Plant Growth Regulation 25: 35–45, 1998. 35 c 1998 Kluwer Academic Publishers. Printed in the Netherlands. Effects of abscisic acid and abscisic acid analogs on the induction of freezing tolerance of winter rye (Secale cereale L.) seedlings Grant C. Churchill 1 , Martin J.T. Reaney 1 , Suzanne R. Abrams 2 & Lawrence V. Gusta 3 1 Crop Science Department, University of Saskatchewan, Saskatoon SK S7N 5A8 (present address: Department of Biochemistry, 140 Gortner Lab, University of Minnesota, 1479 Gortner Avenue, St. Paul, MN 55108); 2 Plant Biotechnology Institute, National Research Council of Canada 110 Gymnasium Place, Saskatoon SK S7N OW4 Canada (present address: Crop Utilization Research Unit, Research Branch, Agriculture and Agri-Food Canada, 118 Veterinary Road, Saskatoon SK, S7N 2R4, Canada); 3 Crop Development Centre, University of Saskatchewan, 51 Campus Drive, Saskatoon, SK S7N 5A8, Canada (corresponding author: tel: 306-966-4974; fax: 306-966-5015) Received 7 May 1997; accepted in revised form 8 December 1997 Key words: abscisic acid, analogs, freezing tolerance, cereals Abstract The ability of abscisic acid (ABA) and abscisic acid analogs to induce freezing tolerance in fall rye (Secale cereale cv Puma) seedlings grown at nonhardening temperatures was investigated. Analogs were constructed with systematic alterations at C-1 (acid replaced with methyl ester, aldehyde or alcohol), at C-4, C-5 (trans double bond replaced with a triple bond), and at C-2 , C-3 (double bond replaced with a single bond so that the side chain and C-2 methyl groups were cis). Freezing tolerance (LT 50 ) was determined 3, 4 and 6 days after the first of two consecutive applications of chemical (100 M) to either the leaves or roots. All analogs were more effective when applied to the plant roots than when applied to the leaves. ABA, acetylenic ABA and 2 ,3 -dihydroacetylenic ABA decreased the LT 50 from 3 C (control) to 9 C. Consistent structure-activity relationships were only detected following root application. No single functional group altered was absolutely required for activity. The effect of any given change to the molecule was modified by the presence of other functional groups. For example, substituting the double bond in the ring with a single bond decreased activity, but concomitant substitution of the trans double bond in the side chain with a triple bond restored activity. In general, analogs with a cis, trans side chain were more active initially but rapidly lost activity, whereas acetylenic analogs maintained or gained activity over the three sampling times. The application of gibberellin biosynthesis inhibitors (100 M; tetcyclacis or mefluidide) did not increase freezing tolerance beyond that induced by ABA, either alone or in combination with ABA. It can be concluded that ABA and certain ABA analogs can induce limited freezing tolerance in whole rye seedlings, and partially substitute for low temperature acclimation. Introduction Untimely frosts cause tremendous economic loss to agricultural crops annually (Caple 1988). This loss could be reduced if it was possible to induce chemically the onset of freezing tolerance. Attempts to increase freezing tolerance with exogenous ABA began soon This work was supported in part by grants to Lawrence V. Gusta from Western Grain Research, Natural Science and Engineering Research Council of Canada. after ABA was suspected to play a role in cold hard- ening (Irving and Lanphear 1967). Exogenous ABA can dramatically harden cultured cells from species capable of acclimation (Chen and Gusta 1983; Reaney and Gusta 1987). Unfortunately, successes appear to be limited to studies in which ABA was applied to the roots under sterile conditions (Rikin et al. 1975; Chen et al. 1979; Chen et al. 1983a; L˚ ang et al. 1989). Moreover, only in plants of limited hardiness potential does ABA-induced tolerance approach low temperature-induced tolerance.