII, 1). 

It has not been clearly established whether extensin is a lectin, but such a lectin property might allow this glycoprotein to bind noncova-lently to polysaccharides within the primary-wall structure. 

Since the discovery of the arabinosyloxy-L-proline-rich glycoprotein, sometimes referred to as extensin, in the primary cell-wall by Lamport,41 and its negative correlation with growth,263 the cell-wall matrix has been regarded by some as an extensin-polysaccharide complex. 

Fig. 8. — Partial Model of Primary Cell-Wall in Lupin Hypocotyl, Proposed by Monro and Coworkers.49 [The half of the Figure labeled (A) represents the extensin-hemicellulose network, and the half labeled (B) represents the separate, pectic network, which is believed not to involve the wall glycoprotein (extensin). Thus, the cellulose microfibrils (M) are separately cross-linked by two networks of polymers, the first (A) being composed of the wall glycoprotein and polysaccharide (probably hemicelluloses), and the second (B) being composed of the pectic polymers. These two networks have been separated in the Figure for clarity. This model is tentative and incomplete, as the nature of the linkages between the polymers in these two networks has not yet been identified. The...

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