Mechanisms regulating desmoglein clustering and hyper-adhesion of desmosomes

Desmosomes provide strong intercellular adhesion and therefore are essential in tissues exposed to constant mechanical stress, e.g. the epidermis or the heart. The molecular composition of desmosomes consists of several protein families, including intercellular adhesion molecules and plaque proteins. Desmosomal cadherins represent the adhesion molecules and are a group of proteins comprising four desmogleins (Dsg1-4) and three desmocollins (Dsc1-3) isoforms. These are transmembrane proteins which interact in a Ca2+-dependent homophilic or heterophilic manner, thereby mediating strong intercellular adhesion with their neighboring cell counterparts. Within the desmosome, plaque proteins serve as linkers of desmosomal cadherins to the intermediate filament cytoskeleton. The plaque proteins consist of armadillo family proteins, plakoglobin (PG) and plakophilins (Pkps), and desmoplakin (DP), a protein of the plakin family. Pkps act as scaffolds in order to mediate adhesion and signaling and further regulate desmosomal turnover and assembly. They exist in three different isoforms (Pkp1-3) which show a tissue-specific expression. Further, Pkps show a wide ranging role in the cell-biological context and their physiological significance is evident in diseases, e.g. mutations in Pkp1 lead to ectodermal dysplasia skin fragility syndrome. However, their role in regulating Dsg binding properties as well as Dsg clustering is not yet elucidated. Murine keratinocytes lacking either Pkp1 or Pkp3 were compared to wild type (wt) cells to characterize the role of Pkp1 and Pkp3 in the regulation of Dsg1- and Dsg3-binding properties and contribution to Dsg3 clustering. Dsg1 and Dsg3 are of particular interest since they are the main targets of autoantibodies in the autoimmune skin blistering disease pemphigus vulgaris (PV). In the first part of the thesis, the roles of Pkp1 and Pkp3 in desmoglein clustering and adhesion were investigated. Characterization of Pkp1- or 3-deficient cell lines revealed reduced cell cohesion and especially in Pkp1-deficient cells loss of intercellular adhesion is strong. Force mapping measurements, performed with atomic force microscopy (AFM), revealed reduced binding frequency of Dsg1 and Dsg3 at the cell borders and displayed low membrane availability of Dsg1 and Dsg3 in Pkp1- or Pkp3-deficient cell lines. This indicates that both proteins are important for proper membrane availability of desmogleins. The single molecule-binding properties of the remaining membrane-bound desmogleins showed Pkp-dependent changes. However, because their numbers were low, presumably these altered binding properties of the remaining cadherins have no strong effect on overall adhesion. Extracellular crosslinking revealed that Pkp1 but not Pkp3 is required for Dsg3 clustering. Overexpression of Dsg3 rescued the reduced number of Dsg3 clusters in Pkp3- but not in Pkp1-deficient cells, as shown by AFM and stimulated emission depletion (STED) microscopy experiments. This demonstrates that Pkp1 and Pkp3 regulate the membrane availability of Dsgs. Furthermore, these data demonstrate that Pkp1 but not Pkp3 is required for the clustering of Dsg3. This novel function of Pkp1 appears to be essential for proper intercellular adhesion. In the second part of the thesis, regulation of desmosomal hyper-adhesion was studied. An interesting property of desmosomes is their ability to occur in two adhesive states, a weaker and a stronger one. Desmosomes in the stronger adhesive state become independent from extracellular Ca2+ and are called hyper-adhesive. However, the roles of Pkps and Dsgs in desmosomal hyper-adhesion is not fully elucidated yet. To address this unsolved issue, the aforementioned cell culture model, a murine keratinocyte Dsg3 knockout cell line and an ex-vivo skin model were used. Murine keratinocytes acquire the hyper-adhesive state 72 hours after exposure to high Ca2+, whereas Pkp- and Dsg3-deficient cell lines fail to reach this state during this time line. AFM force mappings revealed that the hyper-adhesive state correlates with increased Dsg3 single molecule binding strength and prolonged interaction lifetime. Both parameters were unchanged in the Pkp-deficient cells. In parallel, during differentiation, i.e. during the acquisition of hyper-adhesion, the Dsg3 clusters become Ca2+-independent. In contrast, deficiency of Pkp1 prevented Ca2+-independency of Dsg3 clusters whereas lack of Pkp3 led to a reduced amount of Ca2+-independent Dsg3 clusters. Interestingly, Dsg1 clusters remained the same during the investigated time period. In accordance, Dsg1 single molecule binding properties were not altered. This shows that acquisition of hyper-adhesion may not be a state acquired by the entire desmosome but rather reveals an isoform-dependent regulation by Pkps. Ca2+ chelation of ex-vivo human skin samples showed further isoform-specific differences in membrane localization between desmosomal cadherins. Immunostaining for Dsg3 at the cell membrane appeared to be more resistant to Ca2+ chelation compared to Dsg1 staining. This demonstrates that desmosomal cadherins have different roles in the acquisition of hyper-adhesion and that Pkp1-mediated clustering of desmogleins is required for the acquisition of the hyper-adhesive state of desmosomes. In summary, this project revealed that Pkp1 and Pkp3 are important for membrane availability of desmogleins. However, for Pkp3 this phenotype is differentiation dependent and decreases over time. Pkp1, in contrast to Pkp3, plays a crucial role in the clustering of desmogleins. This Pkp1-mediated clustering contributes to acquisition of hyper-adhesion and correlates with altered single molecule binding properties such as binding strength and interaction-lifetime of Dsg3 when becoming hyper-adhesive. Thus, development of the hyper-adhesive state is paralleled by alterations of binding properties of specific Dsg isoforms rather than by the entire desmosome. In a side-project, keratin-dependent regulation of desmoglein-binding was characterized. Keratin filament detachment from the desmosomal plaque occurs as one of the pathomechanism of PV. We found that p38 mitogen-activated protein kinases (p38MAPK) signaling is regulated by keratins. Moreover, this signaling is essential for the regulation of cell adhesion and involves both keratin-dependent and independent mechanisms. Using fluorescence recovery after photobleaching (FRAP) experiments it was observed that desmosomal cadherin uncoupling from the cytoskeleton resulted in higher Dsg3 mobility in the plasma membrane. The studies presented demonstrate that Pkp1-mediated regulation of desmosomal clustering is important for desmosomal adhesion and contributes to hyper-adhesion of desmosomes

