Punktmutationsanalysen bei GLI3-assoziierten Krankheitsbildern: Greig Cephalopolysyndaktylie-Syndrom, Pallister-Hall-Syndrom und isolierte Polydaktylien

Für die Ausbildung der Extremitäten ist eine Vielzahl von Genen verantwortlich. Eine Schlüsselrolle in der Entstehung der anterior-posterioren Polarität der Extremitäten spielt die Sonic Hedgehog-Patched-GLI Signalkaskade (SHH-PTCH-GLI Signalkaskade). Kommt es durch Mutationen zur Fehlregulation eines der beteiligten Faktoren im SHH-Pathway, so resultieren in der Regel, aber nicht ausschließlich, Dysmorphogenesen der Gliedmaßen. Zu diesen zählen auch die in dieser Arbeit untersuchten GLI3- assoziierten Morphopathien. Verschiedene Punktmutationen konnten im humanen GLI3-Gen (7p13), das für einen Transkriptionsfaktor mit Zinkfingermotiven kodiert, detektiert werden. Bisher wurden vier autosomal-dominant vererbte Fehlbildungssyndrome GLI3-Mutationen zugeordnet: 1. Greig Cephalopolysyndaktylie-Syndrom (GCPS) 2. Pallister-Hall-Syndrom (PHS) 3. Postaxiale Polydaktylie Typ A (PAP-A) 4. Präaxiale Polydaktylie Typ IV (PPD-IV) Im Rahmen dieser Arbeit konnten die Fallzahlen der GLI3-assoziierten Morphopathien erhöht werden. Als molekulargenetische Methode kamen für die Untersuchung der Patienten-DNA die nicht-radioaktive Einzelstrang-Konformationsananlyse (SSCA) sowie die Sequenzierung zur Anwendung. Insgesamt konnten 12 Mutationen bei Patienten mit klinisch diagnostiziertem GCPS, 2 Mutationen bei Patienten mit klinisch gesichertem PHS, sowie 2 weitere Mutationen bei Patienten mit PPD-IV im GLI3-Gen identifiziert werden. Dabei betreffen 9 Mutationen den N-Terminus oder die zentralen Zinkfingermotive der DNA-bindenden Domäne des GLI3-Transkriptionsfaktors. Weitere 7 Mutationen betreffen den C-terminalen Bereich. Die Erhöhung der Fallzahlen bestätigt und ergänzt das bekannte Spektrum der GLI3-Mutationen. Mindestens 8 der 16 detektierten Mutationen führen, durch den Einbau eines vorzeitigen Stoppcodons, zu einem verkürzten Protein und somit vermutlich zum Verlust einiger oder aller Funktionen des GLI3-Transkriptionsfaktors. Der Mechanismus der Haploinsuffizienz scheint in diesen Fällen den Phänotyp zu bedingen. Dagegen kann für die 4 identifizierten Missense-Mutationen sowie für die 3 Spleißstellenmutationen keine eindeutige Aussage über die Auswirkung auf Protein-ebene und somit über den molekularen Mechanismus, der den Phänotyp bedingt, getroffen werden. Entgegen der bisher angenommen 100%igen Penetranz von GLI3-Mutationen konnte eine Mutationsübertragung über eine phänotypisch gesunde Probandin im Fall der Missense-Mutation R625W nachgewiesen werden. Vor dem Hintergrund einer nicht vollständigen Penetranz sollten in Zukunft auch seltene Sequenzvarianten auf Proteinebene hinsichtlich veränderter GLI3 Funktionen und möglicher Interaktion mit anderen Faktoren untersucht werden. Die Entstehung der unterschiedlichen Phänotypen durch Mutationen im GLI3-Gen bleibt weiterhin unklar, denn die erhobenen Daten von nunmehr annähernd 50 Mutationen lassen keine eindeutige Genotyp-Phänotyp Korrelation an Hand von Lage und Art der Mutationen zu. Zur Klärung könnten weiterführende Experimente auf funktioneller Ebene (mRNA Transkript- und Proteinebene) beitragen. Drei Punktmutationen konnten mehrfach bei unverwandten Familien identifiziert werden, jedoch ist die Häufigkeit wiederholt auftretender Mutationen zu gering, um von sogenannten Mutations-Hotspots ausgehen zu können. Daher muss auch in Zukunft die molekulare Aufklärung von Defekten das gesamte GLI3-Gen erfassen

