Ribonuclease Activity of Stressed Tomato Leaflets

Desiccation of 8-to 13-day-old seedlings, achieved by withholding nutrient solution from the vermiculite root medium, caused a reduction in nitrate reductase activity of the leaf tissue. Activity declined when leaf water potentials decreased below -2 bars and was 25% of the control at a leaf water potential of -13 bars. Experiments were conducted to determine whether the decrease in nitrate reductase activity was due to reduced levels of nitrate in the tissue, direct inactivation of the enzyme by low leaf water potentials, or to changes in rates of synthesis or decay of the enzyme.Although tissue nitrate content decreased with the onset of desiccation, it did not continue to decline with tissue desiccation and loss of enzyme activity. Nitrate reductase activity recovered when the plants were rewatered with nitrate-free medium, suggesting that the nitrate in the plant was adequate for high nitrate reductase activity. The rate of decay of nitrate reductase activity from desiccated tissue was essentially identical to that of the control, in vivo or in vitro, regardless of the rapidity of desiccation of the tissue. Direct inactivation of the enzyme by the low water potentials was not detected. Polyribosomal content of the tissue declined with the decrease in water potential, prior to the decline in nitrate reductase activity. Changes in ribosomal profiles occurred during desiccation, regardless of whether the tissue had been excised or not and whether desiccation was rapid or slow. Reduction in polyribosomal content did not appear to be associated with changes in ribonuclease activity. Nitrate reductase activity and the polyribosomal content of the tissue recovered upon rewatering, following the recovery in water potential. The increase in polyribosomal content preceded the increase in nitrate reductase activity. Recovery of enzyme activity was prevented by cycloheximide.Based on these results, it appears that nitrate reductase activity was affected primarily by a decrease in the rate of enzyme synthesis at low leaf water potentials.

Section: Resultssupporting

confidence: 60%

“…Among the activity of certain enzymes (1,12,7,22). One of the most severely affected is nitrate reductase which has been shown to decline to 42% of the controls in barley leaves having a T"' of about -11 bars (12) and to 50% of the controls in corn seedlings having a water deficit of 10% (1).…”

mentioning

confidence: 99%

Nitrate Reductase Activity and Polyribosomal Content of Corn (Zea mays L.) Having Low Leaf Water Potentials

Morilla¹,

Boyer²,

Hageman³

1973

“…Prolonged and severe water stress increases the ribonuclease level in plants (6). The possibility existed that the reduction in polysomes in stressed tissue was an artifact caused by increased ribonuclease activity during isolation.…”

Section: Resultsmentioning

confidence: 99%

Rapid Changes in Levels of Polyribosomes in Zea mays in Response to Water Stress

Hsiao

1970

152

The seedlings were rewatered by dipping them in 0.1 mm CaCl2 and then transferring them to paper towels moistened with 0.1 mm CaC12, in covered trays.Polysomes were isolated from the rapidly growing coleoptilar node region. The homogenizing buffer (10 mm tris-CI, 2 mM MgC92) was at pH 8.5. Isolation at pH 8.5 yielded a slightly higher proportion of polysomes and more total ribosomes per unit of tissue than at pH 6.5. High pH prevents the association of endogenous ribonuclease with ribosomes (11) and reduces the activity of maize ribonuclease (unpublished).Segments encompassing the node and the tissue about 4 mm on both sides of the node were cut and immediately buried in pulverized dry ice. Thirty segments, weighing about 0.8 g, were partially thawed and placed in 1.7 ml of ice-cold homogenizing buffer containing 0.81 mg of bentonite (11) in a micro-cup (internally ribbed, 1.4-cm diameter, 5-ml brim capacity) of a Virtis homogenizer. The tissue was homogenized at ice temperature according to a procedure suggested by Dr. J. L. Key (personal communication). The homogenizer was quickly turned to full speed (45,000 rpm) and immediately off again, three times. Surprisingly, homogenizing the tissue in the Virtis tended to yield a slightly higher percentage of polysomes than did pulverizing in powdered Dry Ice. The homogenate was clarified at 17,500 rpm (Sorvall SS-34 rotor) for 10 min. The supernate (0.4 or 0.5 ml) was directly placed on the sucrose gradient without pelleting the ribosomes. A discontinuous gradient, consisting of the "isokinetic" gradient of Noll (15), 15 to 28% of sucrose and totaling 10.3 ml, with 1.0 ml of 7.5%7c sucrose layered on top, was prepared within 3 hr of use. The extra layer allowed good separation of the monoribosomes (monosomes) from the soluble material without prolonged centrifugation, thus minimizing pelleting of the largest polysomes. All sucrose solutions were in the homogenizing buffer. The tube was centrifuged in the SB-283 rotor of an IEC B-60 ultracentrifuge at 41,000 rpm for 1 hr at 1 to 4 C. Plant Physiol. Vol. 46, 1970 The tube content was pumped with an ISCO (Instrumentation Specialties Co., Lincoln, Neb.) fractionator, model 180, through a flow cell with a light path of 2mm. Absorbance at 254 nm was recorded with an ISCO UA-2 ultraviolet analyzer.To determine the proportion of polysomes, the areas under the monomeric and polymeric peaks, arbitrarily divided (dotted lines, Fig. la), were measured with a planimeter. The absorbance base line was assumed to be horizontal. The proportion of monosome was probably overestimated because of contamination by the soluble material.The amount of total ribosomes obtained varied from extraction to extraction, probably because the degrees of homogenization varied. Estimated polysomal percentage in duplicate samples often differed by several points. Hence, small differences in polysomes, when noted herein, were always verified by repeating the experiment several times.Water potential was determined by the procedure of Shardakov (1). Abo...

