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

Abstract

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prevents the association of endogenous ribonuclease with ribosomes (11) and reduces the activity of maize ribonuclease (unpublished).Segments encompassing the node and the tissue about 4 mm on both sides of the node were cut and immediately buried in pulverized dry ice.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...

Water Stress, Rapid Polyribosome Reductions and Growth

“…Electron micrographs obtained by Armstrong and Jones (2) and in our own laboratory of pumpkin cotyledons and pea leaves (Matsuda and Rhodes, unpublished results) suggest that the concentration of 80S cytoplasmic ribosomes found along the rough endoplasmic reticulum is reduced by water loss. Differences between our data obtained with whole roots (Table I) and those obtained with root tips (9,13) may be due to differences in the amount of membrane-associated ribosomes. Polysome Content, Tissue Water Status, and Growth Rate.…”

“…The most rapidly detected alteration in metabolism appears to be that affecting protein synthesis. Hsiao (9) (10). Rapid changes in polyribosome levels are also known to occur in mosses (3,6).…”

Detrimental Effect of Rust Infection on the Water Relations of Bean

“…There is evidence that plant water stress reduces photosynthe- (12), growth (2), and crop yield (9). Because of these reductions and because rust infection can cause unusual levels of water stress to occur throughout the plant, the effect of rust on water relations may well be the most damaging effect on the disease on the host plant.…”

Effects of Water Stress on the Activities of Three Enzymes in Maize Seedlings

“…There is also evidence (19) that water deficit impedes protein synthesis, perhaps at the ribosomal level (10,19). It is now clear that many proteins turn over at different rates (3-6, 17, 24); i.e., some apparently are formed and inactivated quickly whereas others appear to be relatively stable.…”