Pattern of respiration by intact inflorescences of the thermogenic arum lily Philodendron selloum

Inflorescences of the neotropical arum lily, Philodendron selloum, are strongly thermogenic for 2 d during anthesis. Continuous measurements of spadix temperature (Ts) and rate of oxygen consumption (VO2) were made outdoors in whole inflorescences attached to the plants. Some inflorescences were exposed to uncontrolled ambient temperature (Ta) while others were enclosed in clear water-jackets that produced nearly constant ambient conditions. A repeatable, diphasic pattern of heat production appeared, most clearly in water-jacketed inflorescences, and it comprised a short peak phase at sunset followed by a plateau phase that lasted until the following sunset. Regulation of Ta occurred in both phases, but at different levels. Peak phase Ta was regulated in the region of 38-42C, but plateau phase Ta was usually in the range of 25-36C. Both VO2 and total heat produced throughout anthesis increased at lower Ta. The data imply that the short peak phase is related to the enhancement of odour production that attracts a single species of large scarabaeid beetle in its native Brazil, and regulation of maximum Ta may prevent overheating. Thermoregulation in the long plateau phase produces equable temperatures inside the inflorescence that may facilitate the resident beetles' activities as a direct energetic reward.     

Respiration and heat production by the inflorescence of Philodendron selloum Koch

During a 2-d sequence of anthesis, the spadices of the thermogenic arum lily, Philodendron selloum, regulated maximum temperature within a small range (37–44°C) by reversible thermal inhibition of respiratory heat production. This response protects the inflorescence and the attracted insects from thermal damage. Heat production by whole spadices, measured by O₂ respirometry, equalled heat loss, measured by gradient layer calorimetry, which confirmed the heat equivalence of O₂ consumption (20.4 J ml^-1). This also indicated that there was no net phosphorylation during thermogenesis, heat production being the primary function of high rates of respiration. The sterile male florets consumed about 30 ml g^-1 h^-1 and the average 124-g spadix produced about 7 W to maintain a 30°C difference between spadix and ambient temperature. Most of the energy for thermogenesis is present in the florets before anthesis. Despite high respiratory rates, thermogenesis is an energetically inexpensive component of the reproductive process.     

Ubiquitous expression of a gene encoding for uncoupling protein isolated from the thermogenic inflorescence of the dead horse arum Helicodiceros muscivorus

Uncoupling proteins (UCPs) are a family of mitochondrial inner membrane proteins that have been implicated in heat production in mammalian cells. The inflorescences of several members of the arum lily family (Araceae) have also been shown to produce heat during flowering, but the involvement of UCP-mediated heat production in plants is not known. In this work a gene has been isolated termed HmUCPa that encodes for a putative uncoupling protein from Helicodiceros muscivorus, a highly thermogenic arum lily. RT-PCR analysis revealed that the expression of HmUCPa was ubiquitously found, both in thermogenic male florets and appendix, and the non-thermogenic female florets, spathe and club-shaped organs of the spadix. These results suggest that HmUCPa is not primarily involved in organ-specific heat production in H. muscivorus.     

In the heat of the night--alternative pathway respiration drives thermogenesis in Philodendron bipinnatifidum

? Philodendron bipinnatifidum inflorescences heat up to 42 °C and thermoregulate. We investigated whether they generate heat via the cytochrome oxidase pathway uncoupled by uncoupling proteins (pUCPs), or the alternative oxidase (AOX). ? Contribution of AOX and pUCPs to heating in fertile (FM) and sterile (SM) male florets was determined using a combination of oxygen isotope discrimination, protein and substrate analyses. ? Both FM and SM florets thermoregulated independently for up to 30 h ex planta. In both floret types, AOX contributed > 90% of respiratory flux during peak heating. The AOX protein increased fivefold with the onset of thermogenesis in both floret types, whereas pUCP remained low throughout development. These data indicate that AOX is primarily responsible for heating, despite FM and SM florets potentially using different substrates, carbohydrates or lipids, respectively. Measurements of discrimination between O? isotopes in strongly respiring SM florets were affected by diffusion; however, this diffusional limitation was largely overcome using elevated O?. ? The first in vivo respiratory flux measurements in an arum show AOX contributes the bulk of heating in P. bipinnatifidum. Fine-scale regulation of AOX activity is post-translational. We also demonstrate that elevated O? can aid measurement of respiratory pathway fluxes in dense tissues.     

