Papers by Ralf Kaldenhoff
Plant Physiology, Aug 1, 1997
Signaling and communication in plants, 2017
Aquaporins in plants show more abundant and greater diversity than aquaporins in bacteria and ani... more Aquaporins in plants show more abundant and greater diversity than aquaporins in bacteria and animals. This unique characteristic provided versatile tool boxes for plants, dealing with environmental changes, which overcome the disadvantage of immobility. Aquaporins were first known for their function as water channel proteins. Later on, more and more studies showed that other small solutes, i.e., ammonia, glycerol, urea, hydrogen peroxide and metalloids, can also pass through the channel of various aquaporins. Moreover, the function of aquaporins as CO2 gas channels was studied by several groups (Nakhoul et al. Am J Physiol Cell Physiol 43(2):C543–C548, 1998; Yang et al. J Biol Chem 275(4):2686–2692, 2000; Tholen and Zhu Plant Physiol 156(1):90–105, 2011). In parallel, studies on model reconstituted membranes claim that no such type of channel would be needed due to the high permeability of those model membranes (Missner et al. Proc Natl Acad Sci USA 105(52):E123, 2008a; J Biol Chem 283(37):25340–25347, 2008b). However, experimental data showed the physiological significance of CO2-conducting channels, particularly in plants. It is generally accepted that plant science presented the first evidence for the physiological relevance and importance of aquaporins as CO2 transport facilitators (Boron Exp Physiol 95(12):1107–1130, 2010; Terashima and Ono Plant Cell Physiol 43(1):70–78, 2002; Uehlein et al. Nature 425 (6959):734–737, 2003; Heckwolf et al. Plant J 67 (5):795–804, 2011; Uehlein et al. Plant Cell 20(3):648–657, 2008). In this chapter, we discuss the CO2 diffusion across membranes and the role of plant aquaporins during this process.
Journal of Advances in Biology & Biotechnology
Photosynthetic Cyanobacteria can be used as a chassis for different synthetic biology approaches.... more Photosynthetic Cyanobacteria can be used as a chassis for different synthetic biology approaches. However, quantitative comparison of tools for engineering, such as those for heterologous gene expression, is often not available. Here, we report the comparative quantification of heterologous protein production in Synechococcussp.PCC 7002 regarding protein expression cassettes and locations of foreign gene integration using sf-GFP as a reporter. We used promoter cpc560 as reference because it was described as a "super strong" promoter. sf-GFP-expression constructs were integrated into neutral sites NS_1, NS_2, NS_3 and the extrachromosomal plasmid pAQ1. The latter induced a sf-GFP level of approximately 10-fold in comparison to a reference promotor expression. Protein-fusion with 6xHis increased sf-GFP as well as expression of sf-GFP fusion with ß subunit of phycocyanin.
<b>Copyright information:</b>Taken from "The aquaporins"Genome Biology 2006... more <b>Copyright information:</b>Taken from "The aquaporins"Genome Biology 2006;7(2):206-206.Published online 28 Feb 2006PMCID:PMC1431727.Copyright © 2006 BioMed Central Ltd The structure shows six tilted membrane-spanning helices (I-VI) and two pore-forming domains made up of two short α helices entering the membrane from the extracellular and intracellular surfaces (arrows). The two NPA boxes are indicated in green. Amino- and carboxy-terminal domains are oriented to the cytoplasmic side of the membrane. The figure was generated using MODELLER7v7 and Swiss-Pdb Viewer.
Abstract: NOVELTY - Identifying feature-specific genes (A) that in a donor plant (D), provide a p... more Abstract: NOVELTY - Identifying feature-specific genes (A) that in a donor plant (D), provide a predetermined feature comprises: (a) preparing genetic donor material from (D); (b) preparing genetic receptor material from receptor plant (R) lacking the feature; (c) heterologous fusion of the two to form hybrid material (H); (d) culturing H and identifying model plants with the feature; and (e) genetic analysis to identify those genes that are part of (D), as (A). USE - The method is useful for identifying feature-specific genes (A) that in a donor plant (D), provide a predetermined feature (claimed). The method is used to identify feature-specific genes and to generate transgenic or chimeric plants with preselected features for use in forestry, agriculture, environmental protection, pharmaceuticals and production of active compounds. A particular feature is susceptibility to colonization by mycorrhizal fungi, which provide nutrients, trace elements and water, and increase resistance ...
