Stem photosynthesis not pressurized ventilation is responsible for light-enhanced oxygen supply to submerged roots of alder (Alnus glutinosa)

BACKGROUND AND AIMS: Claims that submerged roots of alder and other wetland trees are aerated by pressurized gas flow generated in the stem by a light-induced thermo-osmosis have seemed inconsistent with root anatomy. Our aim was to seek a verification using physical root-stem models, stem segments with or without artificial roots, and rooted saplings. METHODS: Radial O2 loss (ROL) from roots was monitored polarographically as the gas space system of the models, and stems were pressurized artificially. ROL and internal pressurization were also measured when stems were irradiated and the xylem stream was either CO2 enriched or not. Stem photosynthesis and respiration were measured polarographically. Stem and root anatomy were examined by light and fluorescence microscopy. KEY RESULTS: Pressurizing the models and stems to 相似文献   

Refining the statistical parameters for constructing tree-ring chronologies using short-lived species: Alder (Alnus glutinosa Gaertn)

The dendrochronological potential of short-lived species has had varying degrees of success in the past. Where there has been a level of success with short sequenced assemblages, the focus has been on visual comparisons, based on the occurrence of signature rings. Of vital importance to alder’s ability to be cross-correlated is that it produces a significant amount of distinguishable signature rings. Between 2012 and 2013, a large artificial island (crannog) of medieval date was excavated at Drumclay, County Fermanagh, Northern Ireland, and revealed a site of significant longevity, dating from the 9th century AD to the Post-Medieval period. This excavation exposed a vast number of well-preserved waterlogged archaeological features, resulting in the retention of over 9,000 individual wood samples. Oak timbers were used scarcely in the construction of the crannog, with the dominant wood species identified during excavation being alder. While the oak timbers have proved successful in providing spot dates and indicating phases of activity, the full chronological potential of the wood assemblage lies in the ring patterns of the principal species, alder, particularly with respect to understanding construction phases and site evolution. Previous failures to build chronologies using alder have been attributed to the short-lived and site specific nature of the species. Here, we test whether the measurement of large numbers of samples from a single context within a single site overcomes the limitations posed by alder. We measured the ring-widths of 90 alder samples from archaeological features within the crannog’s infrastructure to test if a robust context chronology could be built. The average ring sequence length ranged from 30 to 60 rings, with one timber extending to 108 rings. Visual correlations were used to aid ring pattern matching in conjunction with statistical correlation. We used radiocarbon wiggle-matching to test the robustness of our constructed chronology and to anchor it to an absolute timescale. Our results to date show that problems of autocorrelation can arise when long alder sequences (>100 rings) are used in conjunction with short sequences (30 to 60 rings). Establishing a rigorous protocol for sample selection has enabled us to develop a more statistically refined methodology that has produced t-values as high as 8.3. We show that in order to construct the best possible alder chronology, multiple ring patterns need to be examined from each context. We recommend examining short-lived assemblages on their own merits; the best approach in these cases is not to look for the longest sequences but instead to focus on the those from the mean sequence range.     

Aerenchyma (Gas-space) Formation in Adventitious Roots of Rice (Oryza sativa L.) is not Controlled by Ethylene or Small Partial Pressures of Oxygen

