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Enabling Trustworthy Transition from Centralized to Decentralized Identity Management Systems
No full textAs the digital landscape grows in scale and complexity, identity management has become critical to ensure security and trust. Meanwhile, the growing concerns over privacy, user autonomy, and centralized control have exposed the limitations of traditional identity management systems.
The transition from centralized to decentralized identity management systems is a paradigm shift in how digital identities are issued, verified, and maintained. While centralized systems offer simplicity and control, they often pose risks related to privacy, single points of failure, and limited user involvement. Decentralized identity management addresses these concerns by distributing control among users and trusted entities, enabling more secure, privacy-preserving, and user-centric identity solutions. However, this transition also introduces new challenges, such as establishing trust, especially in the absence of central authorities traditionally responsible for identity assurance.
This dissertation evaluates the critical factors that enable a trustworthy transition from centralized to decentralized identity management systems, focusing on defining and implementing trust through an assurance level approach. It comprehensively analyzes both identity paradigms, outlines the technical and organizational barriers to decentralization, and evaluates existing frameworks and standards. Building on this foundation, the research proposes a novel trust model. The model incorporates contextual parameters and past user behavior to dynamically assess trustworthiness in identity credentials.
To validate the proposed model, a proof-of-concept implementation is developed and tested, demonstrating its potential to enhance trust in decentralized systems. The evaluation highlights the model’s adaptability and effectiveness while maintainng the core principles of decentralization.
Ultimately, this work contributes to the effort of establishing a trustworthy decentralized identity management model. It aims to support the progress in digital ecosystems where trust is not imposed by central authorities but systematically assured through transparent, verifiable, and adaptive mechanisms.2026-08-3
Transgenic fungal expression and characterization of a fluorescently-tagged variant of the Verticillium effector protein TRADEVd, and functional elucidation of its Fusarium homolog TRADEFo, and of another Verticillium effector candidate, LSCE1Vd
No full textVascular wilt fungi have an arsenal of secreted effector proteins, which manipulate the host physiology and immune system for their successful colonization. Among these effectors, Verticillium TRADEVd was previously identified as a key effector candidate responsible for host manipulation and transdifferentiation of bundle sheath cells into novel xylem elements (Subieta et al., 2025).
One of the aims of this study was to explore the expression dynamics, functional relevance and host responses associated with TRADEVd during infection. With the development of fluorescently tagged V. dahliae TRADEVd reporter lines driven by the native TRADEVd and the constitutive fungal gpdA promoters, an in-depth characterization of TRADEVd activity during early-stage colonization was conducted. Host transcript profiling by qRT-PCR during early colonization stages with TRADEVd reporter lines revealed limited induction of vascular reprogramming genes which are typically upregulated in response to TRADEVd activity. This study highlights the critical differences between the native and the constitutive promoter-driven regulation of effector deployment during infection.
Another aim of this study was the functional characterization of a TRADEVd ortholog, TRADEFo, that was identified in the genome of another vascular pathogen, Fusarium oxysporum f. sp. radicis cucumerium (Forc). Comparative analysis revealed its high homology and possible transfer via horizontal gene transfer among these phylogenetically distant fungi. Transgenic expression of these effector proteins in Arabidopsis suggested their conserved function in developmental host plant reprogramming, previously reported in V. dahliae V76 infection (Subieta et al., 2025). While both TRADEVd and TRADEFo trigger host cell identity switches when expressed transgenically in planta, their exogenous application resulted in differential potency to trigger host cell identity switches, possibly due to variable uptake efficiency or their reliance on distinct cellular entry mechanisms.
The final aim of this study was to elucidate the function of another lineage specific effector candidate, LSCE1Vd, in V. dahliae. Through targeted gene knockout and infection experiments, LSCE1Vd was revealed to be essential for core developmental processes in the fungal life cycle. Loss of LSCE1Vd resulted in marked reduction of sporulation and virulence in V. dahliae, underscoring its critical importance in fungal development and pathogenicity. Taken together, our present study provides new advances in molecular, evolutionary and developmental understanding of fungal effector proteins.2026-08-1
The effect of nitrogen and potassium supply on suberization in maize roots
No full textDer radiale Nährstofftransport in Pflanzenwurzeln umfasst die Bewegung von gelösten Stoffen durch mehrere Zellschichten, darunter Rhizodermis, Exodermis, Rinde, Endodermis und das zentrale Leitbündel (Stele). Dieser Transport erfolgt über apoplastische, symplastische und transzelluläre Transportwege. Während Plasmodesmata sowie Nährstofftransporter und -kanäle den symplastischen und transzellulären Transport ermöglichen, wurde der apoplastische Transport, bei dem sich gelöste Stoffe frei im extrazellulären Raum bewegen, lange Zeit vernachlässigt.
Die Suberinlamellen in den Zellwänden der Exo- und Endodermis wirken als apoplastische Barriere und begrenzen aufgrund ihrer hydrophoben Eigenschaften die Nährstoffbewegung. Suberin ist ein komplexes Polymer, das aus aliphatischem und aromatischem Suberin besteht und mithilfe von Glycerin an die primäre Zellwand gebunden ist. Die Zusammensetzung und Menge des Suberins variieren in Abhängigkeit von Pflanzenarten und Umweltbedingungen, was zu Wissenslücken über die Auswirkungen der Suberinisierung auf die Nährstoffaufnahme und den Nährstofftransport führt. Studien mit Arabidopsis-Mutanten deuten darauf hin, dass die suberinisierte Endodermis eine doppelte Funktion erfüllt: Sie reguliert sowohl den Einstrom von Nährstoffen, als auch den unkontrollierten Verlust aus dem Zentralzylinder. Eine frühzeitige Endodermis-Suberinisierung wird mit einer erhöhten Kaliumakkumulation im Spross in Verbindung gebracht. Obwohl über 90% der Bedecktsamer eine suberinisierte Exodermis besitzen, ist ihre genaue Funktion für das Pflanzenwachstum und die Anpassung an Umweltbedingungen bislang wenig erforscht.
