Further examination of abscission zone cells as ethylene target cells in higher plants

Abstract

Background and aims: Two aspects of the competence of abscission zone cells as a specific class of hormone target cell are examined. The first is the competence of these target cells to respond to a remote stele-generated signal, and whether ethylene acts in concert with this signal to initiate abscission of the primary leaf in Phaseolus vulgaris. The second is to extend the concept of dual control of abscission cell competence. Can the concept of developmental memory that is retained by abscission cell of Phaseolus vulgaris post-separation in terms of the inductive/repressive control of beta-1,4-glucan endohydrolase (cellulase) activity exerted by ethylene/auxin be extended to the rachis abscission zone cells of Sambucus nigra?

Methods: Abscission assays were performed using the leaf petiole-pulvinus explants of P. vulgaris with the distal pulvinus stele removed. These (-stele) explants do not separate when treated with ethylene and require a stele-generated signal from the distal pulvinus for separation at the leaf petiole-pulvinis abscission zone. Using these explants, the role of ethylene was examined, using the ethylene action blocker, 1-methyl cyclopropene, as well as the significance of the tissue from which the stele signal originates. Further, leaf rachis abscission explants were excised from the compound leaves of S. nigra, and changes in the activity of cellulase in response to added ethylene and auxin post-separation was examined.

Key results: The use of (-stele) explants has confirmed that ethylene, with the stele-generated signal, is essential for abscission. Neither ethylene alone nor the stelar signal alone is sufficient. Further, in addition to the leaf pulvinus distal to the abscission zone, mid-rib tissue that is excised from senescent or green mid-rib tissue can also generate a competent stelar signal. Experiments with rachis abscission explants of S. nigra have shown that auxin, when added to cells post-separation can retard cellulase activity, with activity re-established with subsequent ethylene treatment.

Conclusions: The triggers that initiate and regulate the separation process are complex with, in bean leaves at least, the generation of a signal (or signals) from remote tissues, in concert with ethylene, a requisite part of the process. Once evoked, abscission cells maintain a developmental memory such that the induction/repression mediated by ethylene/auxin that is observed prior to separation is also retained by the cells post-separation.

Fig. 1.

Recognition, using western analysis, of the pI 9·5 isoform of β-1,4-glucan endohydrolase in extracts of ethylene-treated pulvinus (Pv; 1 µg total protein loaded), ethylene-treated abscission zone (Az; 1 µg total protein loaded) and ethylene-treated green leaf mid-rib (Md; 4 µg total protein loaded) after separation using SDS–PAGE. Antibodies were raised against a recombinant pI 9·5 isoform after expression of pBAC1 in E. coli. Antibody recognition was determined using the Supersignal West Pico Chemiluminescent system.

Fig. 2.

Cellulase activity in extracts of the leaf rachis abscission zone of S. nigra. Explants were maintained in a sealed atmosphere in which endogenous ethylene accumulated and at 24 h were treated with either 2·0-μL droplets of H₂O and enzyme activity measured 24 h later (at 48 h; ▴ — ▴), or with IAA (arrowed) and enzyme activity measured 24 h later (at 48 h; ▴ — •). At 48 h, the H₂O-treated explants were again treated with H₂O and cellulase activity measured 24 h later (at 72 h; ▴ — ▴) and the IAA-treated explants were either treated again with IAA (arrowed) and cellulase activity measured 24 h later (at 72 h; • — •), or with H₂O and cellulase activity measured 24 h later (at 72 h; • — ▪). Values are means ± s.e., n = 5.