Inhibition of Auxin-Induced Cell Elongation of Maize Coleoptiles by Antibodies Specific for Cell Wall Glucanases

Abstract

Polyclonal antibodies were raised in rabbits in response to the administration of purified exo-and endoglucanases extracted from cell walls of maize (Zea mays L. B37 x Mo17) coleoptiles. Since the antibodies formed specific conjugates when challenged with the glucanase antigens in immunoblot assays they were employed to evaluate the participation of glucanases in tissue growth. lndole-3-acetic acid induced cell elongation of abraded coleoptile segments was inhibited when the antibodies were supplied as a short term pretreatment (25-200 microgram/milliliter of serum protein). The extent of inhibition of IAA induced cell elongation was additive when endo-and exoglucanase antibodies were applied together. The results suggest that both enzymes have a role in mediating IAA-induced cell elongation. Pretreatment with exo-and endoglucanases antibodies also inhibited IAA induced degradation of noncellulosic jO-D-glucans and the increased level of cellulosic polymers in maize coleoptiles. Antibodies also inhibited the expression of the autohydrolytic degradation of glucans in isolated cell walls. The extent of inhibition was dependent on the antibody concentration applied. The results support the contention that enzymatic processes mediated by exo-and endoglucanases are responsible for cell wall autolytic reactions and that these reactions are linked to the mechanism for expressing auxin induced cell elongation in maize coleoptiles.applied for this purpose because of its capacity to inhibit specific enzymes (20). Nojirimycin also suppressed both auxin-induced glucan degradation in vivo and cell elongation. However, the development ofantibodies offers an opportunity to conduct experiments that would complement the interpretation of results of experiments where inhibitors act to mimic substrates. Huber and Nevins (10) had first examined the physiological activity of antibodies generated in response to the administration of the LiCl extract of cell wall proteins to rabbits. These antibodies suppressed glucan autolysis and cell elongation ofmaize coleoptiles. An extension ofthat approach using more refined protein fractions reveals that only three maize cell wall proteins generate antibodies with the capability of inhibiting growth and for one of these a capacity for enzymatic activity has yet to be identified. Antibodies raised in response to specific wall glucanases offers an attractive approach for resolving their participation in key steps ofauxin responses (8).We evaluated the role of cell wall glucanases in glucan metabolism and in IAA-induced cell elongation of maize coleoptiles using antibodies raised against purified exo-and endoglucanases. MATERIALS AND METHODS Plant MaterialNoncellulosic ,B-D-glucans of cereal coleoptile cell walls are metabolized during cell extension (16,17,(22)(23)(24)26). While the precise role of glucan metabolism is not clear the process is governed by auxin and the key metabolic steps may relate directly to cell wall changes required for cell elongation ( 18-20). Cell walls isolated ...

How Do Plant Cell Walls Extend?

“…This idea has been further supported in recent years by studies in which selective reagents were used to interfere with wall glycanase activity. Treatment of corn coleoptile segments with polyclonal antibodies against cell wall glycanases interfered with auxininduced growth, wall autolysis, and changes in wall viscoelasticity (Inouhe and Nevins, 1991). In a similar vein, antibodies and lectins that recognize xyloglucans in azuku bean (Hoson and Masuda, 1991) or (1+3, 1+4)-P-~-glucans in maize coleoptiles interfered with auxin-induced growth in excised sections.…”

Structural models of primary cell walls in flowering plants: consistency of molecular structure with the physical properties of the walls during growth

“…Although the data obtained so far are sometimes more confusing than enlightening, antibodies offer a novel approach to probe how the interruption of normal orientation can disturb wall expansion. For example, Nevin's group has prepared antibodies to putative exo-and endo-glucanases purified from the walls of growing seedlings, and these antibodies partially reverse auxin-induced growth (Inouhe and Nevins, 1991). Antibodies to the p-o-glucan itself can also inhibit growth (Hoson and Nevins, 1989).…”

Comparison of the Outer and Inner Epidermis

“…The cause-effect relationship between auxin-induced changes in chemical structure of cell wall polysaccharides and elongation growth has been examined using lectins (4,5), antibodies raised against cell wall polysaccharides (6), and proteins (3,8,11,12,15,22). The results obtained in the present study further confirm the usefulness of antibodies specific for cell wall components for understanding the mechanism by which auxin regulates elongation growth of higher plants.…”

“…Of those wall components that constitute Poaceae cell walls, the (1--3),(1l-4)-f-D-glucans are the prominent polysaccharides undergoing auxin-induced changes. Glucan metabolism has been observed during auxininduced elongation of Avena (16,23), Hordeum (24), Zea (12), and Oryza (26) coleoptiles. Two enzymes, an endo-and an exoglucanase, are involved in the hydrolysis of the polysaccharide in maize (Zea mays L.) coleoptiles (2,9,10).…”

“…Two enzymes, an endo-and an exoglucanase, are involved in the hydrolysis of the polysaccharide in maize (Zea mays L.) coleoptiles (2,9,10). Antibodies raised against (1--3),(1-+4)-f3-D-glucans (6) and glucanases (8,12,15,22) are effective inhibitors of auxininduced elongation, cell wall loosening, and the degradation of the glucans in maize coleoptile segments. The data support the concept that degradation of (1--3),(1 --4)-f3-D-glucans by cell wall glucanases is associated with cell wall loosening responsible for auxin-induced elongation.…”