Respiration,temperature regulation and energetics of thermogenic inflorescences of the dragon lily Dracunculus vulgaris (Araceae)

Dracunculus vulgaris is a protogynous arum lily with thermogenic inflorescences consisting of male and female florets on a spadix within a floral chamber. Above the chamber, an odour-producing appendix and a carrion-coloured spathe attract flying insects. The inflorescence shows a triphasic warming pattern. The floral chamber warms weakly on the first night as the spathe opens. Then the appendix produces a large amount of heat and a powerful scent during the first day. As the appendix cools on the second night, scent production ceases and the floral chamber rewarms. Warming ceases when the pollen is shed on the second day. The heating pattern is associated with attraction of pollinating insects by the appendix on the first day, entrapment in the warm chamber at night and release after pollen shedding. The temperature in the floral chamber is regulated at around 18 °C during the second night. The oxygen consumption rate of the florets is inversely related to the ambient temperature as in other thermoregulatory flowers. Conversely, the oxygen consumption rate of the appendix is directly related to the ambient temperature, indicating that it does not thermoregulate. Thus, temperature regulation is not associated with scent production, but with some activity inside the floral chamber.     

Supramolecular structure of the OXPHOS system in highly thermogenic tissue of Arum maculatum

The protein complexes of the mitochondrial respiratory chain associate in defined ways forming supramolecular structures called respiratory supercomplexes or respirasomes. In plants, additional oxidoreductases participate in respiratory electron transport, e.g. the so-called “alternative NAD(P)H dehydrogenases” or an extra terminal oxidase called “alternative oxidase” (AOX). These additional enzymes were previously reported not to form part of respiratory supercomplexes. However, formation of respiratory supercomplexes might indirectly affect “alternative respiration” because electrons can be channeled within the supercomplexes which reduces access of the alternative enzymes towards their electron donating substrates. Here we report an investigation on the supramolecular organization of the respiratory chain in thermogenic Arum maculatum appendix mitochondria, which are known to have a highly active AOX for heat production. Investigations based on mild membrane solubilization by digitonin and protein separation by blue native PAGE revealed a very special organization of the respiratory chain in A. maculatum, which strikingly differs to the one described for the model plant Arabidopsis thaliana: (i) complex I is not present in monomeric form but exclusively forms part of a I + III₂ supercomplex, (ii) the III₂ + IV and I + III₂ + IV supercomplexes are detectable but of low abundance, (iii) complex II has fewer subunits than in A. thaliana, and (iv) complex IV is mainly present as a monomer in a larger form termed “complex IVa”. Since thermogenic tissue of A. maculatum at the same time has high AOX and I + III₂ supercomplex abundance and activity, negative regulation of the alternative oxidase by supercomplex formation seems not to occur. Functional implications are discussed.     

Cold-induced expression of the VEGF gene in brown adipose tissue is independent of thermogenic oxygen consumption

To examine whether cold-induced vascular endothelial growth factor (VEGF) gene expression in brown adipose tissue involved generation of hypoxic oxygen levels by thermogenic processes, we cold-exposed wild-type mice, as well as uncoupling protein-1 (UCP1)-ablated mice in which no thermogenesis in brown adipocytes can be induced. Cold exposure stimulated VEGF expression in both wild-type and UCP1-ablated mice. Unexpectedly, the effect was 3-fold higher in UCP1-ablated animals, whereas cultured brown adipocytes from both genotypes responded identically to norepinephrine stimulation. These results demonstrate that generation of low oxygen levels does not contribute to cold-induced VEGF expression in brown adipose tissue, but the results are consistent with an adrenergic regulation of expression.     

