Stage-specific inhibition of cotton fiber development by adding α-amanitin to ovule cultures

Abstract

When cotton (Gossypium hirsutum, DPL 90ne) ovules are removed from the carpel on the day of anthesis and placed on an appropriate medium, fiber cells will differentiatein vitro from the ovule epidermis. Alpha-amanitin, an inhibitor of poly (A)+ RNA synthesis was added to ovule cultures at selected times after culture initiation to determine the timing of events in fiber development. Replicate cultures were initiated from day of anthesis, cotton ovules with α-amanitin being added to media at final concentrations ranging from 0.05μg/ml to 5.0 μg/ml. After 21 d in culture, ovules were evaluated for fiber length, percentage of ovules producing fiber, fiber dry weight, and fiber cellulose content. Cultures treated with α-amanitin on the day of anthesis to 2 d postanthesis were prevented from forming normal numbers of fiber cells. Addition of α-amanitin to cultures between 2 and 10 d postanthesis reduced the length that fiber cells attained in culture after 21 d. Addition of α-amanitin to cultures at 4 d postanthesis had as much an effect on the accumulation of cellulose in the fiber cell wall as the later addition of the inhibitor at 8 to 10 d postanthesis. The inhibitory effect of α-amanitin on fiber growth was reversible when ovules were transferred to amanitin-free medium within 2 d after culture initiation. Ovules placed on media containing α-amanitin for 5 d or longer could not overcome the inhibitory effects when transferred to media lacking the inhibitor. These results indicate critical time periods when RNA transcripts important for specific phases of development are being synthesized.