Agronomy
Modern agriculture demands for comprehensive information about the plants themselves. Conventiona... more Modern agriculture demands for comprehensive information about the plants themselves. Conventional chemistry-based analytical methods—due to their low throughput and high associated costs—are no longer capable of providing these data. In recent years, remote reflectance-based characterisation has become one of the most promising solutions for rapid assessments of plant attributes. However, in many cases, expensive equipment is required because accurate quantifications need assessments of the full reflectance spectrum. In this experimental study, we examined the versatility of visible spectral sensors as alternative reflectance measuring devices for biological/biochemical quantifications of sweet basil (Ocimum basilicum). Our results confirm the applicability and scope of visible spectral sensors for analysis and quantification of important plant properties, in particular the contents of valuable substances, such as phenolic compounds and flavonoids.
Microbial Transport Systems
PLoS ONE, 2010
The continuous progress in the structural and functional characterization of aquaporins increasin... more The continuous progress in the structural and functional characterization of aquaporins increasingly attracts attention to study their roles in certain mammalian diseases. Although several structures of aquaporins have already been solved by crystallization, the challenge of producing sufficient amounts of functional proteins still remains. CF (cell free) expression has emerged in recent times as a promising alternative option in order to synthesize large quantities of membrane proteins, and the focus of this report was to evaluate the potential of this technique for the production of eukaryotic aquaporins. We have selected the mouse aquaporin 4 as a representative of mammalian aquaporins. The protein was synthesized in an E. coli extract based cell-free system with two different expression modes, and the efficiencies of two modes were compared. In both, the P-CF (cell-free membrane protein expression as precipitate) mode generating initial aquaporin precipitates as well as in the D-CF (cell-free membrane protein expression in presence of detergent) mode, generating directly detergent solubilized samples, we were able to obtain mg amounts of protein per ml of cell-free reaction. Purified aquaporin samples solubilized in different detergents were reconstituted into liposomes, and analyzed for the water channel activity. The calculated P f value of proteoliposome samples isolated from the D-CF mode was 133 mm/s at 10uC, which was 5 times higher as that of the control. A reversible inhibitory effect of mercury chloride was observed, which is consistent with previous observations of in vitro reconstituted aquaporin 4. In this study, a fast and convenient protocol was established for functional expression of aquaporins, which could serve as basis for further applications such as water filtration.
Plant Physiology, 2006
Strawberry (Fragaria × ananassa) fruit accumulate (hydroxy)cinnamoyl glucose (Glc) esters, which ... more Strawberry (Fragaria × ananassa) fruit accumulate (hydroxy)cinnamoyl glucose (Glc) esters, which may serve as the biogenetic precursors of diverse secondary metabolites, such as the flavor constituents methyl cinnamate and ethyl cinnamate. Here, we report on the isolation of a cDNA encoding a UDP-Glc:cinnamate glucosyltransferase (Fragaria × ananassa glucosyltransferase 2 [FaGT2]) from ripe strawberry cv Elsanta that catalyzes the formation of 1-O-acyl-Glc esters of cinnamic acid, benzoic acid, and their derivatives in vitro. Quantitative real-time PCR analysis indicated that FaGT2 transcripts accumulate to high levels during strawberry fruit ripening and to lower levels in flowers. The levels in fruits positively correlated with the in planta concentration of cinnamoyl, p-coumaroyl, and caffeoyl Glc. In the leaf, high amounts of Glc esters were detected, but FaGT2 mRNA was not observed. The expression of FaGT2 is negatively regulated by auxin, induced by oxidative stress, and by hy...
The Plant Cell, 2002
Leaf-moving organs, remarkable for the rhythmic volume changes of their motor cells, served as a ... more Leaf-moving organs, remarkable for the rhythmic volume changes of their motor cells, served as a model system in which to study the regulation of membrane water fluxes. Two plasma membrane intrinsic protein homolog genes, SsAQP1 and SsAQP2, were cloned from these organs and characterized as aquaporins in Xenopu s laevis oocytes. Osmotic water permeability (P f) was 10 times higher in SsAQP2-expressing oocytes than in SsAQP1-expressing oocytes. SsAQP1 was found to be glycerol permeable, and SsAQP2 was inhibited by 0.5 mM HgCl 2 and by 1 mM phloretin. The aquaporin mRNA levels differed in their spatial distribution in the leaf and were regulated diurnally in phase with leaflet movements. Additionally, SsAQP2 transcription was under circadian control. The P f of motor cell protoplasts was regulated diurnally as well: the morning and/or evening P f increases were inhibited by 50 M HgCl 2 , by 2 mM cycloheximide, and by 250 M phloretin to the noon P f level. Our results link SsAQP2 to the physiological function of rhythmic cell volume changes.
