Nonsteroidal anti-inflammatory drugs as antipyretics and modulators of a molecular clock(s) in the appendix of Sauromatum venosum inflorescence.

The appendix of the Sauromatum senosum inflorescence is a striking example of thermogenesis in plants. On the day of opening, the Sauromatum appendix becomes hot, reaching up to 32 °C. Aspirin, salicylic acid and 2,6-dihydroxybenzoic acid, a subclass of NSAIDs, induce a temperature rise from three mitochondrial sources: alternative oxidase, F1 FO -ATP synthase and adenine nucleotide translocator. This temperature rise is synchronized and compounded under various light/dark regimes. We studied the effect of different subgroups of NSAIDs on the temperature rise. Tissue slices of appendix of Sauromatum and Arum italicum inflorescences at a pre-mature stage were treated with the three inducers in combination with one NSAID under constant light or darkness and under different photoperiods. Temperature rise generated by the three heat sources in the presence of inducers and different non-selective NSAIDs were not compounded and occurred at three different times. Under constant light, DuP-697, ibuprofen, flurbiprofen, acetaminophen and diclofenac suppressed the temperature rise induced by the three salicylates. Desynchronization and delayed temperature rise were detected with 6/42-h light/ dark and 15/33-h light/dark regimes in the presence of celecoxib and ibuprofen. With a 24/24-h light/dark regime, temperature rise was suppressed in the presence of ibuprofen. There were differences in response to individual NSAIDs between appendix tissue of A. italicum and S. venosum. Mitochondrial energy balance is affected by NSAIDs. There is an interaction between light/dark regime and temperature rise and a relationship between timing mechanism and temperature rise.

Oleic Acid and Its Positional Isomer, cis-Vaccinic Acid, in the Appendix of Sauromatum guttatum during Anthesis.

The fatty acid profiles of various organs of the thermogenic inflorescence of Sauromatum guttatum and of the sporophylls of thermogenic male cones of two cycad species (Encephalartos ferox and Dioon edule var edule and var angustifolium) were determined by gas chromatography. During anthesis, palmitate (16:0), oleate [18:1 (9)], cis-vaccinate [18:1 (11)], and linoleate [18:2 (9, 12)] were the most abundant fatty acids in the Sauromatum appendix. cis-Vaccinic acid, a positional isomer of oleic acid, was identified by comparing its retention time on a gas chromatography column and its mass spectrum to an authentic compound. The percentage of oleic acid from total fatty acids dropped from about 9 in the morning 3 d before heat production to 6 in the morning 2 d before heat production. At this time, the percentage of cis-vaccinic acid increased from 3 to 11%, and then remained at this level until the inflorescence dried and died. Palmitoleic acid [16:1 (9)], the common precursor of cis-vaccinic acid, is a minor component of total fatty acids. In six other organs of the Sauromatum inflorescence including thermogenic organs, such as male flowers and lower spadix, palmitate, oleate, and linoleate were prevalent but cis-vaccinate was not. The thermogenic male cones of the two cycad species were rich in palmitic, oleic, and linolenic acids. The level of cis-vaccinic acid in these organs was less than 0.5%.

A development-dependent hemagglutinin from cucumber surfaces: I. Hemagglutinin activity of cotyledon surfaces.

During studies on the chemistry of plant surfaces, we observed that concomitant with the development of cucumber (Cucumis sativus L. cv ;ELEM') cotyledons an agglutinin that agglutinates human erythrocytes appeared on epicuticular surfaces. The agglutinin was released from cotyledon surfaces into distilled water by a 1-minute immersion (or even less). Homogenization of the washed cotyledons released residual agglutinating activity. The surface-located hemagglutinating activity was age-dependent and occurred in dark- and light-grown cotyledons. Agglutinating activities were present in light-grown cotyledons of 2- to 14-day-old seedlings and in dark-grown cotyledons of 3- to 17-day-old seedlings. Agglutinating activities were maximal in cotyledons of 3- to 4-day-old seedlings. No activity could be detected in dry seeds or in seedlings up to 2 days and after 17 days of germination. The hemagglutinating activity was specifically inhibited by N,N',N''-triacetylchitotriose, chitin, and chitosan.

