| 1. |
- Møller, Ian Max, et al.
(författare)
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Isolation of Highly Purified, Intact, and Functional Mitochondria from Potato Tubers Using a Two-in-One Percoll Density Gradient
- 2022
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Ingår i: Plant Mitochondria : Methods and Protocols - Methods and Protocols. - New York, NY : Springer US. - 1940-6029 .- 1064-3745. - 9781071616536 - 9781071616529 ; 2363, s. 39-50
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Bokkapitel (refereegranskat)abstract
- The isolation of mitochondria from potato tubers (Solanum tuberosum L.) is described, but the methodology can easily be adapted to other storage tissues. After homogenization of the tissue, filtration and differential centrifugation, the key step is a Percoll density gradient centrifugation. The Percoll gradient contains two parts: a bottom part containing Percoll in 0.3 M sucrose, and a slightly less dense top part containing Percoll in 0.3 M mannitol. After centrifugation, a density gradient is formed that is almost linear in the central part, and this is where the band containing the purified intact mitochondria is formed. This method makes it possible to process large amounts of plant material (2–6 kg) and saves at least 1.5 h on the preparation time compared to methods where two consecutive purification methods are used. Nonetheless, it yields large amounts of mitochondria (50–125 mg protein) of very high purity, intactness and functionality.
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| 2. |
- Rasmusson, Allan G., et al.
(författare)
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Assessment of Respiratory Enzymes in Intact Cells by Permeabilization with Alamethicin
- 2022
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Ingår i: Plant Mitochondria : Methods and Protocols - Methods and Protocols. - New York, NY : Springer US. - 1940-6029 .- 1064-3745. - 9781071616536 - 9781071616529 ; 2363, s. 77-84
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Bokkapitel (refereegranskat)abstract
- We here describe measurements of respiratory enzymes in situ, which can be done on very small cell samples and make mitochondrial isolation unnecessary. The method is based on the ability of the fungal peptide alamethicin to permeate biological membranes from the net positively charged side, and form nonspecific ion channels. These channels allow rapid transport of substrates and products across the plasma membrane, the inner mitochondrial membrane, and the inner plastid envelope. In this way, mitochondrial enzyme activities can be studied without disrupting the cells. The enzymes can be investigated in their natural proteinaceous environment and the activity of enzymes, also those sensitive to detergents or to dilution, can be quantified on a whole cell basis. We here present protocols for in situ measurement of two mitochondrial enzymatic activities: malate oxidation measured as oxygen consumption by the electron transport chain, which is sensitive to detergents, and NAD+-isocitrate dehydrogenase, a tricarboxylic acid cycle enzyme that dissociates upon dilution.
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| 3. |
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| 4. |
- Agius, Stephanie C, et al.
(författare)
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NAD(P) turnover in plant mitochondria
- 2001
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Ingår i: Australian Journal of Plant Physiology. - 0310-7841. ; 28:6, s. 461-470
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Tidskriftsartikel (refereegranskat)abstract
- An analytical procedure based on alkaline extraction and HPLC analysis was adapted for quantification of pyridine nucleotides in plant mitochondria. The amounts of NAD and NADP extracted from seven different species varied from 1.0 to 3.7 and 0 to 0.5 nmol (mg protein) –1 , respectively. Although NADP was found in four species, its reduced form was in all cases below the detection limit of 0.1 nmol (mg protein) –1 . The NAD pool was mainly oxidized in the absence of substrates. However, oxidation of substrates followed by anaerobiosis caused 50–92% NAD pool reduction, indicating that the majority of the NAD+ was metabolically active. The NAD reduction level in potato tuber mitochondria oxidizing malate varied with assay conditions. The highest level of reduction (>80%) was reached at anaerobiosis, at pH 6.5 and 7.2, conditions favouring malic enzyme (ME), whereas the lowest reduction level (0%) was observed at pH 7.5, conditions favouring malate dehydrogenase (MDH). Mitochondria incubated at 0°C without respiratory substrate showed a loss of endogenous NAD + which correlated with a decline in the rate of oxidation of NAD+ -linked substrates. The lost NAD+ was mainly recovered as breakdown products in both the surrounding medium and the mitochondria. When submitochondrial fractions were incubated with NAD + or NADP + , the highest rate of NAD(P)+metabolism was detected in the outer membrane fraction. The metabolites detected, adenosine monophosphate (AMP), nicotinamide mononucleotide (NMN) and adenosine, imply that several enzymes involved in pyridine nucleotide degradation, including an NAD pyrophosphatase, are localized to the outer membrane.
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| 5. |
- Aidemark, Mari, et al.
(författare)
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Regulation of callose synthase activity in situ in alamethicin-permeabilized Arabidopsis and tobacco suspension cells
- 2009
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Ingår i: BMC Plant Biology. - : Springer Science and Business Media LLC. - 1471-2229. ; 9
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Tidskriftsartikel (refereegranskat)abstract
- Background: The cell wall component callose is mainly synthesized at certain developmental stages and after wounding or pathogen attack. Callose synthases are membrane-bound enzymes that have been relatively well characterized in vitro using isolated membrane fractions or purified enzyme. However, little is known about their functional properties in situ, under conditions when the cell wall is intact. To allow in situ investigations of the regulation of callose synthesis, cell suspensions of Arabidopsis thaliana (Col-0), and tobacco (BY-2), were permeabilized with the channel-forming peptide alamethicin. Results: Nucleic acid-binding dyes and marker enzymes demonstrated alamethicin permeabilization of plasma membrane, mitochondria and plastids, also allowing callose synthase measurements. In the presence of alamethicin, Ca2+ addition was required for callose synthase activity, and the activity was further stimulated by Mg2+ Cells pretreated with oryzalin to destabilize the microtubules prior to alamethicin permeabilization showed significantly lower callose synthase activity as compared to non-treated cells. As judged by aniline blue staining, the callose formed was deposited both at the cell walls joining adjacent cells and at discrete punctate locations earlier described as half plasmodesmata on the outer walls. This pattern was unaffected by oryzalin pretreatment, showing a quantitative rather than a qualitative effect of polymerized tubulin on callose synthase activity. No callose was deposited unless alamethicin, Ca2+ and UDP-glucose were present. Tubulin and callose synthase were furthermore part of the same plasma membrane protein complex, as judged by two-dimensional blue native SDS-PAGE. Conclusion: Alamethicin permeabilization allowed determination of callose synthase regulation and tubulin interaction in the natural crowded cellular environment and under conditions where contacts between the cell wall, the plasma membrane and cytoskeletal macromolecules remained. The results also suggest that alamethicin permeabilization induces a defense response mimicking the natural physical separation of cells (for example when intercellulars are formed), during which plasmodesmata are transiently left open.
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| 6. |
- Aidemark, Mari, et al.
(författare)
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Trichoderma viride cellulase induces resistance to the antibiotic pore-forming peptide alamethicin associated with changes in the plasma membrane lipid composition of tobacco BY-2 cells
- 2010
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Ingår i: Bmc Plant Biology. - : Springer Science and Business Media LLC. - 1471-2229. ; 10
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Tidskriftsartikel (refereegranskat)abstract
- Background: Alamethicin is a membrane-active peptide isolated from the beneficial root-colonising fungus Trichoderma viride. This peptide can insert into membranes to form voltage-dependent pores. We have previously shown that alamethicin efficiently permeabilises the plasma membrane, mitochondria and plastids of cultured plant cells. In the present investigation, tobacco cells (Nicotiana tabacum L. cv Bright Yellow-2) were pre-treated with elicitors of defence responses to study whether this would affect permeabilisation. Results: Oxygen consumption experiments showed that added cellulase, already upon a limited cell wall digestion, induced a cellular resistance to alamethicin permeabilisation. This effect could not be elicited by xylanase or bacterial elicitors such as flg22 or elf18. The induction of alamethicin resistance was independent of novel protein synthesis. Also, the permeabilisation was unaffected by the membrane-depolarising agent FCCP. As judged by lipid analyses, isolated plasma membranes from cellulase-pretreated tobacco cells contained less negatively charged phospholipids ( PS and PI), yet higher ratios of membrane lipid fatty acid to sterol and to protein, as compared to control membranes. Conclusion: We suggest that altered membrane lipid composition as induced by cellulase activity may render the cells resistant to alamethicin. This induced resistance could reflect a natural process where the plant cells alter their sensitivity to membrane pore-forming agents secreted by Trichoderma spp. to attack other microorganisms, and thus adding to the beneficial effect that Trichoderma has for plant root growth. Furthermore, our data extends previous reports on artificial membranes on the importance of lipid packing and charge for alamethicin permeabilisation to in vivo conditions.