Mais stellt aufgrund seiner einzigartigen Wurzelstruktur ein hervorragendes Modell zur Untersuchung der Suberinisierung dar. Im Gegensatz zu vielen anderen Pflanzenarten besitzt Mais sowohl eine Endodermis als auch eine gleichmäßig suberinisierte Exodermis. Gerste hingegen entwickelt unter optimalen Wachstumsbedingungen keine Exodermis, kann aber unter starkem Salzstress eine sogenannte "induzierbare Exodermis" ausbilden. Zwiebelwurzeln weisen eine dimorphe Exodermis auf, die aus langen und kurzen Zellen besteht, wobei sich die Muster der Suberineinlagerung deutlich von denen im Mais unterscheiden. Trotz der weiten Verbreitung der Exodermis und ihrer entscheidenden Rolle bei der Regulation von Nährstoff- und Wasseraufnahme wurde ihre Funktion im Vergleich zur Endodermis bislang deutlich weniger beachtet. Daher wurde in dieser Studie bodengewachsener Mais als Untersuchungsobjekt gewählt, um die Auswirkungen der Stickstoff- und Kaliumverfügbarkeit auf die Suberineinlagerung in den seminalen Wurzeln von Mais zu analysieren. Der Fokus lag dabei auf den spezifischen Beiträgen der Endodermis und Exodermis.
Maiswurzeln setzen unterschiedliche Anpassungsstrategien ein, um auf eine geringe Stickstoffund Kaliumverfügbarkeit zu reagieren. Unter Stickstoffmangel wird die Bildung von Aerenchym als frühe Reaktion priorisiert, noch bevor energieaufwendige Modifikationen der Zellwandsuberinisierung erfolgen. Diese Anpassung senkt die metabolischen Kosten der Wurzeln und verbessert die Effizienz der Nährstoffaufnahme. Im Gegensatz dazu führt Kalium-Mangel in erster Linie zu einer verstärkten Suberinisierung der Endodermis, wodurch die Wurzel ihre Fähigkeit zur Kaliumrückhaltung und -regulation verbessert. Die Reaktion der suberinisierten Exodermis scheint stark vom Schweregrad des Stresses abzuhängen. Entlang der Wurzelachse zeigen die jüngeren Wurzelbereiche die deutlichsten Veränderungen in der Endodermis-Suberinisierung, während die Exodermis-Suberinisierung relativ unempfindlich gegenüber Stickstoff- und Kalium-Mangel erscheint, was weitere Forschung erforderlich macht.
Diese Studie liefert neue Erkenntnisse darüber, wie Maiswurzeln durch Modifikation der Zellwand auf schwankende Nährstoffverfügbarkeiten reagieren. Zudem werden mögliche funktionelle Unterschiede zwischen Endodermis und Exodermis aufgezeigt. Künftige Forschungen sollten sich auf die genetischen und physiologischen Mechanismen des radialen apoplastischen Nährstofftransports konzentrieren. Ein besseres Verständnis dieser Prozesse kann dazu beitragen, die Widerstandsfähigkeit von Kulturpflanzen und deren Nährstoffnutzungseffizienz unter schwierigen Bodenbedingungen zu verbessern.Nutrient radial transport in plant roots involves the movement of solutes across multiple cell layers, including the rhizodermis, exodermis, cortex, endodermis, and stele. This process occurs via apoplastic, symplastic, and transcellular transport pathways. While plasmodesmata and nutrient transporters and/or channels facilitate symplastic and transcellular pathways, apoplastic transport, where solutes move freely through the extracellular space, has historically been overlooked.
The presence of suberin lamellae in the exodermal and endodermal cell walls acts as a critical apoplastic barrier, restricting nutrient movement due to their hydrophobic properties. Suberin, a complex polymer composed of aliphatic suberin and aromatic suberin linked by glycerol to the primary cell walls, exhibits species- and environment-dependent variation in composition and quantity. These variations contribute to knowledge gaps regarding how suberization influences nutrient uptake and transport. The extensive utilization of Arabidopsis and its mutants suggests that the suberized endodermis serves a dual function, regulating both solute influx and preventing uncontrolled leakage from the stele. Notably, early endodermal suberization has been linked to increased potassium accumulation in the shoot. Although over 90% of angiosperms develop a suberized exodermis, its precise role in plant growth and environmental adaptation remains underexplored.
Maize presents a unique model for studying suberization due to its distinct root structure, which includes both an endodermis and a uniformly suberized exodermis. In contrast, barley naturally lacks an exodermis under optimal conditions but can develop an inducible exodermis under severe salinity stress, called "inducible exodermis". Onion roots possess a dimorphic exodermis composed of long and short cells, where suberin deposition patterns differ significantly from those in maize. Despite the widespread presence of the exodermis and its critical role in nutrient and water regulation, its function has received considerably less attention than that of the endodermis. Hence, soil-grown maize was used as the research subject in this thesis to investigate the effects of nitrogen and potassium availability on suberin deposition in maize seminal roots, with a specific focus on the distinct contributions of the endodermis and exodermis.
Maize roots employ distinct adaptive strategies to cope with low N and low K availability. During low nitrogen conditions, aerenchyma formation is prioritized as an early response, occurring before energy-intensive modifications in cell wall suberization. This adaptation reduces root metabolic costs and enhances nutrient foraging efficiency. In contrast, low potassium condition primarily induces increased endodermal suberization, reinforcing the root’s capacity to retain and regulate potassium transport. The response of suberized exodermis appears to be highly dependent on stress severity. Along the root axis, younger root zones exhibit the most pronounced modifications in endodermal suberization. Exodermal suberization appears relatively insensitive to both low nitrogen and low potassium conditions, warranting further research.