“…An increase of RNase activity (1,6,7,9,12,35) and a decrease of RNA level (8,12) was found in tomato and other plants subjected to water stress. The soluble fraction of RNA increased while the ribosomal fraction decreased in sugar beet under water stress (25).…”

Section: Introductionmentioning

confidence: 97%

Abnormal Stomatal Behavior and Hormonal Imbalance in flacca, a Wilty Mutant of Tomato

Puri¹,

Tal²

1977

Plants of the wilty tomato (Lycopersicum esculentum) mutant,flacca, and of the normal cultivar Rheinlands Rubm growing under either "normal" or high humidity were used in this research. Under normal humidity, RNase activity was much higher in mutant plants in which abscisic acid (ABA) and water content were lower than in the normal plant. The mutant also contained less RNA and protein per cell and less soluble RNA relative to ribosomal RNA as compared with the normal genotype. In ABA-treated mutant plants, RNase activity decreased while RNA, protein, the ratio of soluble to ribosomal RNA and water content increased.Under high humidity, RNase activity in mutant plants was decreased, but was still somewhat higher than that in the normal plant, although water saturation deficit was equal in both plant types. Abscisic acid increased RNase activity in the mutant plants. The content of RNA and protein per cell was similar in both types, but the ratio of soluble to ribosomal RNA remained lower in the mutant. In ABA-treated mutant plants, although the content of DNA and RNA per fresh weight was similar to that of control mutant plants, the ratio of RNA to DNA decreased significandy. In addition, ABA caused an increase of the soluble to ribosomal RNA ratio toward the normal value in mutant plants.Contrary to ABA, kinetin increased RNase activity in the mutant under normal humidity and decreased it under high humidity.A similar incorporation of labeled uridine into RNA in normal, mutant, and ABA-treated mutant plants under normal humidity suggests that the difference between mutant and normal plats in respect to total, soluble, and ribosomal RNA results not from a different rate of RNA synthesis but from a different rate of RNA degradation, ie. RNase actvity.An increase of RNase activity (1,6,7,9,12,35) and a decrease of RNA level (8,12) was found in tomato and other plants subjected to water stress. The soluble fraction of RNA increased while the ribosomal fraction decreased in sugar beet under water stress (25). Low relative humidity appears to be associated with slow accumulation and reduced incorporation of labeled uracil into RNA in etiolated leaves of Jack bean (3).An association between RNase activity, RNA level, and hormone balance in the plant was also reported. Srivastava (29) found an increase of RNase activity in detached barley leaves treated with ABA and a decrease of this activity following treatment with kinetin. Similar hormonal effects on RNase activity were reported in other plants (5,13,28,39 The first attempt to elucidate the interrelations among RNase activity, water content, and hormone balance was made by Arad et al. (1). They found that in barley leaves, in which the water balance was not disturbed, ABA increased and kinetin decreased RNase activity. However, these hormonal effects were inverted in leaves subjected to water shortage i.e. ABA decreased and kinetin increased the activity. They interpreted these results to mean that above a certain optimal leaf water content, the specific ho...