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| 7. |
- Björn, Lars Olof, et al.
(författare)
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Photosensitivity in sponge due to cytochrome c oxidase?
- 2009
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Ingår i: Photochemical and Photobiological Sciences. - : Springer Science and Business Media LLC. - 1474-9092 .- 1474-905X. ; 8, s. 755-757
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Tidskriftsartikel (refereegranskat)abstract
- An action spectrum for photosensitivity in sponge larvae published by Leys et al. [J. Comp. Physiol., A, 2002, 188, 199–202] was interpreted by the authors as being due to a combination of light absorption by flavin or carotenoid in the blue region, and another pigment such as pterin in the long-wavelength region. Here we show here that their action spectrum closely matches the absorption spectrum of reduced cytochrome c oxidase that is present in sponges, and compare with other photoreactions which are thought to be due to this chromoprotein.
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| 8. |
- Browse, John A., et al.
(författare)
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Respiration and lipid metabolism
- 2010
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Ingår i: Plant Physiology (5th edition). - 9780878938667 - 0878938664 ; , s. 305-342
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Bokkapitel (refereegranskat)abstract
- Abstract is not available
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| 9. |
- Browse, John A., et al.
(författare)
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Respiration and Lipid Metabolism
- 2014. - 6th ed.
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Ingår i: Plant Physiology and Development. - 9781605352558 - 9781605354354 ; , s. 317-352
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Bokkapitel (refereegranskat)
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| 10. |
- Browse, John, et al.
(författare)
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Respiration and lipid metabolism
- 2006
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Ingår i: Plant Physiology, 4th Ed.. ; , s. 253-288
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Bokkapitel (övrigt vetenskapligt/konstnärligt)
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| 11. |
- Browse, John, et al.
(författare)
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Respiration and lipid metabolism
- 2002
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Ingår i: Plant Physiology, 3rd Ed.. - 0878938230 ; , s. 223-258
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Bokkapitel (övrigt vetenskapligt/konstnärligt)
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| 12. |
- Bykova, Natalia V., et al.
(författare)
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Two separate transhydrogenase activities are present in plant mitochondria
- 1999
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Ingår i: Biochemical and Biophysical Research Communications. - : Elsevier BV. - 0006-291X .- 1090-2104. ; 265:1, s. 106-111
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Tidskriftsartikel (refereegranskat)abstract
- Inside-out submitochondrial particles from both potato tubers and pea leaves catalyze the transfer of hydride equivalents from NADPH to NAD+ as monitored with a substrate-regenerating system. The NAD+ analogue acetylpyridine adenine dinucleotide is also reduced by NADPH and incomplete inhibition by the complex I inhibitor diphenyleneiodonium (DPI) indicates that two enzymes are involved in this reaction. Gel-filtration chromatography of solubilized mitochondrial membrane complexes confirms that the DPI-sensitive TH activity is due to NADH-ubiquinone oxidoreductase (EC 1.6.5.3, complex I), whereas the DPI-insensitive activity is due to a separate enzyme eluting around 220 kDa. The DPI-insensitive TH activity is specific for the 4B proton on NADH, whereas there is no indication of a 4A-specific activity characteristic of a mammalian-type energy-linked TH. The DPI-insensitive TH may be similar to the soluble type of transhydrogenase found in, e.g., Pseudomonas. The presence of non-energy-linked TH activities directly coupling the matrix NAD(H) and NADP(H) pools will have important consequences for the regulation of NADP-linked processes in plant mitochondria.
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| 13. |
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| 14. |
- Catania, Alessia, et al.
(författare)
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Arabidopsis thaliana alternative dehydrogenases : A potential therapy for mitochondrial complex i deficiency? Perspectives and pitfalls
- 2019
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Ingår i: Orphanet Journal of Rare Diseases. - : Springer Science and Business Media LLC. - 1750-1172. ; 14:1
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Tidskriftsartikel (refereegranskat)abstract
- Background: Complex I (CI or NADH:ubiquinone oxidoreductase) deficiency is the most frequent cause of mitochondrial respiratory chain defect. Successful attempts to rescue CI function by introducing an exogenous NADH dehydrogenase, such as the NDI1 from Saccharomyces cerevisiae (ScNDI1), have been reported although with drawbacks related to competition with CI. In contrast to ScNDI1, which is permanently active in yeast naturally devoid of CI, plant alternative NADH dehydrogenases (NDH-2) support the oxidation of NADH only when the CI is metabolically inactive and conceivably when the concentration of matrix NADH exceeds a certain threshold. We therefore explored the feasibility of CI rescue by NDH-2 from Arabidopsis thaliana (At) in human CI defective fibroblasts. Results: We showed that, other than ScNDI1, two different NDH-2 (AtNDA2 and AtNDB4) targeted to the mitochondria were able to rescue CI deficiency and decrease oxidative stress as indicated by a normalization of SOD activity in human CI-defective fibroblasts. We further demonstrated that when expressed in human control fibroblasts, AtNDA2 shows an affinity for NADH oxidation similar to that of CI, thus competing with CI for the oxidation of NADH as opposed to our initial hypothesis. This competition reduced the amount of ATP produced per oxygen atom reduced to water by half in control cells. Conclusions: In conclusion, despite their promising potential to rescue CI defects, due to a possible competition with remaining CI activity, plant NDH-2 should be regarded with caution as potential therapeutic tools for human mitochondrial diseases.
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| 15. |
- Dotson, Bradley R., et al.
(författare)
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The antibiotic peptaibol alamethicin from Trichoderma permeabilises Arabidopsis root apical meristem and epidermis but is antagonised by cellulase-induced resistance to alamethicin
- 2018
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Ingår i: BMC Plant Biology. - : Springer Science and Business Media LLC. - 1471-2229. ; 18:1
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Tidskriftsartikel (refereegranskat)abstract
- Background: Trichoderma fungi live in the soil rhizosphere and are beneficial for plant growth and pathogen resistance. Several species and strains are currently used worldwide in co-cultivation with crops as a biocontrol alternative to chemical pesticides even though little is known about the exact mechanisms of the beneficial interaction. We earlier found alamethicin, a peptide antibiotic secreted by Trichoderma, to efficiently permeabilise cultured tobacco cells. However, pre-treatment with Trichoderma cellulase made the cells resistant to subsequent alamethicin, suggesting a potential mechanism for plant tolerance to Trichoderma, needed for mutualistic symbiosis. Results: We here investigated intact sterile-grown Arabidopsis thaliana seedlings germinated in water or growth medium. These could be permeabilised by alamethicin but not if pretreated with cellulase. By following the fluorescence from the membrane-impermeable DNA-binding probe propidium iodide, we found alamethicin to mainly permeabilise root tips, especially the apical meristem and epidermis cells, but not the root cap and basal meristem cells nor cortex cells. Alamethicin permeabilisation and cellulase-induced resistance were confirmed by developing a quantitative in situ assay based on NADP-isocitrate dehydrogenase accessibility. The combined assays also showed that hyperosmotic treatment after the cellulase pretreatment abolished the induced cellulase resistance. Conclusion: We here conclude the presence of cell-specific alamethicin permeabilisation, and cellulase-induced resistance to it, in root tip apical meristem and epidermis of the model organism A. thaliana. We suggest that contact between the plasma membrane and the cell wall is needed for the resistance to remain. Our results indicate a potential mode for the plant to avoid negative effects of alamethicin on plant growth and localises the point of potential damage and response. The results also open up for identification of plant genetic components essential for beneficial effects from Trichoderma on plants.