This study provides new insights into how maize roots adapt to nutrient availability through regulating cell wall modification and the potential physiological differences between the endodermis and exodermis. Future work should focus on elucidating the genetic and physiological mechanisms governing radial apoplastic nutrient transport in roots, with the aim of improving crop resilience and nutrient-use efficiency in challenging soil conditions.2026-04-2
Fidelity of protein synthesis in vivo
No full textProtein synthesis fidelity is essential for maintaining proteome integrity and cellular fitness. Errors, such as amino acid misincorporations caused by incorrect decoding, result in aberrant proteins that promote proteotoxic stress. Aminoglycoside antibiotics (AGAs) target the ribosome and impair translational fidelity, ultimately disrupting proteostasis. However, key aspects of AGA action – including drug uptake dynamics, the quantitative thresholds of translation errors that trigger specific cellular responses, and the events resulting in proteostasis breakdown – remain poorly understood. Similarly, bacterial resistance mechanisms against AGAs are not fully elucidated.
Using mass spectrometry-based approaches, we investigated AGA action in Escherichia coli. Global proteomic changes upon AGA treatment were quantified using data-independent acquisition (DIA). To quantify ribosomal misreading, we developed a pipeline that combines data-dependent acquisition (DDA) for unbiased detection of missense peptides with targeted mass spectrometry using parallel reaction monitoring (PRM) for validation and quantification. This enabled the identification of over 1,000 unique missense errors across 322 proteins, yielding the most comprehensive AGA-induced misreading profile to date and enabling quantification of rare errors at frequencies as low as 10⁻⁶. To dissect steady-state error levels in full-length proteins shaped by biosynthetic and quality control pathways, we adapted the PUNCH-P protocol to isolate ribosome-nascent chain complexes and enrich nascent peptides for mass spectrometric analysis.
This analysis revealed that AGA uptake kinetics drive dynamic changes in the error landscape and that certain proteins – particularly stress-induced chaperones – show exceptionally high error rates due to synthesis under extreme misreading conditions. Cells respond by activating stress responses and metabolic reprogramming to prioritize survival. At bactericidal AGA concentrations, membrane disruption causes massive drug influx and translational arrest. Error frequencies in nascent chains reach ~10%, while full-length protein synthesis is severely impaired, leading to proteostasis collapse and cell death.
In response to AGA treatment, bacteria preferentially evolve resistance mutations in the fusA gene, encoding elongation factor G. Our collaborative work suggests that fusA mutations selectively silence AGA-corrupted ribosomes, representing the first example of a new resistance strategy. We showed that fusA mutant strains retain low misreading upon AGA exposure, including in the inner membrane proteome. This preserves proteome and membrane integrity at minimal fitness costs, demonstrating that fusA mutations confer resistance early in the AGA uptake cycle.
Overall, this work advances understanding of decoding fidelity under antibiotic pressure, uncovers a resistance mechanism based on selective ribosome silencing, and presents refined proteomic tools to query translational accuracy at scale.2026-09-2
Deriving Forest Structural and Functional Metrics from Airborne and Ground-based Mobile Laser Scanning
No full textDiese Studie untersuchte die Anwendung von LiDAR-basierten Methoden, insbesondere Airborne Laser Scanning (ALS) und bodengestütztem Mobile Laser Scanning (MLS), um Waldstrukturmetriken und ihre ökologischen Funktionen in verschiedenen geografischen Kontexten zu bewerten. Das übergeordnete Ziel war es, die Fähigkeiten dieser Methoden bei der Messung der Waldkomplexität zu bewerten und die Beziehungen zwischen Waldstruktur, Umwelteinflüssen und Ökosystemfunktionen wie der Landoberflächentemperatur zu untersuchen.
Die erste Studie (Kapitel I) konzentrierte sich auf die Bewertung der Vergleichbarkeit von ALS und MLS bei der Ableitung von Waldstrukturmetriken, insbesondere der Box dimension (Db), einem umfassenden Maß für die Waldstrukturkomplexität. Die Studie wurde in einem gemäßigten Buchenwald in Zentraldeutschland durchgeführt, wobei 233 kreisförmige Plots mit einem Durchmesser von 50 Metern verwendet wurden und mehr als 45 Hektar Wald mit beiden ALS- und MLS-Methoden gescannt wurden. Dreizehn Strukturmetriken wurden aus beiden Datensätzen berechnet. Die Ergebnisse zeigten eine moderate bis starke Übereinstimmung zwischen den aus beiden Methoden abgeleiteten Waldstrukturmetriken (mittlerer Pearson-Korrelationskoeffizient ~60%), obwohl die Diskrepanzen durch den Scanwinkel und die Kronenmetriken beeinflusst wurden. ALS war effektiver für Kroneneigenschaften, während MLS feinere Details des Unterwuchses lieferte. Diese Ergebnisse unterstreichen die komplementäre Natur von ALS und MLS und betonen die Notwendigkeit einer sorgfältigen Kalibrierung bei LiDAR-basierten Bewertungen von Waldstrukturmetriken.
