Akiva Apelbaum

Involvement of Polyamines in the Development and Ripening of Avocado Fruits

Summary Polyamine metabolism was studied in avocado fruits, which are known to be typical climacteric fruits. An extract from the fruit mesocarp and the seed coat showed arginine decarboxylase (ADC) activity throughout the period of fruit growth, maturation and ripening, whereas no ornithine decarboxylase could be detected. The specific activity of ADC was 27-fold higher in the extract from the seed coat than in that from the mesocarp. During fruit maturation the seed coat shrivels and ADC activity in the whole fruit was reduced by 25-fold. The levels of putrescine and spermidine in the fruit declined during the course of fruit growth. This decrease was moderate during the period of fruit development until maturation. However, a sharp decline in these polyamines was recorded in fruits undergoing ripening after harvest. Spermine content did not change during development but decreased rapidly during ripening. Addition of putrescine or spermine to avocado fruit tissue inhibited the biosynthesis of the ripening hormone, ethylene, by inhibiting both the activity of 1-aminocyclopropane-1-carboxylic acid (ACC) synthase and the conversion of ACC into ethylene. In view of the above, a possible involvement of polyamines in fruit ripening is discussed.

Influence of ethylene on S-adenosylmethionine decarboxylase activity in etiolated pea seedlings

Summary S-adenosyl-L-methionine decarboxylase was detected in crude extract from the plumular hook of etiolated pea seedlings. The enzyme showed a Km of 0.2mM for S-adenosyl-Lmethionine and a Vmax of 4.76 nmol/mg protein/h. It was not activated by putrescine or by Mg++ and was inhibited by methyl glyoxal-bis-guanyl hydrazone with an I50 of 1µM. Treatment of pea seedlings with ethylene resulted in a 75 % reduction of the enzyme activity with an I50 of 0.3 µl/1 and a maximal effect at 6µl/l. The reduction was observed within 1 hour after exposure to ethylene, reaching a maximum at 18 hours. Upon transferring the ethylenetreated plants to an ethylene-free atmosphere the enzyme gradually recovered from the inhibition, showing a complete recovery within 6 hours. Exposing pea seedlings to treatments that caused a reduction in the endogenous level of ethylene resulted in a 30–48 % increase in the enzyme activity. In view of the results of this study, the possibility that ethylene at the endogenous level might regulate S-adenosyl-L-methionine decarboxylase activity is discussed.

Ethylene formation from 1-aminocyclopropane-1-carboxylic acid in plant mitochondria

Mitochondria isolated from apical parts of etiolated pea seedlings convert ACC to ethylene. This conversion is heat‐sensitive and largely enhanced when the intact mitochondria are sonicated or when the ionophore nigericin is added to the reaction medium. When the sonicated submitochondiral particles are spun down, almost all of the activity is recovered in the pellet. The results suggest that the ethylene‐formation activity is associated with the inner mitochondrial membrane and that transport of ACC into the mitochondrial membrane is a rate limiting step for the reaction. The mechanism of ACC transport across the mitochondrial membrane is discussed.

Interactions between auxin and ethylene in root-knot nematode (Meloidogyne javanica) infected tomato roots†

An increase in the rate of ethylene production occurred in excised tomato roots, whether or not infected with Meloidogyne javanica , when incubated in media containing indole acetic acid (IAA) above 0·6 μ M . However, the overall increase was considerably less pronounced in infected roots than in uninfected ones. Application of IAA to infected cultures resulted in an increase in gall fresh weight which was concentration-dependent and corresponded with the concentration of IAA, which stimulated ethylene production. The hypertrophied cortical parenchyma tissue in galls treated with 6·0 μ M IAA was wider than that of the untreated galls. The levels of free and bound endogenous IAA in infected roots were higher than in uninfected tissues, attaining their highest levels 10 days after inoculation coincident with the time of the highest rate of ethylene production in the infected tissue. The interaction between ethylene and auxin regarding the host—parasite interrelationships of the root-knot nematode is discussed.