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| 16. |
- Escobar, Matthew, et al.
(författare)
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Light regulation of the Arabidopsis respiratory chain. Multiple discrete photoreceptor responses contribute to induction of type II NAD(P)H dehydrogenase genes
- 2004
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Ingår i: Plant Physiology. - : Oxford University Press (OUP). - 1532-2548 .- 0032-0889. ; 136:1, s. 2710-2721
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Tidskriftsartikel (refereegranskat)abstract
- Controlled oxidation reactions catalyzed by the large, proton-pumping complexes of the respiratory chain generate an electrochemical gradient across the mitochondrial inner membrane that is harnessed for ATP production. However, several alternative respiratory pathways in plants allow the maintenance of substrate oxidation while minimizing the production of ATE We have investigated the role of light in the regulation of these energy-dissipating pathways by transcriptional profiling of the alternative oxidase, uncoupling protein, and type II NAD(P)H dehydrogenase gene families in etiolated Arabidopsis seedlings. Expression of the nda1 and ndc1 NAD(P)H dehydrogenase genes was rapidly up-regulated by a broad range of light intensities and qualities. For both genes, light induction appears to be a direct transcriptional effect that is independent of carbon status. Mutant analyses demonstrated the involvement of two separate photoreceptor families in nda1 and ndc1 light regulation: the phytochromes (phyA and phyB) and an undetermined blue light photoreceptor. In the case of the nda1 gene, the different photoreceptor systems generate distinct kinetic induction profiles that are integrated in white light response. Primary transcriptional control of light response was localized to a 99-bp region of the nda1 promoter, which contains an 1-box flanked by two GT-1 elements, an arrangement prevalent in the promoters of photosynthesis-associated genes. Light induction was specific to nda1 and ndc1. The only other substantial light effect observed was a decrease in aox2 expression. Overall, these results suggest that light directly influences the respiratory electron transport chain via photoreceptor-mediated transcriptional control, likely for supporting photosynthetic metabolism.
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| 17. |
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| 18. |
- Escobar, Matthew, et al.
(författare)
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Reorganization of the alternative pathways of the Arabidopsis respiratory chain by nitrogen supply: opposing effects of ammonium and nitrate
- 2006
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Ingår i: Plant Journal. - 1365-313X. ; 45:5, s. 775-788
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Tidskriftsartikel (refereegranskat)abstract
- The mitochondrial oxidative phosphorylation system in plants possesses a variety of alternative pathways that decrease respiratory ATP production. These alternative pathways are mediated by three classes of bypass proteins: the type II NAD(P) H dehydrogenases (which circumvent complex I of the electron transport chain), the alternative oxidases (AOXs; which circumvent complexes III and IV) and the uncoupling proteins ( which circumvent ATP synthase). We have monitored the expression of all genes encoding respiratory bypass proteins in Arabidopsis thaliana growing with different sources of inorganic nitrogen (N). Resupply of nitrate (NO3-) to N-limited seedling cultures caused a decrease in the transcript abundance of several type II NAD(P) H dehydrogenase and AOX genes, while resupply of ammonium (NH4+) led to broad increases in expression in the same gene families. Similar results were observed upon switching between nitrate and ammonium in the absence of N stress. Nitrate signalling was found to be mediated primarily by the nitrate ion itself, whereas ammonium regulation was dependent upon assimilation and affected by changes in apoplastic pH. Corresponding alterations in alternative respiratory pathway capacities were apparent in seedlings supplied with either nitrate or ammonium as an N source and in mitochondria purified from the seedlings. Specifically, AOX capacity and protein abundance, as well as calcium-dependent external NADH oxidation, were substantially elevated after growth on ammonium. The increased capacity of respiratory bypass pathways after switching from nitrate to ammonium was correlated to an overall respiratory increase.
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| 19. |
- Fan, Kaijian, et al.
(författare)
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The pentatricopeptide repeat protein EMP603 is required for the splicing of mitochondrial Nad1 intron 2 and seed development in maize
- 2021
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Ingår i: Journal of Experimental Botany. - : Oxford University Press (OUP). - 0022-0957 .- 1460-2431. ; 72:20, s. 6933-6948
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Tidskriftsartikel (refereegranskat)abstract
- Intron splicing is an essential event in post-transcriptional RNA processing in plant mitochondria, which requires the participation of diverse nuclear-encoded splicing factors. However, it is presently unclear how these proteins cooperatively take part in the splicing of specific introns. In this study, we characterized a nuclear-encoded mitochondrial P-type pentatricopeptide repeat (PPR) protein named EMP603. This protein is essential for splicing of intron 2 in the Nad1 gene and interacts with the mitochondria-localized DEAD-box RNA helicase PMH2-5140, the RAD52-like proteins ODB1-0814 and ODB1-5061, and the CRM domain-containing protein Zm-mCSF1. Further study revealed that the N-terminal region of EMP603 interacts with the DEAD-box of PMH2-5140, the CRM domain of Zm-mCSF1, and OBD1-5061, but not with OBD1-0814, whereas the PPR domain of EMP603 can interact with ODB1-0814, ODB1-5061, and PMH2-5140, but not with Zm-mCSF1. Defects in EMP603 severely disrupt the assembly and activity of mitochondrial complex I, leading to impaired mitochondrial function, and delayed seed development. The interactions revealed between EMP603 and PMH2-5140, ODB1-0814, ODB1-5061, and Zm-mCSF1 indicate a possible involvement of a dynamic 'spliceosome-like' complex in intron splicing, and may accelerate the elucidation of the intron splicing mechanism in plant mitochondria.
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| 20. |
- Florez-Sarasa, Igor, et al.
(författare)
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In vivo cytochrome and alternative pathway respiration in leaves of Arabidopsis thaliana plants with altered alternative oxidase under different light conditions
- 2011
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Ingår i: Plant, Cell and Environment. - : Wiley. - 0140-7791. ; 34:8, s. 1373-1383
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Tidskriftsartikel (refereegranskat)abstract
- The in vivo activity of the alternative pathway (V-alt) has been studied using the oxygen isotope fractionation method in leaves of Arabidopsis thaliana modified for the expression of the AtAOX1a gene by anti-sense (AS-12) or overexpression (XX-2). Under non-stressful conditions, V-alt was similar in all plant lines regardless of its different alternative pathway capacities (V-alt). Total leaf respiration (V-t) and V-alt were directly related to growth light conditions while electron partitioning between the cytochrome pathway (CP) and alternative pathway (AP) was unchanged by light levels. Interestingly, the AP functioned at full capacity in anti-sense plants under both growth light conditions. The role of the AP in response to a high light stress induced by short-term high light treatment (HLT) was also studied. In wild type and XX-2, both CP and AP rates increased proportionally after HLT while in AS-12, where the AP was unable to increase its rate, the CP accommodated all the increase in respiration. The results obtained under high light stress suggest that flexibility in the response of the mitochondrial electron transport chain is involved in sustaining photosynthetic rates in response to this stress while the saturated AP in AS-12 plants may contribute to the observed increase in photoinhibition.
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| 21. |
- Florez-Sarasa, Igor, et al.
(författare)
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Light-responsive metabolite and transcript levels are maintained following a dark-adaptation period in leaves of Arabidopsis thaliana.