In der zweiten Studie (Kapitel II) wurde die Beziehung zwischen Waldstruktur und ökologischer Funktion untersucht, indem ALS-abgeleitete Waldstrukturmetriken mit satellitengestützten Landoberflächentemperatur (LST) in demselben Buchenwald verglichen wurden, in dem die erste Studie durchgeführt wurde. Mittels räumlich ausgerichteter 30 m × 30 m-Plots über eine Fläche von 17 km² wurde festgestellt, dass Wälder mit höherer Strukturkomplexität signifikant niedrigere Oberflächentemperatur hatten. Ein räumliches autoregressives Modell zeigte, dass die Höhe den stärksten Einfluss auf die LST hatte, gefolgt von der Waldstrukturkomplexität, der Neigung und der Ausrichtung. Diese Ergebnisse liefern empirische Beweise dafür, dass strukturkomplexe Wälder eine entscheidende Rolle bei der Regulierung des Mikroklimas spielen, was darauf hindeutet, dass Waldmanagementstrategien, die die Strukturkomplexität erhöhen, zum Klimaschutz beitragen können.
Die dritte Studie (Kapitel III) wandte MLS an, um die Treiber der Waldstrukturkomplexität in den Himalaja-Wäldern des Annapurna-Schutzgebiets in Nepal zu bewerten, wo ALS-Daten nicht verfügbar waren. Insgesamt wurden 69 Plots über Höhen- und Klimagradienten hinweg mittels Hand-MLS gesampelt. Die Studie ergab, dass die Waldstrukturkomplexität signifikant von der Waldhöhe, der Artenvielfalt, der Ausrichtung, dem Boden-pH-Wert und dem Störungsgrad beeinflusst wurde, während Temperatur und Niederschlag keinen messbaren Einfluss hatten. Die Verwendung von MLS ermöglichte die Erfassung detaillierter Daten in anspruchsvollem Gelände und demonstrierte ihren praktischen Wert für die Waldbewertung in abgelegenen Bergregionen.
Zusammen bieten diese Studien eine mehrplattformige Perspektive auf die Anwendung von LiDAR-Technologien zur Bewertung und Interpretation von Waldstruktur und -funktion. Die Ergebnisse unterstreichen das Potenzial von LiDAR, sowohl ALS als auch MLS, als robuste Werkzeuge für die ökologische Überwachung, das Waldmanagement und die klimabezogene Forschung in verschiedenen Waldökosystemen.This study examined the application of LiDAR-based methods, specifically Airborne Laser Scanning (ALS) and ground-based Mobile Laser Scanning (MLS), to evaluate forest structural metrics and their ecological functions across diverse geographic contexts. The overarching aim was to evaluate the capabilities of these methods in measuring forest complexity and to investigate the relationships between forest structure, environmental drivers, and ecosystem functions such as land surface temperature.
The first study (Chapter I) focused on assessing the comparability of ALS and MLS in deriving forest structural metrics, particularly the box dimension (Db), a holistic measure of forest structural complexity. The study was conducted in a temperate beech-dominated forest in central Germany, using 233 circular plots, each with a diameter of 50 meters, and scanning more than 45 hectares of forests using both ALS and handheld MLS methods. Thirteen structural metrics were calculated from both datasets. Results showed moderate to strong agreement between forest structural metrics derived from both methods (mean Pearson correlation ~60%), although discrepancies were influenced by scan angle and canopy metrics. ALS was more effective for canopy-level attributes, while MLS provided finer detail of the understory. These findings highlight the complementary nature of ALS and MLS, emphasizing the need for careful calibration in LiDAR-based assessments of forest structure metrics.
In the second study (Chapter II), the relationship between forest structure and ecological function was examined by linking ALS-derived forest structural metrics with satellite-based land surface temperature (LST) in the same beech-dominated forest where the first study was conducted. Using spatially aligned 30 m × 30 m plots across a 17 km² area, the study found that forests with higher structural complexity had significantly lower surface temperatures. A spatial autoregressive model revealed that elevation had the strongest influence on LST, followed by forest structural complexity, slope, and aspect. These results provide empirical evidence that structurally complex forests play a crucial role in regulating microclimate, suggesting that forest management strategies that enhance structural complexity can contribute to climate change mitigation.
The third study (Chapter III) applied MLS to assess the drivers of forest structural complexity in the Himalayan forests of Nepal’s Annapurna Conservation Area, where ALS data were unavailable. A total of 69 plots were sampled across elevational and climatic gradients using handheld MLS. The study found that forest structural complexity was significantly affected by forest height, species diversity, aspect, soil pH, and disturbance level, whereas temperature and precipitation had no detectable influence. The use of MLS enabled the collection of detailed data in challenging terrain and demonstrated its practical value for forest assessment in remote mountain regions.
Together, these studies provide a multi-platform perspective on how LiDAR technologies can be utilized to assess and interpret forest structure and function. The findings reinforce the potential of LiDAR, both ALS and MLS, as robust tools for ecological monitoring, forest management, and climate-related research across diverse forested landscapes.2026-10-0
The influence of a chronic noise exposure on the degeneration of the organ of Corti of deaf otoferlin knockout mice.
