Enzyme synthesis in the cotyledons of germinating seeds

Abstract. Pea cotyledonary a-amylase increases dramaticallv both in specific activity and total activity between days 7 to 10 when germination occurs in the dark. This enzymatic activity does not seem to appear as a consequence of release or formation of an activator, removal of an inhibitor, dissociation of an inactive amylase complex, or proteolytic decomposition of a zymogen precursor. The possibility remains that the a-amylase is newly synthesized during germination. The preparation and properties of a cell-free protein-synthesizing system from germinating pea cotyledons is described; polyuridylic acid must be added for L-phenylalanine incorporation. Active microsomal preparations can be obtained from cotyledons germinated 10 days.Seed germination is frequently accompanied bv marked increases in the level of activity of certain enzymes (3,4,12,17, 23,32). It has been noted that these often dramatic increases in biocatalytic activity are not necessarily associated only with those parts of the seedling that are actively growing. Rather. the same phenomenon is also observed in the storage tissues of the endosperm or cotyledon (4, 17, 32).We showed in previous reports '(25, 26) that a number of enzvmes required for the degradation of seed carbohydrate reserves are present in the cotvledon of the germinating pea, and at least 2 of these enzymes, an a-amylase and a phosphorylase. increase manyfold in total activity during the time period of germination. Since, in many instances, the cells of plant storage organs neither grow nor divide, the question arises as to what the origin of such new enzymatic activities mlay be. Other workers dernonstrated that activation of latent enzyme occurs in the case of the ,-amylase of wheat (21,22), zymogen activation causes an increase in acid phosphatase and isocitritase activiti-es of a variety of seeds (17), whereas isocitritase (6,10). malate synthetase (10), tyramine methylpherase (12), and barley a-amvlase (5, 28) increase in activity as a result of de novo enzyme synthesis.We report here studies which show that cotvledonary a-amylase of peas most likely does not appear during germination as a consequence of the formation or release of a specific activator, the removal of an inhibitor, or the activation of a latent form (or zymogen precursor) of the enzyme. The possibility that the enzyme is newly synthesized is not eliminated.A cell-free protein-synthesizing system from the cotyledons of germinating peas has been prepared and its properties determined. A catalytically-active microsomal fraction can be prepared from cotyledons at a time during germination when the level of oa-amvlase activity is rapidlv increasilng.

“…Rather. the same phenomenon is also observed in the storage tissues of the endosperm or cotyledon (4,17,32).…”

supporting

confidence: 67%

mentioning

confidence: 99%

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Seed Germination Studies. III. Properties of a Cell-free Amino Acid Incorporating System From Pea Cotyledons; Possible Origin of Cotyledonary α-Amylase

Swain

Dekker

1969

“…The question of whether protein synthesis occurs in the cotyiledon dnring degradation of proteins in 7vivo has been investigated by Youing (24), Cherry (6), and Varner and Chandra (22). The cotyledon, cell poptlulatioil is primarily composed of large parenchyma cellss, whose major role is to suipplyl the embryonic axis with nutrients duirinig the early stages of germination.…”

mentioning

confidence: 99%

De Novo Synthesis of Isocitritase in Peanut (Arachis hypogaea L.) Cotyledons

Gientka-Rychter¹,

Cherry²

1968

Receiv-ed NovTember 20, 1967. A bstract. Germination of peanut seed is accompanied by a rapid increase in isocitritase (isocitrate lyase, EC 4.1.3.1) during the first 4 days. The presence of cycloheximide (50 tg/ml) during water imbibition inhibited the increase in isocitritase aotivity. Actinomycin D conversely did not inhibit isocitritase activity until the second day of imbibition while RNA syn-thesis was inhibited. Germination of peanut seed in 14C-reconstituted amino acids followed by fractionation of a 20 to 35 %c ammonium sulfate preparation on a Sephadex G-200 column (57-fold purification) showed that the active enzymic fraction coincided with a large peak of radioactivity. Germination of peanut seed in 45 %o D^O followed by enzyme purification and CsCI equilibrium cenitrifugation revealed that all the enzyme from D,O seed had a higher density than normal isocitritase. These data indicate that isocitritase in peanut seed is synthesized de novo.