Jackson, M. B., Fenning, T. M., and Jenkins, W. 1985 Aerenchyma(gas-space) formation in adventitious roots of rice (Oryza sativaL.) is not controlled by ethylene or small partial pressuresof oxygen.—J. exp. Bot. 36: 1566–1572. The extent of gas-filled voids (aerenchyma) within the cortexof adventitious roots of vegetative rice plants (Oryza sativaL. cv. RB3) was estimated microscopically from transverse sectionswith the aid of a computer-linked digitizer drawing board. Gas-spacewas detectable in 1-d-old tissue and increased in extent withage. After 7 d, approximately 70% of the cortex had degeneratedto form aerenchyma. The extent of the voids in 1-4-d-old tissuewas not increased by stagnant, poorly-aerated external environmentscharacterized by sub-ambient oxygen partial pressures and accumulationsof carbon dioxide and ethylene. Treatment with small oxygenpartial pressures, or with carbon dioxide or ethylene appliedin vigorously stirred nutrient solution also failed to promotethe formation of cortical gas-space. Furthermore, ethylene productionby rice roots was slowed by small oxygen partial pressures typicalof stagnant conditions. Silver nitrate, an inhibitor of ethylene action, did not retardgas-space formation; similarly when endogenous ethylene productionwas inhibited by the application of aminoethoxyvinylglycine(A VG), aerenchyma development continued unabated. Cobalt chloride,another presumed inhibitor of ethylene biosynthesis, did notimpair formation of the gas in rice roots nor did it decreasethe extent of aerenchyma even if A VG was supplied simultaneously.These results contrast with those obtained earlier using rootsof Zea mays L. We conclude that in rice, aerenchyma forms speedily even inwell-aerated environments as an integral part of ordinary rootdevelopment There seems to be little or no requirement for ethyleneas a stimulus in stagnant root-environments where aerenchymais likely to increase the probability of survival. Key words: Rice (Oryza sativa L.), ethylene, flooding, aeration, aerenchyma, environmental stress     

De novo discovery and multiplexed amplification of microsatellite markers for black alder (Alnus glutinosa) and related species using SSR-enriched shotgun pyrosequencing

Recent developments in sequencing technologies and bioinformatics analyses provide an unprecedented opportunity for cost and time effective high quality microsatellite marker discovery in nonmodel organisms for which no genomic information is available. Here, we use shotgun pyrosequencing of a microsatellite-enriched library to develop, for the first time, microsatellite markers for Alnus glutinosa, a keystone tree species of European riparian woodland communities. From a total of 17?855 short sequences, we identified 590 perfect microsatellites from which 392 had designed primers. A subset of 48 loci were tested for amplification, 12 of which were polymorphic in A. glutinosa. These 12 loci were successfully coamplified in a single multiplex polymerase chain reaction experiment and validated for population genetics applications. In addition, 10 and 8 of these microsatellites were found to be transferable to the related A. incana and A. cordata species. The developed multiplex of 12 microsatellite markers therefore provides new opportunities for experimental evolutionary and forest genetics research in Alnus.     

Carbon Dioxide Fixation in Roots and Nodules of Alnus glutinosa: I. Role of Phosphoenolpyruvate Carboxylase and Carbamyl Phosphate Synthetase in Dark CO(2) Fixation, Citrulline Synthesis, and N(2) Fixation

Detached roots and nodules of the N₂-fixing species, Albus glutinosa (European black alder), actively assimilate CO₂. The maximum rates of dark CO₂ fixation observed for detached nodules and roots were 15 and 3 micromoles CO₂ fixed per gram dry weight per hour, respectively. The net incorporation of CO₂ in these tissues was catalyzed by phosphoenolpyruvate carboxylase which produces organic acids, some of which are used in the synthesis of the amino acids, aspartate, glutamate, and citrulline and by carbamyl phosphate synthetase. The latter accounts for approximately 30 to 40% of the CO₂ fixed and provides carbamyl phosphate for the synthesis of citrulline. Results of labeling studies suggest that there are multiple pools of malate present in nodules. The major pool is apparently metabolically inactive and of unknown function while the smaller pool is rapidly utilized in the synthesis of amino acids. Dark CO₂ fixation and N₂ fixation in nodules decreased after treatment of nodulated plants with nitrate while the percentage of the total ¹⁴C incorporated into organic acids increased. Phosphoenolpyruvate carboxylase and carbamyl phosphate synthetase play key roles in the synthesis of amino acids including citrulline and in the metabolism of N₂-fixing nodules and roots of alder.     

Effect of Glyoxylate on the Sensitivity of Net Photosynthesis to Oxygen (the Warburg Effect) in Tobacco

The addition of glyoxylate to tobacco (Nicotiana tabacum) leaf discs inhibited glycolate synthesis and photorespiration and increased net photosynthetic ¹⁴CO₂ fixation. This inhibition of photorespiration was investigated further by studying the effect of glyoxylate on the stimulation of photosynthesis that occurs when the atmospheric O₂ level was decreased from 21 to 3% (the Warburg effect). The Warburg effect is usually ascribed to the increased glycolate synthesis and metabolism that occurs at higher O₂ concentrations. Photosynthesis in control discs increased from 59.1 to 94.7 micromoles of CO₂ per gram fresh weight per hour (a 60% increase) when the O₂ level was lowered from 21 to 3%, while the rate for discs floated on 15 millimolar glyoxylate increased only from 82.0 to 99.7 micromoles of CO₂ per gram fresh weight per hour (a 22% increase). The decrease in the O₂ sensitivity of photosynthesis in the presence of glyoxylate was explained by changes in the rate of glycolate synthesis under the same conditions.

