Browsing by Subject "Phosphatase"

Now showing 1 - 3 of 3

Funktionelle Charakterisierung der Phosphatase RDGC in Drosophila melanogaster Photorezeptorzellen
(2018) Strauch, Lisa; Huber, Armin
Phosphorylation of important components like rhodopsin and TRP plays a big role in the phototransduction cascade of Drosophila melanogaster. The analyzed phosphatase RDGC is needed for the dephosphorylation of both components. It is yet knwon thet RDGC is expressed in three isoforms which will be named RDGC-S, RDGC-M and RDGC-L. Nothing has been known about the origin of RDGC-M. The present work shows thet RDGC-M is generated by using an alternative translation start codon and an alternative splice site within the RNA of the short isoform. Analysis of the subcellular localization showed membrane assoziation of RDGC-M and -L whereas RDGC-S is found in the soluble fraction. Recominant expression in S2-cells identified acylation of RDGC-M and -L as the source of the membrane association. In addition, acylation of RDGC-L isolated from flies was directly proven by using a biochemical assay. To functionally characterize the three isoforms, mutant flies with different RDGC expression paterns were created and analyzed. As a result, it was shown that rhodopsin hyperphosphorylation that is found in the rdgc nullmutant as well as the associated retinal degeneration is prevented by the expression of any RDGC isoform. Regarding TRP channel phosphorylation none of the three isoforms is mandatory for the dephosphorylation of TRP at Ser936. However, the results revealed thet the total amount of RDGC that is available, in particular RDGC-M, affects the kinetics of the TRP-S936 dephosphorylation. An increased expression of RDGC-M in the absence of RDGC-S leads to a faster dephosphorylation of TRP-S936. Such a change in TRP-S936 dephosphorylation kinetics was not observed in flies overexpressing RDGC-S in an rdgc-nullmutangt backgroundand therefore cannot be attributed to the increased amount of the corresponding protein. Taken together this study shows thet the tgree RDGC isoforms differ in their subcellular localization due to differences in the N-termini. This may be the reason for kinetic differences in the dephosphorylation of TRP-S936 by RDGC-S or RDGC-M. Apart from these findings, all RDGC isoforms are able to dephosphorylate rhodopsin.
Regulation of ammonium transport in Arabidopsis thaliana
(2022) Ganz, Pascal; Ludewig, Uwe
The overarching question of this thesis deals with how plants ensure the selective uptake of ammonium while maintaining ion and pH homeostasis. A key component of this is ammonium transporters (AMTs) with high affinity towards their substrate, which are at the same time part of a multilayered protection system against uncontrolled ammonium influx. Conserved protein sequences inside the transporter were analyzed as well as the regulatory system based on post-translational modification of the transporter.
The functional role of phosphorus-mobilizing bacteria in the rhizosphere of tomato and maize
(2017) Nassal, Dinah; Kandeler, Ellen
Phosphorus (P) is an essential plant nutrient. However, global P reserves are being increasingly exploited and surplus P applied by P fertilization is steadily accumulating in the form of plant-unavailable P compounds in arable soils. Future plant production will therefore require a more effective and sustainable P fertilization regime. One promising approach is the use of phosphorus-mobilizing bacteria (PMB), which are able to mobilize P in soil through mineralization or solubilization so effectively that plant P supply is improved. Increases in plant growth and P uptake by the addition of PMB have been reported several times, but PMB’s functional mechanisms in soils and plants are still poorly understood. However, an understanding of PMB’s functional mechanisms is necessary to evaluate both the potential and limitations of their use as well as to develop practical application recommendations. This thesis aimed to provide a better understanding of PMB’s functional mechanisms in soil; the foci here were mechanisms and interactions of P mineralization with indigenous soil microorganisms. We aimed to identify P mineralization-dependent and -independent as well as direct and indirect mechanisms of PMB on soil and plants. To this end, three rhizobox experiments were performed in the greenhouse using tomato and maize as the test plants and Pseudomonas sp. RU47 (RU47) as the PMB. To identify effective P mineralization beyond the level of endogenous microbial activity, a treatment using unselectively cultivated soil bacteria for inoculation was included. Furthermore, the addition of devitalized RU47 cells provided the opportunity to identify indirect mechanisms. In all three rhizobox experiments the activities of acid and alkaline phosphomonoesterases in rhizosphere and bulk soil were determined, as the latter could be clearly identified as being of microbial origin. Effects on microbial community structure in soil were estimated by denaturing gradient gel electrophoresis (DGGE) and/or phospholipid fatty acid analysis. For deeper investigations of potential effects on microbial population composition and possible dependencies on soil conditions, a fourth experiment was performed using maize, three different Pseudomonas strains possessing PMB abilities, and three different soils varying in parameters which included organic C, pH, and P content. Microbiome shifts in soil were quantitatively determined via quantitative PCR using domain- (bacteria, archaea, fungi) and six bacterial phylum-specific primers. Our experiments showed that tomato plants grown under low P availability soil conditions improved in both growth and P uptake when viable RU47 cells were added. This effect was accompanied by increased alkaline phosphatase activity (PA) in the rhizosphere. We also observed plant growth-promotion effects and a trend of increased PA by the addition of dead RU47 cells. Based on DGGE results, which indicated the promotion of indigenous rhizobacteria, we assume a priming effect induced by the addition of C sources in the form of bacterial residues (dead RU47), which resulted in increased indigenous microbial activity in the rhizosphere. In each rhizobox experiment viable RU47 cells were able to colonize the rhizosphere at high abundances, persisting up to 50 days after sowing. We found indications of phytohormonal influences with the addition of both viable and dead RU47 cells, but this was more pronounced in dead than in viable RU47 treatments. Increasing P availability in soil by mineral P fertilization seemed to improve RU47’s ability to colonize and persist, which was shown by an increased RU47 abundance in both rhizosphere and bulk soils. However, despite an observable slight tendency, strengthened plant growth-promotion that positively correlated with improved RU47 abundance in the rhizosphere could not be detected. In general, colonization by viable RU47 cells did not significantly affect microbial community structure, either in the rhizosphere or in bulk soil. Using three different PMB strains, including RU47, in three contrasting soils, inoculation effects on the microbial community occurred heterogeneously, differing between the strains, soils, and time. Changes at the domain level were due primarily to nutrient availability in the soil, which differed between the soils and over time. Individual shifts in microbial community structure occurred more frequently in the rhizosphere than in bulk soil, but colonizing PMB neither increased bacterial abundance in rhizosphere bacteria, nor displaced copiotrophic rhizobacteria (indicative of C competition).