Diffusion of nitric oxide into low density lipoprotein.

A key early event in the development of atherosclerosis is the oxidation of low density lipoprotein (LDL) via different mechanisms including free radical reactions with both protein and lipid components. Nitric oxide (( small middle dot)NO) is capable of inhibiting LDL oxidation by scavenging radical species involved in oxidative chain propagation reactions. Herein, the diffusion of ( small middle dot)NO into LDL is studied by fluorescence quenching of pyrene derivatives. Selected probes 1-(pyrenyl)methyltrimethylammonium (PMTMA) and 1-(pyrenyl)-methyl-3-(9-octadecenoyloxy)-22,23-bisnor-5-cholenate (PMChO) were chosen so that they could be incorporated at different depths of the LDL particle. Indeed, PMTMA and PMChO were located in the surface and core of LDL, respectively, as indicated by changes in fluorescence spectra, fluorescence quenching studies with water-soluble quenchers and the lifetime values (tau(o)) of the excited probes. The apparent second order rate quenching constants of ( small middle dot)NO (k(NO)) for both probes were 2.6-3.8 x 10(10) m(-1) s(-1) and 1.2 x 10(10) m(-1) s(-1) in solution and native LDL, respectively, indicating that there is no significant barrier to the diffusion of ( small middle dot)NO to the surface and core of LDL. Nitric oxide was also capable of diffusing through oxidized LDL. Considering the preferential partitioning of ( small middle dot)NO in apolar milieu (6-8 for n-octanol:water) and therefore a larger ( small middle dot)NO concentration in LDL with respect to the aqueous phase, a corrected k(NO) value of approximately 0.2 x 10(10) m(-1) s(-1) can be determined, which still is sufficiently large and consistent with a facile diffusion of ( small middle dot)NO through LDL. Applying the Einstein-Smoluchowsky treatment, the apparent diffusion coefficient (D(')NO) of ( small middle dot)NO in native LDL is on average 2 x 10(-5) cm(2) s(-1), six times larger than that previously reported for erythrocyte plasma membrane. Thus, our observations support that ( small middle dot)NO readily traverses the LDL surface accessing the hydrophobic lipid core of the particle and affirm a role for ( small middle dot)NO as a major lipophilic antioxidant in LDL.     

The pathway for the conversion of dihydroagnosterol into cholesterol in rat liver.

Dihydroagnosterol is demethylated by a rat liver homogenate to give 4,4'-dimethylcholesta 7,9-dienol and then cholesta-7,9-dienol. The cholesta-7,9-dienol is isomerized to cholesta-8,14-dienol, which is converted into cholesterol by the normal pathway.     

Release of an acid phosphatase activity during lily pollen tube growth involves components of the secretory pathway

Summary. An acid phosphatase (acPAse) activity was released during germination and tube growth of pollen of Lilium longiflorum Thunb. By inhibiting components of the secretory pathway, the export of the acPase activity was affected and tube growth stopped. Brefeldin A (1 μM) and cytochalasin D (1 μM), which block the production and transport of secretory vesicles, respectively, inhibited the acPase secretion. The Ca²⁺ channel blocker gadolinium (100 μM Gd³⁺) also inhibited acPase secretion and tube growth, whereas 3 mM caffeine, another Ca²⁺ uptake inhibitor, stimulated the acPase release, while tube growth was inhibited. The Yariv reagent (β-D-glucosyl)₃ Yariv phenylglycoside stopped tube growth by binding to arabinogalactan proteins of the tube tip cell wall but did not affect acPase secretion. A strong correlation between tube growth and acPase release was detected. The secreted acPase activity had a pH optimum at pH 5.5, a K _M of 0.4 mM for p-nitrophenyl phosphate, and was inhibited by zinc, molybdate, phosphate, and fluoride ions, but not by tartrate. In electrophoresis gels the main acPase activity was detected at 32 kDa. The conspicuous correlation between activity of the secretory pathway and acPase secretion during tube elongation strongly indicates an important role of the acPase during pollen tube growth and the secreted acPase activity may serve as a useful marker enzyme assay for secretory activity in pollen tubes Received July 25, 2001 Accepted January 15, 2002     