“…Activities of succinate dehydrogenase, cytochrome oxidase, pyruvate dehydrogenase, and isocitrate dehydrogenase from different mitochondrial subfractions of aging cotyledons were determined. In general, the enzyme activities, adenosine diphosphate to oxygen ratios, and the ability of mitochondria to swell and contract followed the same pattern as for respiratory control. Previous research has shown that the cotyledons of germinating seedlings and other plant tissues go through many biochemical changes at the subcellular level during aging (1,4,12,16,21,25,28). Lado and Schwendimann (16) showed that in castor bean endosperm, mitochondrial proteins and enzyme activities decrease rapidly during seed maturation, while other workers (1) observed a rapid increase during the earliest part of germination.…”

mentioning

confidence: 99%

Changes in the Respiratory, Enzymatic, and Swelling and Contraction Properties of Mitochondria from Colytedons of Phaseolus vulgaris L. during Germination

Malhotra

Spencer

1970

Mitochondria isolated from cotyledons of germinating wax beans (Phaseolus vulgaris L.) showed fairly good respiratory control on days 1 and 2 after planting. The respiratory control was completely lost from days 3 to 5. During this period mitochondria were shown to be very leaky, losing about 88% of their total nicotinamide adenine dinucleotide to the suspending medium in a short time. The respiratory control was partially recovered by day 7, after which it completely disappeared again. By the use of differential centrifugation, the mitochondria were divided into subfractions by sequential centrifugation: 10,000g for 5 minutes, 25,000g for 5 minutes, and 40,000g for 5 minutes. The 10,000g subfraction was responsible for the recovery of mitochondrial activity (respiratory control value, adenosine diphosphate to oxygen ratio, and rate of oxygen utilization), on day 7. Activities of succinate dehydrogenase, cytochrome oxidase, pyruvate dehydrogenase, and isocitrate dehydrogenase from different mitochondrial subfractions of aging cotyledons were determined. In general, the enzyme activities, adenosine diphosphate to oxygen ratios, and the ability of mitochondria to swell and contract followed the same pattern as for respiratory control.Previous research has shown that the cotyledons of germinating seedlings and other plant tissues go through many biochemical changes at the subcellular level during aging (1,4,12,16,21,25,28). Lado and Schwendimann (16) showed that in castor bean endosperm, mitochondrial proteins and enzyme activities decrease rapidly during seed maturation, while other workers (1) observed a rapid increase during the earliest part of germination. Opik (21) studied the changes in respiration rate and mitochondrial activity in bean cotyledons. She found that while oxygen uptake of the intact cotyledons rose to a peak between the 3rd and 5th days of germination, mitochondrial activity decreased from 36 hr onwards and in most of the storage cells mitochondrial cristae had become swollen and matrix had darkened by the 3rd day. Since the mitochondria are highly important as centers of energy production, one would expect changes in the physiological state of cotyledons to be reflected in the mitochondrial activity. This report concerns the changes in the bean cotyledon mitochondrial activity with aging. The mitochondrial activity is measured by the ability of mitochondria to oxidize different substrates and to swell and contract, and by the activities of various mitochondrial enzymes. MATERIAIS AND METHODSThe bean seeds (Phaseolus vulgaris L. var. Kinghorn wax) were soaked in running tap water for 4 hr and then planted in vermiculite (horticulture grade). The seedlings were grown in the dark at 27 C. The cotyledons from the germinating seedlings were removed at the required age, washed with deionized water, and used as the source of mitochondria. Mitochondria were prepared from about 50 g of cotyledons by the following method: the cotyledons were gently crushed with a mortar and pestle for 90 sec in...