- 2012
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Ingår i: New Phytologist. - : Wiley. - 1469-8137 .- 0028-646X. ; 195:1, s. 136-148
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Tidskriftsartikel (refereegranskat)abstract
- The effect of previous light conditions on metabolite and transcript levels was investigated in leaves of Arabidopsis thaliana during illumination and after light-enhanced dark respiration (LEDR), when dark respiration was measured. • Primary carbon metabolites and the expression of light-responsive respiratory genes were determined in A. thaliana leaves before and after 30 min of darkness following different light conditions. In addition, metabolite levels were determined in the middle of the night and the in vivo activities of cytochrome and alternative respiratory pathways were determined by oxygen isotope fractionation. • A large number of metabolites were increased in leaves of plants growing in or transiently exposed to higher light intensities. Transcript levels of respiratory genes were also increased after high light treatment. For the majority of the light-induced metabolites and transcripts, the levels were maintained after 30 min of darkness, where higher and persistent respiratory activities were also observed. The levels of many metabolites were lower at night than after 30 min of darkness imposed in the day, but respiratory activities remained similar. • The results obtained suggest that 'dark' respiration measurements, as usually performed, are probably made under conditions in which the overall status of metabolites is strongly influenced by the previous light conditions.
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| 22. |
- Fredlund, Kenneth M., et al.
(författare)
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Oxidation of external NAD(P)H by purified mitochondria from fresh and aged red beetroots (Beta vulgaris L.)
- 1991
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Ingår i: Plant Physiology. - : Oxford University Press (OUP). - 0032-0889 .- 1532-2548. ; 97:1, s. 99-103
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Tidskriftsartikel (refereegranskat)abstract
- Mitochondria were isolated from fresh beetroots (Beta vulgaris L. cvs Rubria and Nina) by differential centrifugation followed by Percoll gradient centrifugation. These purified mitochondria oxidized external NADH, although relatively slowly (20-40 versus 100-120 nanomoles oxygen per minute times milligram protein for NADH and succinate oxidation, respectively), with respiratory control ratios of two to three and ADP/O ratios of 1.2 to 1.6. NADPH was also oxidized, but even more slowly and with little or no coupling. The optimum for both NADH and NADPH oxidation by fresh beetroot mitochondria was pH 6. The rate of external NADH oxidation by isolated mitochondria was enhanced threefold during storage of the intact tubers at 10°C for 12 weeks. The optimum of the induced NADH oxidation was approximately pH 6.8. Succinate and malate oxidation only increased by 30% during the same period and NADPH oxidation was constant. This is strong evidence that NADH and NADPH oxidation are catalyzed by different enzymes at least in beetroots. Activity staining of nondenaturing polyacrylamide gels with NADH and Nitro Blue Tetrazolium did not show differences in banding pattern between mitochondria isolated from fresh and stored beetroots. The induction is discussed in relation to physiological aging processes.
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| 23. |
- Geisler, Daniela, et al.
(författare)
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Antimycin A treatment decreasees respiratory internal rotenone-insensitive NADH oxidation capacity in potato leaves
- 2004
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Ingår i: BMC Plant Biology. - : Springer Science and Business Media LLC. - 1471-2229. ; 4:8, s. 1-10
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Tidskriftsartikel (refereegranskat)abstract
- Background The plant respiratory chain contains several energy-dissipating enzymes, these being type II NAD(P)H dehydrogenases and the alternative oxidase, not present in mammals. The physiological functions of type II NAD(P)H dehydrogenases are largely unclear and little is known about their responses to stress. In this investigation, potato plants (Solanum tuberosum L., cv. Desiree) were sprayed with antimycin A, an inhibitor of the cytochrome pathway. Enzyme capacities of NAD(P)H dehydrogenases (EC 1.6.5.3) and the alternative oxidase were then analysed in isolated leaf mitochondria. Results We report a specific decrease in internal rotenone-insensitive NADH dehydrogenase capacity in mitochondria from antimycin A-treated leaves. External NADPH dehydrogenase and alternative oxidase capacities remained unaffected by the treatment. Western blotting revealed no change in protein abundance for two characterised NAD(P)H dehydrogenase homologues, NDA1 and NDB1, nor for two subunits of complex I. The alternative oxidase was at most only slightly increased. Transcript levels of nda1, as well as an expressed sequence tag derived from a previously uninvestigated closely related potato homologue, remained unchanged by the treatment. As compared to the daily rhythm-regulated nda1, the novel homologue displayed steady transcript levels over the time investigated. Conclusions The internal rotenone-insensitive NADH oxidation decreases after antimycin A treatment of potato leaves. However, the decrease is not due to changes in expression of known nda genes. One consequence of the lower NADH dehydrogenase capacity may be a stabilisation of the respiratory chain reduction level, should the overall capacity of the cytochrome and the alternative pathway be restricted.
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| 24. |
- Geisler, Daniela, et al.
(författare)
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Ca2+-binding and Ca2+-independent respiratory NADH and NADPH dehydrogenases of Arabidopsis thaliana.
- 2007
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Ingår i: Journal of Biological Chemistry. - 1083-351X. ; 282:39, s. 28455-28464
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Tidskriftsartikel (refereegranskat)abstract
- Type II NAD(P)H:quinone oxidoreductases are single polypeptide proteins widespread in the living world. They bypass the first site of respiratory energy conservation, constituted by the type I NADH dehydrogenases. To investigate substrate specificities and Ca2+ binding properties of seven predicted type II NAD(P)H dehydrogenases of Arabidopsis thaliana we have produced them as T7-tagged fusion proteins in Escherichia coli. The NDB1 and NDB2 enzymes were found to bind Ca2+, and a single amino acid substitution in the EF hand motif of NDB1 abolished the Ca2+ binding. NDB2 and NDB4 functionally complemented an E. coli mutant deficient in endogenous type I and type II NADH dehydrogenases. This demonstrates that these two plant enzymes can substitute for the NADH dehydrogenases in the bacterial respiratory chain. Three NDB-type enzymes displayed distinct catalytic profiles with substrate specificities and Ca2+ stimulation being considerably affected by changes in pH and substrate concentrations. Under physiologically relevant conditions, the NDB1 fusion protein acted as a Ca2+-dependent NADPH dehydrogenase. NDB2 and NDB4 fusion proteins were NADH-specific, and NDB2 was stimulated by Ca2+. The observed activity profiles of the NDB-type enzymes provide a fundament for understanding the mitochondrial system for direct oxidation of cytosolic NAD(P)H in plants. Our findings also suggest different modes of regulation and metabolic roles for the analyzed A. thaliana enzymes.
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| 25. |
- Glab, Bartosz, et al.
(författare)
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Cloning of Glycerophosphocholine Acyltransferase (GPCAT) from Fungi and Plants A NOVEL ENZYME IN PHOSPHATIDYLCHOLINE SYNTHESIS
- 2016
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Ingår i: Journal of Biological Chemistry. - : AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC. - 0021-9258 .- 1083-351X. ; 291:48, s. 25066-25076
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Tidskriftsartikel (refereegranskat)abstract
- Glycero-3-phosphocholine (GPC), the product of the complete deacylation of phosphatidylcholine (PC), was long thought to not be a substrate for reacylation. However, it was recently shown that cell-free extracts from yeast and plants could acylate GPC with acyl groups from acyl-CoA. By screening enzyme activities of extracts derived from a yeast knock-out collection, we were able to identify and clone the yeast gene (GPC1) encoding the enzyme, named glycerophosphocholine acyltransferase (GPCAT). By homology search, we also identified and cloned GPCAT genes from three plant species. All enzymes utilize acyl-CoA to acylate GPC, forming lyso-PC, and they show broad acyl specificities in both yeast and plants. In addition to acyl-CoA, GPCAT efficiently utilizes LPC and lysophosphatidylethanolamine as acyl donors in the acylation of GPC. GPCAT homologues were found in the major eukaryotic organism groups but not in prokaryotes or chordates. The enzyme forms its own protein family and does not contain any of the acyl binding or lipase motifs that are present in other studied acyltransferases and transacylases. In vivo labeling studies confirm a role for Gpc1p in PC biosynthesis in yeast. It is postulated that GPCATs contribute to the maintenance of PC homeostasis and also have specific functions in acyl editing of PC (e.g. in transferring acyl groups modified at the sn-2 position of PC to the sn-1 position of this molecule in plant cells).