Cadaverine formation by specific lysine decarboxylation in Pisum sativum seedlings.

Cadaverine was found to be formed in Pisum sativum seedlings via a specific lysine decarboxylation pathway as revealed by specific inhibitor studies. Lysine decarboxylation activity was recorded in the meristems and non-meristematic tissue of the shoots and the roots. In the shoot elongation zone, the specific activity was double that in the other tissues and cadaverine level was 90-fold higher. The results presented in this study suggest possible regulatory control by polyamines of lysine decarboxylase activity in Pisum sativum seedlings.

Antifungal antibiotics and Siba inhibit 1-aminocyclopropane-1-carboxylic acid synthase activity

The antifungal antibiotics Sinefungin and A9145C isolated from Streptomyces griseolus and the synthetic nucleoside Siba, which are analogs of S-adenosylmethionine, inhibit the activity of 1-aminocyclopropane 1-carboxylic acid synthase from tomato fruits. Sinefungin and Siba were shown to be more potent inhibitors than A9145C. In extracts of green fruits, the enzyme activity was inhibited by Sinefungin with an I50 of 1 microM, which was similar to that caused by aminoethoxyvinylglycine, and by Siba with an I50 of 100 microM; in extracts from red tomatoes, the I50s were 25 microM and 100 microM, respectively. The inhibition of ACC synthase by these analogs could be reversed by gel filtration chromatography.

Conversion of 1-aminocyclopropane-1-carboxylic acid to ethylene in submitochondrial particles isolated from plants: Characterization of the system

Conversion of 1‐aminocyclopropane‐1‐carboxylic acid (ACC) to ethylene was studied in submitochondrial particles (SMP) which were prepared by sonication of the mitochondrial fraction isolated from 7‐day‐old etiolated pea (Pisum sativum var. Kelvedon Wonder) seedlings. The reaction was enhanced by the addition of Mn2+ and had a maximal rate at pH 8.0. Conversion of ACC to ethylene was inhibited under anaerobic conditions and by the addition of KCN, EDTA, NaN3, n‐propyl gallate and CoCl2. Addition of the uncouplers 2,4‐DNP, CCCP and FCCP, however, did not inhibit the reaction. Structural analogs of ACC inhibited ACC conversion to enthylene by SMP. The structural analog of methionine, α‐keto‐γ‐methylthiobutyric acid (KMB), was converted to ethylene by SMP at a rate which was only about 2% that of ACC conversion to ethylene.

The Role of Ethylene in the Pathogenic Symptoms Displayed by Meloidogyne Javanica Nematode Infected Tomato Plants

Tomato plants infected with the root-knot nematode (RKN) M. javanica produce ethylene at a rate several fold higher than the uninfected plants (1). This increase coincides with an increase in 1-amino cyclopropane-l-carboxylic acid (ACC) level in the plant’s root and leaves (2). When excised roots were treated with aminoetoxyvinylglicine (AVG) or aminooxyacetic acid the nematode-induced ethylene production was inhibited, indicating that the nematode-infection induced ethylene production by accelerating the rate of ACC formation from S-adenosyl-methionine. Altering the rate of ethylene production in the infected roots did not affect nematode development up to the 3rd stage of juveniles, nor did it affect nematode penetration and initiation of gall formation. However, development of adult nematodes was markedly inhibited by both ethylene stimulators or inhibitors, whereas the rate of gall growth was accelerated by stimulators of ethylene production and suppressed when the production or action of the hormone was inhibited. Examination of fractured galls with SEM revealed that the hypertrophied cortical parenchyma of the galls treated with AVG, was less developed than the untreated control. On the other hand, the parenchyma of galls treated with supraoptimal concentration of auxin was larger in diameter than that of the untreated galls, due to the induction of ethylene production by the auxine. In all treatments the vascular cylinder was not affected in size and structure, nor were the coencytes induced by the RKN M. javanica.