Maize embryos, seedlings and mature plants respond rapidly and dramatically to the stress of a rapid increase to supraoptimal temperature (heat shock; hs) by (1) the new and/or enhanced synthesis of a group of polypeptides, the heat shock polypeptides (hsps) and (2) the reduced synthesis of polypeptides made at a control temperature (Baszczynski MGCNL 56:111,1982).
When seedlings are maintained under temperature stress conditions for prolonged periods, synthesis of hsps gradually decreases and the rate of diminution is hsp specific (Rockwell, Lehman and Nebiolo MGCNL 59:78, 1985; Baszczynski, Ph.D. Thesis, 1985). Reports of the time required for the disappearance of hsp synthesis following prolonged temperature stress vary substantially. P. Cooper and T.-H.D. Ho (Plant Physiol. 71:215,1983) observed the complete disappearance of hsp synthesis in excised roots after 10 hours of temperature stress while Rockwell et al. (1985), working with plumules of intact seedlings (Oh43), observed synthesis of some hsp classes following 18 hours of temperature stress. The disparity may reflect a tissue or genotype specific nature for the response or it may be due to the effects of environmental factors other than temperature. We attempted to determine if altering the humidity level would change the response to prolonged temperature stress.
Intact seedlings (Oh43) were maintained at 25 C (control) under conditions of low humidity (dry; seedlings were maintained on almost dry filter paper) or shifted to 42 C under conditions of high humidity (humid; seedlings were maintained on moist filter paper) or low humidity. At two hour intervals plumules of intact seedlings, incubated in 35S-methionine (0.01 uCi/uL) for two hours, were excised and the proteins extracted in 200 mM Tris-HCI pH 7.4, 5% SDS, 7.5% 2-mercaptoethanol, 1 mM PMSF Polypeptides were subjected to SDS-PAGE and following fluorography the patterns of newly synthesized polypeptides were compared. The identification of hsps was confirmed using antibodies prepared against either the low molecular weight (18kD) or high molecular weight (73 kD - 89 kD) group of hsps (Baszczynski MGCNL 58:134,1984). Following labelling with 35S-methionine as described above, proteins were extracted from plumules in 50 mM Tris, pH 7.2, 0.15 M NaCl, 1% NaDOC, 1% Triton X-100 and 0.1% SDS and immunoprecipitated with either antibody type. lmmunoprecipitates were subjected to SDS-PAGE and fluorography and the presence of hsps among newly synthesized polypeptides was noted.
The level of hsp synthesis was not dramatically reduced during the first 8 hours of temperature stress in plumules maintained under humid or dry conditions. By 10 hours, no further synthesis was observed of most hsp classes in plumules incubated under humid conditions; under dry conditions, plumules continued to synthesize all hsp classes for at least twelve hours, although in reduced amounts. By 24 hours the pattern of polypeptide synthesis in plumules under humid conditions resembled that of control plumules but plumules maintained under dry conditions exhibited a new pattern of polypeptide synthesis which resembled neither that of control nor heat shock plumules.
The difference in the rate of disappearance of heat shock polypeptide synthesis under conditions of low and high humidity may reflect a difference in the physiological condition of the seedlings. We are examining the effect of the humidity level on the ability of seedlings to survive prolonged temperature stress.
Carol A. B. Rees, D. B. Walden, Bruce H. Rockwell1 and Christine M. Nebiolo1
1Allegheny College,
Meadville, Pennsylvania
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