Diffusion and consumption of oxygen in the resting frog sartorius muscle

Adaptations of the method of Takahashi et al. (1966. J. Gen. Physiol. 50:317-333) were used to test the validity of the one-dimensional diffusion equation for O2 in the resting excised frog sartorius muscle. This equation is: (formula: see text) where x is the distance perpendicular to the muscle surface. t is time, P(x, t) is the partial pressure of O2,D and alpha are the diffusion coefficient and solubility for O2 in the tissue, and Q is the rate of O2 consumption. P(O, t), the time-course of PO2 at one muscle surface, was measured by a micro-oxygen electrode. Transients in the PO2 profile of the muscle were induced by two methods: (a) after an equilibration period, one surface was sealed off by a disc in which the O2 electrode was embedded; (b) when PO2 at this surface reached a steady state, a step change was made in the PO2 at the other surface. With either method, the agreement between the measured P(O, t) and that predicted by the diffusion equation was excellent, making possible the calculation of D and Q. These two methods yielded statistically indistinguishable results, with the following pooled means (+/- SEM): (formula: see text) At each temperature, D was independent of muscle thickness (range, 0.67-1.34 mm). The activation energy (EA) for diffusion of oxygen in muscle was -3.85 kcal/mol, which closely matches the corresponding value in water. Together with absolute values of D in water taken from the literature, the present data imply that (Dmuscle/DH2O) is in the range 0.59-0.69. This value, and that of EA, are in agreement with the theory of Wang (1954, J. Am. Chem. Soc. 76:4755-4763), suggesting that with respects to the diffusion of O2, to a useful approximation, frog skeletal muscle may be considered simply as a homogeneous protein solution.     

The introgression of chromosome 6P specifying for increased numbers of florets and kernels from Agropyron cristatum into wheat

A wheat (Triticum aestivum L.) line 4844 with superior numbers of florets and grains per spike was derived from the cross between Fukohokomugi wheat and Agropyron cristatum (L.) Gaertn. In order to determine the genetic control of floret and kernel number per spike in this line, chromosome addition and substitution lines that were derived from line 4844 were characterized by means of in situ hybridization, microsatellite (SSR), and gliadin analyses. Genomic in situ hybridization analysis with biotinylated P genomic DNA of A. cristatum as a probe demonstrated that the increased number of florets and grains in a spike was associated with the introgression of an A. cristatum chromosome. Fluorescence in situ hybridization, using a repetitive sequence, pAs1, derived from Aegilops squarrosa L., indicated the replacement of chromosome 6D of wheat in the wheat-A. cristatum chromosome substitution lines. This was confirmed by microsatellite analyses with wheat SSR markers specific for chromosome 6D, suggesting that the A. cristatum chromosome was homoeologous to group 6 and was therefore designated as 6P. This conclvsion was further confirmed by amplification using EST-SSR markers and gliadin analysis. The increased number of florets and kernels within a spike of the wheat-A. cristatum hybrids thus was controlled by gene(s) located on A. cristatum chromosome 6P.     

Symmorphosis in the proximal pathway for oxygen in the leaf-eared mouse Phyllotis darwini

In this report, we explore the matching of structures to functional needs by comparing previously reported data of maximal oxygen consumption and the development of the lung in the leaf-eared mouse Phyllotis darwini in warm and cold environments. We discuss whether the state of structural design is commensurate with functional needs from regulated morphogenesis as predicted by the hypothesis of symmorphosis. We found a close match between respiratory structures and functional needs during postnatal development, expressed as safety factors close to unity. However, in the adult stage the safety factors were greater than two, which suggests that adult animals acquired a structure greater than that required considering their maximum capacities. A high safety factor in the respiratory system of adult mice may be a consequence of the symmorphosis that operates during ontogeny and does not necessarily support a rejection of this hypothesis.     

