Response of Sugarcane Enzymes to Variable Light: Variable Illumination Studies of Invertase, ATP-ase and Amylase in Plants Experiencing Nitrate and Gibberellic Acid-Induced Stress

Abstract

Prior research has shown that light strongly affects sucrose-enzyme relationships in sugarcane through reactions with the herbicide Paraquat. The present study concerned: 1, Direct light relationships with invertase, amylase, and ATP-ase; and 2, the behavior of light-enzyme relationships in sugarcane having distinct regimes of growth and sugar production. Plants were pretreated with deficient and excessive amounts of nitrate (NO₃) and with foliar gibberellic acid (GA) to produce these regimes. A variable-illumination study was conducted with 16 week old sugarcane of the variety P.R.980. All plants were grown by sand culture in the greenhouse under strict water and nutritional control. Five groups of plants were prepared for study: Two of these were given 0 and 50 meq./l. of nitrate, respectively, beginning at 11 weeks of age; a third group was sprayed with a 0.01 percent GA solution at 14 weeks; a fourth set accompanied the NO₃ and GA as a darkroom control; a fifth group remained in the greenhouse throughout the study as a second control. Randomized block designs were employed with three replicates of each treatment. At 16 weeks groups one to four were placed in a darkroom. They remained without light for 96 hours, and then were reilluminated by transfer back to their original greenhouse positions. Leaf and immature storage tissues were frozen for sucrose and enzyme assay at 0, 2, 8, 24, 48, 96, 102, 120, and 144 hours. Fructose and glucose changes were studied by paper chromatography. The following results were obtained: 1. Five weeks of low and high NO₃ pretreatments succeeded in establishing simulated ripening and pre-ripening regimes, respectively. These included abnormally high sucrose and low enzyme values for N-deficient cane. Typical growth and sugar effects were achieved with high NO₃ and GA. 2. Darkened sugarcane failed to maintain sucrose levels comparable to plants exposed to the usual day-night illumination sequence. Most of the darkroom losses were made up within 6 hours after reexposure to light, indicating no damage to sucrose-forming mechanisms by prolonged darkness. Major sucrose changes were also recorded between morning and afternoon harvests. 3. Evidence of increased sucrose-forming capacity was recorded among all dark-treated plants. GA-treated sugarcane significantly surpassed other plants in improved synthetic potential. The GA effect was temporary, indicating that translocation of sucrose from leaves soon became a limiting factor. Evidence obtained by paper chromatography of leaf extracts suggests that GA also stimulated photosynthetic production of fructose and glucose. It was concluded that GA can significantly improve the potential of cane to form sucrose, but that this potential might not be realized where sugar transport is a limiting factor. 4. Sucrose differentials in NO₃-treated plants were nullified within two hours of darkness. They were not reestablished after transfer of the cane back into light. Both the NO₃-rich and NO₃-deficient treatments failed to counter the sucrose losses experienced in darkness. GA pretreatment also failed to counter sucrose losses in darkness. 5. N-deficient plants were apparently unable to translocate sucrose readily from leaves to immature storage tissue. This was attributed to poor physical condition rather than to factors of N-metabolism. 6. Invertase declined severely in darkness. Activity was fully recovered after reillumination. This supports earlier evidence of light involvement in invertase synthesis. Invertase varied markedly between morning and afternoon. These daily changes also appeared in darkened plants, suggesting that they stem from "endogenous rhythms" rather than direct light exposure. Prolonged darkness eventually ehminated the daily changes. 7. GA increased the sensitivity of invertase to light, but no evidence of a GA effect upon invertase synthesis was recorded. During early hours of darkness GA both stimulated and suppressed invertase. GA failed to arrest invertase decline in darkness, or to promote invertase recovery after reillumination. 8. Invertase in low-NO₃ plants was only slightly affected by variable illumination. Invertase was far more sensitive to light variables in plants given high NO₃, but the darkness-induced decline was not alleviated by high NO₃. 9. Leaf amylase was suppressed in the absence of light. A foliar synthetic mechanism is proposed in which an essential component is destroyed in darkness. In immature storage tissue the enzyme gradually increased activity in response to prolonged darkness. It was suggested that a natural inhibitor of the enzyme is produced in light and is destroyed in darkness. 10. Variable NO₃ modified the dark-induced decline of amylase, but failed to effectively counter the downward trend or to promote major recovery after reillumination. High-NO₃ plants achieved partial amylase recovery. A low-NO₃ suppression recorded at the beginning of light treatment was still in effect at the close of the study. 11. A low-NO₃ suppression of amylase also persisted in immature storage tissue. This behavior of amylase, frequently recorded in the past, appears to be a common denominator of sugarcane under physiological stress. Failure of light variables to alter the preestablished amylase behavior pattern is regarded as a major finding of the study. 12. Leaf amylase suppression by GA disappeared within 2 hours after dark treatment had begun. However, in immature storage tissue, a GA suppression not evident at zero hours gradually developed among darkened plants. This suppression was followed by a massive increase in amylase immediately after reillumination. It was concluded that GA increased the sensitivity of existing amylase to light, interfered with endogenous amylase rhythm, and promoted amylase synthesis. Evidence of increased synthesis of natural amylase inhibitors was also observed. 13. Foliar ATP-ase experienced moderate but persistent suppression in darkened sugarcane. Activity did not decline lower than 50 percent of zerohour values, and no recovery was made after reexposure to light. ATP-ase of darkened plants followed the same endogenous rhythm as that of greenhouse control plants. 14. The initial zero-hour suppression of ATP-ase was retained but not enhanced in darkness. High-NO₃ cane experienced more severe ATP-ase decline than did control or low-NO₃ plants. However, this decline did not pass beyond the critical level established by low NO₃. It was concluded that about half of the apparent ATP-ase work potential is sensitive to light and NO3 variables. It appears that sufficient of the ATP-ase potential is light insensitive to insure accommodation of essential ATP-ase functions. Two mechanisms of ATP-ase synthesis may be present in sugarcane. Two types of ATP-ase having different sensitivities to light is also suggested. 15. ATP-ase in the two groups of NO₃-treated sugarcane responded oppositely to both control-group changes noted immediately after reillumination. Nitrogen stress, regardless of specific NO₃ levels, transcended light and endogenous rhythm effects upon ATP-ase. 16. GA caused ATP-ase decline in immature storage tissue. About 48 hours of darkness were required to fully express the GA effect. Again, ATP-ase resisted decline below the 50-percent level.