Reactions of (-)-sparteine with alkali metal HMDS complexes : conventional meets the unconventional

Conventional (-)-sparteine adducts of lithium and sodium 1,1,1,3,3,3-hexamethyldisilazide (HMDS) were prepared and characterised, along with an unexpected and unconventional hydroxyl-incorporated sodium sodiate, [(-)-sparteine·Na(-HMDS)Na·(-)-sparteine]+[Na4(-HMDS)4(OH)]--the complex anion of which is the first inverse crown ether anion

Investigating the trimethylaluminium/water ALD process on mesoporous silica by in situ gravimetric monitoring

A low amount of AlOx was successfully deposited on an unordered, mesoporous SiO2 powder using 1–3 ALD (Atomic Layer Deposition) cycles of trimethylaluminium and water. The process was realized in a self-built ALD setup featuring a microbalanceand a fixed particle bed. The reactor temperature was varied between 75, 120, and 200 °C. The self-limiting nature of the deposition was verified by in situ gravimetric monitoring for all temperatures. The coated material was further analyzed by nitrogen sorption, inductively coupled plasma-optical emission spectroscopy, powder X-ray diffraction, high-resolution transmission electron microscopy, attenuated total reflection Fourier transformed infrared spectroscopy, and elemental analysis. The obtained mass gains correspond to average growth between 0.81–1.10 Å/cycle depending on substrate temperature. In addition, the different mass gains during the half-cycles in combination with the analyzed aluminum content after one, two, and three cycles indicate a change in the preferred surface reaction of the trimethylaluminium molecule from a predominately two-ligand exchange with hydroxyl groups to more single-ligand exchange with increasing cycle number. Nitrogen sorption isotherms demonstrate (1) homogeneously coated mesopores, (2) a decrease in surface area, and (3) a reduction of the pore size. The experiment is successfully repeated in a scale-up using a ten times higher substrate batch size.DFG, 325093850, Open Access Publizieren 2017 - 2018 / Technische Universität Berli

A structurally versatile nickel phosphite acting as a robust bifunctional electrocatalyst for overall water splitting

The design and development of economical and highly efficient electrocatalysts for the hydrogen evolution reaction (HER) and oxygen evolution reaction (OER) under alkaline conditions are vital in lowering the overall energy losses in alkaline water electrolysis. Here we present a nickel phosphite, Ni11(HPO3)8(OH)6, belonging to the unique class of phosphorus-based inorganic materials with striking structural features that have been explored for the first time in the reaction of electrocatalytic overall water splitting with a profound understanding of the system using in situ and ex situ techniques. When electrophoretically deposited, the nickel phosphite exhibited remarkable electrocatalytic activity, yielding considerably low overpotentials for both the OER and HER with extreme structural stability and enhanced durability in alkaline media. Apart from the attractive structural merits, the higher activity of nickel phosphite is mainly attributed to the formation of oxidized nickel species in the catalytic OER process, while subtle experimental evidence of the participation of phosphite anions for the acceleration of the HER with the support of Ni2+ cations as catalytically active sites is identified, which is highly compelling and has never been previously discovered. Finally, the bifunctionality of nickel phosphite was demonstrated by constructing an alkaline water electrolyzer with a low cell voltage and over 4 days of undiminishing stability. This work offers an appealing cost-effective system based on earth-abundant metals for water electrolysis and can be extended to other transition metal based homo- or hetero-bimetallic phosphites