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| 26. |
- Gomez, Federico, et al.
(författare)
-
Metabolomic evaluation of pulsed electric field-induced stress on potato tissue
- 2009
-
Ingår i: Planta. - : Springer Science and Business Media LLC. - 0032-0935 .- 1432-2048. ; 230:3, s. 469-479
-
Tidskriftsartikel (refereegranskat)abstract
- Metabolite profiling was used to characterize stress responses of potato tissue subjected to reversible electroporation, providing insights on how potato tissue responds to a physical stimulus such as pulsed electric fields (PEF), which is an artificial stress. Wounded potato tissue was subjected to field strengths ranging from 200 to 400 V/cm, with a single rectangular pulse of 1 ms. Electroporation was demonstrated by propidium iodide staining of the cell nucleae. Metabolic profiling of data obtained through GC/TOF-MS and UPLC/TOF-MS complemented with orthogonal projections to latent structures clustering analysis showed that 24 h after the application of PEF, potato metabolism shows PEF-specific responses characterized by the changes in the hexose pool that may involve starch and ascorbic acid degradation.
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| 27. |
- Gomez, Federico, et al.
(författare)
-
Plant Stress Physiology: Opportunities and Challenges for the Food Industry.
- 2007
-
Ingår i: Critical Reviews in Food Science and Nutrition. - : Informa UK Limited. - 1040-8398 .- 1549-7852. ; 47:8, s. 749-763
-
Forskningsöversikt (refereegranskat)abstract
- We review and analyze the possible advantages and disadvantages of plant-stress-related metabolic and structural changes on applications in the fruit and vegetable processing industry. Knowledge of the cellular and tissue transformations that result from environmental conditions or industrial manipulation is a powerful means for food engineers to gain a better understanding of biological systems in order to avoid potential side effects. Our aim is to provide an overview of the understanding and implementation of physiological and biochemical principles in the industrial processing of fruits and vegetables.
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| 28. |
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| 29. |
- Hao, Meng-Shu, et al.
(författare)
-
The Ca2+-Regulation of the Mitochondrial External NADPH Dehydrogenase in Plants Is Controlled by Cytosolic pH
- 2015
-
Ingår i: PLOS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 10:9
-
Tidskriftsartikel (refereegranskat)abstract
- NADPH is a key reductant carrier that maintains internal redox and antioxidant status, and that links biosynthetic, catabolic and signalling pathways. Plants have a mitochondrial external NADPH oxidation pathway, which depends on Ca2+ and pH in vitro, but concentrations of Ca2+ needed are not known. We have determined the K-0.5(Ca2+) of the external NADPH dehydrogenase from Solanum tuberosum mitochondria and membranes of E. coli expressing Arabidopsis thaliana NDB1 over the physiological pH range using O-2 and decylubiquinone as electron acceptors. The K-0.5(Ca2+) of NADPH oxidation was generally higher than for NADH oxidation, and unlike the latter, it depended on pH. At pH 7.5, K-0.5(Ca2+) for NADPH oxidation was high (approximate to 100 mu M), yet 20-fold lower K-0.5(Ca2+) values were determined at pH 6.8. Lower K-0.5(Ca2+) values were observed with decylubiquinone than with O-2 as terminal electron acceptor. NADPH oxidation responded to changes in Ca2+ concentrations more rapidly than NADH oxidation did. Thus, cytosolic acidification is an important activator of external NADPH oxidation, by decreasing the Ca2+-requirements for NDB1. The results are discussed in relation to the present knowledge on how whole cell NADPH redox homeostasis is affected in plants modified for the NDB1 gene.
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| 30. |
- Hao, Mengshu, et al.
(författare)
-
The evolution of substrate specificity-associated residues and Ca2+-binding motifs in EF-hand-containing type II NAD(P)H dehydrogenases
- 2016
-
Ingår i: Physiologia Plantarum. - : WILEY. - 0031-9317 .- 1399-3054. ; 157:3, s. 338-351
-
Tidskriftsartikel (refereegranskat)abstract
- Most eukaryotic organisms, except some animal clades, have mitochondrial alternative electron transport enzymes that allow respiration to bypass the energy coupling in oxidative phosphorylation. The energy bypass enzymes in plants include the external type II NAD(P)H dehydrogenases (DHs) of the NDB family, which are characterized by an EF-hand domain for Ca2+ binding. Here we investigate these plant enzymes by combining molecular modeling with evolutionary analysis. Molecular modeling of the Arabidopsis thalianaAtNDB1 with the yeast ScNDI1 as template revealed distinct similarities in the core catalytic parts, and highlighted the interaction between the pyridine nucleotide and residues correlating with NAD(P)H substrate specificity. The EF-hand domain of AtNDB1 has no counterpart in ScNDI1, and was instead modeled with Ca2+-binding signal transducer proteins. Combined models displayed a proximity of the AtNDB1 EF-hand domain to the substrate entrance side of the catalytic part. Evolutionary analysis of the eukaryotic NDB-type proteins revealed ancient and recent reversions between the motif observed in proteins specific for NADH (acidic type) and NADPH (non-acidic type), and that the clade of enzymes with acidic motifs in angiosperms derives from non-acidic-motif NDB-type proteins present in basal plants, fungi and protists. The results suggest that Ca2+-dependent external NADPH oxidation is an ancient process, indicating that it has a fundamental importance for eukaryotic cellular redox metabolism. In contrast, the external NADH DHs in plants are products of a recent expansion, mirroring the expansion of the alternative oxidase family.
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| 31. |
- Heiser, Volker, et al.
(författare)
-
Antisense repression of the mitochondrial nadh-binding subunit of complex I in transgenic potato plants affects male fertility
- 1997
-
Ingår i: Plant Science. - 0168-9452. ; 127:1, s. 61-69
-
Tidskriftsartikel (refereegranskat)abstract
- Mitochondrial respiratory chain complex I is a large multi-subunit enzyme composed of both organellar and nuclear encoded proteins. To investigate the role of the nuclear encoded components, expression of the gene for the 55 kDa NADH-binding subunit of complex I was disturbed by antisense repression in transgenic potato plants. The antisense construct driven by the CaMV 35S promoter decreases the steady-state mRNA levels of transcripts for the 55 kDa subunit to 33% of the wild type levels. Quantities of the 55 kDa protein in mitochondrial protein extracts are lowered to about 50% in these plants. Transgenic plants show normal vegetative growth and tuber formation, but pollen maturation is found to be disturbed. The reduced male fertility of the transgenic 55 kDa antisense plants may be caused by an insufficient mitochondrial respiratory chain, impaired by the decreased expression of the NADH-binding component of mitochondrial complex I.
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| 32. |
- Johansson, Fredrik I, et al.
(författare)
-
Oxidation and reduction of pyridine nucleotides in alamethicin-permeabilized plant mitochondria
- 2004
-
Ingår i: Biochemical Journal. - 0264-6021. ; 380:1, s. 193-202
-
Tidskriftsartikel (refereegranskat)abstract
- The inner mitochondrial membrane is selectively permeable, which limits the transport of solutes and metabolites across the membrane. This constitutes a problem when intramitochondrial enzymes are studied. The channel-forming antibiotic AlaM (alamethicin) was used as a potentially less invasive method to permearbilize mitochondria and study the highly branched electron-transport chain in potato tuber (Solanum tuberosum) and pea leaf (Pisum sativum) mitochondria. We show that AlaM permeabilized the inner membrane of plant mitochondria to NAD(P)H, allowing the quantification of internal NAD(P)H dehydrogenarses as well as matrix enzymes in situ. AlaM was found to inhibit the electron-tran sport chain at the external Ca2+-dependent rotenone-insensitive NADH dehydrogenase and around complexes III and IV. Nevertheless, under optimal conditions, especially complex I-mediated NADH oxidation in AlaM-treated mitochondria was much higher than what has been previously measured by other techniques. Our results also show a difference in substrate specificities for complex I in mitochondria as compared with inside-out submitochondrial particles. AlaM facilitated the passage of cofactors to and from the mitochondrial matrix and allowed the determination of NAD(+) requirements of malate oxidation in situ. In summary, we conclude that AlaM provides the best method for quantifying NADH dehydrogenase activities and that AlaM will prove to be an important method to study enzymes under conditions that resemble their native environment not only in plant mitochondria but also in other membrane-enclosed compartments, such as intact cells, chloroplasts and peroxisomes.
