A comparison of the response of seedlings to heat shock, cadmium chloride and lannate

The response of plumules and radicles of etiolated maize seedlings (5 days old) to heavy metal treatment involves both a general reduction in protein synthesis and the induction of a group of polypeptides with Mr's and pI's similar to those of heat shock proteins (HSPs) (MNL 60:92, 1986). The carbamate insecticide lannate (active ingredient:methomyl, C5HN2OS2) has a similar impact on protein synthesis in plumules but not radicles of etiolated maize seedlings.

Fluorographic analysis of PAGE separations of proteins extracted from plumules and radicles treated with varying doses of cadmium chloride or lannate (3 hours) and labelled with 35S-methionine (during the last 2 hours of treatment) revealed that cadmium chloride-induced polypeptides are synthesized following treatment with 0.2 mM but not after treatment with 0.02 mM cadmium chloride. Lannate-induced polypeptides are synthesized in plumules of maize seedlings following treatment with 8.5 ml/100 ml H2O but not following treatment with 3.4 ml/100 ml H2O. The recommended field dose for lannate is 1.7 ml/100 ml H2O.

Protein synthesis (monitored as incorporation of 35S-methionine into TCA precipitable material, cpm/ug protein) is reduced approximately 40% following treatment with 0.2 mM cadmium chloride but is not altered significantly by treatment with 0.02 mM cadmium chloride. Following treatment with lannate (8.5 ml/100) protein synthesis in plumules is reduced approximately 60%. It is interesting to note that exposing intact maize seedlings to a heat shock (42.5 C, 3 hours) does not markedly reduce protein synthesis in plumules and radicles. Raising the concentration of cadmium chloride to 2 mM results in a 90% reduction in protein synthesis of treated plumules and radicles and synthesis of cadmium chloride-induced polypeptides remains evident. Following treatment of plumules with 17 ml/100 ml H2O lannate, protein synthesis is reduced by at least 90% while synthesis of lannate-induced polypeptides remains evident.

Fluorographic analysis of PAGE separations of immunoprecipitates obtained using polyclonal antibodies raised against low Mr (18 Kd) HSPs has revealed the presence of a single polypeptide band (Mr 18 Kd) immunoprecipitated from proteins synthesized in cadmium chloride treated (5 mM; 3 hours) plumules and radicles and in lannate treated (8.5 ml/100 ml H2O; 3 hours) plumules. Cadmium chloride treatment administered to plumules and radicles of maize seedlings therefore induces the synthesis of a group of polypeptides with the same Mrs and pIs as 18 Kd HSPs and which are recognized by antibodies raised against these HSPs. Lannate treatment, however, only induces synthesis of these polypeptides in treated plumules.

Following treatment with lannate (8.5 ml/100 ml H2O; 3 hours) protein synthesis in radicles of intact maize seedlings is reduced by at least 90%. HSP-like polypeptide synthesis in radicles is not apparent following treatment with any concentration of lannate so far tested. Antibodies raised against 18 Kd HSPs do not precipitate a polypeptide from proteins extracted from lannate treated radicles. Lannate is a solution (1.48 M) of methomyl in methanol (68%) (Dupont). Methanol treatment alone does not induce the synthesis of HSP-like polypeptides in plumules or radicles at any of the concentrations tested (1.2%, 2.4%, 5.0% or 12.0%; these concentrations represent the amount of methanol in 1.7, 3.4, 8.5, 17.0 ml/100 ml H2O lannate respectively). General protein synthesis decreases as the concentration of methanol increases but the decrease is not as rapid as following treatment with increasing concentrations of lannate. It remains to be determined whether methomyl alone will induce the synthesis of HSP-like polypeptides or whether this effect is synergistic, requiring the presence of methanol.

Methomyl is selectively toxic to maize lines carrying the male sterile (T) cytoplasm. The effect of methomyl on plants with the T cytoplasm is similar to that of T-toxin isolated from Helminthosporium maydis race T (HMT) (Humaydan and Scott, 1977; Hortscience 12:312-313). Both methomyl and T-toxin block oxidative phosophorylation in mitochondria isolated from plants with the T-cytoplasm (Berville et al. 1984, Plant Physiol. 76:508-519). A preliminary comparison of the response of plants (Oh51A) with a normal male fertile cytoplasm (N or carrying the male sterile cytoplasms T, S or C) to heat shock and lannate does not reveal any major differences in the polypeptide synthesis patterns (after ID-PAGE) in plumules of the four maize lines. Currently the response of isolated mitochondria from these four lines to heat shock and lannate are being compared.

Carol A.B. Rees, Annette M. Gullons and D.B. Walden
 
 


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