Evergreen Foliage Contributions to the Spring Growth of Taxus

Taxus media cv. Hicksii plants were grown one season under a low and high level of nitrogen fertilization. Before growth in the spring the plants were divided into two groups, one of which was defoliated and the other left intact. The growth and spring utilization of the nitrogen and carbohydrate reserves of defoliated plants were compared to the intact plants 0, 2, 4 and 6 weeks after growth started in the spring. The plants were separated into buds (all new growth), roots and stems and analyzed for changes in total nitrogen, basic and non-basic amino acids, hemicelluloses, soluble sugars, organic acids and chlorophyll. The older evergreen needles from plants grown under low nitrogen levels contain 20 % of the carbohydrate and 24% of the nitrogen used in spring growth. The needles from plants grown under high nitrogen levels contained 56% of the carbohydrate and 49% of the nitrogen used in spring growth. Removal of the old needles before spring growth removed this nitrogen and carbohydrate reserve and reduced the total plant chlorophyll content after 6 weeks of growth to 50% of that found in intact plants, with the result that defoliated plants did not show a growth response to nitrogen. Amino acids accumulated in the stems and buds of defoliated plants as carbohydrates became limiting. The defoliated plants removed 25% more available carbohydrates from the roots and stems than intact plants and their buds contained 50% less available carbohydrates. Plants without old needles showed similar growth rates under low and high nitrogen regimes and produced 33% of the dry weight of intact plants grown under high nitrogen levels and 66% of the dry weight of intact plants grown under low nitrogen levels. The old needles of taxus plants contain substantial amounts of reserve nitrogen and carbohydrate and these needles greatly influence the extent and rapidness of growth in the spring. When the needles are removed, the other tissues can supply an adequate amount of nitrogen but the carbohydrate supply becomes limiting for spring growth.     

Glutamine synthesis in germinating seeds of Cucurbita moschata

During germination, an increase in glutamine and glutamine synthetase[L-glutamate: ammonia ligase (ADP), EC 6.3.1.2 [EC] ] occurred inthe cotyledons reaching a maximum at 4 to 6 days. The enzymehad a Km of 4.5 nun for L-glutamate, and 0.67 mu for hydroxylamine.Hydroxylamine exhibited substrate inhibition kinetics. The enzymewas inhibited by calcium ion, fluoride ion and p-hydroxymercuribenzoatebut not by EDTA, sodium ion or chloride ion. The sulf hydrylinhibition was reversed by dithiothreitol. In vivo synthesisof glutamine-¹⁴C from glutamate-¹⁴C was found to parallel theincrease in glutamine content and the in vitro glutamine synthetaseactivity during germination. ¹ Present address: Department of Biology, Mercyhurst College,Erie, Pennsylvania 16501, U.S.A. (Received June 12, 1971; )     

Changes in Urease Activity in Apple Trees as Related to Urea Applications

Changes in urease (E.C.3.5.1.5.) were followed during the growth of 1-year-old MM 106 and 9-year-old Golden Delicious apple trees (Malus pumila Rehd.). Urease was found in leaves, roots, and bark with actively growing tissues containing more activity than senescing tissues. The urease activity in the leaves declined steadily during leaf senescence but abscised leaves still contained about half of their initial urease activity. In the bark the urease activity changed only slightly. Urease activities in the leaves and bark of apple trees were always greater in those trees which had received an application of urea. In senescing apple leaves, urea induced a rapid increase in urease activity. The changes in total activity and specific activity of urease were parallel and suggests that urease was synthesized de novo. After urease activity reached a maximum, a rapid decline occurred. Urease was inhibited by low concentrations of ammonia and this decline may be due to product inhibition.     

The Utilization of Carbohydrate and Nitrogen Reserves by Taxus During Its Spring Growth Period

The spring growth and the utilization of carbohydrate and nitrogen reserves in this growth was studied in Taxus media cv. Hicksii plants 0, 2, 4 and 6 weeks after the plants started growing in the spring. The effect of nitrogen applied the previous season on the storage and utilization of the carbohydrate and nitrogen reserves during spring growth was determined. The plants were separated into buds (all new growth), stems, needles (those produced the previous season) and roots and analyzed for changes in total nitrogen, basic and non-basic amino acids, total available carbohydrate, sugars, hemicelluloses, organic acids and chlorophyll. The bulk of the soluble nitrogen reserves were stored as arginine in the stems and old needles. With the onset of spring growth, arginine nitrogen was converted to other amino acids which accumulated in the new growth (buds). The roots, stems and needles of plants grown under high nitrogen levels always contained more total nitrogen than those grown under low nitrogen levels. The bulk of the carbohydrate reserves were stored as hemicelluloses. The plants grown under high nitrogen levels utilized the bulk of the carbohydrate reserves from the roots and smaller amounts from the stems and old needles, while plants grown under low nitrogen levels used only the reserves in the roots. In the low nitrogen plants, carbohydrates accumulated in the needles and stems. Both the carbohydrate and nitrogen reserves were important in the dry weight increase due to spring growth. However, the nitrogen reserves were the limiting factor and the high nitrogen plants grew twice as much, produced more chlorophyll, and utilized more nitrogen and carbohydrate reserve in spring growth than low nitrogen plants. The additional chlorophyll allowed the production of more carbohydrates and these additional carbohydrates were used in increased growth rates, while in the low nitrogen plants the carbohydrate produced was less and accumulated within the plant.     