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| 33. |
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| 34. |
- Kjell, Jonas, et al.
(författare)
-
Protein complexes of the plant plasma membrane resolved by Blue Native PAGE
- 2004
-
Ingår i: Physiologia Plantarum. - : Wiley. - 0031-9317 .- 1399-3054. ; 121:4, s. 546-555
-
Tidskriftsartikel (refereegranskat)abstract
- With the characterization of the total genomes of Arabidopsis thaliana and Oryza sativa, several putative plasma membrane components have been identified. However, a lack of knowledge at the protein level, especially for hydrophobic proteins, have hampered analyses of physiological changes. To address whether protein complexes may be present in the native membrane, we subjected plasma membranes isolated from Spinacia oleracea leaves to blue-native polyacrylamide gel electrophoresis (BN-PAGE). BN-PAGE is well established in the separation of functional membrane protein complexes from mitochondria and chloroplasts, but a resolved protein complex pattern from PM of eukaryotic cells has previously not been reported. Using this method, protein complexes from Spinacia oleracea PM could be efficiently solubilized and separated, including the highly hydrophobic aquaporin (apparent molecular mass 230 kDa), a putative tetramer of H+-ATPase, and several less abundant complexes with apparent masses around or above 750 kDa. After denaturation and separation of the complexes into their subunits in a second dimension (SDS-PAGE), several of the complexes were identified as hydrophobic membrane proteins. Large amounts of protein (up to 1 mg) can be resolved in each lane, which suggests that the method could be used to study also low-abundance protein complexes, e.g. under different physiological conditions.
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| 35. |
- Kwao, Stephen, et al.
(författare)
-
Effect of guard cells electroporation on drying kinetics and aroma compounds of Genovese basil (Ocimum basilicum L.) leaves
- 2016
-
Ingår i: Innovative Food Science and Emerging Technologies. - : Elsevier BV. - 1466-8564. ; 38, s. 15-23
-
Tidskriftsartikel (refereegranskat)abstract
- In this study the effect of the application of pulsed electric fields as pre-treatment before air drying of basil leaves at 50 °C was investigated. The parameters of the electric treatment were designed in such a way that they either (i) electroporated the tissue reversibly (tissue remains viable) and without electroporating the guard cells in the stomata complex (65 pulses of 600 V/cm, 120 μs pulse width, 500 μs between pulses), (ii) electroporated the tissue reversibly causing the guard cells to irreversibly open the stomata (65 pulses of 600 V/cm, 150 μs pulse width, 760 μs between pulses), or (iii) irreversibly electroporated the tissue leading to cell death (65 pulses of 1500 V/cm, 150 μs pulse width, 760 μs between pulses). Results demonstrate that dehydration of basil leaves at 50 °C can be aided by the application of pulsed electric fields (PEF). If the applied PEF conditions are such that stomata are irreversibly opened while the rest of the tissue remains viable, the drying process is faster and the product keeps a better colour, is richer in aroma compounds and has better rehydration capacity than the untreated control. Irreversible tissue damage provoked by high intensity PEF drastically decreased the drying time but the measured quality characteristics of the dried leaves were inferior compared to those of the dried product when the stomatal function of the fresh leaf was selectively targeted. Industrial relevance This paper addresses the ever-increasing global demand from consumers for high-quality foods, reporting a novel methodology which improves the quality of dried aromatic herbs using pulsed electric fields.
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| 36. |
- Lager, Ida, et al.
(författare)
-
Changes in external pH rapidly alter plant gene expression and modulate auxin and elicitor responses
- 2010
-
Ingår i: Plant, Cell and Environment. - : Wiley. - 0140-7791 .- 1365-3040. ; 33:9, s. 1513-1528
-
Tidskriftsartikel (refereegranskat)abstract
- pH is a highly variable environmental factor for the root, and plant cells can modify apoplastic pH for nutrient acquisition and in response to extracellular signals. Nevertheless, surprisingly few effects of external pH on plant gene expression have been reported. We have used microarrays to investigate whether external pH affects global gene expression. In Arabidopsis thaliana roots, 881 genes displayed at least twofold changes in transcript abundance 8 h after shifting medium pH from 6.0 to 4.5, identifying pH as a major affector of global gene expression. Several genes responded within 20 min, and gene responses were also observed in leaves of seedling cultures. The pH 4.5 treatment was not associated with abiotic stress, as evaluated from growth and transcriptional response. However, the observed patterns of global gene expression indicated redundancies and interactions between the responses to pH, auxin and pathogen elicitors. In addition, major shifts in gene expression were associated with cell wall modifications and Ca2+ signalling. Correspondingly, a marked overrepresentation of Ca2+/calmodulin-associated motifs was observed in the promoters of pH-responsive genes. This strongly suggests that plant pH recognition involves intracellular Ca2+. Overall, the results emphasize the previously underappreciated role of pH in plant responses to the environment.
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| 37. |
- Liu, Yunjun, et al.
(författare)
-
A Redox-Mediated Modulation of Stem Bolting in Transgenic Nicotiana sylvestris Differentially Expressing the External Mitochondrial NADPH Dehydrogenase.
- 2009
-
Ingår i: Plant Physiology. - : Oxford University Press (OUP). - 1532-2548. ; 150, s. 1248-1259
-
Tidskriftsartikel (refereegranskat)abstract
- Cytosolic NADPH can be directly oxidized by a calcium-dependent NADPH dehydrogenase, NDB1, present in the plant mitochondrial electron transport chain. However, little is known regarding the impact of modified cytosolic NADPH reduction levels on growth and metabolism. Nicotiana sylvestris plants overexpressing potato NDB1 displayed early bolting whereas sense-suppression of the same gene led to delayed bolting, with consequential changes in flowering time. The phenotype was dependent on light irradiance, but not linked to any change in biomass accumulation. Whereas the leaf NADPH/NADP(+)-ratio was unaffected, the stem NADPH/NADP(+)-ratio was altered following the genetic modification and strongly correlated to the bolting phenotype. Metabolic profiling of the stem displayed that the NADP(H) change affected relatively few, albeit central, metabolites, including 2-oxoglutarate, glutamate, ascorbate, sugars and hexose phosphates. Consistent with the phenotype, the modified NDB1 level also affected expression of putative floral meristem identity genes of the SQUAMOSA and LEAFY types. Further evidence for involvement of the NADPH redox in stem development was seen in the distinct decrease in the stem apex NADPH/NADP(+)-ratio during bolting. Additionally, the potato NDB1 protein was specifically detected in mitochondria, and a survey of its abundance in major organs revealed that the highest levels are present in green stems. The results thus strongly suggest that NDB1 in the mitochondrial electron transport chain can, by modifying cell redox levels, specifically affect developmental processes.
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| 38. |
- Liu, Yunjun, et al.
