Domänenanalyse des Cyclase-assoziierten Proteins (CAP) und Charakterisierung von Filactin, einem neuartigen actinähnlichen Protein aus Dictyostelium discoideum

The cyclase-associated protein (CAP) from Dictyostelium discoideum was studied in detail regarding its structure and function relationships. The second part of the thesis describes the characterization of the novel actin-related protein filactin from D. discoideum. CAP homologs are multifunctional proteins: they are involved in signal dependent changes in the actin cytoskeleton, in vesicle transport and cell development. The binding of monomeric actin through the C-terminal domain represents a common feature of all CAPs. From four highly conserved regions in this domain the verprolin homology region of the D. discoideum CAP was analyzed in this work. Loss of this region led to a clear decrease, but not suppression of the actin-sequestering activity. In agreement to these data, stable complexes of the modified CAP-C with G-actin could be identified in chemical crosslinking experiments, and it could be shown that CAP-C is able to dimerize. Considering the high conservation of cyclase-associated proteins and their importance for cell biological processes it is remarkable that no structure of this protein has been elucidated so far. Therefore, in the frame of this thesis the structure of the membrane-associated N-terminal domain of D. discoideum CAP was to be determined in cooperation with the group of T. Holak at the MPI f. Biochemistry (Martinsried). For this purpose numerous constructs from the N-terminal domain had to be cloned and expressed, to be purified and examined for their stability and threedimensional folding. It turned out that the stable core of the aminoterminal domain covers the amino acids 51-226. The structure was determined by nuclear magnetic resonance spectroscopy and X-ray crystallography. Six antiparallel a-helices are connected by loop elements and form a cylindrical core domain which can form a dimer in the crystal, dimerization occurs through Mg2+ ions. The clarification of the N-terminal structure of D. discoideum CAP will now simplify the research on its interaction with phospholipids and CAP binding proteins The second part of this work dealt with the characterization of filactin. The unique 105 kDa protein contains two filamin homologous regions in its N-terminal, and a clearly actin-related domain within the C-terminal part. While in resting cells the endogenous filactin shows cytoplasmic distribution and interaction with protein aggregates as well, the GFP construct of the actin-related domain displays an actin-like behavior during cell movement or phagocytosis. A stimulus-induced colocalization of actin and filactin was observed in experiments with chemotactically stimulated cells. The alignment of its C-terminal amino acid sequence with the structure of muscle actin predicts a globular, actin-related structure containing all residues that are important for ATP/ADP binding.In dieser Arbeit standen zwei Aufgaben im Mittelpunkt: Zum einen wurde das Cyclase-assoziierte Protein (CAP) aus Dictyostelium discoideum hinsichtlich seiner Struktur-Funktionsbeziehungen eingehend untersucht, zum zweiten erfolgte eine erste Charakterisierung des neuartigen actinähnlichen Proteins Filactin aus D. discoideum. CAP-homologe Proteine sind multifunktionell und entscheidend an signalabhängigen Veränderungen des Actin-Cytoskeletts, am Vesikeltransport und an der Zellentwicklung beteiligt. Die Bindung von monomerem Actin über die carboxyterminale Domäne stellt wahrscheinlich eine gemeinsame Eigenschaft aller Homologen dar. Von vier hochkonservierten Regionen im C-terminalen Abschnitt, die für eine spezifische Bindung in Frage kommen, wurde in dieser Arbeit der Verprolin-homologe Bereich des D. discoideum CAPs untersucht. Der Verlust dieser Domäne führte zwar zu einer deutlichen Senkung, nicht aber Unterdrückung der Actinbindung durch den carboxyterminalen Proteinbereich. Dementsprechend wurden in chemischen Quervernetzungsexperimenten stabile Komplexe des verkürzten CAP-C mit G-Actin erhalten. Durch das Auftreten von CAP-C-Dimeren wurde hier außerdem eine weitere Funktion des Proteins nachgewiesen. Angesichts der weiten Verbreitung der Cyclase-assoziierten Proteine und ihrer Bedeutung für zellbiologische Prozesse ist es bemerkenswert, daß bisher noch keine Struktur dieses Proteintyps veröffentlicht ist. Es sollte daher in Kooperation mit der Gruppe von T. Holak am MPI f. Biochemie (Martinsried) die Struktur der membranassoziierten N-terminalen Domäne von D. discoideum CAP erarbeitet werden. Zu diesem Zweck mußten zunächst zahlreiche Proteinabschnitte aus dem N-terminalen Bereich kloniert und exprimiert, gereinigt und auf ihre Stabilität und dreidimensionale Faltung hin untersucht werden. Es stellte sich heraus, daß der stabile Bereich der aminoterminalen Domäne die Aminosäuren 51-226 umfasst. Nach NMR-spektroskopischen und röntgenkristallographischen Analysen liegt nun die Struktur vor. Danach bilden sechs antiparallel angeordnete und über Loop-Elemente miteinander verbundene a-Helices eine zylindrische Domäne aus, welche im Kristall dimerisiert vorliegt. Pro Dimer wird dabei ein Mg2+ komplexiert. Die Aufklärung der N-terminalen Struktur von D. discoideum CAP erlaubt nun gezielte Untersuchungen hinsichtlich der Interaktion mit Phospholipiden und CAP-Bindeproteinen. Im zweiten Teil der Arbeit stand die Charakterisierung von Filactin im Mittelpunkt. Das bisher einzigartige und etwa 105 kDa große Protein verfügt über zwei Filamin-homologe Bereiche in der N-terminalen Region und eine stark ausgeprägte actinähnliche Domäne im C-terminalen Bereich. Während das endogene Filactin in der ruhenden Zelle sowohl eine cytoplasmatische Verteilung als auch eine Anbindung an bisher unbestimmte Proteinaggregate zeigt, verhält sich das GFP-Konstrukt der actinähnlichen Domäne im Zuge der Zellbewegung oder Phagocytose übereinstimmend mit Actin. Experimente mit chemotaktisch stimulierten Zellen zeigen darüber hinaus auch eine Stimulus-induzierte Kolokalisation von Actin und Filactin. Die Anpassung seiner carboxyterminalen Aminosäurensequenz an die Struktur von Muskelactin sagt für das Protein eine globuläre, actinähnliche Struktur voraus, in der alle für eine ATP/ADP-Bindung erforderlichen Aminosäurenreste vorhanden sind