Yttrium complexes of Arsine, Arsenide, and Arsinidene Ligands

Deprotonation of the yttrium–arsine complex [Cp′3Y{As(H)2Mes}] (1) (Cp′=η5-C5H4Me, Mes=mesityl) by nBuLi produces the μ-arsenide complex [{Cp′2Y[μ-As(H)Mes]}3] (2). Deprotonation of the As[BOND]H bonds in 2 by nBuLi produces [Li(thf)4]2[{Cp′2Y(μ3-AsMes)}3Li], [Li(thf)4]2[3], in which the dianion 3 contains the first example of an arsinidene ligand in rare-earth metal chemistry. The molecular structures of the arsine, arsenide, and arsinidene complexes are described, and the yttrium–arsenic bonding is analyzed by density functional theory

Theoretical studies of 31P NMR spectral properties of phosphanes and related compounds in solution

Selected theoretical methods, basis sets and solvation models have been tested in their ability to predict 31P NMR chemical shifts of large phosphorous-containing molecular systems in solution. The most efficient strategy was found to involve NMR shift calculations at the GIAO-MPW1K/6-311++G(2d,2p)//MPW1K/6-31G(d) level in combination with a dual solvation model including the explicit consideration of single solvent molecules and a continuum (PCM) solvation model. For larger systems it has also been established that reliable 31P shift predictions require Boltzmann averaging over all accessible conformations in solution

Daily Negative Work Events and Employees' Physiological and Psychological Reactions

Scholars have accumulated an abundant amount of knowledge on the association between work stressors and employees’ health and well-being. However, notions of the complex interplay of physiological and psychological components of stress reactions are still in their infancy. Building on the Allostastic Load (AL) model, the present study considers short-term within-person effects of negative work events (NWEs) on indicators of both physiological (i.e., salivary cortisol) and psychological distress responses (i.e., negative affect and emotional exhaustion). Multilevel findings from an experience sampling study with 83 healthcare professionals suggest that reported NWEs predict employees’ psychological but not endocrine stress responses. Results contribute to a more comprehensive understanding of employees’ daily response patterns to occupational stressors

Spectroscopic Characterization, Computational Investigation, and Comparisons of ECX

Three newly synthesized [Na+(221-Kryptofix)] salts containing AsCO–, PCO–, and PCS– anions were successfully electrosprayed into a vacuum, and these three ECX– anions were investigated by negative ion photoelectron spectroscopy (NIPES) along with high-resolution photoelectron imaging spectroscopy. For each ECX– anion, a well-resolved NIPE spectrum was obtained, in which every major peak is split into a doublet. The splittings are attributed to spin–orbit coupling (SOC) in the ECX• radicals. Vibrational progressions in the NIPE spectra of ECX– were assigned to the symmetric and the antisymmetric stretching modes in ECX• radicals. The electron affinities (EAs) and SO splittings of ECX• are determined from the NIPE spectra to be AsCO•: EA = 2.414 ± 0.002 eV, SO splitting = 988 cm–1; PCO•: EA = 2.670 ± 0.005 eV, SO splitting = 175 cm–1; PCS•: EA = 2.850 ± 0.005 eV, SO splitting = 300 cm–1. Calculations using the B3LYP, CASPT2, and CCSD(T) methods all predict linear geometries for both the anions and the neutral radicals. The calculated EAs and SO splittings for ECX• are in excellent agreement with the experimentally measured values. The simulated NIPE spectra, which are based on the calculated Franck–Condon factors, and the SO splittings nicely reproduce all of the observed spectral peaks, thus allowing unambiguous spectral assignments. The finding that PCS• has the greatest EA of the three triatomic molecules considered here is counterintuitive based upon simple electronegativity considerations, but this finding is understandable in terms of the movement of electron density from phosphorus in the HOMO of PCO– to sulfur in the HOMO of PCS–. Comparisons of the EAs of PCO• and PCS• with the previously measured EA values for NCO• and NCS• are made and discussed.National Science Foundation (U.S.) (Grant CHE-1362118