The rate of metabolism of the added glyoxylate by tobacco leaf discs was about 1.35 micromoles per gram fresh weight per hour and was not dependent on the O₂ concentration in the atmosphere. This rate of metabolism is about 10% the amount of stimulation in the rate of CO₂ fixation caused by the glyoxylate treatment on a molar carbon basis. Glyoxylate (10 millimolar) had no effect on the carboxylase/oxygenase activity of isolated ribulose diphosphate carboxylase. Although the biochemical mechanism by which glyoxylate inhibits glycolate synthesis and photorespiration and thereby decreases the Warburg effect is still uncertain, these results show that cellular metabolites can regulate the extent of the Warburg effect.

     

Oxygen Inhibition of Photosynthesis: I. Temperature Dependence and Relation to O(2)/CO(2) Solubility Ratio

The magnitude of the percentage inhibition of photosynthesis by atmospheric levels of O₂ in the C₃ species Solanum tuberosum L., Medicago sativa L., Phaseolus vulgaris L., Glycine max L., and Triticum aestivum L. increases in a similar manner with an increase in the apparent solubility ratio of O₂/CO₂ in the leaf over a range of solubility ratios from 25 to 45. The solubility ratio is based on calculated levels of O₂ and CO₂ in the intercellular spaces of leaves as derived from whole leaf measurements of photosynthesis and transpiration. The solubility ratio of O₂/CO₂ can be increased by increased leaf temperature under constant atmospheric levels of O₂ and CO₂ (since O₂ is relatively more soluble than CO₂ with increasing temperature); by increasing the relative levels of O₂/CO₂ in the atmosphere at a given leaf temperature, or by increased stomatal resistance. If the solubility ratio of O₂/CO₂ is kept constant, as leaf temperature is increased, by varying the levels of O₂ or CO₂ in the atmosphere, then the percentage inhibition of photosynthesis by O₂ is similar. The decreased solubility of CO₂ relative to O₂ (decreased CO₂/O₂ ratio) may be partly responsible for the increased percentage inhibition of photosynthesis by O₂ under atmospheric conditions with increasing temperature.     

Internal CO(2) Supply during Photosynthesis of Sun and Shade Grown CAM Plants in Relation to Photoinhibition

Leaves of Kalanchoë pinnata were exposed in the dark to air (allowing the fixation of CO₂ into malic acid) or 2% O₂, 0% CO₂ (preventing malic acid accumulation). They were then exposed to bright light in the presence or absence of external CO₂ and light dependent inhibition of photosynthetic properties assessed by changes in 77 K fluorescence from photosystem II (PSII), light response curves and quantum yields of O₂ exchange, rates of electron transport from H₂O through Q_B (secondary electron acceptor from the PSII reaction center) in isolated thylakoids, and numbers of functional PSII centers in intact leaf discs. Sun leaves of K. pinnata experienced greater photoinhibition when exposed to high light in the absence of CO₂ if malic acid accumulation had been prevented during the previous dark period. Shade leaves experienced a high degree of photoinhibition when exposed to high light regardless of whether malic acid had been allowed to accumulate in the previous dark period or not. Quantum yields were depressed to a greater degree than was 77 K fluorescence from PSII following photoinhibition.     