Cbp3–Cbp6 interacts with the yeast mitochondrial ribosomal tunnel exit and promotes cytochrome b synthesis and assembly

A complex specifically required for the biogenesis of the respiratory chain component cytochrome b binds to the tunnel exit of yeast mitochondrial ribosomes to coordinate protein synthesis and assembly

Orbital Evolution of Compact White Dwarf Binaries

The new-found prevalence of extremely low mass (ELM, Mhe<0.2 Msun) helium white dwarfs (WDs) in tight binaries with more massive WDs has raised our interest in understanding the nature of their mass transfer. Possessing small (Menv~1e-3 Msun) but thick hydrogen envelopes, these objects have larger radii than cold WDs and so initiate mass transfer of H-rich material at orbital periods of 6-10 minutes. Building on the original work of D'Antona et al., we confirm the 1e6 yr period of continued inspiral with mass transfer of H-rich matter and highlight that the inspiraling direct-impact double WD binary HM Cancri likely has an ELM WD donor. The ELM WDs have less of a radius expansion under mass loss, thus enabling a larger range of donor masses that can stably transfer matter and become a He mass transferring AM CVn binary. Even once in the long-lived AM CVn mass transferring stage, these He WDs have larger radii due to their higher entropy from the prolonged H burning stage.Comment: 9 pages, 8 figures. Accepted for publication in the Astrophysical Journa

Identification and characterization of two novel primate-specific histone H3 variants, H3.X and H3.Y