Page switching in mixed-device web surveys: prevalence and data quality

As a self-administered survey mode, web surveys allow respondents to temporarily leave the survey page and switch to another web page in a different browser tab or to another window/app. This form of sequential multitasking has the potential to disrupt the response process and reduce data quality if respondents become distracted (Krosnick, 1991; Sendelbah et al., 2016). Browser data indicating respondents leaving the survey page allow non-reactive measurement of their multitasking. We investigated the prevalence of page switching, number of switching events and time spent absent per event with respect to respondents’ characteristics and devices used. Furthermore, we analysed the association with data quality (item missing, differentiation in grid questions and number of characters to open-ended questions). The results indicate that the prevalence of page switching is relatively low and the durations of page switching events are rather short. Also, respondents using a PC/tablet are more likely to leave the survey page than those using a smartphone. As to data quality, we did not find any correlation between page switching and the quality of the answers. Thus, this study provides no evidence that multitasking poses a threat to data quality. The findings are discussed with respect to the delimitations of multitasking using browser paradata

Active Learning for SAT Solver Benchmarking

Benchmarking is a crucial phase when developing algorithms. This also applies to solvers for the SAT (propositional satisfiability) problem. Benchmark selection is about choosing representative problem instances that reliably discriminate solvers based on their runtime. In this paper, we present a dynamic benchmark selection approach based on active learning. Our approach predicts the rank of a new solver among its competitors with minimum runtime and maximum rank prediction accuracy. We evaluated this approach on the Anniversary Track dataset from the 2022 SAT Competition. Our selection approach can predict the rank of a new solver after about 10 % of the time it would take to run the solver on all instances of this dataset, with a prediction accuracy of about 92 %. We also discuss the importance of instance families in the selection process. Overall, our tool provides a reliable way for solver engineers to determine a new solver’s performance efficiently