Developmental changes in the ultrastructure of the Sauromatum guttatum (Araceae) mitochondria.

Electron microscopic studies show that the electron density of the mitochondria of the thermogenic appendix of Sauromatum guttatum inflorescences changes during development. Two to 3 days before heat-production, the mitochondria accumulate osmiophilic, electron-dense material between the inner and outer membranes. During heat-production and the release of volatiles compounds the osmiophilic material disappears from the inter membrane space. Deposits with the same electron density are also found in the endoplasmic reticulum. It is concluded that the mitochondria may also have the capacity to accumulate that material in the inter membrane space.

Further studies of the glandular tissue of the Sauromatum guttatum (Araceae) appendix.

Electron microscopic studies showed that the trans-Golgi network (trans indicates the polarity of cisternae within the Golgi apparatus; it is opposite to the cis-face that is adjacent to the rough endoplasmic reticulum) was involved in the processing of the osmiophilic material present in the appendix of the inflorescence of Sauromatum guttatum. This material accumulated in the rough endoplasmic reticulum and in special pockets of the plasma membrane prior to heat production. Associations between the endoplasmic reticulum and trans-Golgi network were observed. The Golgi apparatus was composed of 5-6 dictyosomes on one side and one or two somewhat detached cisternae on the other side. Various nonosmiophilic Golgi-derived vesicles were observed: small ones covered with spike-like material, large ones with a smooth surface, and irregularly shaped ones. These electron-translucent vesicles seemed to accumulate in specific localities at the plasma membrane surface in the vicinity of the osmiophilic material; they were not found when the aroma was released. During heat production, the Golgi structures shrank and the activity of the trans-Golgi network seemed to be reduced. At the same time, coated pits were seen at the plasma membrane surface. In some cells, hypertrophic Golgi apparatuses were seen with only 2-3 dictyosomes that contained granulated material in their lumens. Finally, the osmiophilic material was also found in the plasmodesmata.

Cytochemical evidence for the presence of actin in the nucleus of the voodoo lily appendix.

Immunoflorescence microscopy of sections of the voodoo lily Sauromatum guttatum appendix stained with monoclonal antibodies against alpha-smooth muscle actin and cytoplasmic actin revealed different staining intensity of different parts of the cell. The anti-cytoplasmic-actin recognized antigens present mainly in the cytoplasm, and the anti-alpha-smooth muscle-actin recognized more intensively antigens present in the nuclei. A positive staining of the nucleus was also obtained with FITC-phalloidin confirming the presence of actin in its filamenous form in the nucleus. The presence of a nuclear alpha-smooth muscle-actin-like protein was further confirmed by confocal laser microscopy. On Western blots, the two anti-actins labelled a protein band that comigrated with standard actin at the approximate molecular weight of 43 kDa. Several other proteins interacted with the two antibodies to a different degree. The monoclonal antibodies against beta-tubulin subunit stained only the periphery of the cytoplasm and anti-pan cytoplasmic myosin stained the cytoplasm weakly. On a Western blot, anti-beta-tubulin subunit primarily recognized a protein band at the appropriate molecular weight of 50 kDa. This is the first cytochemical evidence for the presence of alpha-smooth muscle-actin-like protein in the plant nucleus.

Oxidation of Proline and Glutamate by Mitochondria of the Inflorescence of Voodoo Lily (Sauromatum guttatum).