The expression of a new histone variant H3.Y increases during cellular stress to regulate cell cycle progression and gene expression

Long-COVID Symptoms in Individuals Infected with Different SARS-CoV-2 Variants of Concern:A Systematic Review of the Literature

The association of SARS-CoV-2 variants with long-COVID symptoms is still scarce, but new data are appearing at a fast pace. This systematic review compares the prevalence of long-COVID symptoms according to relevant SARS-CoV-2 variants in COVID-19 survivors. The MEDLINE, CINAHL, PubMed, EMBASE and Web of Science databases, as well as the medRxiv and bioRxiv preprint servers, were searched up to 25 October 2022. Case-control and cohort studies analyzing the presence of post-COVID symptoms appearing after an acute SARS-CoV-2 infection by the Alpha (B.1.1.7), Delta (B.1.617.2) or Omicron (B.1.1.529/BA.1) variants were included. Methodological quality was assessed using the Newcastle–Ottawa Scale. From 430 studies identified, 5 peer-reviewed studies and 1 preprint met the inclusion criteria. The sample included 355 patients infected with the historical variant, 512 infected with the Alpha variant, 41,563 infected with the Delta variant, and 57,616 infected with the Omicron variant. The methodological quality of all studies was high. The prevalence of long-COVID was higher in individuals infected with the historical variant (50%) compared to those infected with the Alpha, Delta or Omicron variants. It seems that the prevalence of long-COVID in individuals infected with the Omicron variant is the smallest, but current data are heterogeneous, and long-term data have, at this stage, an obviously shorter follow-up compared with the earlier variants. Fatigue is the most prevalent long-COVID symptom in all SARS-CoV-2 variants, but pain is likewise prevalent. The available data suggest that the infection with the Omicron variant results in fewer long-COVID symptoms compared to previous variants; however, the small number of studies and the lack of the control of cofounders, e.g., reinfections or vaccine status, in some studies limit the generality of the results. It appears that individuals infected with the historical variant are more likely to develop long-COVID symptomatology

The Central X-Ray Point Source in Cassiopeia A

The spectacular first light observation by the Chandra X-Ray Observatory revealed an X-ray point source near the center of the 300 yr old Cas A supernova remnant. We present an analysis of the public X-ray spectral and timing data. No coherent pulsations were detected in the Chandra/HRC data. The 3-sigma upper limit on the pulsed fraction is 20 ms. The Chandra/ACIS spectrum of the point source may be fit with an ideal blackbody (kT=0.5 keV), or with BB models modified by the presence of a NS atmosphere (kT=0.25-0.35 keV), but the temperature is higher and the inferred emitting area lower than expected for a 300 yr old NS according to standard cooling models. The spectrum may also be fit with a power law model (photon index 2.8-3.6). Both the spectral properties and the timing limits of the point source are inconsistent with a young Crab-like pulsar, but are quite similar to the properties of the anomalous X-ray pulsars. The spectral parameters are also very similar to those of the other radio-quiet X-ray point sources in the supernova remnants Pup A, RCW 103, and PKS 1209-52. Current limits on an optical counterpart for the Cas A point source rule out models that invoke fallback accretion onto a compact object if fallback disk properties are similar to those in quiescent low-mass X-ray binaries. However, the optical limits are marginally consistent with plausible alternative assumptions for a fallback disk. In this case, accreting NS models can explain the X-ray data, but an accreting BH model is not promising.Comment: 17 pages including 2 figs. To appear in ApJ, Vol. 546 (Jan 10, 2001). Minor revisions per referee. Pulsation limits revised in light of HRC wiring problem. Typos correcte

The Miocene mammal record of the Vallès-Penedès Basin (Catalonia)

