Cytokinin antagonists and antibiotics related to nalidixic acid : effects on Phaseolus and Nicotiana callus tissues Public Deposited

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  • The effects of five cytokinin antagonists on the growth of cytokinin-autonomous lines of Phaseolus lunatus cv. Kingston, Phaseolus vulgaris cv. Great Northern, and Nicotiana tabacum cv. Wisconsin 38 callus tissues have been compared. The antagonists tested included four N⁴-substituted 4-amino-2-methylthiopyrrolo[2,3-d]pyrimidines bearing the following N⁴ side chains, 4-n-pentyl- (ms²pnN⁴Prp), 4-n-hexyl- (ms²HxN⁴Prp), 4-cyclopentyl- (ms²cPnN⁴prp), and 4-cyclohexyl- (ms²cHxN⁴prp) and one 7- amino-3-methylpyrazolo[4,3-d]pyrimidine with the N⁷ side chain, 7-n-pentyl- (m³pnN⁷pzp). All antagonists tested inhibited Nicotiana and Phaseolus callus tissues growth. Inhibition of the Phaseolus callus tissues required higher antagonist concentrations than required to inhibit Nicotiana callus tissue. With all three tissues, the most potent of the antagonists was m³PnN⁷Pzp. However, the antagonist structure/activity relationships observed in tests with the pyrrolopyrimidine derivatives were not identical for all three tissues. The inhibitory effects of ms²PnN⁴Prp and m³PnN⁷Pzp on the growth of cytokinin-autonomous N. tabacum and P. lunatus callus tissues were partially reversed by the addition of exogenous N⁶-(Δ²-isopentenyl)adenine (i⁶Ade), as evidenced by the effect of i⁶Ade in increasing the concentrations of antagonists required to produce 50% inhibition of callus growth (I₅₀ values). The interactions of cytokinin antagonists and exogenous cytokinins in the cytokinin-autonomous Nicotiana callus tissue appear to be more complex than in the corresponding cytokinin-dependent Nicotiana callus tissue. The exogenously supplied cytokinins are themselves inhibitory to the growth of the cytokinin-autonomous line of Nicotiana callus tissue. Nalidixic acid, a compound reported to inhibit mammalian cell cultures in a manner reversible by cytokinin treatment (Quesney-Huneeus et al., (1980), Proc. Natl. Acad. Sci. USA., 77 5842.), was tested along with the related antibiotics, oxolinic acid and novobiocin, for effects on the growth of cytokinin-autonomous and cytokinin-dependent P. lunatus cv. Kingston and N. tabacum cv. Wisconsin 38 callus tissues. This investigation was undertaken to determine whether interactions of cytokinins and nalidixic acid similar to those observed in mammalian cell cultures could be detected in plant cell cultures. All three antibiotics inhibited growth of the callus tissues. The Nicotiana callus tissues were more sensitive than the Phaseolus tissues. In all tissues, oxolinic acid was the most effective of the antibiotics in inhibiting callus growth, and novobiocin was the least effective of the three antibiotics. The inhibitory effects of nalidixic acid and oxolinic acid on cytokinin-dependent Phaseolus callus tissue appeared to be partially alleviated by exogenous i⁶Ade treatment as evidenced by the effect of i⁶Ade in increasing the I₅₀ values for nalidixic acid and oxolinic acid. By this criterion, the inhibitory effects of novobiocin on callus growth did not appear to be reversed by treatment with exogenous i⁶Ade. Interestingly, in tests with the cytokinin-autonomous Nicotiana callus tissue, a low concentration (0.01 μM) of nalidixic acid appeared to reverse the inhibitory effects of high i⁶Ade concentrations. A similar effect with this tissue was observed in tests of the antagonist m³PnN⁷Pzp. The results of this study raise the possibility that some part of the effects of nalidixic acid and oxolinic acid on plant tissues may be due to effects on processes specifically involving cytokinin metabolism or function. However, in no case was i⁶Ade able to completely reverse the effects of antibiotic treatment.
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