Binding of red form of Orange Carotenoid Protein (OCP) to phycobilisome is not sufficient for quenching

The Orange Carotenoid Protein (OCP) is responsible for photoprotection in many cyanobacteria. Absorption of blue light drives the conversion of the orange, inactive form (OCP^O) to the red, active form (OCP^R). Concomitantly, the N–terminal domain (NTD) and the C–terminal domain (CTD) of OCP separate, which ultimately leads to the formation of a quenched OCP^R–PBS complex. The details of the photoactivation of OCP have been intensely researched. Binding site(s) of OCP^R on the PBS core have also been proposed. However, the post–binding events of the OCP^R–PBS complex remain unclear. Here, we demonstrate that PBS–bound OCP^R is not sufficient as a PBS excitation energy quencher. Using site–directed mutagenesis, we generated a suite of single point mutations at OCP Leucine 51 (L51) of Synechocystis 6803. Steady–state and time–resolved fluorescence analyses demonstrated that all mutant proteins are unable to quench the PBS fluorescence, owing to either failed OCP binding to PBS, or, if bound, an OCP–PBS quenching state failed to form. The SDS–PAGE and Western blot analysis support that the L51A (Alanine) mutant binds to the PBS and therefore belongs to the second category. We hypothesize that upon binding to PBS, OCP^R likely reorganizes and adopts a new conformational state (OCP^3rd) different than either OCP^O or OCP^R to allow energy quenching, depending on the cross–talk between OCP^R and its PBS core–binding counterpart.     

Oxygen is not required for the browning and crosslinking of protein by pentoses: relevance to Maillard reactions in vivo

The chemical modification and crosslinking of proteins by the Maillard or browning reaction contributes to the aging of tissue proteins, and acceleration of this reaction during hyperglycemia is implicated in the pathogenesis of diabetic complications. Metal-catalyzed autoxidation reactions catalyze the browning of proteins by glucose, a process known as autoxidative glycosylation, but the effects of oxidative conditions on browning of proteins by smaller sugars has not been reported. In this work we studied the browning and crosslinking of the model protein, RNase A, by pentoses. Although antioxidative conditions inhibited the formation of glyoxal and the advanced glycation end-product, N epsilon-(carboxymethyl)lysine from arabinose, browning and crosslinking, and formation of the fluorescent crosslink pentosidine proceeded at comparable rates under oxidative and antioxidative conditions. These studies and other work on smaller dicarbonyl compounds indicate that Maillard reactions of simpler carbohydrates proceed efficiently in the absence of oxygen and suggest that antioxidant therapy for treatment of diabetic complications may have limited clinical efficacy.     

Alpha-lactalbumin unfolding is not sufficient to cause apoptosis, but is required for the conversion to HAMLET (human alpha-lactalbumin made lethal to tumor cells)

HAMLET (human alpha-lactalbumin made lethal to tumor cells) is a complex of human alpha-lactalbumin and oleic acid (C18:1:9 cis) that kills tumor cells by an apoptosis-like mechanism. Previous studies have shown that a conformational change is required to form HAMLET from alpha-lactalbumin, and that a partially unfolded conformation is maintained in the HAMLET complex. This study examined if unfolding of alpha-lactalbumin is sufficient to induce cell death. We used the bovine alpha-lactalbumin Ca(2+) site mutant D87A, which is unable to bind Ca(2+), and thus remains partially unfolded regardless of solvent conditions. The D87A mutant protein was found to be inactive in the apoptosis assay, but could readily be converted to a HAMLET-like complex in the presence of oleic acid. BAMLET (bovine alpha-lactalbumin made lethal to tumor cells) and D87A-BAMLET complexes were both able to kill tumor cells. This activity was independent of the Ca(2+)site, as HAMLET maintained a high affinity for Ca(2+) but D87A-BAMLET was active with no Ca(2+) bound. We conclude that partial unfolding of alpha-lactalbumin is necessary but not sufficient to trigger cell death, and that the activity of HAMLET is defined both by the protein and the lipid cofactor. Furthermore, a functional Ca(2+)-binding site is not required for conversion of alpha-lactalbumin to the active complex or to cause cell death. This suggests that the lipid cofactor stabilizes the altered fold without interfering with the Ca(2+)site.     

