Identifizierung und funktionelle Charakterisierung der SUMOylierung des Tight Junction-Proteins Zona occludens-2 (ZO-2)

Die Aufrechterhaltung und Integrität von Tight Junctions (TJs) ist von zentraler Bedeutung für die Zellpolarität in epithelialen und endothelialen Geweben. Als TJ-assoziiertes Adapter-Protein verbindet Zona occludens-2 (ZO-2) die integralen Tight Junction-Proteine mit dem Aktin-Cytoskelett und trägt dort zur Ausbildung des cytoplasmatischen Plaques bei. ZO-2 ist ein dual-lokalisiertes Nacos (nuclear and adhesion complexes)-Protein, das in konfluenten Zellen an der Plasmamembran und in dünn ausgesäten Zellen vorrangig im Kern zu finden ist. Die intrazelluläre Lokalisation von ZO-2 wird durch mehrere konservierte Kernimport- und Kernexport-Signale sowie posttranslationale Modifikationen u.a. innerhalb dieser Motive reguliert. Im Rahmen dieser Dissertation wurden im humanen ZO-2 mehrere evolutionär hochkonservierte potentielle SUMOylierungsstellen identifiziert und die Interaktion von ZO-2 mit Komponenten der SUMOylierungsmaschinerie nachgewiesen. Die SUMOylierung von ZO-2 konnte in vivo und in vitro bestätigt werden. Mit Hilfe von Mutationsanalysen wurde Lysin 730 als mögliche SUMO-Akzeptorstelle identifiziert. Die Mutation dieses Lysins führt zu einer vorrangig nukleären Lokalisation, wohingegen ein SUMO-ZO-2-Fusionsprotein, das ein konstitutiv SUMOyliertes ZO-2-Protein nachahmt, vermehrt im Cytoplasma zu finden ist. ZO-2 wurde in früheren Studien als Repressor des Wnt/β-Catenin-Signalwegs beschrieben. Das SUMO-ZO-2-Fusionsprotein besitzt keine transkriptionsreprimierende Aktivität, während die SUMOylierungs-defiziente ZO-2-K730R-Mutante die β-Catenin/TCF-4-vermittelte Transkriptionsaktivität sogar etwas stärker reprimiert. In diesem Zusammenhang wurde eine Interaktion von ZO-2 mit β-Catenin und GSK3β nachgewiesen. Aufgrund dieser Beobachtungen kann festgestellt werden, dass die intrazelluläre Lokalisation und die Signalfunktion von ZO-2 durch posttranslationale SUMOylierung reguliert wird

The Angular Momentum Problem in Cosmological Simulations of Disk Galaxy Formation

We conduct a systematic study of the angular momentum problem in numerical simulations of disk galaxy formation. We investigate the role of numerical resolution using a semi-cosmological setup which combines an efficient use of the number of particles in an isolated halo while preserving the hierarchical build-up of the disk through the merging of clumps. We perform the same simulation varying the resolution over 4 orders of magnitude. Independent on the level of resolution, the loss of angular momentum stays the same and can be tied to dynamical friction during the build-up phase. This is confirmed in a cosmological simulation. We also perform simulations including star formation and star formation and supernova feedback. While the former has no influence on the angular momentum problem, the latter reduces the loss to a level potentially in agreement with observations. This is achieved through a suppression of early star formation and therefore the formation of a large, slowly rotating bulge. We conclude that feedback is a critical component to achieve realistic disk galaxies in cosmological simulations. Numerical resolution is important, but by itself not capable of solving the angular momentum problem.Comment: 13 pages, 13 figures, submitted to MNRA

Impact of Partial Volume Correction on [18F]GE-180 PET Quantification in Subcortical Brain Regions of Patients with Corticobasal Syndrome.

Corticobasal syndrome (CBS) is a rare neurodegenerative condition characterized by four-repeat tau aggregation in the cortical and subcortical brain regions and accompanied by severe atrophy. The aim of this study was to evaluate partial volume effect correction (PVEC) in patients with CBS compared to a control cohort imaged with the 18-kDa translocator protein (TSPO) positron emission tomography (PET) tracer [18F]GE-180. Eighteen patients with CBS and 12 age- and sex-matched healthy controls underwent [18F]GE-180 PET. The cortical and subcortical regions were delineated by deep nuclei parcellation (DNP) of a 3D-T1 MRI. Region-specific subcortical volumes and standardized uptake values and ratios (SUV and SUVr) were extracted before and after region-based voxel-wise PVEC. Regional volumes were compared between patients with CBS and controls. The % group differences and effect sizes (CBS vs. controls) of uncorrected and PVE-corrected SUVr data were compared. Single-region positivity in patients with CBS was assessed by a >2 SD threshold vs. controls and compared between uncorrected and PVE-corrected data. Smaller regional volumes were detected in patients with CBS compared to controls in the right ventral striatum (p = 0.041), the left putamen (p = 0.005), the right putamen (p = 0.038) and the left pallidum (p = 0.015). After applying PVEC, the % group differences were distinctly higher, but the effect sizes of TSPO uptake were only slightly stronger due to the higher variance after PVEC. The single-region positivity of TSPO PET increased in patients with CBS after PVEC (100 vs. 83 regions). PVEC in the cortical and subcortical regions is valuable for TSPO imaging of patients with CBS, leading to the improved detection of elevated [18F]GE-180 uptake, although the effect sizes in the comparison against the controls did not improve strongly

Translocator protein (18kDA) (TSPO) marks mesenchymal glioblastoma cell populations characterized by elevated numbers of tumor-associated macrophages

TSPO is a promising novel tracer target for positron-emission tomography (PET) imaging of brain tumors. However, due to the heterogeneity of cell populations that contribute to the TSPO-PET signal, imaging interpretation may be challenging. We therefore evaluated TSPO enrichment/expression in connection with its underlying histopathological and molecular features in gliomas. We analyzed TSPO expression and its regulatory mechanisms in large in silico datasets and by performing direct bisulfite sequencing of the TSPO promotor. In glioblastoma tissue samples of our TSPO-PET imaging study cohort, we dissected the association of TSPO tracer enrichment and protein labeling with the expression of cell lineage markers by immunohistochemistry and fluorescence multiplex stains. Furthermore, we identified relevant TSPO-associated signaling pathways by RNA sequencing.We found that TSPO expression is associated with prognostically unfavorable glioma phenotypes and that TSPO promotor hypermethylation is linked to IDH mutation. Careful histological analysis revealed that TSPO immunohistochemistry correlates with the TSPO-PET signal and that TSPO is expressed by diverse cell populations. While tumor core areas are the major contributor to the overall TSPO signal, TSPO signals in the tumor rim are mainly driven by CD68-positive microglia/macrophages. Molecularly, high TSPO expression marks prognostically unfavorable glioblastoma cell subpopulations characterized by an enrichment of mesenchymal gene sets and higher amounts of tumor-associated macrophages.In conclusion, our study improves the understanding of TSPO as an imaging marker in gliomas by unveiling IDH-dependent differences in TSPO expression/regulation, regional heterogeneity of the TSPO PET signal and functional implications of TSPO in terms of tumor immune cell interactions