Relationship between Freezing Tolerance of Root-Tip Cells and Cold Stability of Microtubules in Rye (Secale cereale L. cv Puma)

Abstract

The response of cortical microtubules to low temperature and freezing was assessed for root tips of cold-acclimated and non-acclimated winter rye (Secale cereale L. cv Puma) seedlings using indirect immunofluorescence microscopy with antitubulin antibodies. Roots cooled to 0 or -3 degrees C were fixed for immunofluorescence microscopy at these temperatures or after an additional hour at 4 degrees C. Typical arrays of cortical microtubules were present in root-tip cells of seedlings exposed to the cold-acclimation treatment of 4 degrees C for 2 days. Microtubules in these cold-acclimated cells were more easily depolymerized by a 0 degrees C treatment than microtubules in root-tip cells of nonacclimated, 22 degrees C-grown seedlings. Microtubules were still present in some cells of both nonacclimated and cold-acclimated roots at 0 and -3 degrees C; however, the number of microtubules in these cells was lower than in controls. Microtubules remaining during the -3 degrees C freeze were shorter than microtubules in unfrozen control cells. Repolymerization of microtubules after both the 0 and -3 degrees C treatments occurred within 1 h. Root tips of nonacclimated seedlings had an LT-50 of -9 degrees C. Cold acclimation lowered this value to -14 degrees C. Treatment of 22 degrees C-grown seedlings for 24 h with the microtubule-stabilizing drug taxol caused a decrease in the freezing tolerance of root tips, indicated by a LT-50 of -3 degrees C. Treatment with D-secotaxol, an analog of taxol that was less effective in stabilizing microtubules, did not alter the freezing tolerance. We interpret these data to indicate that a degree of depolymerization of microtubules is necessary for realization of maximum freezing tolerance in root-tip cells of rye.