(författare)
-
The mitochondrial external NADPH dehydrogenase modulates the leaf NADPH/NADP+ ratio in transgenic Nicotiana sylvestris
- 2008
-
Ingår i: Plant and Cell Physiology. - : Oxford University Press (OUP). - 1471-9053 .- 0032-0781. ; 49:1, s. 251-263
-
Tidskriftsartikel (refereegranskat)abstract
- Plant mitochondria contain alternative external NAD(P)H dehydrogenases, which oxidise cytosolic NADH or NADPH and reduce ubiquinone without inherent linkage to proton pumping and ATP production. In potato, St-NDB1 is an external Ca2+-dependent NADPH dehydrogenase. The physiological function of this enzyme was investigated in homozygous Nicotiana sylvestris lines overexpressing St-ndb1 and co-suppressing St-ndb1 and an N. sylvestris ndb1. In leaf mitochondria isolated from the overexpressor lines, higher activity of alternative oxidase (AOX) was detected. However, the AOX induction was substantially weaker than in the complex I deficient CMSII mutant, previously shown to contain elevated amounts of NAD(P)H dehydrogenases and AOX. An aox1b and an aox2 gene were up-regulated in CMSII, but only aox1b showed a response, albeit smaller, in the transgenic lines, indicating differences in AOX activation between the genotypes. As in CMSII, the increase of AOX in the overexpressing lines was not due to a general oxidative stress. The lines overexpressing St-ndb1 had consistently lowered leaf NADPH/NADP+ ratios in the light and variably decreased levels in darkness, but unchanged NADH/NAD+ ratios. CMSII instead had similar NADPH/NADP+ and lower NADH/NAD+ ratios than wildtype. These results demonstrate that St-NDB1 is able to modulate the cellular balance of NADPH and NADP+ at least in the day and that reduction of NADP(H) and NAD(H) is independently controlled. Similar growth rates, chloroplast malate dehydrogenase activation and xanthophyll ratios indicate that the change in reduction does not communicate to the chloroplast, and that the cell tolerates significant changes in NADP(H) reduction without deleterious effects.
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| 39. |
- Matic, Sandra, et al.
(författare)
-
Alamethicin permeabilizes the plasma membrane and mitochondria but not the tonoplast in tobacco (Nicotiana tabacum L. cv Bright Yellow) suspension cells
- 2005
-
Ingår i: Biochemical Journal. - 0264-6021. ; 389:Pt 3, s. 695-704
-
Tidskriftsartikel (refereegranskat)abstract
- The ion channel-forming peptide AlaM (alamethicin) is known to permeabilize isolated mitochondria as well as animal cells. When intact tobacco (Nicotiana tabacum L.) Bright Yellow-2 cells were treated with AlaM, the cells became permeable for low-molecular-mass molecules as shown by induced leakage of NAD(P)(+). After the addition of cofactors and substrates, activities of cytosolic as well as mitochondrial respiratory enzymes could be directly determined inside the permeabilized cells. However, at an AlaM concentration at which the cytoplasmic enzymes were maximally accessible, the vacuole remained intact, as indicated by an unaffected tonoplast proton gradient. Low-flux permeabilization of plasma membranes and mitochondria at moderate AlaM concentrations was reversible and did not affect cell vigour. Higher AlaM concentrations induced cell death. After the addition of catalase that removes the H2O2 necessary for NADH oxidation by apoplastic peroxidases, mitochondrial oxygen consumption could be measured in permeabilized cells. Inhibitor-sensitive oxidation of the respiratory substrates succinate, malate and NADH was observed after the addition of the appropriate coenzymes (ATP, NAD(+)). The capacities of different pathways in the respiratory electron-transport chain could thus be determined directly. We conclude that AlaM permeabilization provides a very useful tool for monitoring metabolic pathways or individual enzymes in their native proteinaccous environment with controlled cofactor concentrations. Possible uses and limitations of this method for plant cell research are discussed.
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| 40. |
- Michalecka, Agnieszka, et al.
(författare)
-
Arabidopsis genes encoding mitochondrial type II NAD(P)H dehydrogenases have different evolutionary orgin and show distinct responses to light.
- 2003
-
Ingår i: Plant Physiology. - : Oxford University Press (OUP). - 1532-2548 .- 0032-0889. ; 133:2, s. 642-652
-
Tidskriftsartikel (refereegranskat)abstract
- In addition to proton-pumping complex I, plant mitochondria contain several type II NAD(P)H dehydrogenases in the electron transport chain. The extra enzymes allow the nonenergy-conserving electron transfer from cytoplasmic and matrix NAD(P)H to ubiquinone. We have investigated the type II NAD(P)H dehydrogenase gene families in Arabidopsis. This model plant contains two and four genes closely related to potato (Solanum tuberosum) genes nda1 and ndb1, respectively. A novel homolog, termed ndc1, with a lower but significant similarity to potato nda1 and ndb1, is also present. All genes are expressed in several organs of the plant. Among the nda genes, expression of nda1, but not nda2, is dependent on light and circadian regulation, suggesting separate roles in photosynthesis-associated and other respiratory NADH oxidation. Genes from all three gene families encode proteins exclusively targeted to mitochondria, as revealed by expression of green fluorescent fusion proteins and by western blotting of fractionated cells. Phylogenetic analysis indicates that ndc1 affiliates with cyanobacterial type II NADH dehydrogenase genes, suggesting that this gene entered the eukaryotic cell via the chloroplast progenitor. The ndc1 should then have been transferred to the nucleus and acquired a signal for mitochondrial targeting of the protein product. Although they are of different origin, the nda, ndb, and ndc genes carry an identical intron position.
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| 41. |
- Michalecka, Agnieszka, et al.
(författare)
-
Identification of a mitochondrial external NADPH dehydrogenase by overexpression in transgenic Nicotiana sylvestris
- 2004
-
Ingår i: Plant Journal. - 1365-313X. ; 37:3, s. 415-425
-
Tidskriftsartikel (refereegranskat)abstract
- The plant respiratory chain contains a complex setup of non-energy conserving NAD(P)H dehydrogenases, the physiological consequences of which are highly unclear. An expression construct for the potato (Solanum tuberosum L., cv. Desiree) ndb1 gene, a homologue of bacterial and fungal type II NAD(P)H dehydrogenases, was introduced into Nicotiana sylvestris. Transgenic lines with high transcript and protein levels for St-NDB1 had up to threefold increased activity of external NADPH dehydrogenase in isolated mitochondria as compared to the wild type (WT). In two lines, the external NADPH dehydrogenase activity was instead 10-fold decreased, indicating that the corresponding N. sylvestris gene had been suppressed. Activities of external and internal rotenone-insensitive NADH dehydrogenases were unchanged in the transgenic lines. The results demonstrate that the St-ndb1 encodes an external dehydrogenase specific for NADPH and dependent on calcium for activity. Transgenic lines overexpressing St-ndb1 had specifically increased protein levels for alternative oxidase and uncoupling protein, as compared to the WT and one co-suppressing line. This indicates cross-talk in the expressional control, or metabolic conditions influencing it, for the different categories of energy-dissipating proteins that bypass oxidative phosphorylation. The potential effects of external NADPH oxidation on other cellular processes are discussed.
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| 42. |
- Møller, Ian M., et al.
(författare)
-
Isolation of Mitochondria
- 2010
-
Ingår i: A Companion to Plant Physiology, 5th edition. ; Online 2010, s. 1-11
-
Bokkapitel (refereegranskat)abstract
- Although it is possible to measure respiration by intact tissues it is often useful to be able to isolate uncontaminated, intact and functional mitochondria. Here, we will consider some of the principles for isolating cell organelles. This discussion is not meant to be exhaustive nor will it be a cookbook with recipes to be followed step-by-step. Useful general references are Douce 1985, Møller et al. 1996, and a number of chapters in Volume 148 of Methods in Enzymology from 1987.
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| 43. |
- Møller, Ian M, et al.
(författare)
-
Isolation of mitochondria
- 2006
-
Ingår i: A companion to Plant Physiology, 4th Ed..
-
Bokkapitel (övrigt vetenskapligt/konstnärligt)
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| 44. |
- Møller, Ian M., et al.