PSII supercomplex disassembly is not needed for the induction of energy quenching (qE)

Photoprotection by non-photochemical quenching is important for optimal growth and development, especially during dynamic changes of the light intensity. The main component responsible for energy dissipation is called qE. It has been proposed that qE involves the reorganization of the photosynthetic complexes and especially of Photosystem II. However, despite a number of studies, there are still contradictory results concerning the structural changes in PSII during qE induction. The main limitation in addressing this point is the very fast nature of the off switch of qE, since the illumination is usually performed in folio and the preparation of the thylakoids requires a dark period. To avoid qE relaxation during thylakoid isolation, in this work quenching was induced directly on isolated and functional thylakoids that were then solubilized in the light. The analysis of the quenched thylakoids in native gel showed only a small decrease in the large PSII supercomplexes (C₂S₂M₂/C₂S₂M) which is most likely due to photoinhibition/light acclimation since it does not recover in the dark. This result indicates that qE rise is not accompanied by a structural disassembly of the PSII supercomplexes.

     

The C terminus of sigma(32) is not essential for degradation by FtsH

A key step in the regulation of heat shock genes in Escherichia coli is the stress-dependent degradation of the heat shock promoter-specific sigma(32) subunit of RNA polymerase by the AAA protease, FtsH. Previous studies implicated the C termini of protein substrates, including sigma(32), as degradation signals for AAA proteases. We investigated the role of the C terminus of sigma(32) in FtsH-dependent degradation by analysis of C-terminally truncated sigma(32) mutant proteins. Deletion of the 5, 11, 15, and 21 C-terminal residues of sigma(32) did not affect degradation in vivo or in vitro. Furthermore, a peptide comprising the C-terminal 21 residues of sigma(32) was not degraded by FtsH in vitro and thus did not serve as a recognition sequence for the protease, while an unrelated peptide of similar length was efficiently degraded. The truncated sigma(32) mutant proteins remained capable of associating with DnaK and DnaJ in vitro but showed intermediate (5-amino-acid deletion) and strong (11-, 15-, and 21-amino-acid deletions) defects in association with RNA polymerase in vitro and biological activity in vivo. These results indicate an important role for the C terminus of sigma(32) in RNA polymerase binding but no essential role for FtsH-dependent degradation and association of chaperones.     

Cortical Air Spaces (Aerenchyma) in Roots of Corn Subjected to Oxygen Stress: STRUCTURE AND INFLUENCE ON UPTAKE AND TRANSLOCATION OF RUBIDIUM IONS

When the seminal root system of 14-day-old corn (Zea mays cv. Dekalb 202) was subjected to O₂ stress, nodal roots with well developed cortical air spaces (aerenchyma) grew into the deoxygenated solution. Microscopic examination showed that there was extensive breakdown of cells in the midcortex of these roots, while the stele, endodermis, and inner layer of cortical cells remained complete, as did the outer layers of the cortex and the epidermis. Occasional files of intact cells, and the wall residues of collapsed cells, crossed the space between inner and outer cortex. Experiments with short, intact root segments with and without air spaces showed that in the presence of O₂ the ability to absorb and translocate ⁸⁶Rb⁺, per unit volume or length of root, was little affected by cortical degeneration. The distribution across root sections of recently supplied strontium and rubidium, determined by electron microprobe analysis, indicated that in roots with air spaces the strands of wall residues bridging the cortex could be involved in maintaining the conduction of ions from the outer cortex up to the endodermis.     

RDH13L,an Enzyme Responsible for the Aldehyde-Alcohol Redox Coupling Reaction (AL-OL Coupling Reaction) to Supply 11-cis Retinal in the Carp Cone Retinoid Cycle

Cone photoreceptors require effective pigment regeneration mechanisms to maintain their sensitivity in the light. Our previous studies in carp cones suggested the presence of an unconventional and very effective mechanism to produce 11-cis retinal, the necessary component in pigment regeneration. In this reaction (aldehyde-alcohol redox coupling reaction, AL-OL coupling reaction), formation of 11-cis retinal, i.e. oxidation of 11-cis retinol is coupled to reduction of an aldehyde at a 1:1 molar ratio without exogenous NADP(H) which is usually required in this kind of reaction. Here, we identified carp retinol dehydrogenase 13-like (RDH13L) as an enzyme catalyzing the AL-OL coupling reaction. RDH13L was partially purified from purified carp cones, identified as a candidate protein, and its AL-OL coupling activity was confirmed using recombinant RDH13L. We further examined the substrate specificity, subcellular localization, and expression level of RDH13L. Based on these results, we concluded that RDH13L contributes to a significant part, but not all, of the AL-OL coupling activity in carp cones. RDH13L contained tightly bound NADP⁺ which presumably functions as a cofactor in the reaction. Mouse RDH14, a mouse homolog of carp RDH13L, also showed the AL-OL coupling activity. Interestingly, although carp cone membranes, carp RDH13L and mouse RDH14 all showed the coupling activity at 15–37 °C, they also showed a conventional NADP⁺-dependent 11-cis retinol oxidation activity above 25 °C without addition of aldehydes. This dual mechanism of 11-cis retinal synthesis attained by carp RDH13L and mouse RDH14 probably contribute to effective pigment regeneration in cones that function in the light.     