Genome Biology, Feb 28, 2006
Water is the major component of all living cells, and efficient regulation of water homeostasis i... more Water is the major component of all living cells, and efficient regulation of water homeostasis is essential for many biological processes. The mechanism by which water passes through biological membranes was a matter of debate until the discovery of the aquaporin water channels. Aquaporins are intrinsic membrane proteins characterized by six transmembrane helices that selectively allow water or other small uncharged molecules to pass along the osmotic gradient. In addition, recent observations show that some aquaporins also facilitate the transport of volatile substances, such as carbon dioxide (CO2) and ammonia (NH3), across membranes. Aquaporins usually form tetramers, with each monomer defining a single pore. Aquaporin-related proteins are found in all organisms, from archaea to mammals. In both uni- and multicellular organisms, numerous isoforms have been identified that are differentially expressed and modified by post-translational processes, thus allowing fine-tuned tissue-s...
Plant Aquaporins, 2017
Aquaporins in plants show more abundant and greater diversity than aquaporins in bacteria and ani... more Aquaporins in plants show more abundant and greater diversity than aquaporins in bacteria and animals. This unique characteristic provided versatile tool boxes for plants, dealing with environmental changes, which overcome the disadvantage of immobility. Aquaporins were first known for their function as water channel proteins. Later on, more and more studies showed that other small solutes, i.e., ammonia, glycerol, urea, hydrogen peroxide and metalloids, can also pass through the channel of various aquaporins. Moreover, the function of aquaporins as CO2 gas channels was studied by several groups (Nakhoul et al. Am J Physiol Cell Physiol 43(2):C543–C548, 1998; Yang et al. J Biol Chem 275(4):2686–2692, 2000; Tholen and Zhu Plant Physiol 156(1):90–105, 2011). In parallel, studies on model reconstituted membranes claim that no such type of channel would be needed due to the high permeability of those model membranes (Missner et al. Proc Natl Acad Sci USA 105(52):E123, 2008a; J Biol Chem 283(37):25340–25347, 2008b). However, experimental data showed the physiological significance of CO2-conducting channels, particularly in plants. It is generally accepted that plant science presented the first evidence for the physiological relevance and importance of aquaporins as CO2 transport facilitators (Boron Exp Physiol 95(12):1107–1130, 2010; Terashima and Ono Plant Cell Physiol 43(1):70–78, 2002; Uehlein et al. Nature 425 (6959):734–737, 2003; Heckwolf et al. Plant J 67 (5):795–804, 2011; Uehlein et al. Plant Cell 20(3):648–657, 2008). In this chapter, we discuss the CO2 diffusion across membranes and the role of plant aquaporins during this process.
The possible involvement of the methylglyoxal and proline accumulation in leaves and roots of thr... more The possible involvement of the methylglyoxal and proline accumulation in leaves and roots of three pistachio rootstocks, cv. Sarakha, Abareqi and Bane baghi, pre-inoculated with arbuscular mycorrhizal fungus (Glomus mosseae) in response to salt stress was studied during a greenhouse experiment in 2013. Six months old pistachio seedlings were exposed to four salinity levels of irrigation water (EC of 0.5 as control, 5, 10 and 15 dS m-1) for 70 days. Methylglyoxal and proline of the roots and leaves were increased by increasing salt stress. The highest concentrations of proline in leaves and roots were recorded in Abareqi rootstock while the lowest concentration was observed in Sarakhs. In general, a negative relationship was obtained between proline and methylglyoxal concentrations in both tissues especially at two highest levels of salinity. A very strong relationship between salinity and measured biochemical markers were found. The level of both biomarkers were reduced in both tissues and in all rootstocks as the effect of mycorrhizal symbiosis. Root colonization percentage was declined as the effect of salinity in Abareqi and Bane baghi and not in Sarakhs.
