Inhibition of Auxin-induced Deoxyribonucleic Acid Synthesis and Chromatin Activity by 5-Fluorodeoxyuridine in Soybean Hypocotyl

Holm, Robert E.; Key, Joe L.

doi:10.1104/pp.47.5.606

Cited by 13 publications

(5 citation statements)

References 11 publications

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“…Since no effect of ethylene on RNA metabolism could be detected during 18 (14,16). However, when data of these studies are expressed as RNA/g fresh weight, the ethylene effect becomes insignificant, as shown with isotope experiments in the same tissue (15). Possibly the change in RNA synthesis induced by ethylene in these long term studies may be due to a prolongation of the growth phase (2).…”

Section: Discussionmentioning

confidence: 88%

Ethylene-induced Pea Internode Swelling

Eisinger

Burg

1972

Plant Physiol.

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Exposure of etiolated pea (Pisum sativum L. var. Alaska) subapical sections to ethylene disrupts normal polar cell expansion, but fresh weight increase is little affected and the tissue expands radially, swelling. Ethylene has no effect on gross ribonucleic acid metabolism before or during the period when swelling occurs, but incorporation of "4C-proline and leucine into wall-associated protein is markedly inhibited after an initial 3-hour lag period which precedes swelling. Ethylene affects the composition of this protein, altering the prolinehydroxyproline ratio. The gas also alters the optical birefringence pattern of the cell wall, indicating that the cellulose microfibrillar orientation has been changed.The normal growth pattern of excised subapical sections from the third internode of etiolated pea seedlings is a highly polar expansion. When ethylene is applied, linear expansion is markedly inhibited, but fresh weight increase is hardly affected and the tissue expands radially. Since no cell divisions occur in this internodal tissue of either intact seedlings or excised sections in the presence or absence of ethylene, the ethyleneinduced shift to nonpolar, radial expansion is due solely to cellular swelling (2). Ethylene induces subapical swelling in excised sections by a process resembling that which occurs in intact seedlings; for example, the same concentrations of ethylene are effective (6, 7). In vivo, however, ethylene reduces the rate, but greatly prolongs the duration of cellular expansion (3, 5), whereas excised sections are not so affected and invariably stop growing in 12 to 18 hr. When excised sections swell in response to ethylene, their total dry weight, weight of wall material, respiration, content of various sugars, and permeability are not altered (6, 7). Slight stimulation of RNA synthesis in response to ethylene has been noted in certain intact vegetative tissues (14, 16), but does not occur in isolated soybean hypocotyl (15) and has not previously been studied in pea internodal sections. During ethylene-induced swelling the optical birefringence pattern of the cell wall is altered (7) MATERIALS AND METHODSSeeds of Pisum sativum L. (var. Alaska) were soaked in tap water for 6 hr, planted in vermiculite, and grown in darkness at 25 C. After 7 days, 1-cm sections were cut from the third internode of the epicotyl immediately below the hook and lots of 10 or 15 were incubated with or without 10 ,d/liter of ethylene in sealed 125-ml flasks with 10 ml of medium containing 5 mM potassium phosphate buffer (pH 6.8), 2% sucrose, S M cobalt chloride, and varying amounts of IAA and 'H-or "4C-labeled substrates (6). Following isotope incubations, tissue was rinsed with distilled water and floated in ice water for 15 min to clear residual isotope from the free space. Under these conditions, sections exposed to "4C-proline (0.05 ,uc/ ml) for about 1 sec retained less than twice background count.Effects of Ethylene on Uptake. Short term uptake of "4C-proline by pea internode tissue was studied ...

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Section: Discussionmentioning

confidence: 88%

Ethylene-induced Pea Internode Swelling

Eisinger

Burg

1972

Plant Physiol.

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“…From measurements made at the end of the experimental period, 6-methylpurine (Table I, Fig. 10), actinomycin D (Tables I and II, Figs. 10 and 11), and puromycin (Table III, Fig. 13) all appeared to affect total nitrate uptake relatively less than synthesis of nitrate reductase.…”

Section: Discussionmentioning

confidence: 91%

Nitrate Uptake by Dark-grown Corn Seedlings

Jackson¹,

Flesher²,

Hageman³

1973

Plant Physiol.

212

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Five-or six-day old seedlings of corn (Zea inays L.) were exposed to 0.25 mM Ca (NO3) reveal that prior exposure to ammonium ion did not eliminate the initial lag phase. The same pattern also has been observed with cotton and tobacco depleted of nitrate prior to the experimental period and with 10-day-old corn grown exclusively with ammonium ion as the nitrogen source (11). The latter observation contrasted to the nitrate uptake pattern of similar corn plants grown with nitrate in which case nitrate uptake did not exhibit the initial lag phase. A similar increase in nitrate uptake rate has been reported recently for tobacco cell suspension cultures (8) which had been grown previously with urea and in cultures which were nitrogen starved.The present investigation with dark-grown corn seedlings was initiated to characterize more fully this nitrate uptake pattern by root tissue of higher plants. It is shown that a substance from the endosperm was essential for full development of the accelerated rate of nitrate uptake, and that this development was restricted by certain inhibitors of RNA and protein synthesis. MATERIALS AND METHODSAll experiments were conducted with corn hybrid (Zea mays L.) OH43 x B14, except that DeKalb XL45 hybrid was used to obtain part of the data of Figure 5. DeKalb XL45 and OH43 X B14 show similar patterns of nitrate uptake. Corn seeds were rinsed for 1 min in 5.25% NaOCl, placed in running tap water for 3 hr, rinsed again for 1 min in the NaOCl solution, rinsed thoroughly in deionized water, and about 150 seeds were spread on stainless steel screens (14 X 14 cm) over 4-liter containers of vigorously aerated 0.1 mM CaSO4. The seeds were covered with one layer of filter paper kept moist by wick action. Germination took place at 25 C in darkness. Four days later the seedlings were removed, and all roots except the primary root were excised. The primary roots of approximately 100 seedlings were then threaded through a stainless steel screen into a 4-liter beaker containing 0.5 mM (NH,),SO4, 0.6 mM KSO4, 0.4 mM KH,PO4, 1.6 mM MgSO,, 0.8 mm CaSO,, and 0.125 g of CaCO,/l. Iron was supplied at 4 mg/l as monosodium hydrogen ferric diethylenetriaminepentaacetate, and the remaining trace elements were at one-fifth the concentration of Hoagland's solution (9). The containers were again placed in the dark germinator at 25 C, and the solutions were aerated vigorously.After a further 24 or 48 hr, the etiolated seedlings were removed, and the roots were rinsed thoroughly in deionized water. The seedlings were then floated in trays on 0.5 mM CaSO, while unifrom seedlings were selected, assembled in groups of seven, and supported in plastic frames. Roots of each group were then blotted lightly with paper towels and placed 120 www.plantphysiol.org on May 12, 2018 -Published by Downloaded from