(författare)
-
Isolation of Mitochondria
- 2015
-
Ingår i: Plant Physiology and Development 6e Companion Website.
-
Bokkapitel (refereegranskat)
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| 45. |
- Møller, Ian Max, et al.
(författare)
-
Matrix Redox Physiology Governs the Regulation of Plant Mitochondrial Metabolism through Posttranslational Protein Modifications
- 2020
-
Ingår i: The Plant cell. - : Oxford University Press (OUP). - 1040-4651 .- 1532-298X. ; 32:3, s. 573-594
-
Forskningsöversikt (refereegranskat)abstract
- Mitochondria function as hubs of plant metabolism. Oxidative phosphorylation produces ATP, but it is also a central high-capacity electron sink required by many metabolic pathways that must be flexibly coordinated and integrated. Here, we review the crucial roles of redox-associated posttranslational protein modifications (PTMs) in mitochondrial metabolic regulation. We discuss several major concepts. First, the major redox couples in the mitochondrial matrix (NAD, NADP, thioredoxin, glutathione, and ascorbate) are in kinetic steady state rather than thermodynamic equilibrium. Second, targeted proteomics have produced long lists of proteins potentially regulated by Cys oxidation/thioredoxin, Met-SO formation, phosphorylation, or Lys acetylation, but we currently only understand the functional importance of a few of these PTMs. Some site modifications may represent molecular noise caused by spurious reactions. Third, different PTMs on the same protein or on different proteins in the same metabolic pathway can interact to fine-tune metabolic regulation. Fourth, PTMs take part in the repair of stress-induced damage (e.g., by reducing Met and Cys oxidation products) as well as adjusting metabolic functions in response to environmental variation, such as changes in light irradiance or oxygen availability. Finally, PTMs form a multidimensional regulatory system that provides the speed and flexibility needed for mitochondrial coordination far beyond that provided by changes in nuclear gene expression alone.
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| 46. |
- Møller, Ian M., et al.
(författare)
-
NAD(P)H-ubiquinone oxidoreductases in plant mitochondria
- 1993
-
Ingår i: Journal of Bioenergetics and Biomembranes. - 0145-479X. ; 25:4, s. 377-384
-
Tidskriftsartikel (refereegranskat)abstract
- Plant (and fungal) mitochondria contain multiple NAD(P)H dehydrogenases in the inner membrane all of which are connected to the respiratory chain via ubiquinone. On the outer surface, facing the intermembrane space and the cytoplasm, NADH and NADPH are oxidized by what is probably a single low-molecular-weight, nonproton-pumping, unspecific rotenone-insensitive NAD(P)H dehydrogenase. Exogenous NADH oxidation is completely dependent on the presence of free Ca2+ with a K0.5 of about 1 μM. On the inner surface facing the matrix there are two dehydrogenases: (1) the proton-pumping rotenone-sensitive multisubunit Complex I with properties similar to those of Complex I in mammalian and fungal mitochondria. (2) a rotenone-insensitive NAD(P)H dehydrogenase with equal activity with NADH and NADPH and no proton-pumping activity. The NADPH-oxidizing activity of this enzyme is completely dependent on Ca2+ with a K0.5 of 3 μM. The enzyme consists of a single subunit of 26 kDa and has a native size of 76 kDa, which means that it may form a trimer.
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| 47. |
- Møller, Ian Max, et al.
(författare)
-
Plant mitochondria – past, present and future
- 2021
-
Ingår i: Plant Journal. - : Wiley. - 0960-7412 .- 1365-313X. ; 108:4, s. 912-959
-
Tidskriftsartikel (refereegranskat)abstract
- The study of plant mitochondria started in earnest around 1950 with the first isolations of mitochondria from animal and plant tissues. The first 35 years were spent establishing the basic properties of plant mitochondria and plant respiration using biochemical and physiological approaches. A number of unique properties (compared to mammalian mitochondria) were observed: (i) the ability to oxidize malate, glycine and cytosolic NAD(P)H at high rates; (ii) the partial insensitivity to rotenone, which turned out to be due to the presence of a second NADH dehydrogenase on the inner surface of the inner mitochondrial membrane in addition to the classical Complex I NADH dehydrogenase; and (iii) the partial insensitivity to cyanide, which turned out to be due to an alternative oxidase, which is also located on the inner surface of the inner mitochondrial membrane, in addition to the classical Complex IV, cytochrome oxidase. With the appearance of molecular biology methods around 1985, followed by genomics, further unique properties were discovered: (iv) plant mitochondrial DNA (mtDNA) is 10–600 times larger than the mammalian mtDNA, yet it only contains approximately 50% more genes; (v) plant mtDNA has kept the standard genetic code, and it has a low divergence rate with respect to point mutations, but a high recombinatorial activity; (vi) mitochondrial mRNA maturation includes a uniquely complex set of activities for processing, splicing and editing (at hundreds of sites); (vii) recombination in mtDNA creates novel reading frames that can produce male sterility; and (viii) plant mitochondria have a large proteome with 2000–3000 different proteins containing many unique proteins such as 200–300 pentatricopeptide repeat proteins. We describe the present and fairly detailed picture of the structure and function of plant mitochondria and how the unique properties make their metabolism more flexible allowing them to be involved in many diverse processes in the plant cell, such as photosynthesis, photorespiration, CAM and C4 metabolism, heat production, temperature control, stress resistance mechanisms, programmed cell death and genomic evolution. However, it is still a challenge to understand how the regulation of metabolism and mtDNA expression works at the cellular level and how retrograde signaling from the mitochondria coordinates all those processes.
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| 48. |
|
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| 49. |
- Møller, Ian M., et al.
(författare)
-
The role of NADP in the mitochondrial matrix
- 1998
-
Ingår i: Trends in Plant Science. - 1360-1385. ; 3:1, s. 21-27
-
Tidskriftsartikel (refereegranskat)abstract
- Many diverse metabolic processes are coupled to the turnover of the coenzyme NADP in the matrix of plant mitochondria. NADPH can be produced via the NADP-specific isocitrate dehydrogenase as well as via enzymes like NAD-malic enzyme, NAD-malate dehydrogenase and Δ1-pyrroline-5-carboxylate dehydrogenase. Although not NADP-specific, the latter enzymes can all catalyse the reduction of NADP+ at appreciable rates. The NADPH produced can be used in folate metabolism, by glutathione reductase for protection against oxidative damage, and by thioredoxin reductase in the (putative) regulation of metabolic pathways via thiol-group reduction. It can also be oxidized by the respiratory chain via a Ca2+-dependent NADPH dehydrogenase - this is a potential way of regulating the NADP reduction level in the matrix and thus, indirectly, the other processes. It is now possible to present an integrated picture of NADP turnover inside the mitochondrion.
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| 50. |
- Møller, Ian M., et al.
(författare)
-
Ubiquinone-1 induces external deamino-NADH oxidation in potato tuber mitochondria
- 1996
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Ingår i: Plant Physiology. - 0032-0889. ; 112:1, s. 75-78
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Tidskriftsartikel (refereegranskat)abstract
- The addition of ubiquinone-1 (UQ-1) induced Ca2+-independent oxidation of deamino-NADH and NADH by intact potato (Solanum tuberosum L. cv Bintje) tuber mitochondria. The induced oxidation was coupled to the generation of a membrane potential. Measurements of NAD+-malate dehydrogenase activity indicated that the permeability of the inner mitochondrial membrane to NADH and deamino-NADH was not altered by the addition of UQ-1. We conclude that UQ-1-induced external deamino-NADH oxidation is due to a change in specificity of the external rotenone-insensitive NADH dehydrogenase. The addition of UQ-1 also induced rotenone-insensitive oxidation of deamino-NADH by inside-out submitochondrial particles, but whether this was due to a change in the specificity of the internal rotenone-insensitive NAD(P)H dehydrogenase or to a bypass in complex I could not be determined.
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