The effects of kinetin and gibberellic acids (GA3, GA4, GA7, GA9, and a mixture of GA4,7,9) on cold hardening, dehardening, and growth of smooth bromegrass (Bromus inermis Leyss. cv. Manchar) suspension cultures treated with abscisic acid (ABA) were determined. Bromegrass cells treated with 75 μM racemic ABA for 7 days at 25 °C tolerated −37 °C, whereas cells treated with both racemic ABA (75 μM) and a mixture of GA4, GA7, and GA9 (total gibberellic acid concentration 100 μM) were similar in hardiness to the controls (LT50, −10 °C). GA4,7,9 at concentrations greater than 10 μM inhibited the growth of cells. Although 400 μM GA4,7,9 was lethal to cells, 75 μM ABA overcame the lethal effect but did not overcome growth inhibition. The twofold reduction in cell water content due to 75 μM ABA treatment for 7 days was partially overcome by GA4,7,9 at concentrations greater than 400 μM. GA4, GA7, and GA9 were equally effective at limiting growth and inhibiting freezing tolerance induced by ABA, whereas GA3 had little effect on cold hardiness, growth, and water content. During the first 4 days, kinetin at concentrations greater than 100 μM inhibited growth of both control cells and cells treated with ABA. Kinetin (> 100 μM) also inhibited freezing tolerance induced by abscisic acid after 4 days, but had no effect after 8 days. Bromegrass cells treated with 75 μM ABA for 7 days were hardened to −37 °C but dehardened to −12 °C after transfer to fresh medium minus ABA after 12 days at 10 °C. GA4,7,9 (40 μM) had no effect on the rate of dehardening, whereas kinetin increased the rate of dehardening.
Effects of Abscisic Acid Metabolites and Analogs on Freezing Tolerance and Gene Expression in Bromegrass (Bromus inermis Leyss) Cell Cultures