In appendices of Sauromatum guttatum that are developing thermogenicity, mitochondria isolated from successive developmental stages of the inflorescence show an increase in the oxidation rates of proline and glutamate. A similar rise in the oxidation rates of these compounds is observed in mitochondria obtained from the spathe, a nonthermogenic organ of the inflorescence. Changes in oxidative metabolism were also observed in mitochondria isolated from sections of immature appendix treated with salicylic acid (SA) at 0.69 microgram per gram fresh weight indicating that they are induced by SA. At that concentration, however, SA has no effect on oxygen consumption by mitochondria in the presence of glutamate, proline, or malate. Furthermore, oxygen uptake by mitochondria in the presence of proline or glutamate is partially sensitive to salicylhydroxamic acid (SHAM) at concentrations greater than 2 millimolar when in the presence of 1 millimolar KCN. For NADH, succinate, and malate a high capacity of the alternative (cyanide-resistant) pathway is found that is completely sensitive to SHAM at 1.5 to 4 millimolar. The increase in the mitochondrial capacity to oxidize either amino acid is also found in four other Araceae species including both thermogenic and nonthermogenic ones. After anthesis, the rates of proline and glutamate oxidation decline.

Heat Production in the Voodoo Lily (Sauromatum guttatum) as Monitored by Infrared Thermography.

The pattern of surface temperatures of the inflorescence of Sauromatum guttatum was investigated by using an infrared camera. The male flowers are weakly thermogenic on the first day of inflorescence opening (D-day) as well as on the next day (D + 1), reaching 0.5 to 1 degrees C above ambient temperature. The appendix (the upper sterile part of the inflorescence) is highly thermogenic on D-day, reaching 32 degrees C, and is faintly thermogenic on D + 1, reaching 1 degrees C above ambient temperature. The lower part of the spadix, close to the female flowers, is also thermogenic on D-day and D + 1, reaching a temperature similar to that of the appendix only on D + 1. Salicylic acid does not induce heat production in the lower part of the spadix, as it does in the appendix. Respiration of tissue slices obtained from the appendix shows that the capacity for cyanide-insensitive respiration is present in young and mature appendices. This alternative respiratory pathway is not, however, utilized in young appendix tissue, but is engaged during the maturation of that tissue.

Infrared thermography ofArum lily inflorescences.

The infrared radiation emitted from the surface of inflorescences of 12 aroid species was monitored with an infrared camera, capable of 0.1°C resolution, and the data were converted to temperature values by means of temperature reference standards. Images representing surface temperatures were obtained forAmorphophallus bulbifer Blume,A. campanulatus Blume,A. forbesii Engl. et Gehrm.,A. rivieri Dur.,Philodendron selloum Koch,Monstera deliciosa Liebm.,Dracunculus vulgaris Schott,Arum italicum Mill.,A. dioscoridis Sibth.,A. creticum Boiss et Heldr.,Caladium sp., andRemusatia vivipara Schott. These images were different among species with respect to temperature, duration of detectable heat development, and organ type (male and female flowers, spathe and appendix) found to be thermogenic. All these species, however, exhibited three common characteristics: 1) production of heat by the male flowers; 2) pollen-shedding immediately after heat production had ceased; and 3) when male flowers were some distance away from female flowers along the spadix, heat was not detected in female flowers. Heat emission was associated with the alternative, cyanide-insensitive pathway that was fully operative.

Pathway of terpene excretion by the appendix of Sauromatum guttatum.

Electron microscopy of the cells of the thermogenic appendix of Sauromatum guttatum has revealed a fusion event between pocket-like structures of the rough endoplasmic reticulum (rER) and the plasma membrane. As a result of the fusion event, many regions of the plasma membrane have paired unit membranes (four leaflets instead of two). The fusion allows the transfer of osmiophilic material from the rER pockets to the plasma membrane, where the osmiophilic material is confined to bilayer, pocket-like structures. A clear correlation is found between the presence of the osmiophilic compound and sesquiterpenes. Prior to heat production, the rER- and plasma-membrane pockets are electron dense, and sesquiterpenes are detectable only in tissue extracts. On the day of heat production, electron-translucent pockets are subsequently found and the stored sesquiterpenes are released to the atmosphere. Three sesquiterpenes have been identified by gas chromatography-mass spectrometry as alpha-copaene and beta- and alpha-caryophyllene.