No full textThis study investigated the age-related and noise-associated degeneration of the organ of Corti in otoferlin knockout mice. This knockout causes a complete loss of otoferlin expression in the organ of Corti in mice and thus represents a model for researching mutations in the DFNB9 locus. The phenotype of DFNB9 is often characterized by prelingual, non-syndromic, autosomal recessive hearing loss. The cause is a synaptic disorder of the inner hair cells of the inner ear. Since the sound-amplifying outer hair cells are primarily intact, the disease may remain undetected during functional testing of the outer hair cells as part of newborn hearing screening. In addition to being treated with a power hearing aid or a cochlear implant, genetic repair of the gene locus for the expression of otoferlin by insertion of adeno-associated viruses is currently being tested. To obtain a better understanding of age-related degeneration and noise susceptibility of the inner ear in DFNB9 patients, we not only examined otoferlin knockout mice during aging, but also placed these mice in a self-configured noise setup with chronic sound exposure (40 hours/week) for three months at an intensity of 83 ± 1.5 dB. We hypothesized that deafness could trigger increased susceptibility to noise trauma due to the failure of the body's own protective reflexes, such as the middle ear reflex or the middle olive complex reflex. In addition to in vivo hearing tests to examine hearing function, we used immunofluorescence staining to quantify cell damage. Analysis of early auditory evoked potentials showed that already the non-exposed otoferlin knockout mice were unable to produce signals exceeding the summation potential. We thus confirmed the deafness of these animals. We used the comparison of auditory brainstem responses in exposed and unexposed wild-type animals to assess the severity of noise exposure with regard to the hearing damage caused. The exposed wild-type animals had significantly increased hearing thresholds in many frequency regions and the amplitude of the summation potential and the first wave in the auditory brainstem responses also decreased as a result of exposure. We could thus demonstrate the significant influence of chronic noise exposure on the degeneration of the organ of Corti in wild-type animals. By analyzing cell counts, we were able to detect a more pronounced age-related degeneration of the hair cells required for cochlear amplification, transduction, and transmission in otoferlin knockout mice. At 24 weeks, the number of inner hair cells across the entire length of the basilar membrane was already considerably reduced compared to 8-week-old animals, showing a large difference to wild-type animals, which had much more stable cell counts even at 34 weeks. In addition to the number of inner hair cells being reduced, the average length of these cells was also reduced. This difference was particularly prominent in regions where many hair cells had been lost. By examining the afferent synapses, we were able to show that the number of afferent synapses in otoferlin knockout mice was reduced by between 44 and 65% in the regions examined compared to wild-type animals. Using Imaris-supported analysis of the afferent synapses, we were able to demonstrate an enlargement of the presynapses and postsynapses in older otoferlin knockout mice, as already described in the literature for Vglut3-/- mice. The outer hair cells were also reduced in otoferlin knockout mice as a result of age-related accelerated degeneration, without any age-related degeneration of the efferent synapses. Noise trauma caused more severe hair cell loss in the sensory epithelium of otoferlin knockout mice than in wild-type animals. Analysis of the inner hair cells showed that apical hair cell density in particular was reduced. For the first time, we were able to demonstrate that the outer hair cells in otoferlin knockout mice degenerate even more severely than the inner hair cells as a result of chronic exposure to noise. We concluded that the decrease in otoacoustic emissions frequently observed in DFNB9 patients during treatment with a power hearing aid may be attributed to the pronounced degeneration of the outer hair cells during acoustic stimulation. Surprisingly, the number of afferent synapses in otoferlin knockout mice was very resilient to acoustic overstimulation. In contrast to wild-type animals, we did not observe any tendency toward afferent synapse loss in otoferlin knockout mice. In addition, sound exposure reduced the volume of postsynapses in otoferlin knockout mice. This is interesting since there is none or only minimal exocytosis, suggesting that the size of postsynapses must be regulated independently of glutamate. Furthermore, we observed a larger decrease in efferent synapses as a result of noise exposure in otoferlin knockout mice. In combination with the already increased age-related degeneration of the sensory epithelium, our data suggests that initial treatment with power hearing aids in DFNB9 patients should be critically questioned and long-term chronic sound exposure should be prevented with particular care. In addition, genetic treatment should be carried out as early as possible so that previous degeneration of the sensory epithelium does not severely limit hearing performance.2026-12-0
The arrhythmogenic effect of cardiac troponin C expression knockdown in atrial induced pluripotent stem cell derived cardiomyocytes
No full textVorhofflimmern ist die häufigste Herzrhythmusstörung, trotzdem ist die Pathophysiologie noch unzureichend verstanden. Verfügbare Therapieverfahren haben eine hohe Rezidivquote und die Komplikationen sowohl des Vorhofflimmerns, wie z. B. thromboembolische Ereignisse oder negative Auswirkungen auf die Herzleistung, als auch der begleitenden Therapie, wie z. B. ein erhöhtes Blutungsrisiko, sind nicht zu vernachlässigen.
In atrialen Kardiomyozyten von Patient*innen mit persistierendem Vorhofflimmern konnte eine reduzierte Expression des kardialen Troponin C sowie eine erniedrigte Ca2+-Pufferung nachgewiesen werden. Die zytosolische Ca2+-Pufferung wird wesentlich von Troponin C vermittelt. Da physiologisch etwa 99 % des zytosolischen Ca2+ an Puffer gebunden vorliegt, ist davon auszugehen, dass bereits kleine Veränderungen der Ca2+-Puffer erhebliche Auswirkungen auf die Funktion der Kardiomyozyten haben. Der Fokus dieser Arbeit lag darauf, die Auswirkungen des Verlusts von kardialem Troponin C auf den Ca2+-Haushalt der Kardiomyozyten zu untersuchen und das arrhythmogene Potenzial dieser Veränderungen zu evaluieren.
Dazu wurden mit Hilfe von small interfering Ribonukleinsäuren von induzierten pluripotenten Stammzellen abgeleitete atriale Kardiomyozyten mit reduzierter Troponin-C-Expression modelliert. Der Erfolg des so vermittelten Knockdowns wurde mit der quantitativen Polymerase-Kettenreaktion mit reverser Transkription sowie mit Western-Blots überprüft. Als Kontrolle dienten gleichartige Zellen, die mit nicht-spezifischen small interfering Ribonukleinsäuren transfiziert wurden.
Die modellierten Zellen wurden an einem Patch-Clamp-Messplatz simultan elektrophysiologisch und mit Hilfe von Ca2+-Indikatoren epifluoreszenzmikroskopisch untersucht. Außerdem wurden epifluoreszenzbasierte Messungen der Aktionspotenziale und der diastolischen Ca2+-Leckage des sarkoplasmatischen Retikulums der Zellen sowie Messungen von Ca2+-Sparks mit Hilfe konfokaler Lasermikroskopie durchgeführt. In weiteren Western-Blots wurden die Expression von Junctophilin 2 sowie die Expression und der Phosphorylierungsstatus des Ryanodinrezeptors 2 und der Ca2+-Calmodulin-abhängigen Proteinkinase II untersucht.