Formation of highly reactive gamma-ketoaldehydes (neuroketals) as products of the neuroprostane pathway.

Neuroprostanes are prostaglandin-like compounds produced by free radical-induced peroxidation of docosahexaenoic acid, which is highly enriched in the brain. We previously described the formation of highly reactive gamma-ketoaldehydes (isoketals) as products of the isoprostane pathway of free radical-induced peroxidation of arachidonic acid. We therefore explored whether isoketal-like compounds (neuroketals) are also formed via the neuroprostane pathway. Utilizing mass spectrometric analyses, neuroketals were found to be formed in abundance in vitro during oxidation of docosahexaenoic acid and were formed in greater abundance than isoketals during co-oxidation of docosahexaenoic and arachidonic acid. Neuroketals were shown to rapidly adduct to lysine, forming lactam and Schiff base adducts. Neuroketal lysyl-lactam protein adducts were detected in nonoxidized rat brain synaptosomes at a level of 0.09 ng/mg of protein, which increased 19-fold following oxidation in vitro. Neuroketal lysyl-lactam protein adducts were also detected in vivo in normal human brain at a level of 9.9 +/- 3.7 ng/g of brain tissue. These studies identify a new class of highly reactive molecules that may participate in the formation of protein adducts and protein-protein cross-links in neurodegenerative diseases and contribute to the injurious effects of other oxidative pathologies in the brain.     

Hochachka's "Hypoxia Defense Strategies" and the development of the pathway for oxygen

Hochachka's "Hypoxia Defense Strategies" identify oxygen signalling, metabolic arrest, channel arrest and coordinated suppression of ATP turnover rates as key factors that determine the ability of organisms to survive exposure to chronic hypoxia. In this review, I assess the developmental role played by these phenomena in the morphogenesis of the gas exchange tissues that define the pathway for oxygen transport to cytochrome c oxidase. Key areas of regulation lie in: (I) the suppression of fetal mitochondrial oxidative function in hand with mitochondrial biogenesis (metabolic arrest), (II) the role of hypoxia-driven oxygen signalling pathways in directing the scope of non-differentiated stem cell proliferation in placenta and lung development and (III) the regulation of epithelial fluid secretion/absorption in the lung through the oxygen-dependent modulation of Na+ conductance pathways. The identification of developmental roles for Hochachka's "Hypoxia Defense Strategies" in directing the morphogenesis of gas exchange structures bears with it the implication that these strategies are fundamental to establishing the scope for aerobic metabolic performance throughout life.     

Craniometric Data Supports Demic Diffusion Model for the Spread of Agriculture into Europe

Background

The spread of agriculture into Europe and the ancestry of the first European farmers have been subjects of debate and controversy among geneticists, archaeologists, linguists and anthropologists. Debates have centred on the extent to which the transition was associated with the active migration of people as opposed to the diffusion of cultural practices. Recent studies have shown that patterns of human cranial shape variation can be employed as a reliable proxy for the neutral genetic relationships of human populations.

Methodology/Principal Findings

Here, we employ measurements of Mesolithic (hunter-gatherers) and Neolithic (farmers) crania from Southwest Asia and Europe to test several alternative population dispersal and hunter-farmer gene-flow models. We base our alternative hypothetical models on a null evolutionary model of isolation-by-geographic and temporal distance. Partial Mantel tests were used to assess the congruence between craniometric distance and each of the geographic model matrices, while controlling for temporal distance. Our results demonstrate that the craniometric data fit a model of continuous dispersal of people (and their genes) from Southwest Asia to Europe significantly better than a null model of cultural diffusion.

Conclusions/Significance

Therefore, this study does not support the assertion that farming in Europe solely involved the adoption of technologies and ideas from Southwest Asia by indigenous Mesolithic hunter-gatherers. Moreover, the results highlight the utility of craniometric data for assessing patterns of past population dispersal and gene flow.