Abstract: NOVELTY - A transgenic crop plant (T1) comprises at least one root cell and at least on... more Abstract: NOVELTY - A transgenic crop plant (T1) comprises at least one root cell and at least one leaf cell transformed with a DNA construct having a polynucleotide encoding Nicotiana tabacum aquaporin-1 (NtAQP1), where the plant has increase yield compared to a corresponding non-transgenic plant. USE - As a transgenic crop plant (where the plant is selected from a plant producing fruit, flower and ornamental plant, grain producing plant (i.e. wheat, oats, barely, rye, rice, maize), legumes (i.e. peanuts, peas soybean lentil), plant producing forage, plant producing fiber (including cotton and flax), a tree for wood industry, plant producing tuber or root crop, sugar beet, sugar came, plant producing oil (i.e. canola, sunflower, sesame) and tomato plant useful for generating tissue culture (claimed). ADVANTAGE - The transgenic crop plant is grown under optimal water availability conditions or abiotic stress conditions selected from water stress (drought) (i.e. water content of less...
Algal Research, 2020
Abstract The unicellular green alga Chlamydomonas reinhardtii has been used as a model organism i... more Abstract The unicellular green alga Chlamydomonas reinhardtii has been used as a model organism in different research fields for the last 70 years. A variety of methods have been developed for its genetic modification, ranging from classical mutagenesis to the introduction, integration, and expression of foreign genes as well as cutting-edge genome editing technology. Nevertheless, the unique biology of this species also presents different challenges and technical issues that prevent the application of its complete potential. Here, we highlight the most relevant achievements in C. reinhardtii genetic engineering as well as latest strategies to overcome the remaining challenges.
Lighting Research & Technology, 2020
To meet the increasing demand for mass-produced microalgae, production processes must be optimise... more To meet the increasing demand for mass-produced microalgae, production processes must be optimised and monitored. The herein described optical sensors provide an instantaneous and direct opportunity to monitor biological and biochemical information for microalgae cultivation. Combination of expertise in natural science and engineering, as well as the application of novel data acquisition and analysis methods, are required to develop an appropriate system for industrial purpose. Prior to this, the correlation between biological and physical parameters must be determined. In this work, we combined colour science with the experimentally determined dataset from the cultivation of Microchloropsis salina. Our results indicate a resilient correlation between algal biomass and its specific pigment concentration with the determined reflection spectrum. Data evaluation allowed us to establish identification models for contactless quantification of the respective biological parameter in large-...
Scientific Reports, 2020
In Chlamydomonas reinhardtii, ketocarotenoid biosynthesis is limited to the diploid zygospore sta... more In Chlamydomonas reinhardtii, ketocarotenoid biosynthesis is limited to the diploid zygospore stage. In this study, we attempted to engineer the ketocarotenoid pathway into Chlamydomonas haploid vegetative green cells by overexpressing the key enzyme ß-carotene ketolase (CrBKT). We chose strain CC-4102 for the approach; competitive pathways, α-carotene biosynthesis and xanthophyll cycle are silenced in this strain. Driven by the strong constitutive HSP70/RBCS2 promoter CrBKT overexpression resulted in the production of canthaxanthin, the ketolation product from ß-carotene as well as a drastic reduction in the chlorophyll concentration. Intriguingly, these phenotypes could only be detected from lines transformed and grown heterotrophically in the dark. Once exposed to light, these transformants lost the aforementioned phenotypes as well as their antibiotic resistance. This phenomenon is in agreement with the fact that we were unable to recover any canthaxanthin-producing line among l...
Cells, 2019
Aquaporins are important and well-studied water channel membrane proteins. However, being membran... more Aquaporins are important and well-studied water channel membrane proteins. However, being membrane proteins, sample preparation for functional analysis is tedious and time-consuming. In this paper, we report a new approach for the co-translational insertion of two aquaporins from Escherichia coli and Nicotiana tabacum using the CFPS system. This was done in the presence of liposomes with a modified procedure to form homogenous proteo-liposomes suitable for functional analysis of water permeability using stopped-flow spectrophotometry. Two model aquaporins, AqpZ and NtPIP2;1, were successfully incorporated into the liposome in their active forms. Shifted green fluorescent protein was fused to the C-terminal part of AqpZ to monitor its insertion and status in the lipid environment. This new fast approach offers a fast and straightforward method for the functional analysis of aquaporins in both prokaryotic and eukaryotic organisms.
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Papers by Ralf Kaldenhoff