Changes in Amino Acid Content and the Metabolism of γ-Aminobutyrate in Cucurbita moschata Seedlings

Ungerminated pumpkin (Cucurbita moschata Poir.) cotyledons contained 30 % of their dry weight as lipid and 26 % as protein, of which 93 % was globulin. There was a rapid degradation of these reserves 4 to 8 days after planting when the cotyledons had their maximum metabolic activity. About half of the mole percent of amino acids found in the globulin reserve was in arginine, glutamate, aspartate, and their amides. The cotyledons had a large soluble pool of arginine, glutamine, glutamate, and leucine. Most amino acids increased steadily in amount in the cotyledons during germination, except glutamine, ornithine, alanine, serine, glycine, and γ-aminobutyrate and these appeared in large amounts in the translocation stream to the axis tissue. Little arginine or proline was translocated. By 10 days, when translocation had decreased, amino acids accumulated. Ornithine, γ-aminobutyrate, and aspartate were rapidly utilized in the hypocotyl, while glutamine, glycine, and alanine accumulated there. Cysteine and methionine levels were low in the reserve, trans-location stream and soluble fractions. γ-Aminobutyrate-U^?14C injected into cotyledons or incubated with hypocotyls was utilized in a similar fashion. The label appeared in citric acid cycle acids and in the amino acids closely related to this cycle, but the bulk of the label appeared in CO₂. The labeling pattern suggests that γ-aminobutyrate was utilized via succinate, and thus entered the citric acid cycle. A close relationship between arginine, ornithine, glutamate, and γ-aminobutyrate exists in the cotyledon with all but arginine being translocated rapidly to the axis tissue where these amino acids are rapidly metabolized.     

Characterization of aspartate aminotransferase in germinating pumpkin cotyledon

During germination an increase in aspartate aminotransferase(E.C. 2.6.1.1 [EC] .) occurs in the cotyledons of pumpkin seedlingsgrown either in the light or dark with the bulk of the enzymeactivity in the soluble fraction. The soluble and particulateenzymes had a pH optimum of 8.0, an -ketoglutarate optimum of0.02 mM and a broad aspartate optimum from 0.05 to 0.2 mM. Thevelocity of the reaction was proportional to the enzyme concentrationover a wide range. The soluble enzyme was shown to require manganeseand pyridoxal-5-phosphate and an active sulfhydryl group formaximum activitiy. The soluble enzyme was easily destroyed bypepsin while the particulate enzyme lost activity upon washing.It is suggested that aspartate aminotransferase plays a rolein nitrogen metabolism during protein hydrolysis in germinatingpumpkin cotyledons. (Received January 6, 1970; )     

Proline-Dehydrogenase from Pumpkin (Cucurbita moschata) Cotyledons

A NAD specific proline-dehydrogenase was found in pumpkin (Cucurbita moschata Poir. cv. Dickinson Field) which oxidized proline to Δ¹-pyrroline-5-carboxylate. NADP did not substitute for NAD and L-proline-methyl-ester and thiazolidine-4-carboxylate were substrates in the reaction, at a rate of 107% and 33% respectively, of the rate with L-proline. Pumpkin cotyledons contained the bulk of the enzyme activity with 90% of the activity being in the soluble fraction. Proline-dehydrogenase, which was not treated at high temperature, was stable at –10°C for 4 months in the presence of high ammonium sulfate concentration. The Michaelis constant for NAD was 2.2 mM and for L-proline was 2.5 mM. At 5 mM NADP, a 40% non-competitive inhibition of proline-dehydrogenase was obtained, while 50 μM NADP was sufficient to induce 20% inhibition.     

Arginine metabolism by pumpkin seedlings. Separation of plant extracts by ion exchange resins

Arginine-U-¹⁴C was injected into the cotyledons of 7-day oldpumpkin seedlings. At most, 24% of the administered ¹⁴C wastransported to the axis tissue. The amounts of arginine incorporatedinto cotyledonary protein suggests that turnover was occurringat a rapid rate. Arginine was extensively metabolized, and after96 hr 50% of the administered ¹⁴C had been released as ¹⁴CO₂.The remaining label was primarily in unmetabolized arginine,protein or transported to the axis tissue with little labelin other amino acids. The results suggest that the carbon fromarginine is incorporated into protein or catabolized to CO₂while the carbon for new amino acid skeletons is derived fromsugar. A simple, reproducible method for the quantitative fractionationof plant extracts or hydrolysates of insoluble plant materialinto basic amino acids, acidic amino acids, neutral amino acids,organic acids and sugars was reported. (Received September 10, 1968; )