Freezing tolerance, protein composition, and abscisic acid localization and content of pea epicotyl, shoot, and root tissue in response to temperature and water stress

The freezing tolerance of many plants, such as pea (Pisum sativum), is increased by exposure to low temperature or abscisic acid treatment, although the physiological basis of this phenomenon is poorly understood. The freezing tolerance of pea shoot tips, root tips, and epicotyl tissue was tested after cold acclimation at 2 °C, dehydration/rehydration, applications of 10-4 M abscisic acid (ABA), and deacclimation at 25 °C. Tests were conducted using the cultivar 'Alaska', an ABA-deficient mutant 'wil', and its 'wild-type'. Freezing injury was determined graphically as the temperature that caused 50% injury (T50) from electrical conductivity. Endogenous ABA was measured using an indirect enzyme-linked immunosorbant assay, and novel proteins were detected using 2-dimensional polyacrylamide gel electrophoresis. The maximum decrease in T50 for root tissue was 1 °C for all genotypes, regardless of treatment. For 'Alaska' shoot tips and epicotyl tissue, exogenous ABA increased the freezing tolerance by -1.5 to -4.0 °C, while cold treatment increased the freezing tolerance by -7.5 to -14.8 °C. Cold treatment increased the freezing tolerance of shoot tips by -9 and -15 °C for 'wil' and 'wild-type', respectively. Cold acclimation increased endogenous ABA concentrations in 'Alaska' shoot tips and epicotyls 3- to 4-fold. Immunogold labeling increased noticeably in the nucleus and cytoplasm of the epicotyl after 7 d at 2 °C and was greatest after 30 d at the time of maximum freezing tolerance and soluble ABA concentration. Cold treatment induced the production of seven, three, and two proteins in shoot, epicotyl, and root tissue of 'Alaska', respectively. In 'Alaska' shoot tissue, five out of seven novel proteins accumulated in response to both ABA and cold treatment. However, only a 24 kDa protein was produced in 'wil' and 'wild-type' shoot and epicotyl tissues after cold treatment. Abscisic acid and cold treatment additively increased the freezing tolerance of pea epicotyl and shoot tissues through apparently independent mechanisms that both resulted in the production of a 24 kDa protein.