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“…The chromatin activity (based on fresh weight) is about 12-fold greater after a 24-hr auxin treatment; however, the specific activity (based on DNA or protein) increases only up to 12 hr, resulting in about a 5-to 7-fold increase in specific activity. Increases in total RNA polymerase activity above 5-to 7-fold presumably result from cell divisions in the mature hypocotyl (2,12 RNA Polymerase Solubilized from the Chromatin Supernatant. Since only a small amount of a-amanitin-sensitive enzyme is associated with the chromatin and no distinct RNA polymerase II peak is observed on DEAE-cellulose chromatography of solubilized chromatin proteins, high-speed supernatant fractions were examined for RNA polymerase II activity.…”

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confidence: 99%

Enhancement of soybean RNA polymerase I by auxin.

Guilfoyle¹,

Lin²,

Chen³

et al. 1975

Proc. Natl. Acad. Sci. U.S.A.

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When etiolated soybean seedlings are treated with the synthetic auxin, 2,4-dichlorophenoxyacetic acid, cells of the mature hypocotyl become swollen and proliferate abnormally. This abnormal growth induced by auxin coincides with a 5-to 8-fold increase in the a-amanitin-insensitive RNA polymerase associated with isolated chromatin or nuclei. The a-amanitin-sensitive RNA polymerase activity of the auxin-treated hypocotyl was similar to that of control tissue. The increase in RNA polymerase I activity of chromatin and nuclei was maintained after solubilization and fractionation on DEAEcellulose. Auxin thus appears to enhance RNA synthetic activity (i.e., ribosomal RNA) in mature soybean tissue by altering RNA polymerase I directly rather than by altering the chromatin template.When the synthetic auxin, 2,4-dichlorophenoxyacetic acid (2,-4-D), is applied to young etiolated soybeans, cells of the mature hypocotyl enlarge radially and proliferate, while the normal growing point or apical meristem becomes quiescent. The abnormal proliferation of the mature hypocotyl is preceded by a large increase in RNA, especially ribosomal RNA (1, 2). This large accumulation of RNA in the mature tissue is associated with an enhanced chromatin-directed RNA synthetic activity (3). The auxin-enhanced chromatin activity is insensitive to a-amanitin (4), indicating that the increased RNA synthetic activity relates to a nucleolar type RNA polymerase as described in animals (e.g., ref. 5). Hybridization studies also indicate that the chromatin-bound RNA polymerase is transcribing primarily ribosomal RNA (6). Recent studies indicate that soybean chromatin isolated by conventional methodology (7) in fact possesses only RNA polymerase I activity, while nearly all RNA polymerase II is found in high-speed supernatant fractions (8).In the present work the effects of the synthetic auxin, 2,4-D, on RNA polymerase I and II activities from both chromatin and nuclei were studied. The enzymes were solubilized and fractionated on DEAE-cellulose. RNA polymerase I activity increased several-fold after auxin treatment, while RNA polymerase II activity was at most only slightly affected. MATERIALS AND METHODSPlant Material. Soybean seeds (Glycine max, var, Wayne) that had been pretreated with 10% Chlorox were planted in Abbreviations: 2,4-D, 2,4-dichlorophenoxyacetic acid; buffer A, 50 mM Tris-HCl (pH 8.0), 10 mM 2-mercaptoethanol, 1 mM MgCl2, 250 mM sucrose, 0.1 mM EDTA, and 0.5 mM phenylmethylsulfonylfluoride; buffer B, 50 mM Tris -HCl (pH 8.0), 10 mM dithiothreitol, 5 mM MgC12, 25% glycerol, 0.1 mM EDTA, and 0.5 mM phenylmethylsulfonyliluoride. (8).Preparation of Soybean Nuclei. Nuclei were isolated and purified by a modification of the procedure described by Stern (9). The details of this procedure will be described in a forthcoming publication.Preparation of Soybean RNA Polymerases. RNA polymerase I was solubilized by stirring chromatin suspensions for 5 hr at 00 in the presence of buffer B containing 500 mM ammonium sulfate. The solubiliz...

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Inhibition of Auxin-induced Deoxyribonucleic Acid Synthesis and Chromatin Activity by 5-Fluorodeoxyuridine in Soybean Hypocotyl

Cited by 13 publications

References 11 publications

Ethylene-induced Pea Internode Swelling

Ethylene-induced Pea Internode Swelling

Nitrate Uptake by Dark-grown Corn Seedlings

Enhancement of soybean RNA polymerase I by auxin.

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