Sowohl die Menge der mRNA als auch die Proteinmenge des kardialen Troponin C konnten in den Zellen erfolgreich reduziert werden.
In den Zellen mit reduzierter Troponin-C-Expression zeigte sich eine erhöhte Amplitude des systolischen Ca2+-Transienten sowie des koffeininduzierten Ca2+-Transienten trotz unverändertem Ca2+-Gehalt des sarkoplasmatischen Retikulums. Die Auswertung der aus dem Na+-Ca2+-Austauscher-Strom abgeleiteten Ca2+-Gesamtmenge in Relation zur Menge des freien Ca2+, die dem koffeininduzierten Ca2+-Transienten entspricht, zeigte eine reduzierte Ca2+-Pufferung in den Zellen mit reduzierter Troponin-C-Expression.
In den modellierten Zellen konnten ein Auftreten von Alternanz-Phänomenen des Aktionspotenzials bei niedrigeren Stimulationsfrequenzen sowie eine erhöhte Frequenz diastolischer Ca2+-Sparks beobachtet werden. Beides sind bekannte Mechanismen atrialer Arrhythmien. Die Untersuchungen zur diastolischen Ca2+-Leckage des sarkoplasmatischen Retikulums und die Western-Blots von Ryanodinrezeptor 2, Ca2+-Calmodulin-abhängiger Proteinkinase II und Junctophilin 2 zeigen, dass in den modellierten Kardiomyozyten die reduzierte Ca2+-Pufferung für diese arrhythmogenen Ereignisse verantwortlich ist.
Die Behandlung der Zellen mit dem Ca2+-Sensitizer EMD57033 konnte die Frequenz des Auftretens der Ca2+-Sparks normalisieren, sodass eine Modulation der verbliebenen Ca2+-Puffer ein mögliches therapeutisches Ziel bei der Behandlung von persistierendem Vorhofflimmern sein könnte.
Der Verlust von Troponin C bei Vorhofflimmern scheint Teil langfristiger Remodeling-Prozesse zu sein und könnte so zu einem arrhythmogenen Substrat beitragen, das die Aufrechterhaltung von Vorhofflimmern unterstützt.
Diese Arbeit konnte erstmals zeigen, dass eine reduzierte Troponin-C-Expression eine reduzierte Ca2+-Pufferung bedingt, die u. a. zu einer erhöhten Ca2+-Sparks-Frequenz führt. Diese arrhythmogenen Veränderungen könnten eine Rolle im Rahmen der Entstehung und Aufrechterhaltung von Vorhofflimmern spielen und neue therapeutische Ansätze ermöglichen.Atrial fibrillation is the most frequent cardiac arrhythmia; nevertheless, its pathophysiology is still insufficiently understood. Available therapeutic approaches have high recurrence rates, and complications of both atrial fibrillation itself - such as thromboembolic events - and of the accompanying therapy - such as an increased bleeding risk - must not be neglected.
In atrial cardiomyocytes from patients with persistent atrial fibrillation, reduced expression of cardiac troponin C as well as decreased Ca2+ buffering have been demonstrated. Cytosolic Ca2+ buffering is largely mediated by troponin C. Since physiologically about 99% of cytosolic Ca2+ is bound to buffers, even small changes in Ca2+ buffers are likely to have considerable effects on cardiomyocyte function. The focus of this work was to investigate the effects of the loss of cardiac troponin C on cardiomyocyte Ca2+ homeostasis and to evaluate the arrhythmogenic potential of these changes.
Atrial cardiomyocytes derived from induced pluripotent stem cells were modeled with reduced troponin C expression using small interfering ribonucleic acids. The success of the knockdown was verified by quantitative reverse transcription polymerase chain reaction as well as by Western blots. As a control, identical cells transfected with non-specific small interfering ribonucleic acids were used.
The modeled cells were examined electrophysiologically with patch-clamp and by epifluorescence microscopy using Ca2+ indicators simultaneosly. In addition, epifluorescence-based measurements of action potentials and of diastolic Ca2+ leak from the sarcoplasmic reticulum were performed, as well as measurements of Ca2+ sparks using confocal laser microscopy. Further Western blots assessed the expression of junctophilin 2 as well as the expression and phosphorylation status of ryanodine receptor 2 and Ca2+/calmodulin-dependent protein kinase II.
Both the mRNA amount and the protein amount of cardiac troponin C were successfully reduced in the cells.
In cells with reduced troponin C expression, an increased amplitude of the systolic Ca2+ transient and of the caffeine-induced Ca2+ transient was observed despite an unchanged Ca2+ content of the sarcoplasmic reticulum. Analysis of the total Ca2+ amount derived from the Na+/Ca2+ exchanger current in relation to the amount of free Ca2+ corresponding to the caffeine-induced Ca2+ transient showed reduced Ca2+ buffering in cells with reduced troponin C expression.
In the modeled cells, the occurrence of action potential alternans at lower stimulation frequencies as well as an increased frequency of diastolic Ca2+ sparks could be observed. Both are known mechanisms of atrial arrhythmias. Investigations of diastolic Ca2+ leak from the sarcoplasmic reticulum and the Western blots for ryanodine receptor 2, Ca2+/calmodulin-dependent protein kinase II, and junctophilin 2 indicate that the reduced Ca2+ buffering is responsible for these arrhythmogenic events in the modeled cardiomyocytes.
Treatment of the cells with the Ca2+ sensitizer EMD57033 was able to normalize the frequency of Ca2+ spark occurrence, suggesting that modulation of the remaining Ca2+ buffers could be a possible therapeutic target in the treatment of persistent atrial fibrillation.