Autorschaft und Materialität der Literatur zwischen 1765 und 1815

Autorschaft erfordert mehr als das Schreiben eines Textes. Um 1800 wird Autorschaft als bürgerliche Subjektform durch das gedruckte Buch konstituiert. Deshalb untersucht die vorliegende Studie die Praktiken des Publizierens hinsichtlich ihrer materialen Bedingtheit. Der Bogen reicht von handgeschriebenen Büchern über gedruckte Artefakte bis zur Makulatur. Die Materialität von Literatur wird dabei in ihren ästhetischen, merkantilen, poetologischen, rechtlichen sowie wissensgeschichtlichen Dimensionen betrachtet. Anhand von Jean Paul, aber auch am Beispiel von Georg Christoph Lichtenberg und Friedrich Nicolai werden Publizierende als „Büchermacher“ in den Blick genommen. Dabei liegt ein besonderes Augenmerk auf dem Publizieren als einem sozialen Phänomen des ausgehenden 18. Jahrhundert

Оценка динамики состояния растений-биоиндикаторов атмосферного загрязнения на основе данных дистанционного зондирования Земли

Рассмотрены вопросы использования данных дистанционного зондирования Земли из космоса для оценки воздействия химического загрязнения атмосферы на состояние сосны сибирской кедровой. Исследования проведены в зоне влияния факелов для сжигания попутного газа на территории Приобского нефтяного месторождения. Оценка состояния растений-биоиндикаторов производилась с использованием вегетационных индексов NDVI и ARVI, рассчитанных по космическим снимкам со спутника Landsat. Для обработки и анализа данных использован инструментарий, предоставляемый программным пакетом ERDAS Imagine и геоинформационной системой ArcGIS. Показано, что негативные изменения в состоянии растений-биоиндикаторов, нарастающие по мере приближения к факелам и выявленные методами биоиндикации, обнаруживаются на космических снимках среднего пространственного разрешения (30 м)

Heat transfer challenge and design evaluation for a multi-stage temperature swing adsorption (TSA) process

Functionalized solid amine-based temperature swing adsorption (TSA) processes have recently been proposed as a potential way to reduce the energy-penalty of post-combustion carbon capture processes (1). If TSA is to be carried out at large scale and with high energy-efficiency, continuously operated counter-current contactors are required for thermodynamic reasons. This could, generally, be achieved by using moving bed contactors. However, the heat exchange requirement of TSA is significant and heat transfer is poor in fixed and moving beds. Therefore, multi-stage fluidized bed contactors with counter-current flow of solids and gas phase and immersed heat exchange surfaces may solve the heat transfer challenge while maintaining the thermodynamic process requirements. Experiments have shown that adsorption and desorption kinetics of suitable functionalized amine sorbents are fast and equilibrium loadings are practically reached in the stages (1). Thus, heat exchange is the dominant limiting factor for a practical stage design in multi-stage fluidized bed TSA. The present work rationally develops design requirements for TSA stages based on the necessary heat exchange rates. The considered particles are Geldart Type B (diameter 200-300 µm, particle density 1000-1500 kg/m3). Scalability of the design proposal is considered and vertically orientated heat exchanger tubes are compared to horizontal tube bundles. The net movement and mixing of particles within the bubbling bed stage must be maintained in spite of the emulsified heat exchangers (possible dead zones in the area of the tube bundles). It is shown that the pressure drop of multi-stage fluidized bed TSA units for flue gas CO2 capture is practically determined by the heat exchange requirement and not by the space-time of the solids for the adsorption. Future work will employ a bubbling fluidized bed heat exchange testing device for optimization of the heat exchanger geometry with respect to heat transfer rates and particle residence time distribution in the stage. Heat exchange measurement devices have been presented recently in literature for horizontal tube bundles and Geldart Type A particles (2), but the importance of the heat exchanger issue in continuous fluidized bed TSA requires the detailed investigation for the Geldart B range, potentially considering the macroscopic particle movement relative to the heat exchangers within each individual TSA stage. Please click Additional Files below to see the full abstract