The land mammal record of the Vallès-Penedès Basin (Catalonia, NE Spain) ranges from the early Miocene (Ramblian) to the late Miocene (Turolian), that is from about 20 to 7 Ma. Here we present an updated review of the mammal succession focusing on biochronology as well as on environmental and faunal changes. Based on faunal similarities with central Europe, we interpret this basin as a transitional zone between the forested environments of northern regions and the more arid landscapes of the inner Iberian Peninsula. The quality of the Vallès-Penedès record and its chronostratigraphic control is clearly better for the late Aragonian and the Vallesian (between 12.6-9.0 Ma), especially for small mammals. Therefore, we analyze small mammal diversity dynamics during this interval. Contrary to previous analyses, which found an abrupt extinction event coinciding with the early/late Vallesian boundary (the Vallesian Crisis), our results show that this pattern is due to uneven sampling. Instead, taxonomic richness slowly decreased since the late Vallesian as a result of a series of extinctions that mostly affected forest-dwelling taxa

Synergy between CD26/DPP-IV Inhibition and G-CSF Improves Cardiac Function after Acute Myocardial Infarction

SummaryIschemic cardiomyopathy is one of the main causes of death, which may be prevented by stem cell-based therapies. SDF-1α is the major chemokine attracting stem cells to the heart. Since SDF-1α is cleaved and inactivated by CD26/dipeptidylpeptidase IV (DPP-IV), we established a therapeutic concept—applicable to ischemic disorders in general—by combining genetic and pharmacologic inhibition of DPP-IV with G-CSF-mediated stem cell mobilization after myocardial infarction in mice. This approach leads to (1) decreased myocardial DPP-IV activity, (2) increased myocardial homing of circulating CXCR-4+ stem cells, (3) reduced cardiac remodeling, and (4) improved heart function and survival. Indeed, CD26 depletion promoted posttranslational stabilization of active SDF-1α in heart lysates and preserved the cardiac SDF-1-CXCR4 homing axis. Therefore, we propose pharmacological DPP-IV inhibition and G-CSF-based stem cell mobilization as a therapeutic concept for future stem cell trials after myocardial infarction

STRATIGRAPHIC FRAMEWORK FOR ZECHSTEIN CARBONATES ON THE UTSIRA HIGH, NORWEGIAN NORTH SEA

The preserved Zechstein succession on the Utsira High in the NE part of the Norwegian North Sea is 25-100 m thick and is dominated by shelf carbonates. Internal subdivision of the succession is based on the recognition of key surfaces in petrophysical logs and cores, and suggests that the carbonates mainly consist of ZS2 and ZS3 deposits and that younger ZS4 and ZS5 deposits are only locally preserved. The carbonates have undergone early, syn-depositional dolomitization followed by later dolomite recrystallization and calcitization. Calcitization, interpreted as dedolomitization, is restricted to the upper part of the ZS3 carbonate unit and based on U/Pb dating took place during the Triassic, with a later phase of recrystallization linked to mid-Jurassic uplift. Both dedolomitization and dolomite recrystallization relate to fresh-water infiltration with the resetting of δO18 values prior to the Late Jurassic drowning of the Utsira High. The reservoir quality of the carbonates is directly linked to post-depositional meteoric diagenesis, and the best reservoir properties are recorded in intervals dominated by recrystallized dolomites in ZS2 and lower ZS3 carbonates. Dedolomitization significantly reduced porosity in the upper ZS3 carbonates

The Aragonian and Vallesian high-resolution micromammal succession from the Calatayud-Montalbán Basin (Aragón, Spain)

We present an updated taxonomy and faunal distribution of the micromammal fossil record from the Aragonian and lower Vallesian of the Calatayud-Montalbán Basin. The analysed record includes the orders Rodentia, Eulipotyphla, and Lagomorpha. The pattern of species turnover showsseven major faunal events,which are correlated to major climate changes based on marine stable oxygen and carbon isotope records. The episodesδ 18OMi-2 and Mi-3 are significantly correlated with major micromammal turnover at the boundaries between lower and middle Aragonian and middle and upper Aragonian, respectively. Our resultssupport the existence of a selective turnover during the Aragonian and lower Vallesian