Expression of Genes for Photosynthesis and the Relationship to Salt Tolerance of Alfalfa (Medicago sativa) Cells

The basis for the salt tolerant phenotype of a line of Medicagosativa (alfalfa) cells (HG2-N1) derived by selection from asalt sensitive line (HG2) was studied. The salt tolerant HG2-N1cell line shows eleven fold elevated chlorophyll content overthat of the parent salt sensitive HG2 cell line, with an additionaltwo fold increase in chlorophyll levels when the cells are grownin 1% NaCl. In this study, we demonstrate that the chlorophyllaccumulation and response to salt was associated with largeincreases in the two photosynthesis related mRNAs, rbcL (ribulose-l,5-bis-phosphatecarboxylase [Rubisco] large subunit) and rbcS (Rubisco smallsubunit) and a substantial increase in the activity of the holoenzyme.The salinity-induced increase in catalytically competent Rubiscoprotein in the salt tolerant cell line was highly responsiveto light and correlated with the salt tolerant phenotype. Inaddition, NaCl stimulated rbcL and rbcS mRNA and Rubisco accumulationin dark grown salt tolerant cells, indicating that salt couldsubstitute to some degree for light in stimulating increasesin specific mRNA and protein concentrations. Increased photosyntheticcompetence associated with these increased protein levels wasapparently important in contributing to the salt tolerant phenotypeof HG2-N1, since PS II electron transport inhibitors (DCMU,cyanazine) were found to significantly reduce the growth ofthis cell line in the presence of salt, but not in the absenceof salt. These results suggest that the salt-induced increasein mRNA and protein accumulation involved in photosynthesismay play a significant role in the salt tolerant capabilityof HG2-N1 alfalfa cells. (Received April 2, 1990; Accepted September 10, 1990)     

Binding to the high-affinity M-type receptor for secreted phospholipases A(2) is not obligatory for the presynaptic neurotoxicity of ammodytoxin A

R180, isolated from porcine brain cortex, is a high-affinity membrane receptor for ammodytoxin A (AtxA), a secreted phospholipase A(2) (sPLA(2)) and presynaptically active neurotoxin from venom of the long-nosed viper (Vipera ammodytes ammodytes). As a member of the M-type sPLA(2) receptors, present on the mammalian plasma membrane, R180 has been proposed to be responsible for one of the first events in the process of presynaptic neurotoxicity, the binding of the toxin to the nerve cell. To test this hypothesis, we prepared and analyzed three N-terminal fusion proteins of AtxA possessing a 12 or 5 amino acid residue peptide. The presence of such an additional "propeptide" prevented interaction of the toxin with the M-type receptor but not its lethality in mouse and neurotoxic effects on a mouse phrenic nerve-hemidiaphragm preparation. In addition, antibodies raised against the sPLA(2)-binding C-type lectin-like domain 5 of the M-type sPLA(2) receptor were unable to abolish the neurotoxic action of AtxA on the neuromuscular preparation. The specific enymatic activities of the fusion AtxAs were two to three orders of magnitude lower from that of the wild type, yet resulting in a similar but less pronounced neurotoxic profile on the neuromuscular junction. This is in accordance with other data showing that a minimal enzymatic activity suffices for presynaptic toxicity of sPLA(2)s to occur. Our results indicate that the interaction of AtxA with the M-type sPLA(2) receptor at the plasma membrane is not essential for presynaptic activity of the toxin. Interaction of AtxA with two intracellular proteins, calmodulin and the R25 receptor, was affected but not prevented by the presence of the N-terminal fusion peptides, implying that these proteins may play a role in the sPLA(2) neurotoxicity.