Loss of troponin C in atrial fibrillation appears to be part of long-term remodeling processes and could thus contribute to an arrhythmogenic substrate that supports the maintenance of atrial fibrillation.
This work was the first to show that reduced troponin C expression causes reduced Ca2+ buffering, which, among other effects, leads to an increased Ca2+ spark frequency. These arrhythmogenic changes could play a role in the development and maintenance of atrial fibrillation and enable new therapeutic approaches.2026-03-1
ECM recycling as a remodeling mechanism of brain plasticity
No full textAlthough long understudied, the neural extracellular matrix (ECM) has emerged as a key regulator of synaptic function and plasticity in research conducted over the past two decades. This development has led to the expansion of the classical concept of the chemical synapse toward the tetrapartite synapse, which includes not only the presynaptic and postsynaptic terminals but also the surrounding ECM and astrocytes. The neural ECM is composed of extremely long-lived molecules and is therefore regarded as the most stable structure in the brain. As such, it provides ideal conditions for long-term storage of structural information, prompting some authors to refer to it as “template” for long-term memory. However, this well-established view has been challenged by recent findings that suggest synapses undergo constant morphological change. Such continuous structural fluctuations require rapid restructuring processes, which are unlikely to be accomplished by the currently known remodeling mechanisms (proteolysis and de novo synthesis). In addition to high metabolic costs, the long lifespan of ECM molecules is also a major contradiction. This work presents comprehensive evidence for the existence of a novel mechanism based on the recycling of ECM molecules, which may resolve the paradox of an ECM that is both long-lived and adaptable.
Various fluorescence microscopy assays indicate that a dynamic pool of the glycoprotein tenascin-R (TN-R), which is expressed exclusively in the central nervous system, is regularly internalized by a specific neuron population and, after approximately 3 days, returns to the cell surface in an activity-dependent manner, predominantly in the vicinity of synapses. Furthermore, super-resolution microscopy (stimulated emission depletion) has revealed that the recycled TN-R molecules are reglycosylated upon endocytosis. Thus, the comparatively slow TN-R recycling could serve as a repair mechanism for damaged sugar moieties of surface-exposed molecules. Using a blocking labeling assay, it was shown that other long-lived ECM molecules also appear on the cell surface in significantly greater quantities than would be expected based on their exceptionally long half-life. Additionally, the ECM receptor subunit β1 integrin was found to likely play a functional role in the recycling of TN-R.
Future studies will need to further investigate whether the emerging analogy between TN-R and other ECM components is indeed an indication of recycling processes and whether ECM recycling represents a previously unexplored global remodeling mechanism in brain plasticity. Corresponding projects may benefit from an optimized labeling protocol that enables the use of nanoscale secondary ion mass spectrometry (NanoSIMS) to examine the protein turnover of ECM molecules in neuron populations cultivated directly on silicon chips, without requiring additional methods. This technique could prove useful, for example, in tracking the metabolic age of recycling ECM molecules.
A more detailed knowledge of ECM recycling will help to improve our understanding of neurophysiological processes in the brain (e.g., regulation of ECM, neuroplasticity, and homeostasis). Since many diseases of the central nervous system (CNS) are accompanied by changes in ECM, ECM recycling—as a potential modulator of a highly dynamic ECM—would be expected to have considerable neuropathological relevance as well. This hypothesis is supported by the observation that manipulations of TN-R recycling lead to impaired synaptic transmission and spine motility. The pharmacological modulation of ECM recycling and trafficking therefore holds promising therapeutic potential for the treatment of CNS disorders associated with ECM abnormalities.2026-03-1
Investigating the role of BRD4-mediated epigenetic programming in the development of pancreatic ductal adenocarcinoma
No full textPancreatic ductal adenocarcinoma (PDAC) is a highly lethal cancer, with a 5-year survival rate of only 13%. It is projected to become the second leading cause of cancer-related death by 2040. Over 80% of PDACs arise from non-invasive precursor lesions, primarily pancreatic intraepithelial neoplasia (PanIN), through a stepwise accumulation of genetic and epigenetic changes. Activating mutations in KRAS gene, present in over 90% of cases, are recognized as the initiating events in PDAC. However, KRAS alone is inadequate to drive full malignant progression; it requires essential epigenetic and transcription factors to promote PDAC growth and aggressiveness. Among these factors, MYC and the AP-1 transcription factor complex are crucial for mediating KRAS-driven oncogenic programs. Importantly, the epigenetic reader BRD4 plays a significant role by binding to acetylated histones, maintaining high-levels of MYC and AP-1 target gene expression. To investigate how BRD4-mediated epigenetic regulation downstream of mutant KRAS influences PDAC initiation and progression, we performed a pancreas-specific deletion of the epigenetic reader BRD4 in the established KPC (LSL-KrasG12D/+; LSL-Trp53R172H/+; Pdx1-cre) mouse model. Surprisingly, the loss of BRD4 redirected tumorigenesis from the canonical PanIN-PDAC progression route to the intraductal papillary mucinous neoplasm (IPMN) route, leading to invasive PDAC. This finding highlights BRD4 as a critical regulator of pancreatic epithelial lineage under KRAS-driven PDAC malignancy. Histological and transcriptomic analyses revealed that invasive PDACs exhibited hyperactivation of TGF-β1 signaling compared to IPMN lesions in the context of BRD4 loss. Mechanistically, TGF-β1 promoted an epithelial-mesenchymal transition (EMT)-like plasticity and reprogrammed human IPMN cells toward a partially mesenchymal ductal-like state, characterized by co-induction of lineage factors such as SOX9 and aggressive markers like KRT17. Notably, both mouse and human spatial transcriptomic data showed co-upregulation of TGF-β1 and its receptor, TGF-βR2. Pharmacological inhibition or genetic silencing of TGF-βR2 suppressed TGF-β1 expression as well as TGF-β1-mediated EMT plasticity and the invasive phenotype. Together, these findings support a “compensatory overshoot” model, where the loss of BRD4 disrupts the homeostatic regulation of TGF-β1, triggering a self-amplifying TGF-βR2–TGF-β1 autocrine signaling loop that redirects the transition from IPMN to invasive PDAC in KRAS-mediated PDAC models. Our study identifies BRD4 as a gatekeeper of pancreatic epithelial fate, links epigenetic dysregulation to TGF-β1-driven plasticity, and highlights TGF-βR2 as a potential therapeutically actionable target to prevent malignant progression of pre-invasive pancreatic lesions during PDAC development.2026-12-1
Structure and functioning of soil animal communities in beech forests mixed with Douglas-fir, silver fir and spruce
No full textIncreasing temperatures and recurring drought events in the face of climate change are challenging European temperate forests. Notably, large-scale diebacks have been reported in monoculture Norway spruce stands. In this context, more drought-resistant alternative tree species are being promoted and implemented in silvicultural practices of tree species mixtures. Non-native Douglas fir as well as silver fir native to southern Europe are considered suitable options, especially when establishing mixed stands with European beech. However, consequences of implementing non-native Douglas-fir remain relatively unknown, particularly regarding the belowground food web. The soil fauna are a fundamental part of forest ecosystems, linking above-and belowground food webs and providing key ecosystem functions, such as nutrient cycling, nutrient availability and soil formation. Changes in their community composition and functionality are therefore important to understand to infer effects of the implementation of the non-native Douglas-fir into European forest ecosystem on the soil food web.
Here we study soil fauna communities and their ecosystem functions in Douglas-fir - beech forests compared with beech and the native conifers silver fir and spruce, including different proportions of conifer-beech mixtures, across three regions in Germany: northern Bavaria, southern, and northern Lower Saxony.
First we investigated macrofauna communities and trophic niches based on stable isotope analysis of 13C and 15N of predators, primary and secondary decomposers in Lower Saxony with two contrasting site conditions of nutrient-rich and nutrient-poor soils (Chapter 2). When comparing Douglas-fir forests with spruce and beech as well as the conifer - beech mixtures, Douglas-fir had a negative impact on soil macrofauna in nutrient-poor sites. However, Douglas-fir - beech mixtures mitigated the effects on density and diversity as well as the detrital shift, which was noticeable in the conifer forests compared to pure beech. The community composition of macrofauna in the conifer mixtures was in between the respective monocultures. In general, trophic groups differed more between regions than forest types, while trophic positions remained robust between regions. Forest type in loamy sites generally had little influence on soil macrofauna. Nevertheless, primary decomposers benefited from Douglas-fir and especially Douglas-fir - mixtures compared to spruce forests, profiting from lower acidification and higher quality of Douglas-fir leaf litter.
To further investigate the influence of conifer-beech proportions, we studied the soil mesofauna (Oribatida, Collembola and Mesostigmata) in increasing proportions of Douglas-fir in beech dominated forests compared with silver fir - beech forests in the Spessart region of northern Bavaria (Chapter 3). Here, Oribatida communities shifted towards increasing proportions of families associated as primary decomposers in forests with higher Douglas-fir proportions, confirming a positive effect of Douglas-fir on primary decomposers for the soil fauna (Chapter 2 & 3). The
community composition of Oribatida responded sensitively to the differences in forest types for Douglas-fir beech, silver fir - beech and pure beech stands on species and family - level, while the community composition of Collembola and Mesostigmata varied little. With only Collembola diversity increasing in Douglas-fir - beech forests compared to silver fir –beech mixtures, there were few differences in density and diversity between the forest types. Therefore the soil mesofauna further highlights the general robustness of the soil food web between different forest types and with increasing conifer proportions.
Finally we looked into the energy fluxes of the soil food web of decomposers and predators comprising macrofauna and mesofauna with increasing proportions of conifers in beech stands, including Douglas-fir and silver fir (Chapter 4). Mesofauna dominated in biomass and energy fluxes, highlighting their importance towards ecosystem functions in acidic forest types as occurring in the Spessart. Energy fluxes of decomposers profited from moderate Douglas-fir proportions compared to silver - fir, while energy fluxes of predators increased in higher conifer proportions, with responses being conifer tree species specific. The diversity of macrofauna was driven by soil pH, with total energy fluxes positively driven by mesofauna diversity. Group specific energy fluxes increased with greater diversity of decomposers and predators, indicating a positive biodiversity ecosystem functioning relationship and the importance of maintaining a diverse soil food web.
To conclude, the results of this thesis show the robustness of soil animal communities and food webs under the introduction of Douglas-fir into European temperate forest systems. However, soil conditions should be considered when introducing Douglas-fir, but the soil food web and the ecosystem functions are resilient to changes in forest types, especially by mixing Douglas-fir with beech. Douglas-fir - beech mixtures can positively influence primary decomposers and decomposition of soil fauna communities compared to native conifer tree species, especially in forests with lower Douglas-fir proportions (Chapter 5). Therefore, Douglas-fir can be considered an alternative to the climate vulnerable Norway spruce, regarding the influence on soil fauna. In general, by conserving biodiversity and promoting soil fauna communities favored by soil conditions, planting Douglas-fir in temperate beech forests can maintain key ecosystem functions and can contribute to the silviculture of the future. Further research needs to focus on further environmental conditions and basal resource distribution in Douglas-fir forests, to understand changes in abiotic and biotic factors such as resource shifts under Douglas-fir.2026-02-0